Application of pyruvate in the treatment of lipodystrophy syndrome

Abstract

The present invention provides the use of pyruvate in the treatment of lipodystrophy syndrome, belonging to the technical field of endocrine and metabolic disease treatment. This invention proposes for the first time the use of pyruvate, such as sodium pyruvate and potassium pyruvate, in the treatment of lipodystrophy syndrome. It has been found that pyruvate can restore atrophic adipose tissue and improve the progression of diabetic complications and acute pancreatitis associated with hypertriglyceridemia, thereby increasing patient survival and improving overall prognosis.

Description

Technical Field

[0001] The present invention belongs to the technical field of endocrine and metabolic disease treatment, and in particular relates to the application of pyruvate in treating lipodystrophy syndrome. Background Art

[0002] Lipodystrophy syndrome is a heterogeneous group of diseases characterized by complete or partial loss of adipose tissue. Due to mutations in key genes for lipid synthesis or viral infection, the patient's adipocyte lipid synthesis is impaired, resulting in a loss of body fat. The degree of adipose tissue loss can vary from systemic (involving almost all body fat), to partial (affecting only the limbs or upper body), to localized (limited to a small area). Hereditary lipodystrophy may manifest at birth, or symptoms of lipoatrophy may appear later in life. Acquired lipodystrophy usually occurs in childhood, but may also appear later. Patients with systemic or partial lipodystrophy are more likely to develop metabolic complications such as insulin resistance, diabetes, hypertriglyceridemia and non-alcoholic fatty liver disease (NAFLD). The severity of metabolic complications is usually related to the extent of lipoatrophy.

[0003] The current 2021 JES practice guidelines for lipodystrophy syndrome point out that the treatment of lipodystrophy syndrome mainly includes diet therapy, exercise therapy and drug therapy. In the diet, the energy intake of patients with lipodystrophy syndrome should be determined based on 25-35kcal / kg × standard weight (kg) calculated according to the patient's height, and factors such as age, gender and physical activity should be taken into account. In addition, patients with hypertriglyceridemia need to limit fat intake. Exercise therapy refers to improving metabolic disorders and other diseases by maintaining skeletal muscle mass and promoting energy consumption. For patients who are effectively improved by the above treatments, the guidelines recommend the use of drug therapy, which mainly includes three categories. The first category is antidiabetic drugs (thiazolidinediones, sodium-glucose co-transporter 2 (SGLT2) inhibitors, glucagon-like peptide-1 (GLP-1) receptor agonists, etc.) and antilipidemic drugs (fibrates, statins, etc.) for diabetes and hypertriglyceridemia. The second category is insulin-like growth factor 1 (IGF-1) treatment. IGF-1 has been reported to be effective for diabetic patients with severe insulin resistance (including lipodystrophy), but it lacks an appetite-boosting effect and carries the potential risk of exacerbating hypertrophic cardiomyopathy associated with lipodystrophy. Because leptin levels are absolutely deficient in patients due to adipose tissue loss, a third class of drugs is leptin replacement. Currently, the FDA has approved the use of a leptin analog, metreleptin, for the treatment of lipodystrophy, alleviating fatty liver disease, blood lipids, and blood sugar levels in patients.

[0004] However, exercise and diet therapy have limited efficacy in treating patients with lipodystrophy syndrome, and patients with generalized or partial lipodystrophy syndrome often require drug therapy. Current drug treatments still have significant limitations. Antidiabetic drugs, such as thiazolidinediones, can improve hyperglycemia or hypertriglyceridemia in some patients with lipodystrophy syndrome but are less effective in patients with generalized lipodystrophy syndrome. The insulin sensitizer metformin, while effective in HIV-associated lipodystrophy, is not suitable for other forms of lipodystrophy syndrome. Sodium-glucose cotransporter 2 (SGLT2) inhibitors, due to their insulin-independent mechanism of action, are expected to exert some glycemic control. Hyperlipidemia associated with lipodystrophy syndrome can be treated with drugs such as fibrates, statins, eicosapentaenoic acid (EPA), or intestinal cholesterol transporter inhibitors, but achieving a stable and significant therapeutic effect is generally difficult. Although IGF-1 has been reported to be effective in patients with lipodystrophy syndrome who have severe insulin resistance, IGF-1 lacks appetite-enhancing effects and carries the potential risk of developing hypertrophic cardiomyopathy associated with lipodystrophy syndrome. Therefore, IGF-1 has not been widely used to treat lipodystrophy syndrome. Although metreleptin can alleviate patients' complications such as fatty liver, blood lipids and blood sugar, it still cannot restore atrophic adipose tissue.

