CN103550242B - Pharmaceutical composition for treating hepatic fibrosis and preparation method thereof - Google Patents

Abstract

The invention relates to a pharmaceutical composition for treating liver fibrosis and a preparation method thereof. It is a preparation prepared by taking pirfenidone and inosine as main components and pharmaceutically acceptable auxiliary materials, and its main component ratio is as follows: 100-300 parts of pirfenidone and 50-150 parts of inosine. The pirfenidone and inosine preparation of the invention is stable in quality, controllable, safe and effective. This preparation is mainly used clinically in the treatment of liver fibrosis, liver cirrhosis, liver cancer and other diseases.

Description

A pharmaceutical composition for treating liver fibrosis and its preparation method

technical field

The invention relates to a pharmaceutical composition for treating liver fibrosis and a preparation method thereof, belonging to the technical field of medicine.

Background technique

Hepatic fibrosis refers to the pathological process of chronic liver damage caused by various disease factors. During the repair process, the synthesis and degradation of extracellular matrix such as collagen are out of balance, resulting in the abnormal increase and excessive deposition of ECM in the liver. Liver fibrosis is the only way to develop into cirrhosis, and cirrhosis is an important cause of death in chronic liver diseases. Research suggests that liver fibrosis can be reversed. The etiology of liver fibrosis is complex and difficult to detect early. Drugs targeting liver fibrosis, such as interferon, colchicine, and traditional Chinese medicine, may have side effects or have inaccurate curative effects. Currently, no drug has been approved by the U.S. Food and Drug Administration ( FDA) approved for the treatment of liver fibrosis.

Pirfenidone PFD is a new pyridone compound with spectral anti-fibrosis effect that has been widely studied abroad in recent years. It has a good effect on fibrosis in lungs, heart, liver and other organs. PFD is a small molecular compound synthesized in 1970. Initial research found that it has a certain anti-inflammatory effect. Later, it was found that PFD has anti-fibrosis effect and has little adverse reactions. In 1995, Lyer et al. first reported that PFD could significantly prevent the formation of bleomycin-induced liver fibrosis in hamsters, and then PFD was proved to have a good anti-fibrosis effect in various organ fibrosis. However, pirfenidone has many adverse reactions, including nausea, dyspepsia, vomiting, anorexia, photosensitivity, rash and dizziness.

Inosine can directly enter the body cells through the cells and activate pyruvate oxidase, so that the cells in the low-energy and hypoxic state can continue to metabolize smoothly, and participate in the energy metabolism and protein synthesis of the human body. It is currently used clinically for white blood cells or platelets Decrease syndrome, various acute and chronic liver diseases, heart diseases such as pulmonary heart disease, central retinitis, optic atrophy and other diseases.

Contents of the invention

The invention provides a pharmaceutical composition for treating liver fibrosis, which contains pirfenidone and inosine as main components.

The medicine of the invention has stable quality, and the compatible use of pirfenidone and inosine can exert a synergistic drug effect, is convenient for clinical application, and provides a new choice for clinical practice.

The invention provides a pharmaceutical composition for treating hepatic fibrosis, which contains pirfenidone and inosine as main components and a preparation prepared from pharmaceutically acceptable auxiliary materials:

100-300 parts of pirfenidone, 50-150 parts of inosine.

Further preferably, it contains a preparation prepared from pirfenidone and inosine as main components:

200 parts of pirfenidone, 100 parts of inosine.

Wherein, the preparations are ordinary tablets, film-coated tablets, hard capsules, granules, enteric-coated tablets, and effervescent tablets.

Pharmaceutically acceptable auxiliary materials can be fillers, binders, disintegrants, lubricants and coating agents.

Further, the filler includes, but is not limited to, lactose, sucrose, glucose, mannitol, sorbitol, microcrystalline cellulose, pregelatinized starch, carboxymethyl starch, and hydroxypropyl cellulose.

Further, the binder includes but is not limited to starch, gelatin, dextrin, povidone, maltodextrin, sucrose, methylcellulose, carboxymethylcellulose, and hydroxypropylcellulose.

Further, the disintegrants include but are not limited to pregelatinized starch, microcrystalline cellulose, alginic acid, sodium carboxymethyl starch, cross-linked polyvinylpyrrolidone, low-substituted hydroxypropyl cellulose or effervescent disintegrants .

