YBX1 protein degradation agent fisetin and application thereof in preparation of medicine for preventing and treating obesity or fatty liver

Abstract

The invention discloses a YBX1 protein degradation agent fisetin and application thereof in preparation of medicines for preventing and treating obesity or fatty liver, and belongs to the technical field of medicines. The embodiment of the invention proves that fisetin as a YBX1 protein degradation agent can relieve obesity induced by high-fat diet and reduce the accumulation of triglyceride in the liver of a mouse with high-fat diet. Meanwhile, the fisetin improves the glucose tolerance of the mouse induced by the high fat diet and improves the insulin resistance of the mouse induced by the high fat diet. In addition, fisetin can also reduce the liver weight and fat content of mice induced by high fat diet, and relieve liver injury and ferroptosis. Fisetin belongs to polyphenol flavonoids, exists in various fruits and vegetables, is low in price, wide in source, smaller in toxic and side effects, extremely easy to obtain and lower in cost, and has a wide application prospect in preparation of medicines for preventing and treating obesity or fatty liver.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, and in particular relates to a YBX1 protein degrader, Fisetin, and its application in the preparation of drugs for the prevention and treatment of obesity or fatty liver. Background Technology

[0002] In recent years, with the accelerated pace of life and the increasing prevalence of high-fat diets, the incidence of obesity and overweight has risen sharply. Obesity is a chronic metabolic disease caused by energy intake exceeding energy expenditure, leading to excessive fat accumulation that endangers health and can cause many related diseases. The prevalence of obesity has significantly increased the incidence of metabolic dysfunction-associated steatotic liver disease (MASLD). Studies have shown that the prevalence of MASLD is higher in overweight and obese individuals than in the general population. Furthermore, MASLD can also affect obesity through various mechanisms, including liver dysfunction, oxidative stress, glucotoxicity, and lipotoxicity.

[0003] Metabolic dysfunction-associated steatotic liver disease (MASLD) is a clinicopathological syndrome characterized primarily by hepatic steatosis, excluding alcohol and other clearly defined liver injury factors. MASLD is a spectrum of diseases, ranging from simple hepatic steatosis to metabolic-associated steatohepatitis (MASH), liver fibrosis, cirrhosis, and hepatocellular carcinoma (HCC), seriously impacting people's health. In recent years, the incidence of MASLD has been rapidly increasing globally, now surpassing viral hepatitis to become the leading cause of liver disease worldwide. Furthermore, increasing evidence suggests that MASLD is a complex disease affecting multiple systems, significantly increasing the risk of type 2 diabetes, cardiovascular disease, and chronic kidney disease. Due to the complexity of the disease, there are currently no widely used and effective treatments in clinical practice; lifestyle interventions remain the first-line treatment for MASLD, with weight loss interventions (diet and exercise) targeting obesity being a reasonable option for its management. However, MASLD patients often find it difficult to maintain improved lifestyles. Therefore, finding safe and effective drugs for the prevention and treatment of MASLD is of significant practical importance.

[0004] Like all chronic diseases, treating obesity requires a long-term, multimodal approach. Currently, modern medicine treats obesity in a relatively singular way, relying solely on lifestyle changes and medication, which suffers from slow efficacy, high relapse rates, and significant side effects. In recent years, increasing research has shown that natural small molecules have safe and effective alleviating effects on obesity and MASLD. Therefore, screening for drugs to prevent and treat obesity and MASLD from natural small molecules with virtually no toxic side effects holds great promise.

[0005] Y-box binding protein 1 (YBX1) is a member of the cold shock protein superfamily and is a multifunctional RNA-binding protein involved in various biological processes such as cell proliferation, differentiation, stress response, and malignant cell transformation. In tumors, YBX1 promotes malignant disease progression by stabilizing the expression of key oncogenes (such as MYC and BCL2 mRNA).