[0005] In summary, although currently known treatments can improve metabolic diseases such as diabetes, hyperlipidemia, and fatty liver in some patients, there is no treatment that can restore atrophic adipose tissue. Summary of the Invention

[0006] In view of this, the purpose of the present invention is to provide the use of pyruvate in the preparation of a drug for treating lipodystrophy syndrome. Pyruvate can restore atrophic adipose tissue, treat lipodystrophy syndrome from the root and improve subsequent metabolic complications.

[0007] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0008] The present invention provides the use of pyruvate in preparing a medicine for treating lipodystrophy syndrome.

[0009] Preferably, the pyruvate increases the expression levels of lipid synthesis-related genes Acaca and Fasn, and increases the expression level of the glucose transporter gene Glut4.

[0010] Preferably, the pyruvate increases the activation level of the insulin signaling pathway.

[0011] Preferably, the pyruvate increases the acetyl-CoA content.

[0012] Preferably, the pyruvate increases the acetylation level of the histone site H3K14 in the promoter region of the downstream target gene of the insulin signaling.

[0013] Preferably, the pyruvate increases body fat content.

[0014] Preferably, the pyruvate increases subcutaneous fat weight.

[0015] Preferably, the effective dose of pyruvate is 0.02 mg / kg to 2 mg / kg body weight.

[0016] Preferably, the pyruvate includes sodium pyruvate and potassium pyruvate.

[0017] The present invention also provides a medicine for treating lipodystrophy syndrome, wherein the active ingredient of the medicine comprises an effective dose of pyruvate, and the pyruvate comprises sodium pyruvate and potassium pyruvate.

[0018] Beneficial effects of the present invention:

[0019] This study proposes for the first time that pyruvates, such as sodium pyruvate and potassium pyruvate, can treat lipodystrophy syndrome. By upregulating the expression of key genes for lipid synthesis, pyruvate can promote adipocyte differentiation, restore adipose tissue volume, and rejuvenate atrophic adipose tissue, thereby addressing the root causes of lipodystrophy syndrome and subsequent metabolic diseases. Pyruvate can also improve the progression of diabetic complications such as acute pancreatitis associated with hypertriglyceridemia, thereby increasing patient survival and improving overall prognosis, potentially making a complete cure for lipodystrophy syndrome possible. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 This is an oil red staining image of primary inguinal white adipose tissue (iWAT) cells in a mouse model of lipoatrophy after sodium pyruvate intervention in the early stage of induction;

[0021] Figure 2 This is the quantitative graph of acetyl-CoA after sodium pyruvate intervention in the early stage of induction of iWAT primary cells in the lipoatrophy mouse model;

[0022] Figure 3 To investigate the expression changes of lipid synthesis-related genes and glucose transporter genes after early sodium pyruvate intervention in primary WAT cells of the lipoatrophy mouse model;

[0023] Figure 4 To investigate the expression changes of lipid synthesis-related genes Acaca and Fasn regulatory proteins after early sodium pyruvate intervention in primary WAT cells of the lipoatrophy mouse model;

[0024] Figure 5 After adding sodium pyruvate and histone acetyltransferase inhibitor A485 to adipocytes, the expression of histone acetylation site H3K14 protein changes;

[0025] Figure 6 For lipoatrophy model (PPARγ - / - ) Changes in the expression of lipid synthesis genes and glucose transporter genes in adipose tissue of mice treated with sodium pyruvate;

[0026] Figure 7 For lipoatrophy model (PPARγ - / - ) Changes in body fat content in mice treated with sodium pyruvate;

[0027] Figure 8 For lipoatrophy model (PPARγ - / - ) Changes in subcutaneous fat weight in mice treated with sodium pyruvate;

[0028] Figure 9 Figure 3 shows the weight changes of control mice after treatment with sodium pyruvate.

[0029] Figure 10 For lipoatrophy model (PPARγ - / - ) Changes in the expression of lipid synthesis genes and glucose transporter genes in adipose tissue of mice treated with potassium pyruvate;

[0030] Figure 11 For lipoatrophy model (PPARγ - / - ) Changes in body fat content in mice treated with potassium pyruvate;

[0031] Figure 12 For lipoatrophy model (PPARγ - / - ) Changes in subcutaneous fat weight in mice treated with potassium pyruvate;

[0032] Figure 13 Figure 3 shows the weight changes of control mice after treatment with potassium pyruvate. DETAILED DESCRIPTION

[0033] The present invention provides the use of pyruvate in the preparation of a medicament for treating lipodystrophy syndrome. The pyruvate described herein includes sodium pyruvate, potassium pyruvate, and the like. The present invention does not particularly limit the specific source of the pyruvate; any commercially available product in the art can be used.

