CN105796563A - New application of lopinavir to medicines - Google Patents

Abstract

The invention provides a new drug application of lopinavir, in particular to the application of lopinavir in drugs for preventing or treating obesity, type II diabetes, diabetic nephropathy and non-alcoholic fatty liver. In the above application, the oral dosage range is 50 mg to 2000 mg; preferably 50 to 1000 mg.

Description

New medicinal uses of lopinavir

technical field

The invention belongs to the field of chemical medicine, and relates to a new medical application of lopinavir; in particular, it relates to preventing or treating obesity, type II diabetes, Diabetic nephropathy and non-alcoholic fatty liver and other related metabolic disorders.

Background technique

Type 2 diabetes is adult-onset diabetes, accounting for more than 90% of diabetic patients. People with type 2 diabetes are obese and develop insulin resistance, resulting in decreased sensitivity to insulin. Diabetic nephropathy is an important complication of diabetes, and nearly 40% of patients with late diabetes will develop diabetic nephropathy. Obesity is an unhealthy state of body fat accumulation and weight gain due to calorie intake exceeding consumption. The incidence of obesity is increasing rapidly in both developed countries and developing countries, and it is showing a trend of younger age. It has become a global epidemic that seriously affects health. At present, there are at least 200-300 million overweight people in my country, and a breakthrough in obesity patients 70 million people. Obesity not only affects people's daily life, but also is the main factor leading to type 2 diabetes, coronary heart disease, hypertension, dyslipidemia, cholecystitis and other diseases. Therefore, the prevention and treatment of obesity has important clinical significance.

High mobility group protein B1 (high mobility group protein, HMGB1) is a highly conserved nuclear protein widely distributed in mammalian cells. With the discovery of its late pro-inflammatory effect, HMGB1 has become a hot target in critical care medicine research in recent years. Hyperglycemia can promote the expression of HMGB1 [VolzHC1, SeidelC, LaohachewinD, KayaZ, MüllerOJ, PlegerST, LasitschkaF, BianchiME, RemppisA, BierhausA, KatusHA, AndrassyM.HMGB1:themissinglinkbetweendiabetesmellitusandheartfailure.BasicResCardiol.2010:105-8]5(6) . HMGB1 is closely related to obesity, especially the high expression of HMGB1 in macrophages in adipose tissue [WeisbergSP, McCannD, DesaiM, RosenbaumM, LeibelRL, FerranteAW, Jr. Obesity is associated with macrophageaccumulationinadiposetissue.JClinInvest.2003; .Adangerous duoinadipose issue: high-mobility group box 1 protein and macrophages. Yale J Biol Med. 2014;87(2):127-133.].

Clinically, the combination of lopinavir and ritonavir (trade name Kaletra) is used for the treatment of HIV/AIDS infection, and it can also be used in combination with other antiretroviral drugs for the treatment of HIV infection in adults and children over 6 months old , but the pharmacological effects of lopinavir in the prevention or treatment of obesity, type II diabetes, diabetic nephropathy and non-alcoholic fatty liver have not been reported. The inventor found through a large number of studies that lopinavir can prevent or treat obesity, type II diabetes, diabetic nephropathy and non-alcoholic fatty liver by inhibiting the expression of HMGB1. Based on this, the inventors have discovered that lopinavir inhibits the development of obesity, type II diabetes, diabetic nephropathy and non-alcoholic fatty liver by inhibiting the expression of IHMGB1, so as to prevent or treat obesity, type II diabetes, diabetic nephropathy and Nonalcoholic fatty liver.

Contents of the invention

The invention provides the application of lopinavir in the preparation of medicines for preventing or treating obesity, type II diabetes, diabetic nephropathy and non-alcoholic fatty liver.

The invention provides the application of lopinavir in the preparation of medicaments for preventing or treating obesity.

The invention provides the application of lopinavir in the preparation of medicaments for preventing or treating type II diabetes.

The invention provides the application of lopinavir in the preparation of medicaments for preventing or treating diabetic nephropathy.

The invention provides the application of lopinavir in the preparation of medicaments for preventing or treating non-alcoholic fatty liver.

The invention provides the application of lopinavir in medicines for obesity, type II diabetes, diabetic nephropathy and non-alcoholic fatty liver by inhibiting the expression of HMGB1.

The lopinavir provided by the present invention can be a medicine composed of a pharmaceutically acceptable carrier or adjuvant, and the medicine can be prepared into tablets, capsules, and drop pills by conventional pharmaceutical methods, preferably in the form of capsules.