[0006] Therefore, screening for small molecules with low toxicity and side effects that can bind to the YBX1 protein and inhibiting hepatocyte ferroptosis by degrading the YBX1 protein is of significant practical importance for alleviating metabolic-related fatty liver disease. This strategy may not only provide new targets for the treatment of MASLD and MASH, but also offer new therapeutic ideas for other metabolic disorders.

[0007] Fisetin, found in various fruits and vegetables, is a fat-soluble polyphenolic flavonoid with diverse biological activities, including antitumor, antioxidant, anti-inflammatory, anti-angiogenic, lipid-lowering, and neuroprotective effects. These multifaceted properties make Fisetin an excellent anticancer drug. Meanwhile, studies have shown that MASLD is associated with increased liver inflammation and senescent cell burden. Acute or intermittent treatment of prematurely aging and naturally aging mice with Fisetin can reduce aging markers in various tissues. These studies indicate that Fisetin plays an important role in preventing aging and treating various diseases such as cancer and kidney disease.

[0008] To date, there are no reports in the existing technology regarding the prevention and treatment of metabolic-related fatty liver disease and obesity. Summary of the Invention

[0009] The purpose of this invention is to provide a YBX1 protein degrader, fisetinone, and its application in the preparation of drugs for the prevention and treatment of obesity or fatty liver, aiming to solve the current problem of lacking effective low-toxicity natural small molecule drugs for the prevention and treatment of metabolic-related fatty liver disease or obesity. The research of this invention found that fisetinone, as a YBX1 degrader, can improve hepatocyte ferroptosis and alleviate lipid accumulation, and has broad prospects in the preparation of drugs for the prevention and treatment of metabolic-related fatty liver disease or obesity.

[0010] The objective of this invention is achieved through the following technical solution:

[0011] The embodiments of this invention demonstrate both in vitro and in vivo experiments:

[0012] In in vitro experiments, fisetin was the molecule with the highest score when docking with YBX1 protein, as determined by LibDock virtual screening. Cell heat transfer assays confirmed the direct interaction between fisetin and YBX1. Treatment with fisetin and the protein translation inhibitor actinomycin (CHX) resulted in a dose-dependent decrease in YBX1 protein levels, indicating that fisetin promotes YBX1 degradation. The YBX1 protein level in the fisetin combined with MG132 treatment group was significantly higher than that in the fisetin-only treatment group. This suggests that fisetin-induced YBX1 degradation does indeed depend on the proteasome pathway. Simultaneously, fisetin dose-dependently reduced triglyceride levels in human hepatocellular carcinoma cells (HepG2), with a more significant effect at 100 μM. Fisetin also reduced lipid content in HepG2 cells, alleviating lipid accumulation. Furthermore, fisetin significantly reduced YBX1-induced intracellular Fe... 2+ These findings collectively suggest that fisetin reduces intracellular ferroptosis and lipid accumulation in hepatocytes.

[0013] In in vivo experiments, fisetin can alleviate high-fat diet-induced obesity and reduce triglyceride accumulation in the liver of mice fed a high-fat diet. Simultaneously, fisetin improves glucose tolerance and insulin sensitivity in high-fat diet-induced mice, and also reduces liver weight and fat content, alleviating liver damage. Furthermore, compared to mice fed a high-fat diet, mice supplemented with fisetin showed increased total iron and iron content in their liver tissue. 2+ The levels were significantly reduced. In conclusion, fisetin can improve glucose and lipid metabolism disorders, alleviate liver damage and ferroptosis caused by a high-fat diet, and has a significant effect in the prevention and treatment of metabolic-related fatty liver disease and obesity induced by a high-fat diet.

[0014] The results of in vitro and in vivo experiments in the embodiments of the present invention confirm the significant role of fisetin as a YBX1 protein degrader in the prevention and treatment of obesity and metabolic-related fatty liver disease, thus supporting the application of fisetin in the preparation of drugs for the prevention and treatment of obesity or fatty liver.

[0015] Specifically, the present invention discovers a YBX1 protein degrader, wherein the YBX1 protein degrader is fisetin.

[0016] The application of fisetin in the preparation of drugs for the prevention and treatment of obesity or fatty liver. More preferably, from the perspective of mechanism of action, the above application is based on the YBX1 protein degrader function of fisetin.