[0034] The pyruvate of the present invention can increase the expression levels of lipid synthesis-related genes Acaca and Fasn, the expression level of the glucose transporter gene Glut4, and the activation level of the insulin signaling pathway.

[0035] Pyruvate of the present invention can increase acetyl-CoA content, improve histone site H3K14 acetylation level. Fatty acid de novo synthesis is to convert citric acid into acetyl-CoA by ATP-citrate lyase (ACLY), and the acetyl-CoA produced is carboxylated to propionate-CoA by acetyl-CoA carboxylase (ACACA), and then fatty acid synthase (FASN) realizes the conversion of propionate-CoA to palmitic acid, and finally through a series of reactions, palmitic acid is converted into complex fatty acid. In addition, in the nucleus, the rising of acetyl-CoA concentration can increase lysine acetyltransferase (KAT) activity to enhance histone acetylation, thereby enhancing gene expression. Therefore, pyruvate, by raising the content of acetyl-CoA, promotes the generation of de novo synthesis of fatty acids, and simultaneously promotes the generation of protein acetylation modification of lipid synthesis-related genes, finally realizing the regeneration of fat.

[0036] The pyruvate of the present invention can increase body fat content and subcutaneous fat weight.

[0037] The effective dosage of the sodium pyruvate of the present invention is 0.02 mg / kg to 2 mg / kg body weight, preferably 0.2 mg / kg to 1.5 mg / kg body weight.

[0038] The present invention also provides a drug for treating lipodystrophy syndrome, wherein the active ingredient of the drug includes an effective dose of a pyruvate salt, such as sodium pyruvate or potassium pyruvate. The active ingredient in the drug for treating lipodystrophy syndrome of the present invention may be sodium pyruvate or potassium pyruvate or a pyruvate salt as the sole active ingredient, or sodium pyruvate or potassium pyruvate or a pyruvate salt may be used in combination with other medicinally active ingredients that have the effect of treating lipodystrophy syndrome.

[0039] The drug of the present invention also includes a pharmaceutically acceptable carrier. The present invention does not specifically limit the other excipients and dosage forms contained in the drug; commonly used excipients and dosage forms in the art may be used. The content of pyruvate, such as sodium pyruvate or potassium pyruvate, in the drug of the present invention is 0.1 to 99.9 wt%.

[0040] The technical solutions provided by the present invention are described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0041] In the following examples, unless otherwise specified, all methods are conventional.

[0042] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0043] In a specific embodiment of the present invention, sodium pyruvate and potassium pyruvate were purchased from Sangon Biotech (Shanghai) Co., Ltd.

[0044] Example 1

[0045] This example conducts in vitro experiments on primary iWAT cells of a mouse model:

[0046] Three- to four-week-old male control mice and lipoatrophic mice were sacrificed by cervical dislocation. Inguinal white adipose tissue (iWAT) was isolated, digested, and centrifuged to obtain primary adipocytes. Primary adipocytes were cultured in F12 / DMEM supplemented with 10% FBS and recombinant basic fibroblast growth factor (rmFGF) (10 ng / ml). Two days after cells reached confluence, the cell culture medium was changed to adipocyte induction medium A for two days. Cells were then cultured in adipocyte induction medium B for six days. Medium A: F12 / DMEM supplemented with 10% FBS, insulin (8.4 μg / mL), isobutylmethylxanthine (IBMX, 0.5 mmol / L), and dexamethasone (1 μmol / L); Medium B: F12 / DMEM supplemented with 10% FBS and 8.4 μg / mL insulin. On days 2 to 8 of induction, sodium pyruvate (10 mmol / L) was added to the experimental group, while an equal volume of distilled water was added to the control group. The culture medium was changed every 2 days until differentiation was complete.

[0047] After induction, oil red staining was performed; acetyl-CoA content was detected using an acetyl-CoA detection kit; RNA was extracted for qPCR detection of related gene expression; protein was extracted for Western blotting to detect related protein expression. Figures 1 to 4 .

[0048] Oil red staining Scale bar: 250 μm. All data are mean ± SEM, p < 0.05 = *, p < 0.01 = **, p < 0.001 = ***, Figure 2 and Figure 3 The results were analyzed using Two-way ANOVA.

