CN108524501A - Purposes of the Deferasirox in preparing treatment flesh and lacking disease drug - Google Patents

Abstract

The present invention relates to a new use of deferasirox, in particular to the use of deferasirox in the preparation of medicines for treating sarcopenia, which belongs to the technical field of new uses of medicine; at present, there is no clinical treatment for sarcopenia involving iron accumulation factors It has been proved by the present invention that deferasirox can significantly inhibit skeletal muscle atrophy and oxidative stress damage under the pathological state of sarcopenia, and is suitable for being developed as a preventive drug for sarcopenia.

Description

Use of deferasirox in the preparation of medicines for the treatment of sarcopenia

technical field

The invention relates to the use of deferasirox, in particular to the use of deferasirox in the preparation of medicines for treating sarcopenia, and belongs to the technical field of medicine.

Background technique

The chemical name of deferasirox (also known as defersino, DFS) is 4-[3,5-bis(2-hydroxyphenyl)-1,2,4-triazol-1-yl]benzoic acid, The molecular formula is C21H15N3O4, and the molecular weight is 373.36200. Deerasirox is a new type of oral triligand iron chelator, which binds Fe ^{3 +} at a ratio of 2:1, can effectively reduce the iron load in the body, and is clinically applied to chronic iron accumulation in children with thalassemia over 2 years old And patients with iron accumulation caused by other transfusion-dependent diseases. Clinical studies have confirmed that compared with deferoxamine and deferiprone, other iron chelators, deferasirox is safer to use, with higher patient tolerance and better compliance.

Sarcopenia is a progressive, age-related, systemic decrease in muscle mass and/or muscle strength or decreased muscle physiological function. It is one of the important causes and manifestations of the gradual decline in physiological function in the elderly. The occurrence of sarcopenia is considered to be related to various factors such as aging, nutrition, oxidative damage, apoptosis, etc., and its pathogenesis is complex and overlapping. At present, its prevention and treatment methods mainly include nutritional supplementation, sports rehabilitation training, vitamin D supplementation, and hormone replacement therapy. Consider other factors.

In recent years, studies have found that serum ferritin in people with sarcopenia is significantly elevated, and sarcopenia may be related to iron accumulation. However, there is no relevant report on reducing iron accumulation to prevent sarcopenia, and there is no report on the iron chelator deferasirox reducing iron accumulation in the treatment of sarcopenia.

Contents of the invention

The problem solved by the present invention is to overcome the lack of drugs for sarcopenic diseases in the prior art, and provide a use of deferasirox in the preparation of drugs for treating sarcopenic diseases.

The invention provides a use of deferasirox in the preparation of medicines for treating sarcopenia.

According to the preclinical research results of deferasirox interfering with skeletal muscle cells and the drug method of deferasirox for iron accumulation diseases in the blood system, the present invention draws that when the present invention treats sarcopenia, preferably, said deferasirox The dose of rosi is 10-20 mg/kg per day.

In the present invention, deferasirox is used to intervene skeletal muscle cells in a state of simulating sarcopenia, and it is found that it has the effect of reversing skeletal muscle atrophy or alleviating skeletal muscle damage. This discovery will have a major impact on the research of sarcopenic diseases, and has clinical practical value, opening up new clinical uses for deferasirox. It proves that deferasirox is not only used for the treatment of blood system iron accumulation diseases, but also can be used as a tool drug in other system diseases with iron accumulation.

The administration of deferasirox for humans can be administered alone or in any convenient pharmaceutical form, including making tablets containing the compound, and the tablets can be ordinary tablets or dispersible tablets. Oral administration is preferred. Pharmaceutical carriers suitable for formulating the compound, including lactose, starch, magnesium stearate, microcrystalline cellulose, povidone, etc., can be used in the tablet.

The advantage that the present invention compares with prior art is:

At present, there is no drug for the treatment of sarcopenia that involves iron accumulation factors in clinical practice. The iron chelators currently used in clinical practice mainly include deferoxamine, deferiprone and deferasirox; deferoxamine is poorly absorbed orally and has a short half-life. It requires slow subcutaneous or intravenous infusion and continuous administration, and the long-term treatment compliance of patients is poor; although deferiprone can be taken orally, its clinical use is limited due to serious side effects such as granulocytopenia and agranulocytosis ; As a new type of oral iron chelator, deferasirox has been proved to be less toxic, safe for clinical use, and patient compliance is good, so it is suitable for development as a preventive drug for sarcopenia. The present invention has proved that deferasirox can significantly inhibit skeletal muscle atrophy and oxidative stress damage in the pathological state of sarcopenia.