[0017] The prevention and treatment include one or more of prevention, relief or treatment, preferably prevention, relief or treatment in the process of obesity caused by a high-fat diet and metabolic-related fatty liver disease.

[0018] The obesity mentioned is preferably obesity induced by a high-fat diet.

[0019] The fatty liver is preferably metabolic-associated fatty liver disease; more preferably, the fatty liver is metabolic-associated fatty liver disease induced by a high-fat diet.

[0020] The drug for preventing or treating obesity or fatty liver is preferably an oral formulation.

[0021] The oral preparation is one of the following: capsule preparation, soft capsule preparation, oral liquid preparation, drop pill preparation, or tablet preparation.

[0022] The oral formulation further contains pharmaceutically acceptable excipients and / or carriers. The excipients and / or carriers are one or more selected from saline, sterile water, Ringer's solution, buffered saline, glucose solution, maltodextrin solution, glycerol, ethanol, lactose, glucose, sucrose, sorbitol, mannitol, starch, gum arabic, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, syrup, methylcellulose, methylparaben, propylparaben, talc, magnesium stearate, and mineral oil.

[0023] Compared with the prior art, the present invention has the following advantages:

[0024] This invention, through both in vitro and in vivo experiments, has found that fisetin can prevent and treat obesity and metabolic-associated fatty liver disease induced by a high-fat diet, and its application in the preparation of drugs for the prevention and treatment of these conditions. Furthermore, since its market launch, there have been no reports on the prevention and treatment of obesity and metabolic-associated fatty liver disease with fisetin. This invention also reveals that fisetin, as a YBX1 protein degrader, can improve hepatocyte ferroptosis and lipid accumulation. This study is the first to discover that YBX1 also plays an important role in metabolic-associated fatty liver disease (MASLD); specifically, YBX1 can promote MASLD by regulating ferroptosis-related pathways. YBX1 protein degraders alleviate metabolic-associated fatty liver disease and obesity induced by a high-fat diet. Moreover, compared to currently available high-cost weight-loss drugs, fisetin, belonging to the polyphenol flavonoid class, is found in various fruits and vegetables, is inexpensive, widely available, has fewer toxic side effects, is easily obtained, and has a lower cost. Attached Figure Description

[0025] Figure 1 The image shows the molecule with the highest docking score with the YBX1 protein obtained from the LibDock virtual screening.

[0026] Figure 2 The image shows the results of cell heat transfer assays for Fisetin and YBX1.

[0027] Figure 3 Figure showing the expression level of YBX1 protein in HepG2 human liver cancer cells after treatment with different concentrations of Fisetin.

[0028] Figure 4 The image shows the expression level of YBX1 protein in HepG2 human liver cancer cells after co-treatment with the protein translation inhibitor actinomycin (CHX) and different concentrations of Fisetin.

[0029] Figure 5 The figure shows the time-series analysis results of YBX1 protein expression levels in HepG2 human liver cancer cells after co-treatment with the protein translation inhibitor actinomycin (CHX) and fisetin.

[0030] Figure 6 The image shows the expression level of YBX1 protein in HepG2 human liver cancer cells after co-treatment with the protein translation inhibitor actinomycin (CHX), fisetin, and the proteasome inhibitor MG132.

[0031] Figure 7 The figure shows the effect of different concentrations of Fisetin on the activity of HepG2 human liver cancer cells.

[0032] Figure 8The image shows the results of TG concentration detection in HepG2 human liver cancer cells after treatment with different concentrations of Fisetin.

[0033] Figure 9 The image shows the fluorescence detection results of HepG2 human liver cancer cells BODIPY 493 / 503 after staining in OA / PA and Fisetin culture media.

[0034] Figure 10 The total iron and Fe content in YBX1-induced HepG2 human liver cancer cells after Fisetin treatment. 2+ Horizontal test results diagram.

[0035] Figure 11 Figure shows the weight of mice after high-fat diet and Fisetin treatment.