Detailed ways:

Embodiment 1 The preparation of pharmaceutical tablet of the present invention

prescription:

Pirfenidone 200 g

Inosine 100 g

Lactose 250 g

Microcrystalline Cellulose 50 g

Sodium Carboxymethyl Starch 30 g

Povidone K30 20 g

Made into 1000 pieces

Operation: Take pulverized and sieved 80-mesh pirfenidone, inosine, lactose, and microcrystalline cellulose according to the prescription amount, mix thoroughly with equal volume incremental mixing, and then pulverize and sieve. Take povidone K30, add distilled water to heat it to make a 5% solution of povidone K30, add it to the mixed powder to make a soft material, granulate it with a 20-mesh sieve, dry it at 60°C for 4 hours, collect the granules, add carboxymethyl Sodium starch, mixed evenly, granulated with a 18-mesh sieve to make a suitable preparation.

Embodiment 2 The preparation of pharmaceutical capsule of the present invention

prescription:

Pirfenidone 250 g

Inosine 80 g

Sodium carboxymethyl starch 25g

5% starch slurry Appropriate amount

Magnesium stearate 3 g

Made into 1000 capsules

Operation: Weigh crushed and sieved 80-mesh pirfenidone and inosine according to the prescription; add 5% starch slurry to make a suitable soft material, granulate with a 24-mesh sieve, dry at 60°C for 4 hours, and granulate Add sodium carboxymethyl starch and magnesium stearate, mix well, and fill into hollow capsules.

Example 3 Preparation of orally disintegrating tablets of the present invention

prescription:

Pirfenidone 180 g

Inosine 120 g

Sodium carboxymethyl starch 15g

Croscarmellose Sodium 15g

Microcrystalline Cellulose 30g

Pregelatinized starch 26g

Sucralose 4g

Lemon essence 1g

Magnesium stearate 3 g

Made into 1000 pieces

The raw materials and auxiliary materials of pirfenidone and inosine were pulverized respectively, and passed through an 80-mesh sieve. Mix pirfenidone and inosine with pregelatinized starch, microcrystalline cellulose, and carboxymethyl starch sodium evenly, use water as a binder to make a soft material, granulate with a 24-mesh sieve, and sieve through a 24-mesh sieve after drying grain. Add croscarmellose sodium, sucralose, lemon essence and magnesium stearate, mix well, and press into tablets to obtain the product.

Experimental example 1

1.1 Materials

1.1.1 Animals A total of 75 2-month-old male Wister rats, weighing 150-170 g, were purchased from the Animal Experiment Center of West China Hospital.

1.1.2 Reagents PFD was purchased from Zhejiang Warner Pharmaceutical Co., Ltd.; inosine was purchased from Jinan Mingxin Pharmaceutical Co., Ltd.; DMN (molecular formula C2H6N20, relative molecular weight 74.08) was purchased from Tianjin Institute of Chemical Reagents; laminin (LN) , hyaluronic acid (HA), type Ⅲ procollagen (PCⅢ), type Ⅳ collagen (Ⅳ-C) enzyme-linked immunoassay assay kits were purchased from Shanghai Haiyan Medical Biotechnology Center. Transforming growth factor β1 (TGF-β1) kit was purchased from Zhejiang University Biogenetic Engineering Co., Ltd.

1.2 Method

1.2.1 Establishment and grouping of rat models of liver fibrosis A rat model of liver fibrosis was established by intraperitoneal injection of DMN. Rats were fed normally for 1 week, and were randomly divided into 5 groups: normal group, model group, PFD group, inosine group, and combined drug group. Among them, rats in PFD group, inosine group, combination drug group and model group were injected with 1% DMN 10 mg/(kg d) (diluted in normal saline) continuously 3 days a week for a total of 4 weeks. The PFD group was given PFD 500 mg/(kg d) at the same time, the inosine group was given inosine 200 mg/(kg d) at the same time, and the combined drug group was given PFD 200 mg/(kg d) and inosine 100 mg/( kg·d) gavage, once a day, for 4 weeks in total, the model group and the normal group were gavaged with an equal volume of normal saline as a control. At the end of the 4th week, the rats were anesthetized by intraperitoneal injection of 8 mL of 3% pentobarbital sodium, and the body weight was weighed; the abdominal cavity was opened to observe changes in the liver and spleen, and heart blood was collected, centrifuged at 15,000 g for 3 min, and the supernatant was removed and stored at -20°C.