Description of drawings

Figure 1 shows the effect of deferasirox (DFS) on the proliferation and apoptosis of myoblast C2C12 in a cell culture environment simulating skeletal muscle nutritional deficiency in vivo. In the figure, A is the effect of DFS on the proliferation of myoblast C2C12 under the condition of serum-free culture; B is the effect of DFS on the apoptosis of myoblast C2C12 under the condition of serum-free culture. Compared between groups, * P ﹤0.05.

Figure 2 shows the effect of deferasirox (DFS) on the number of myotubes of skeletal muscle cells under the simulated in vivo skeletal muscle nutrition deficiency culture environment. In the figure, A is the cell morphology of the control group (×10); B is the serum-free medium group Cell morphology (×10); C is the cell morphology of serum-free medium plus DFS co-incubation group (×10); D is the software counting result. Compared between groups, * P ﹤0.05.

Figure 3 is the effect of deferasirox (DFS) on the content of malondialdehyde (MDA) and glutathione peroxidase (GSH-Px) in skeletal muscle cells under the simulated in vivo skeletal muscle oxidative damage environment. Figure A is the effect of DFS on MDA in skeletal muscle cells under the condition of tumor necrosis factor-α (TNF-α) intervention; B is the effect of DFS on GSH-Px of skeletal muscle cells under the condition of TNF-α intervention. Compared between groups, * P ﹤0.05.

Figure 4 shows the effect of deferasirox (DFS) on the expression of skeletal muscle cell myogenic genes myosin heavy chain (MyHC) and myogenin (MyOG) mRNA under the simulated in vivo skeletal muscle oxidative damage environment. Figure A is the effect of DFS on the expression of MyHCmRNA in skeletal muscle cells under the condition of TNF-α intervention; B is the effect of DFS on the expression of MyOG mRNA in skeletal muscle cells under the condition of TNF-α intervention. Compared between groups, * P ﹤0.05.

Figure 5 shows the effect of deferasirox (DFS) on the number of myotubes in skeletal muscle cells under the simulated in vivo skeletal muscle oxidative damage environment. In the figure, A is the cell morphology of the control group (×10); B is the cell morphology of the TNF-α group (×10); C is the cell morphology of the TNF-α plus DFS co-incubation group (×10); D is the software counting result. Compared between groups, * P ﹤0.05.

The "+" and "-" signs in the above figure indicate whether the substance is added to the control group or the experimental group.

Detailed ways

The present invention will be further described below in conjunction with the accompanying drawings.

C2C12 cells are a myoblast cell line derived from the immortalization of mouse skeletal muscle satellite cells, and are generally used in the research of skeletal muscle; the present invention uses C2C12 cells as the research object to establish skeletal muscle nutrition in a simulated state of sarcopenia in vivo Two cell culture models of deficiency and oxidative damage were then validated by the following experiments.

Example 1:

Establishment of a cell culture model simulating skeletal muscle trophic deficiency in vivo. The well-cultured C2C12 cells (Shanghai Cell Bank, Chinese Academy of Sciences) were divided into 3 groups, the control group (the culture medium was DMEM (high glucose), containing 10% fetal bovine serum, penicillin 100 μg/mL and streptomycin 100 μg/mL ), serum-free medium group (the medium is DMEM (high glucose), containing penicillin 100 μg/mL and streptomycin 100 μg/mL) and serum-free medium plus deferasirox (DFS) group (the medium is DMEM ( High glucose), containing penicillin 100 μg/mL, streptomycin 100 μg/mL, DFS 10 μmol/L), cultured in 5% CO ₂ , 37°C incubator respectively, after 48 hours of intervention, CCK-8 detection reagent (Dojindo Company, Japan) was used to detect the growth activity of cells, and the apoptosis kit (Invitrogen Company, USA) was used to detect cell apoptosis.