[0036] Figure 12 The figure shows the results of fat quality testing in mice after high-fat diet and Fisetin treatment.

[0037] Figure 13 The figure shows the results of glucose tolerance (GTT) tests in mice after a high-fat diet and fisetin treatment.

[0038] Figure 14 Figure 1 shows the results of insulin sensitivity (ITT) tests in mice after a high-fat diet and Fisetin treatment.

[0039] Figure 15 The image shows the results of liver tissue weight detection in mice after high-fat diet and Fisetin treatment.

[0040] Figure 16 This diagram illustrates the results of Oil Red O staining, HE staining, and Masson staining after a high-fat diet and Fisetin treatment.

[0041] Figure 17 The figure shows the results of detecting TG, aspartate aminotransferase (AST), and alanine aminotransferase (ALT) levels in the liver of mice after a high-fat diet and fisetin treatment.

[0042] Figure 18 Total iron and Fe content in the liver of mice after high-fat diet and Fisetin treatment 2+ Horizontal test results diagram. Detailed Implementation

[0043] 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 a part of the embodiments of the present invention, and not all of them. 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. In addition, the technical solutions of the various embodiments can be combined with each other, but this must be based on the ability of those skilled in the art to implement them. When the combination of technical solutions is contradictory or cannot be implemented, it should be considered that such combination of technical solutions does not exist and is not within the scope of protection claimed by the present invention.

[0044] Currently, there is a lack of effective, low-toxicity natural small molecules for the prevention and treatment of metabolic-related fatty liver disease and obesity, and existing drugs suffer from significant side effects and high prices. To address these technical problems, this invention proposes a YBX1 protein degrader, Fisetin, and its application in the preparation of drugs for the prevention and treatment of obesity or fatty liver.

[0045] Example 1

[0046] Technical Solution: In this embodiment, the regulatory effect of fisetin on hepatic lipid metabolism disorders was investigated using an oleic acid / palmitic acid (OA / PA)-induced HepG2 cell fat accumulation model and a high-fat diet-induced C57BL / 6 mouse obesity model, respectively. In vitro experiments: HepG2 cells were passaged and cultured in DMEM containing fetal bovine serum for 24 hours. An oleic acid / palmitic acid-induced fat accumulation model was used, and cells were treated with fisetin for 24 hours, with a control group not treated with fisetin. The effects of different concentrations of fisetin on YBX1 protein levels, ferroptosis, and lipid accumulation in HepG2 cells were explored. In vivo experiments: 24 eight-week-old C57BL / 6 mice were randomly divided into three groups (n=8 per group): (1) normal diet group (10% Kcal fat); (2) high-fat diet group (60% Kcal fat); (3) Fisetin-treated high-fat diet group (60% Kcal fat + Fisetin, Fisetin dosage 100 mg / Kg). The experiment lasted for eight weeks. The ambient temperature was 25±2℃ and the relative humidity was 50±5%. Incandescent lamps were used for 12 hours of light and 12 hours of darkness alternately. During the feeding period, the mice had free access to water and food. After the feeding period, the mice were sacrificed and tissue samples were collected to detect the effects of Fisetin on liver lipid metabolism, fat deposition, ferroptosis and liver damage in mice.

[0047] (a) The specific in vitro experiments are as follows:

[0048] 1. Detection of small molecules that can directly bind to YBX1 protein

[0049] A virtual screening was performed on 17,931 commercially available natural product molecules from the ZINC15 database. LibDock was used to identify the molecules that scored highest when docking with the YBX1 protein. Cellular thermal transfer assay (CETSA) was then used to determine if there was a direct interaction between this molecule and YBX1.

[0050] 2. Effect of fisetin on YBX1 protein levels in HepG2 cells.

[0051] Cell samples were treated with different concentrations of fisetin, and the expression level of YBX1 protein in HepG2 cells was detected by Western blot. Subsequently, the protein translation inhibitor actinomycin (CHX) was combined with the proteasome inhibitor MG132 to study protein stability and degradation pathways.