1.2..2 Determination of liver and spleen index in rats Cut out the intact liver and spleen, take out fascia and connective tissue, wash, drain the blood, weigh the liver and spleen wet weight (g), calculate the liver and spleen index, liver/spleen Spleen index = liver or spleen wet weight/body weight (g/100g).

1.2.3 Histopathological observation Liver tissue specimens were fixed with 10% neutral formaldehyde, dehydrated, paraffin-embedded and sectioned 5 μm, and slided routinely. Hepatic pathological changes were observed by conventional HE staining, and liver fibrosis was observed by Masson collagen staining. In accordance with the viral hepatitis prevention and treatment plan jointly revised by the Infectious Diseases and Parasitic Diseases Branch and the Hepatology Branch of the Chinese Medical Association in 2000, the liver tissue inflammation activity was graded and the fibrosis stage was staged.

1.2.4 Observation of ultramicrostructure Liver tissues with a size of about 1 mm ³ were fixed with 2.5% glutaraldehyde, and samples were prepared under a conventional electron microscope. Changes in liver ultrastructure were observed under a H-800 transmission electron microscope.

1.2.5 Serological index detection HA, LN, PCⅢ, Ⅳ-C and TCF-β1 concentrations were detected according to the instructions of the enzyme-linked immunoassay kit for glutamine transaminase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL ) and albumin (ALB) levels were detected by an automatic biochemical analyzer.

2.1 Changes in appearance and morphology of rats During the experiment, the hair color of the normal group was white and smooth, the luster was natural and soft, and the response was sensitive; 2 weeks after the start of the experiment, the hair color of the model group gradually turned yellow, the color became dim, and the food intake decreased. , the rats gradually became thinner, their urine turned yellow, their stools were loose, their spirits were low, and their reactions became slow; compared with the model group, the PFD group, inosine group, and combined drug group had better mental state, better appetite, and whiter and more shiny fur. , but worse than that of the normal group, and the appearance of the combined drug group was better than that of the PFD group and the inosine group. At the end of the course of treatment, there was no statistical difference in the mortality rate among the 5 groups.

2.2 Comparison of body weight, wet weight of liver and spleen, and liver and spleen index of rats in each group Compared with the normal group, there was no significant change in body weight, wet weight of liver and spleen, and liver index of rats in the PFD group, inosine group, and combined drug group; Compared with the model group, the body weight of the rats in the PFD group, the inosine group, and the combination drug group increased significantly, and the liver and spleen index decreased significantly, as shown in Table 1.

Table 1 The weight of rats in each group, the wet weight of liver and spleen, and the changes of liver and spleen index (x±s)

group no Body mass (m/g) Liver wet weight (m/g) Liver index (g/100g) Spleen wet weight (m/g) Spleen index (g/100g) normal group 15 324.61±13.90 12.17±0.51 3.75±0.05 0.92±0.07 0.28±0.02 model group 13 271.30±14.00 11.42±0.93 4.20±0.18 1.14±0.09 0.42±0.03 PFD group 14 304.90±26.50 11.96±0.41 3.92±0.07 1.00±0.03 0.32±0.11 Inosine group 16 312.20±20.10 12.05±0.84 3.86±0.07 0.99±0.08 0.32±0.08 Combined drug group 17 322.49±18.20 12.12±0.63 3.77±0.05 0.93±0.02 0.29±0.03

2.3 Determination results of serum HA, LN, PCⅢ, Ⅳ-C and TCF-β1 in each group of rats Serum HA, LN, PCⅢ, Ⅳ-C and TCF-β1 in the model group were significantly higher than those in the normal group. Rat serum HA, LN, PCⅢ, Ⅳ-C, PCⅢ and TCF-β1 in PFD group, inosine group and combined drug group were all decreased compared with model group, HA, LN, PCⅢ, Ⅳ-C and TCF in combined drug group The measurement results of -β1 content were similar to those of the normal group, and compared with the PFD group and the inosine group, they all decreased significantly, as shown in Table 2.