Figure 1 shows the effect of deferasirox on the proliferation and apoptosis of C2C12 cells in this culture environment. It was found that deferasirox can increase the viability of C2C12 cells under serum-free culture conditions and reduce the viability of C2C12 cells under serum-free culture conditions. of apoptosis.

Example 2:

Establishment of a cell culture model simulating skeletal muscle trophic deficiency in vivo. The well-cultured C2C12 cells were differentiated into mature skeletal muscle cells under the induction of horse serum, and the cells were divided into three groups, the control group (the medium was DMEM (high glucose), containing 10% fetal bovine serum, penicillin 100 μ/mL and Streptomycin 100 μg/mL), serum-free medium group (the medium is DMEM (high glucose), containing penicillin 100 μg/mL and streptomycin 100 μg/mL) and serum-free medium plus deferasirox (DFS ) group (the culture medium is DMEM (high glucose), containing 100 μg/mL penicillin, 100 μg/mL streptomycin, 10 μmol/L DFS), cultured in 5% CO ₂ , 37°C incubator, intervened 72 After one hour, 10 fields of view under the microscope were randomly observed and the number of skeletal muscle myotubes was counted using Image-Pro Plus 6.0 software.

Figure 2 shows the effect of deferasirox on the number of myotubes of skeletal muscle cells in this culture environment. In the figure, A is the cell morphology of the control group (×10); B is the cell morphology of the serum-free medium group (×10); C Cell morphology in serum-free medium plus DFS co-incubation group (×10); D is software counting results. It was found that deferasirox increased the number of myotubes formed by skeletal muscle cells in serum-free culture conditions.

Example 3:

Establishment of a cell culture model that mimics oxidative damage to skeletal muscle in vivo. The well-cultured C2C12 cells were differentiated into mature skeletal muscle cells under the induction of horse serum, and the cells were divided into three groups, the control group (the medium was DMEM (high glucose), containing 10% fetal bovine serum, penicillin 100 μ/mL and streptomycin 100 μg/mL), tumor necrosis factor-α (TNF-α) group (the culture medium is DMEM (high glucose) containing 10% fetal bovine serum, penicillin 100 μg/mL, streptomycin 100 μg/ mL, TNF-α 10 ng/mL) and tumor necrosis factor-α (TNF-α) plus deferasirox (DFS) co-incubation group (the medium is DMEM (high glucose), containing 10% fetal bovine serum, penicillin 100 μ/mL, streptomycin 100 μg/mL, TNF-α 10 ng/mL, DFS 10 μmol/L), after 72 h of intervention, the malondialdehyde (MDA) kit (Shanghai Beyond Company) detected intracellular MDA level; glutathione peroxidase (GSH-Px) kit (Beijing Suolaibao Company) was used to detect the level of GSH-Px in cell culture supernatant.

Figure 3 shows the effect of deferasirox on the content of MDA and GSH-Px in skeletal muscle cells in this environment. In the figure A is the effect of deferasirox on MDA in skeletal muscle cells (simulating skeletal muscle oxidative damage model) under TNF-α intervention culture conditions; B is the effect of deferasirox on skeletal muscle cells under TNF-α intervention culture conditions Effects of GSH-Px in cells (simulating skeletal muscle oxidative damage model). The results show that deferasirox can reduce the level of MDA in skeletal muscle cells cultured with TNF-α intervention, and increase the level of GSH-Px in skeletal muscle cells cultured with TNF-α intervention, which proves that deferasirox can reduce oxidative stress Oxidative damage to skeletal muscle cells in the environment.

Example 4:

Establishment of a cell culture model that mimics oxidative damage to skeletal muscle in vivo. C2C12 cells were differentiated into mature skeletal muscle cells under the induction of horse serum, and the cells were divided into 3 groups: the control group (the medium was DMEM (high glucose), containing 10% fetal bovine serum, penicillin 100 μ/mL and streptomycin 100 μg/mL), tumor necrosis factor-α (TNF-α) group (the culture medium is DMEM (high glucose), containing 10% fetal bovine serum, penicillin 100 μg/mL, streptomycin 100 μg/mL, TNF -α 10 ng/mL) and tumor necrosis factor-α (TNF-α) plus deferasirox (DFS) co-incubation group (the culture medium is DMEM (high glucose), containing 10% fetal bovine serum, penicillin 100μ/mL , streptomycin 100 μg/mL, TNF-α 10 ng/mL, DFS 10 μmol/L), cultured in 5% CO ₂ , 37°C incubator, after 72 hours of intervention, real-time quantitative PCR method was used to detect the myogenic genes of the cells Expression levels of myosin heavy chain (MyHC) mRNA and myogenin (MyOG) mRNA.