[0052] 3. Detection of the effect of fisetin on HepG2 cell viability

[0053] Cells were seeded in 96-well plates and incubated for 24 h. Then, they were treated with 0, 20, 40, 60, 80, and 100 μM fisetin for 24 h. After co-incubation with the cells using the Cell Counting Kit-8 (Solepro) for 1 h, the effect of fisetin on hepatocyte activity was detected.

[0054] 4. Detection of the effect of fisetin on lipid metabolism in HepG2 cells

[0055] Cell samples were treated with different concentrations of fisetin and the changes in intracellular and extracellular triglycerides (TG) were detected using a triglyceride assay kit. Cells in the OA / PA and fisetin groups were stained with BODIPY 493 / 503 dye, photographed using a fluorescence microscope, and the mean fluorescence intensity (MFI) was calculated.

[0056] 5. Detection of the effects of fisetin on total iron and Fe content in HepG2 cells. 2+ The influence of level

[0057] Using the FerroFarRed ferrous ion fluorescent probe and total iron / ferrous (Fe) ratio 2+ The ion detection kit was used to detect the effects of fisetin on intracellular total iron and Fe content in cells of PCDNA3.1+DMSO, PCDNA3.1+Fisetin, YBX1+DMSO, and YBX1+Fisetin groups. 2+ The influence of level.

[0058] (II) The specific details of the live animal experiments are as follows:

[0059] 6. To investigate the effect of fisetin on obesity induced by a high-fat diet.

[0060] During the in vivo experiment, the body weight of three groups of mice was measured weekly, and the body fat percentage of the mice in the three groups was measured by MRI. After the feeding period, adipose tissue was collected, photographed, and weighed to investigate the effect of fisetin on obesity induced by a high-fat diet.

[0061] 7. Detection of the effect of fisetin on glucose metabolism in mice on a high-fat diet

[0062] In week 10 of the experiment, after fasting for 16 hours, fasting blood glucose was measured by blood collected from the tail vein of mice. Glucose solution (2 g / kg) was then injected intraperitoneally, and blood glucose levels were measured at 15, 30, 60, 90, and 120 minutes after injection. In week 11 of the experiment, after fasting for 4 hours, fasting blood glucose was measured by blood collected from the tail vein of mice. Insulin (0.75 U / kg) was then injected intraperitoneally, and blood glucose levels were measured at 15, 30, 60, 90, and 120 minutes after injection.

[0063] 8. Effects of fisetin on fatty liver and liver damage in mice fed a high-fat diet

[0064] After the feeding experiment, liver tissue was collected, photographed, weighed, and subjected to experiments such as H&E staining, Oil Red O staining, Masson staining, TG detection, AST detection, and ALT detection to investigate the effects of fisetin on fatty liver and liver damage induced by a high-fat diet.

[0065] 9. Detection of the effect of fisetin on hepatocyte ferroptosis in the livers of mice fed a high-fat diet.

[0066] After the feeding experiment, liver tissue was collected, photographed, weighed, and the total iron content and Fe content of the liver tissue were analyzed. 2+ The effects of fisetin on ferroptosis in hepatocytes of mice fed a high-fat diet were investigated by measuring the levels of fisetin.

[0067] The experimental results show that:

[0068] 1. Fisexinone was the molecule with the highest docking score when docking with the YBX1 protein.

[0069] Virtual screening of 17,931 commercially available natural product molecules in the ZINC15 database revealed that fisetin was the molecule with the highest score when docking with the YBX1 protein (results are shown below). Figure 1 (As shown). The direct interaction between this molecule and YBX1 was then confirmed using a cell heat transfer assay (CETSA) (results shown in Figure 1). Figure 2 (As shown).

[0070] 2. Fisexin promotes the degradation of YBX1 protein.