[0049] Figure 1 This is an oil red staining image of primary inguinal white adipose tissue (iWAT) cells from a mouse model of lipoatrophy after sodium pyruvate intervention in the early stage of induction. Figure 1 It can be seen that after sodium pyruvate intervention in the early stage of induction, the lipid droplets increased significantly compared with the group without sodium pyruvate intervention, indicating that sodium pyruvate can restore the inguinal white fat (iWAT) of the fat atrophy mouse model.

[0050] Figure 2 This is the quantitative chart of acetyl-CoA in primary cells of iWAT mouse model after sodium pyruvate intervention in the early stage of induction. Figure 2 It can be seen that the acetyl-CoA content in primary cells of iWAT from the fat atrophy mouse model was significantly increased after intervention with sodium pyruvate.

[0051] Figure 3 This study examines the expression changes of lipid synthesis-related genes and glucose transporter genes in primary WAT cells of a mouse model of lipoatrophy after early sodium pyruvate intervention. Figure 3 It can be seen that after sodium pyruvate intervention, the expression of lipid synthesis-related genes Acaca, Fasn and glucose transporter gene Glut4 in primary cells of iWAT mouse model was significantly increased.

[0052] Figure 4 This study investigates the expression changes of lipid synthesis-related genes Acaca, Fasn and other regulatory proteins after early sodium pyruvate intervention in primary cells of iWAT mouse model. Figure 4 It can be seen that after sodium pyruvate intervention in the primary cells of iWAT in the fat atrophy mouse model, the levels of insulin signaling pathway and lipid synthesis-related gene regulatory proteins significantly increased.

[0053] Example 2

[0054] This example uses the adipocyte cell line 3T3 / L1 to perform an experiment to detect histone acetylation levels after sodium pyruvate intervention:

[0055] The levels of histone acetylation sites were measured using the adipocyte cell line 3T3 / L1. After 3T3 / L1 adipocytes reached confluency for 2 days, the cells were switched to medium A for 2 days. The cells were then cultured in medium B for 6 days. Medium A consisted of DMEM supplemented with 10% FBS, insulin (8.4 μg / mL), isobutylmethylxanthine (IBMX, 0.5 mmol / L), dexamethasone (1 μmol / L), and rosiglitazone (1 μmol / L); medium B consisted of DMEM supplemented with 10% FBS and 8.4 μg / mL insulin. Sodium pyruvate (10 mmol / L) was added on days 2 to 8 of induction, while an equal amount of distilled water was added to the control group. On day 7 of induction, the histone acetyltransferase inhibitor A485 was added, while an equal amount of DMSO was added to the control group. The medium was changed every 2 days until differentiation was complete.

[0056] After induction, protein was extracted and Western blotting was performed to detect the expression of histone acetylation site H3K14. Figure 5 .

[0057] Figure 5 After adding sodium pyruvate and histone acetyltransferase inhibitor A485 to adipocytes, the expression of histone acetylation site H3K14 protein changes. Figure 5 It can be seen that after adding sodium pyruvate, the acetylation level of histone site H3K14 was significantly increased, and after adding the histone acetyltransferase inhibitor A485, the increase effect declined.

[0058] Example 3

[0059] This example uses a lipoatrophy model (PPARγ - / - ) In vivo experiments with sodium pyruvate in mice:

[0060] An 8-week-old male lipoatrophy model (PPARγ - / - In vivo sodium pyruvate experiments were conducted on mice treated with pyruvate and normal control mice. Multiple injections were performed at 48-hour intervals into the subcutaneous fat of the mice using an in situ injection method. The experimental group received 0.2 mg / kg body weight of sodium pyruvate, while the control group received an equal volume of distilled water. This experiment was repeated five times over a total of 10 days.

[0061] After the 10-day experiment, the body fat of the mice was measured using an animal fat analyzer (Echo MRI); the mice were killed by cervical dislocation, and subcutaneous fat was separated and weighed; and RNA was extracted from some tissues for qPCR detection of related gene expression. Figures 6-9 .

[0062] All data are mean ± SEM, p < 0.05 = *, p < 0.01 = **, p < 0.001 = ***, Figure 6 、 Figure 7 and Figure 8 The results were analyzed using Two-way ANOVA.

[0063] Figure 6 For lipoatrophy model (PPARγ - / - ) mice treated with sodium pyruvate, the expression of lipid synthesis genes and glucose transporter genes (Glut4, Acaca, Fasn) in adipose tissue. Figure 6 It can be seen that the fat atrophy model (PPARγ - / - ) mice were treated with sodium pyruvate, and the expression of lipid synthesis-related genes Acaca, Fasn and glucose transporter gene Glut4 increased significantly.