Figure 4 shows the effect of deferasirox on the expression of MyHC and MyOG mRNA in skeletal muscle cells in this culture environment. Figure A is the effect of deferasirox on the expression of MyHC mRNA in skeletal muscle cells (simulating skeletal muscle oxidative damage model) under TNF-α intervention culture conditions; B is the effect of deferasirox on skeletal muscle cells under TNF-α intervention culture conditions Effects on MyOG mRNA expression in cells (simulated skeletal muscle oxidative damage model). The results showed that deferasirox could reverse the decline in the expression of myogenic genes MyHC and MyOG mRNA in C2C12 cells under the condition of TNF-α intervention culture, which proved that deferasirox can improve the expression of skeletal muscle under TNF-α intervention culture. Intracellular myogenic genes MyHC and MyOG mRNA expression levels.

Example 5:

Establishment of a cell culture model that mimics oxidative damage to skeletal muscle in vivo. C2C12 cells were differentiated into mature skeletal muscle cells under the induction of horse serum, and the cells were divided into 3 groups: the control group (the medium was DMEM (high glucose), containing 10% fetal bovine serum, penicillin 100 μ/mL and streptomycin 100 μg/mL), tumor necrosis factor-α (TNF-α) group (the culture medium is DMEM (high glucose), containing 10% fetal bovine serum, penicillin 100 μg/mL, streptomycin 100 μg/mL, TNF -α 10 ng/mL) and tumor necrosis factor-α (TNF-α) plus deferasirox (DFS) co-incubation group (the culture medium is DMEM (high glucose), containing 10% fetal bovine serum, penicillin 100μ/mL , Streptomycin 100 μg/mL, TNF-α 10 ng/mL, DFS 10 μmol/L), cultured in 5% CO ₂ , 37°C incubator, after 72 hours of intervention, randomly observe 10 visual fields under the microscope and use Image-Pro Plus 6.0 software counted the number of skeletal muscle myotubes.

Figure 5 shows the effect of deferasirox on the number of myotubes of skeletal muscle cells in this culture environment. Among them, A is the cell morphology of the control group (×10); B is the cell morphology of the TNF-α group (×10); C is the cell morphology of the TNF-α plus DFS co-incubation group (×10); D is the software counting result. The results showed that deferasirox could reverse the reduction of myotube formation in C2C12 cells under the condition of TNF-α intervention, which proved that deferasirox could increase the number of myotube formation of skeletal muscle cells under TNF-α intervention culture.

The above examples are only to illustrate the basic ideas and technical principles of the present invention, and its purpose is to allow those familiar with this technology to understand the content of the present invention and implement it accordingly, and cannot limit the protection scope of the present invention. All equivalent changes or modifications made according to the spirit of the present invention shall fall within the protection scope of the present invention.

Claims (8)

1. The use of deferasirox in the preparation of medicines for the treatment of sarcopenia. 2. The use according to claim 1, characterized in that the sarcopenic disease is a sarcopenic disease accompanied by iron accumulation. 3. The use according to claim 1, characterized in that the sarcopenic disease is skeletal muscle atrophy and oxidative stress damage under the pathological state of sarcopenia. 4. The use according to claim 1, characterized in that the dosage of deferasirox is 10-20 mg/kg per day. 5. The use according to claim 1, characterized in that said deferasirox is administered alone or in any convenient pharmaceutical dosage form. 6. The use according to claim 5, wherein the pharmaceutical dosage form is a tablet comprising the compound. 7. The use according to claim 5, characterized in that, the administration method is oral administration. 8. A medicament for treating sarcopenia, characterized in that the medicament contains an active ingredient of deferasirox.