[0071] Treatment of HepG2 cells with 0-100 μM felsone significantly reduced intracellular YBX1 protein levels, with the effect being more pronounced at 100 μM (results are shown in Figure 1). Figure 3 As shown); after co-treatment with fisetin and the protein inhibitor actinomycin (CHX), the YBX1 protein level decreased in a dose-dependent manner (results are shown in Figure 1). Figure 4 As shown in the figure), time-history analysis showed that the YBX1 protein level decreased in a time-dependent manner after fisetin treatment (results are shown in the figure). Figure 5 As shown above, these results indicate that fisetin promotes the degradation of YBX1 protein. Subsequently, the proteasome inhibitor MG132 was added to determine whether this degradation occurred via the proteasome pathway. The results showed that the YBX1 protein level in the fisetin combined with MG132 treatment group was significantly higher than that in the fisetin alone treatment group (results are shown in Figure 1). Figure 6 As shown in the figure, this indicates that fisetin-induced YBX1 degradation does indeed depend on the proteasome pathway.

[0072] 3. Fesedon had no significant effect on the viability of HepG2 cells.

[0073] To investigate the effect of fisetin on cell viability, cells were treated with different concentrations (0 μM, 20 μM, 40 μM, 60 μM, 80 μM, and 100 μM) of fisetin. It was found that fisetin had no toxic effect on HepG2 cells up to 100 μM (results are shown in Figure 1). Figure 7 (As shown).

[0074] 4. Fesedon alleviates lipid metabolism disorders in HepG2 cells.

[0075] Treatment of HepG2 cells with 0-100 μM fenestrone resulted in a dose-dependent decrease in intracellular triglyceride levels, with the effect being more significant at 100 μM (results are shown in Figure 1). Figure 8 (As shown). The mean fluorescence intensity of cells in the Fisetin-treated group was significantly lower than that in the OA / PA group (results are shown in Figure 1). Figure 9 (As shown in the image). The above results indicate that fisetin can alleviate lipid dysregulation and lipid accumulation in HepG2 cells.

[0076] 5. Fesedon significantly reduced total iron and Fe content in HepG2 cells. 2+ level

[0077] Using the FerroFarRed ferrous ion fluorescent probe and total iron / ferrous (Fe) ratio 2+An ion detection kit was used to detect cells in the PCDNA3.1+DMSO, PCDNA3.1+Fisetin, YBX1+DMSO, and YBX1+Fisetin groups. The results showed that the red fluorescence intensity was significantly weaker in the YBX1+Fisetin group compared to the YBX1+DMSO group (see results below). Figure 10 As shown in the figure, fisetin significantly reduces total iron content and Fe2+ in HepG2 cells. 2+ level.

[0078] 6. Fisesone alleviates obesity induced by a high-fat diet.

[0079] Twenty-four 8-week-old C57BL / 6 mice were randomly divided into three groups of eight each: a standard diet group (CK), a high-fat diet group (HFD), and a high-fat diet plus fisetin group (HFD+Fisetin). Treatment lasted for 8 weeks. The study found that mice in the HFD group experienced rapid weight gain. In contrast, the addition of fisetin to the high-fat diet significantly reduced the weight gain induced by the high-fat diet (results are shown in Figure 1). Figure 11 As shown), and reduced body fat percentage (results as shown). Figure 12 (As shown in the image). The above results indicate that fisetin can alleviate fat deposition induced by a high-fat diet in mice and reduce obesity.

[0080] 7. Fesetone can alleviate glucose metabolism disorders induced by a high-fat diet.

[0081] To further evaluate glucose and insulin levels in mice under different treatment conditions, GTT and ITT tests were performed at weeks 10 and 11 of treatment. GTT results are as follows: Figure 13 As shown, mice in the HFD group exhibited decreased glucose tolerance after glucose injection. Compared to the HFD group, the HFD+Fisetin group significantly reduced the rise in blood glucose levels after glucose injection and promoted the recovery of blood glucose to normal levels. ITT results are as follows... Figure 14 As shown, after insulin injection, the blood glucose level in the HFD group decreased slowly, indicating insulin resistance. In contrast, the blood glucose decrease in the HFD+Fisetin group was significantly lower than that in the HFD group, suggesting that adding fisetin to the diet can improve insulin resistance induced by HFD. The experimental results indicate that fisetin can alleviate glucose metabolism disorders caused by high-fat diets.