[0064] Figure 7 For lipoatrophy model (PPARγ - / - ) mice treated with sodium pyruvate and changes in body fat content. Figure 7 It can be seen that the fat atrophy model (PPARγ - / - ) mice showed a significant recovery in fat content after sodium pyruvate intervention.

[0065] Figure 8 For lipoatrophy model (PPARγ - / - ) mice treated with sodium pyruvate and the weight changes of inguinal subcutaneous fat (iWAT). Figure 8It can be seen that the fat atrophy model (PPARγ - / - ) mice also showed a significant increase in inguinal subcutaneous fat weight after sodium pyruvate intervention.

[0066] Figure 9 Figure 2 shows the body weight of control mice after treatment with sodium pyruvate. Figure 9 It can be seen that the body weight of the two-week sodium pyruvate intervention group had no significant change compared with the non-sodium pyruvate injection group, proving that sodium pyruvate intervention has no effect on the growth of mice.

[0067] Example 4

[0068] This example uses a lipoatrophy model (PPARγ - / - ) In vivo experiments with potassium pyruvate in mice:

[0069] After observing a range of effects of sodium pyruvate, another common pyruvate salt, pyruvate, was chosen for in vivo experiments in mice.

[0070] The 8-week-old male lipoatrophy model (PPARγ - / - In vivo potassium pyruvate experiments were conducted on mice treated with pyruvate and normal control mice. Multiple in situ injections were administered into the subcutaneous fat of the mice every 48 hours. The experimental group received 0.2 mg / kg of potassium pyruvate, while the control group received an equal amount of distilled water. Five injections were administered over a total of 10 days.

[0071] After the 10-day experiment, the body fat of the mice was measured using an animal fat analyzer (Echo MRI); the mice were killed by cervical dislocation, and subcutaneous fat was separated and weighed; and RNA was extracted from some tissues for qPCR detection of related gene expression. Figures 10-13 .

[0072] All data are mean ± SEM, p < 0.05 = *, p < 0.01 = **, p < 0.001 = ***, Figure 10 、 Figure 11 and Figure 12 The results were analyzed using Two-way ANOVA.

[0073] Figure 10 For lipoatrophy model (PPARγ - / - ) mice treated with potassium pyruvate, the expression of lipid synthesis genes and glucose transporter genes (Glut4, Acaca, Fasn) in adipose tissue. Figure 10 It can be seen that the fat atrophy model (PPARγ - / - ) mice were treated with potassium pyruvate, and the expression of lipid synthesis-related genes Acaca, Fasn and glucose transporter gene Glut4 increased significantly.

[0074] Figure 11 For lipoatrophy model (PPARγ - / - ) mice treated with potassium pyruvate and changes in body fat content. Figure 11 It can be seen that the fat atrophy model (PPARγ - / - ) mice showed a significant recovery in fat content after potassium pyruvate intervention.

[0075] Figure 12 For lipoatrophy model (PPARγ - / - Changes in inguinal subcutaneous fat (iWAT) weight in mice treated with potassium pyruvate. Figure 12 It can be seen that the fat atrophy model (PPARγ - / - ) mice also showed a significant increase in inguinal subcutaneous fat weight after potassium pyruvate intervention.

[0076] Figure 13 The weight of control mice after treatment with potassium pyruvate. Figure 13 It can be seen that the body weight of the two-week potassium pyruvate intervention group had no significant change compared with the non-potassium pyruvate injection group, proving that potassium pyruvate intervention has no effect on the growth of mice.

[0077] These experimental results demonstrate that both sodium pyruvate and potassium pyruvate can effectively promote fatty acid synthesis and lipid droplet formation without affecting normal growth in mice. Their improved insulin signaling pathways can also alleviate the insulin resistance associated with lipodystrophy. Furthermore, the addition of sodium pyruvate increases acetyl-CoA levels by increasing KAT activity and enhancing histone acetylation. This demonstrates that pyruvates, such as sodium pyruvate and potassium pyruvate, can be used in the treatment of lipodystrophy by restoring atrophic adipocytes and adipose tissue.

[0078] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as within the scope of protection of the present invention.

Claims

1. The use of pyruvate in the preparation of a drug for treating lipodystrophy syndrome, characterized in that: The pyruvate salts are sodium pyruvate and potassium pyruvate; the effective dosage of the pyruvate salts is 0.02 mg / kg to 2 mg / kg of body weight.

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