When the lopinavir provided by the invention is used for obesity, type II diabetes, diabetic nephropathy and non-alcoholic fatty liver, its oral dosage range is 50mg-2000mg; preferably 50-1000mg.

detailed description

Example 1 Preparation of Lopinavir Capsules

Weigh 50.0g of lopinavir and 150.0g of sodium carboxymethyl starch, mix well and pass through a 100-mesh sieve, add an appropriate amount of 3% PVP _K30 aqueous solution to make a soft material, granulate with a 20-mesh sieve, and dry at 60°C for 3 hours , 18-mesh sieve, add 2.0g of magnesium stearate, mix evenly, pack into capsules, adjust the weight of the capsules to about 200mg, and get final product.

specific embodiment

Test Example 1 Effect of lopinavir on ob/ob obese mice

1.1 Experimental materials

Glycosylated hemoglobin assay kit, batch number: 0590072; HMGB1 primary antibody was purchased from Sigma; lopinavir (purity 99.5%, purchased from Dalian Meilun Biotechnology Co., Ltd.); high-density lipoprotein (HDL) kit (Beijing Zhong Biotechnology Company, batch number: 130706), low-density lipoprotein (LDL) kit (Beijing Zhongsheng Technology Company, batch number: 130806) and triglyceride (TG) kit (Beijing Zhongsheng Technology Company, batch number: 130406).

ob/ob mice, 50 males, 10 weeks old, weighing 50-55 g, were purchased from Nanjing Institute of Model Animals.

1.2 Experimental methods and results

50 ob/ob mice were randomly divided into 5 groups, namely model group, lopinavir 10mg/kg, 20mg/kg, 200mg/kg, 400mg/kg group by intragastric administration. In addition, 10 normal ob/ob mice were taken as the control group. The corresponding drugs were given to each group for 4 consecutive weeks. Weigh and calculate food intake, measure glycated hemoglobin (HbA1c), blood glucose, high-density lipoprotein (HDL), low-density lipoprotein (LDL) and triglyceride (TG). Get adipose tissue, homogenize, after BCA protein assay kit (Pierce company) measures protein concentration, get 50 μ g sample protein and carry out 10% sodium dodecylsulfonate polyacrylamide (SDS-PAGE) gel denaturation electrophoresis, then Transfer to polyvinylidene fluoride (PVDF) membrane, add HMGB1 primary antibody, β-actin as an internal reference, and detect the expression of HMGB1 protein in adipose tissue by Western blot. The difference between each administration group and the model group was compared, and the T test was carried out between the groups.

Table 1 The effect of lopinavir administration on ob/ob mice for 4 weeks (n=10)

^* , p<0.05, ^** , p<0.01, compared with the model group

Table 2 The effect of lopinavir administration on the blood biochemical indicators of ob/ob mice for 4 weeks (n=10)

^* , p<0.05, ^** , p<0.01, compared with the model group

The results in Table 1 and Table 2 show that the lopinavir 10mg/kg, 20mg/kg, 200mg/kg and 400mg/kg groups significantly reduced body weight and food intake, reduced plasma HbA1c, LDL and TG levels, and inhibited blood sugar elevation. High, inhibit the decrease of HMGB1 expression in adipose tissue, and increase HDL level (p<0.05 or p<0.01). The lopinavir 200mg/kg group reduced body weight and food intake, decreased plasma HbA1c, LDL and TG levels, inhibited the increase in blood sugar, inhibited the decrease in the expression of HMGB1 in adipose tissue, and increased HDL levels compared with the lopinavir 400mg/kg group. Significant difference.

The influence of test example 2 lopinavir on db/db type II diabetic mice

2.1 Experimental materials

Glycosylated hemoglobin assay kit, batch number: 0590072; HMGB1 primary antibody purchased from sigma; lopinavir (purity 99.5%, purchased from Dalian Meilun Biotechnology Co., Ltd.); db/db mice, 60 males, 10 weeks age, weight 50-55 g; purchased from Nanjing Institute of Model Animals.

2.2 Experimental methods and results

Fifty db/db mice were randomly divided into 5 groups, namely the model group and the lopinavir 10mg/kg, 20mg/kg, 200mg/kg, 400mg/kg groups by intragastric administration. In addition, 10 normal db/db mice were taken as the control group. The corresponding drugs were given to each group for 4 consecutive weeks. Weighing, determination of glycosylated hemoglobin (HbA1c), blood sugar,. Adipose tissue was taken, homogenized, and after the BCA protein assay kit (Pierce company) was used to measure the protein concentration, 50 μg of sample protein was taken to carry out 10% sodium dodecylsulfonate 2 polyacrylamide (SDS-PAGE) gel denaturing electrophoresis, Then transfer to polyvinylidene fluoride (PVDF) membrane, add HMGB1 primary antibody, β-actin as an internal reference, and Western blot to detect the expression of HMGB1 protein in adipose tissue. The difference between each administration group and the model group was compared, and the T test was carried out between the groups.