Table 2 Determination results of serum HA, LN, PCⅢ, Ⅳ-C and TCF-β1 in each group of rats (x±s)

group no HA (ρ/μg·L ^-1 ) LN (ρ/μg·L ^-1 ) PCⅢ (ρ/μg·L ^-1 ) Ⅳ-C (ρ/μg·L ^-1 ) TCF-β1 (ρ/ng·L ^-1 ) normal group 15 98.40±8.93 72.10±12.28 21.30±4.20 37.40±3.65 3.45±0.24 model group 13 178.90±20.46 124.10±19.82 40.40±3.23 82.50±10.56 11.90±1.50 PFD group 14 145.30±14.12 97.80±7.22 38.10±4.37 52.80±4.62 5.98±0.92 Inosine group 16 150.50±16.20 100.12±13.35 40.07±3.89 60.45±8.76 7..11±1.08 Combined drug group 17 110.10±9.27 80.07±10.23 30.27±2.78 39.17±2.45 3.89±0.46

2.4 Test results of serum ALT, AST, TBLL and ALB of rats in each group The serum of the model group decreased significantly; the serum ALS, AST and TBLL of the rats in the PFD group, inosine group, and the combined drug group decreased significantly compared with the model group, and ALB was significantly lower than that of the model group increased, see Table 3.

Table 3 Comparison of serum ALS, AST, TBLL and ALB of rats in each group (x±s)

group no ALT(U/L) AST(U/L) TBLL (c/μmol·L ^-1 ) ALB (ρ/g·L ^-1 ) normal group 15 60.24±7.68 76.27±13.60 1.86±0.67 35.10±2.27 model group 13 210.83±50.95 262.61±67.35 3.33±0.78 28.83±2.73 PFD group 14 130.36±31.29 151.52±25.07 1.64±0.39 33.74±3.70 Inosine group 16 146.42±58.17 120.35±5.04 2.45±0.84 32.17±2.59 Combined drug group 17 65.10±12.11 83.43±10.38 1.92±0.64 34.25±3.04

2.5 Results of MASSON staining The liver lobules of rats in the normal group had a clear structure without proliferation of collagen fibers. In the model group, the normal structure of the hepatic lobule disappeared, the intrahepatic vascular structure was disordered, the degeneration and necrosis of stem cells were severe, debris necrosis and bridging necrosis were seen, and a large number of inflammatory cells infiltrated in the portal area. In the PFD group and the inosine group, the structure of the hepatic lobule was basically normal, the liver cells were slightly swollen, a small amount of inflammatory cell infiltration was seen in the portal area, and the collagen fibers were slightly hyperplastic. In the combined treatment group, the structure of the hepatic lobules was clear, and the collagen fibers proliferated slightly.

Claims (8)

1. A pharmaceutical composition for the treatment of hepatic fibrosis, characterized in that: it contains pirfenidone and inosine as the preparation prepared from main components and pharmaceutically acceptable adjuvants: 100-300 parts of pirfenidone, 50-150 parts of inosine. 2.2. The pharmaceutical composition for treating liver fibrosis according to claim 1, characterized in that: it contains pirfenidone and inosine as the main components of the preparation: 200 parts of pirfenidone, 100 parts of inosine. 3.3. The pharmaceutical composition for treating liver fibrosis according to claim 1 or 2, characterized in that: said preparation is ordinary tablet, film-coated tablet, hard capsule, granule, enteric-coated tablet, effervescent tablet. 4.4． The pharmaceutical composition for treating liver fibrosis according to claim 1, characterized in that: the pharmaceutically acceptable auxiliary materials can be fillers, binders, disintegrants, lubricants and coating agents. 5.5． The pharmaceutical composition for treating liver fibrosis according to claim 4, characterized in that: the filler is lactose, sucrose, glucose, mannitol, sorbitol, microcrystalline cellulose, pregelatinized starch, carboxymethyl Starch, hydroxypropyl cellulose. 6.6. The pharmaceutical composition for treating liver cellulose according to claim 4, wherein the binder is starch, gelatin, dextrin, maltodextrin, sucrose, methylcellulose, carboxymethylcellulose , Hydroxypropyl cellulose. 7.7. The pharmaceutical composition for treating liver cellulose according to claim 4, wherein the disintegrating agent is pregelatinized starch, microcrystalline cellulose, alginic acid, sodium carboxymethyl starch, cross-linked polyethylene Pyrrolidone, low-substituted hydroxypropyl cellulose or effervescent disintegrant. 8.8． The preparation according to claim 1, characterized in that the preparation is mainly used clinically for the treatment of liver fibrosis, liver cirrhosis and liver cancer.