[0082] 8. Fesedon alleviates high-fat diet-induced hepatic steatosis.

[0083] After the mice were sacrificed, their liver tissues were collected. Compared with the CK group, the HFD group showed a significant increase in liver tissue weight, while the HFD+Fisetin group significantly reduced the increase in liver tissue weight (results are shown in Figure 1). Figure 15(As shown). Liver tissue was stained with H&E, Oil Red O, and Masson stain (results shown). Figure 16 As shown in the figure), Oil Red O staining results showed that compared with the HFD group, the number of liver lipid droplets was significantly reduced and lipid accumulation was significantly alleviated in the Fisetin group; HE staining results showed that compared with the HFD group, the Fisetin group had no obvious vacuoles and the number of liver lipid droplets was significantly reduced; Masson staining results showed that compared with the HFD group, the Fisetin group had almost no blue collagen fibers and no obvious collagen fiber hyperplasia and fibrosis. Further analysis of liver tissue for TG, ALT, and AST levels was performed (results are shown in the figure). Figure 17 As shown in the figure, the results showed that the levels of TG, ALT, and AST in the HFD+Fisetin group were significantly lower than those in the HFD group, indicating that fisetin can effectively alleviate fat accumulation and liver damage in mouse liver tissue and has a significant effect on the prevention and treatment of metabolic-related fatty liver disease induced by high-fat diet. This further verifies the application of fisetin in the preparation of drugs for the prevention and treatment of metabolic-related fatty liver disease induced by high-fat diet.

[0084] 9. Fiseridone significantly reduced hepatocyte ferroptosis in the livers of mice fed a high-fat diet.

[0085] After the mice were sacrificed, their liver tissue was collected, and the total iron content and Fe content of the liver tissue were analyzed. 2+ The levels were measured, and the results showed that the total iron content and Fe content in the liver of mice in the HFD+Fisetin group were... 2+ The levels were significantly reduced (results as follows). Figure 18 As shown in the figure, fisetin can effectively alleviate hepatocyte ferroptosis caused by a high-fat diet.

[0086] In summary, the experimental results in this embodiment of the invention demonstrate, from both in vitro and in vivo experimental perspectives, that:

[0087] In in vitro experiments, fisetin promoted the degradation of YBX1 protein via the proteasome pathway; fisetin dose-dependently reduced triglyceride levels in HepG2 human liver cancer cells, with more significant effects at 100 μM treatment; fisetin reduced lipid content in HepG2 human liver cancer cells, effectively alleviating lipid accumulation; fisetin significantly reduced total iron and Fe content in HepG2 cells. 2+ level.

[0088] In in vivo experiments, MRI scans of mice revealed that fisetin significantly slowed the increase in body weight and fat percentage induced by a high-fat diet, indicating its ability to improve obesity induced by a high-fat diet. Fisetin also reduced significant traits of metabolic-related fatty liver disease, such as triglyceride accumulation in the liver, demonstrating its significant efficacy in preventing and treating high-fat diet-induced metabolic-related fatty liver disease. Detection of glucose and insulin levels in mice under different treatment conditions showed that fisetin enhanced glucose tolerance and improved insulin resistance induced by a high-fat diet, indicating that fisetin can alleviate glucose metabolism disorders caused by a high-fat diet. Compared with the HFD group, the total iron content and Fe content in the liver of mice treated with fisetin were significantly higher. 2+ The levels were significantly reduced, indicating that fisetin can effectively alleviate hepatocyte ferroptosis induced by a high-fat diet. These results suggest that fisetin can improve liver damage, ferroptosis, and glucose and lipid metabolism disorders caused by a high-fat diet.

[0089] The results of the above examples indicate that fisetin is a YBX1 protein degrader.

[0090] Furthermore, fisetin is used in the preparation of drugs for the prevention and treatment of obesity or metabolic-related fatty liver disease.