Table 3 The effect of lopinavir administration on db/db diabetic mice for 4 weeks (n=10)

^* , p<0.05, ^** , p<0.01, compared with the model group

The results in Table 3 show that the 10mg/kg, 20mg/kg, 200mg/kg and 400mg/kg groups of intragastric administration of lopinavir significantly reduce body weight, reduce plasma HbA1c levels, inhibit blood sugar elevation, and inhibit the decline of HMGB1 expression in adipose tissue (p <0.05 or p<0.01). Compared with the lopinavir 400mg/kg group, the lopinavir 200mg/kg group reduced body weight, decreased plasma HbA1c levels, inhibited blood sugar rise, and inhibited the decrease of HMGB1 expression in adipose tissue compared with the lopinavir 400mg/kg group.

Experimental Example 3 Effect of Lopinavir on Diabetic Nephropathy Rats

3.1 Experimental materials

Lopinavir (purity 99.5%, purchased from Dalian Meilun Biotechnology Co., Ltd.)

Benazepril, produced by Beijing Novartis Pharmaceutical Co., Ltd., batch number: 130106; streptozotocin (Sigma Company), blood glucose determination kit (Beijing Zhongsheng Reagent Co., Ltd. batch number: 130503); creatinine and urea nitrogen kit (Beijing Zhongsheng Technology Company, lot number: 130606).

Experimental animals: SPF grade Sprague Dawley rats, male, weighing 150g-200g, provided by the Experimental Animal Center of Shandong Luye Pharmaceutical Co., Ltd., the animal qualification certificate number is: SYXK (Lu) 20030020.

3.2 Test methods and results

100 male SD rats, after adaptive feeding for 1 week, all rats were anesthetized in 10% chloral hydrate (0.35mL/100g, ip), left nephrectomy was performed, and a single intravenous injection of 1% chain Ureazotocin solution 60mg/kg, streptozotocin with 0.1mol/L, pH is 4.5 citrate buffer solution dissolving, orbital blood collection after 72h, blood glucose measurement kit is measured, and blood glucose ≥ 16.7mmol/L is determined to be made The model was successful, and after 4 weeks, they were randomly divided into 6 groups, namely the model group, the benazepril (2mg/kg) group, the lopinavir 5mg/kg group, the lopinavir 10mg/kg group, and the lopinavir group. The pinavir 100mg/kg group, the lopinavir 200mg/kg group, and another 10 rats were taken as the normal control group. Each group was administered continuously for 8 weeks, blood was collected to measure serum creatinine and blood urea nitrogen, and 24h urinary protein excretion of rats in each group.

Get kidney tissue, homogenize, after BCA protein assay kit (Pierce company) measures protein concentration, take 50 μ g sample protein and carry out 10% sodium dodecylsulfonate 2 polyacrylamide (SDS-PAGE) gel denaturation electrophoresis, Then transfer to polyvinylidene fluoride (PVDF) membrane, add HMGB1 primary antibody, β-actin as internal reference, detect the expression of HMGB1 protein in kidney tissue by Western blot, compare the difference between each administration group and the model group, and make pathological sections of the kidney Perform microscopic examination.

Table 4 The effect of lopinavir administration on diabetic nephropathy rats for 8 weeks (n=10)

^* , p<0.05, ^** , p<0.01, compared with the model group

Table 5 The effect of lopinavir administration on renal coefficient and HMGB1 in rats with diabetic nephropathy for 8 weeks (n=10)

^* , p<0.05, ^** , p<0.01, compared with the model group

The results of Table 4 and Table 5 show that the lopinavir administrated 5mg/kg group, the lopinavir administrated 10mg/kg group, the lopinavir administrated 100mg/kg group, and the lopinavir administrated 200mg/kg group The 100mg/kg lopinavir intragastric administration group significantly reduced the serum creatinine and urea nitrogen 24h urinary protein excretion, and reduced the expression of HMGB1 in renal tissue (compared with the model control group, p<0.05 or 0.01); Creatinine, blood urea nitrogen, and 24h urinary protein excretion, decreased the expression of HMGB1 in renal tissue compared with the lopinavir 200mg/kg intragastric administration group, there was no significant difference.

Pathological examination: The glomerular basement membrane of rats in the normal control group was normal without widening, and the foot processes were arranged neatly. In the model control group, the glomerular basement membrane was significantly widened, the foot processes were disordered and fused, and the mesangial cells and matrix were mildly segmental hyperplasia. The lesions in the lopinavir administration groups were milder than those in the model group, and the pathology was gradually alleviated with the increase of the dose.

Test Example 4 Effect of Lopinavir on Nonalcoholic Fatty Liver Rats

4.1 Experimental materials

Lopinavir (purity 99.5%, purchased from Dalian Meilun Biotechnology Co., Ltd.)

Metformin tablets (Shanghai Xinyi Pharmaceutical Co., Ltd., batch number: 1307231); free fatty acid (FFA) detection kit, purchased from Nanjing Jiancheng Bioengineering Co., Ltd., batch number 20130715; triglyceride TG (enzymatic endpoint method) detection kit , Wittmann Biotechnology (Nanjing) Co., Ltd., batch number ZGY13307; total cholesterol (TC kit (Beijing Zhongsheng Technology Company, batch number: 130506)

Experimental animals: SPF grade Sprague Dawley rats, male, weighing 150g-200g, provided by the Experimental Animal Center of Shandong Luye Pharmaceutical Co., Ltd., the animal qualification certificate number is: SYXK (Lu) 20030020.

3.2 Test methods and results

After one week of adaptive feeding, the rats were numbered according to body weight and randomly divided into a normal group of 10 rats and a high-fat model group of 50 rats. The normal group was fed with common feed, and the basic feed formula was: 20% flour, 10% rice flour, 20% corn, 25% drum skin, 20% bean material, 2% fish meal, and 2% bone meal. The high-fat model group was fed with high-fat feed, and the formula of high-fat feed was: 77.6% of basic feed, 10% of lard, 2.0% of cholesterol, 0.2% of bile salt, 0.2% of propylthiosulfate, 5% of egg yolk powder, and 5% of sucrose. %. All had free access to drinking water. Modeling continued for 8 weeks, and rats were divided into model group, metformin 10mg/kg, and lopinavir 5mg/kg, 10mg/kg, 100mg/kg, 200mg/kg groups by intragastric administration according to body weight, and each group continued to receive high At the same time, the above-mentioned corresponding drugs were administered with fat feed, and the rats were killed at 16 weeks, the body weight was measured, the liver coefficient was calculated, blood samples were collected, and serum ALT, AST, TG, total cholesterol (TC), and free fatty acid (FFA) were measured. Get liver tissue, homogenate, BCA protein assay kit (Pierce company) after measuring protein concentration, take 50 μ g sample protein and carry out 10% sodium dodecylsulfonate 2 polyacrylamide (SDS-PAGE) gel denaturation electrophoresis, Then transfer to polyvinylidene fluoride (PVDF) membrane, add HMGB1 primary antibody, β-actin as internal reference, detect the expression of HMGB1 protein in liver tissue by Western blot, and compare the differences between each administration group and the model group.

Table 6 The effect of lopinavir administration on non-alcoholic fatty liver rats for 8 weeks (n=10)

^* , p<0.05, ^** , p<0.01, compared with the model group

Table 7 The effect of lopinavir administration on non-alcoholic fatty liver rats for 8 weeks (n=10)

^* , p<0.05, ^** , p<0.01, compared with the model group

The results of Table 6 and Table 7 show that metformin 10mg/kg group, lopinavir intragastric administration 5mg/kg group, lopinavir intragastric administration 10mg/kg group, lopinavir intragastric administration 100mg/kg group, lopinavir intragastric administration 100mg/kg group, The administration of 200mg/kg Weiwei group for 8 weeks significantly reduced body weight, liver coefficient and HMGB1 expression in liver tissue, and decreased serum ALT, AST, TG, TC and FFA (compared with the model control group, p<0.05 or 0.01); Navir gavage 100mg/kg group decreased body weight, liver coefficient and liver tissue HMGB1 expression, decreased serum ALT, AST, TG, TC and FFA compared with lopinavir 200mg/kg gavage group, there was no significant difference.

Claims (7)

1. A new medical application of lopinavir, specifically related to the application of lopinavir in the medicine for preventing or treating obesity, type II diabetes, diabetic nephropathy and non-alcoholic fatty liver. 2. The application according to claim 1, characterized in that lopinavir can prevent or treat obesity. 3. The application according to claim 1, characterized in that lopinavir can prevent or treat type II diabetes. 4. The application according to claim 1, characterized in that lopinavir can prevent or treat diabetic nephropathy. 5. The application according to claim 1, characterized in that lopinavir can prevent or treat non-alcoholic fatty liver. 6. The application according to any one of claims 2-5, characterized in that the dosage range of lopinavir is 50mg-2000mg. 7. The application according to claim 6, characterized in that the dosage range of lopinavir is preferably 50-1000 mg.