[0091] The aforementioned prevention and treatment include one or more of prevention, relief, or treatment, preferably prevention, relief, or treatment in the process of obesity and fatty liver caused by a high-fat diet.

[0092] The obesity mentioned is preferably obesity induced by a high-fat diet.

[0093] The fatty liver mentioned is metabolic-associated fatty liver disease, preferably metabolic-associated fatty liver disease induced by a high-fat diet.

[0094] The drug for preventing and treating obesity or fatty liver is preferably an oral formulation.

[0095] The oral preparation is one of the following: capsule preparation, soft capsule preparation, oral liquid preparation, drop pill preparation, or tablet preparation.

[0096] The oral formulation further contains pharmaceutically acceptable excipients and / or carriers. The excipients and / or carriers are one or more selected from saline, sterile water, Ringer's solution, buffered saline, glucose solution, maltodextrin solution, glycerol, ethanol, lactose, glucose, sucrose, sorbitol, mannitol, starch, gum arabic, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, syrup, methylcellulose, methylparaben, propylparaben, talc, magnesium stearate, and mineral oil. This invention has the following advantages compared to the prior art:

[0097] This invention, through both in vitro and in vivo experiments, has found that fisetin can prevent and treat obesity and metabolic-related fatty liver disease induced by a high-fat diet, and can be used in the preparation of drugs for the prevention and treatment of these conditions. Furthermore, since its market launch, there have been no reports on fisetin's effectiveness in preventing obesity or improving metabolic-related fatty liver disease. This invention also found that fisetin, as a YBX1 protein degrader, can improve hepatocyte ferroptosis and lipid accumulation, alleviating metabolic-related fatty liver disease and obesity induced by a high-fat diet. Moreover, compared to currently available high-cost weight-loss drugs, fisetin, belonging to the polyphenol flavonoid class, is found in various fruits and vegetables, is inexpensive, widely available, has fewer toxic side effects, is easily obtained, and has a lower cost.

[0098] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A YBX1 protein degrading agent, characterized in that: The YBX1 protein degrading agent is fisetin.

2. Application of fisetin in the preparation of drugs for the prevention and treatment of obesity or fatty liver.

3. The use of phenothione according to claim 2 in the preparation of drugs for preventing and treating obesity or fatty liver, characterized in that: The aforementioned prevention and treatment include one or more of prevention, relief, or treatment.

4. The use of phenothione according to claim 1 in the preparation of drugs for preventing and treating obesity or fatty liver, characterized in that: The obesity mentioned refers to obesity induced by a high-fat diet.

5. The use of phenothione according to claim 1 in the preparation of drugs for preventing and treating obesity or fatty liver, characterized in that: The fatty liver mentioned refers to metabolic-associated fatty liver disease.

6. The use of phenothione according to claim 1 in the preparation of drugs for preventing and treating obesity or fatty liver, characterized in that: The fatty liver mentioned refers to metabolic-related fatty liver disease induced by a high-fat diet.

7. The use of phenothione according to claim 1 in the preparation of drugs for preventing and treating obesity or fatty liver, characterized in that: The drug in question is an oral preparation.

8. The use of phenothione according to claim 7 in the preparation of drugs for preventing and treating obesity or fatty liver, characterized in that: The oral preparation is one of the following: capsule preparation, soft capsule preparation, oral liquid preparation, drop pill preparation, or tablet preparation.

9. The use of phenothione according to claim 7 in the preparation of drugs for preventing and treating obesity or fatty liver, characterized in that: The oral formulation also contains pharmaceutically acceptable excipients and / or carriers.

10. The use of phenothione according to claim 9 in the preparation of drugs for preventing and treating obesity or fatty liver, characterized in that: The excipients and / or carriers are one or more of the following: saline, sterile water, Ringer's solution, buffered saline, glucose solution, maltodextrin solution, glycerol, ethanol, lactose, glucose, sucrose, sorbitol, mannitol, starch, gum arabic, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, syrup, methylcellulose, methylparaben, propylparaben, talc, magnesium stearate, and mineral oil.

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