WO2013113141A1 - New applications of β3 adrenergic receptor agonist - Google Patents

Abstract

Disclosed are new applications of the β3 adrenergic receptor agonist. The β3 adrenergic receptor agonist is an important factor for regulating and controlling SIRT1, mTOR, and p53. The β3 adrenergic receptor agonist can be used for preparing medicine for treating diseases related to signaling pathways such as SIRT1, mTOR, and p53, for example, treating the diseases such as malignancy, hypertrophic cardiomyopathy, pulmonary hypertension, cirrhosis, rheumatoid arthritis, renal failure, and Alzheimer's disease.

Description

 New use of β 3 adrenergic receptor agonists

 Technical field

 The present invention relates to the field of signaling pathways, and in particular to the novel use of β 3 adrenergic receptor agonists in the preparation of a medicament for treating new diseases.

 Background technique

 β 3 in the human body Adrenergic receptors are mainly expressed in fat cells, especially visceral fat cells, which promote fat breakdown and energy supply. A variety of agonists and blockers have been discovered that selectively act on the receptor. For β 3 The drug research of the receptor is mainly based on its lipolysis, and the β 3 receptor agonist is used as a weight loss drug. However, for β 3 There are no reports on the role of receptors in the development and progression of aging-related diseases and their potential therapeutic effects.

 Tumor suppressor gene TP53 may be a key gene regulating aging and cell cycle rearrangement, TP53 It is a aging-related gene, and its anti-cancer effect is closely related to aging. It controls cell senescence by regulating DNA damage, free radical generation and clearance, and p53 overactivity causes premature senescence of stem cells, p53 The modification itself is the main reason for maintaining its function. Numerous studies have shown that Sirtuin 1 (SIRT1) can be deacetylated by p53 in p53 A key role in mediated aging and anti-tumor responses. Yeast Information Regulator 2 (SIR2) Is a histone deacetylase that is dependent on nicotinamide adenine dinucleotide, SIRT1 is the most homologous family member of mammalian Sir2, SIRT1 Proteins are present in mammalian cytoplasm and nucleus and play important regulatory roles in cell survival, aging and oxidative stress. SIRT1 deacetylates p53 and inhibits its transcriptional activity. In addition SIRT1 Can be recruited to the p53 target promoter, resulting in transcriptional silencing of the hypoacetylated nucleosome.

Summary of the invention

The present invention discloses that the adrenergic beta-3-receptor (ADRB3) regulates SIRT1, p53, mTOR (mammalian target of rapamycin, mammalian target of rapamycin), MIF and microRNA-16 signaling pathways. An important receptor. The present invention discloses the use of a β 3 adrenergic receptor agonist for the treatment of diseases associated with the mTOR /SIRT1/p53 signaling pathway, such as cardiac hypertrophy, hypertrophic cardiomyopathy, myocardial fibrosis, pulmonary hypertension, cirrhosis, kidney Dysfunction, Alzheimer's disease, and malignant tumors. The present invention discloses the use of a β 3 adrenergic receptor agonist for the preparation of an anti-aging drug. The present invention discloses the use of a β 3 adrenergic receptor agonist for preventing stem cell senescence. The invention also discloses the use of a β 3 adrenergic receptor agonist as a medicament for increasing the efficiency of induced pluripotent stem cells (iPS ) . The present inventors have found that β 3 receptor blockers can reduce SIRT1 expression in tumors and peripheral adipose tissue, leading to p53 acetylation, increasing p53 transcriptional activity, increasing p53 in tumor tissues, promoting tumor cell senescence, apoptosis, and blocking Transfer from a distance. 3 3 receptor agonists can increase SIRT1 expression, reduce p53, activate mTOR, down-regulate miR-16-1 and miR-15a expression, and play an anti-aging role. The inventors have also discovered that agonizing the β 3 receptor can reduce mTOR protein expression, particularly the mTOR-Rictor complex. The present inventors have also found that agonizing the β 3 receptor can reduce mitochondrial autophagy, and the β 3 receptor agonist can be used as an autophagy antagonist.

 The β 3 receptor agonist which can be used in the present invention is not particularly limited, and all compounds capable of exciting the β 3 receptor are suitable for use in the present invention. Representative Examples of β 3 receptor agonists include, but are not limited to, BRL 37344, SR58611A, TAK2677, N25984, and the like.

 β 3 of the present invention The dosage form and preparation method of the receptor agonist are not particularly limited, and various preparation forms such as tablets, capsules, granules, sustained release agents, and injections can be prepared by a method conventionally used in the art.

 The β 3 receptor agonist of the present invention can be used for treating cardiac hypertrophy, myocardial fibrosis, pulmonary hypertension, cirrhosis, Alzheimer's disease and malignant tumor Drug. The β 3 receptor agonist can also be used as an agonist of SIRT1 for anti-aging.

 The β 3 receptor agonist of the present invention can increase induced pluripotent stem cells (Induced pluripotent stem) Cells , iPS ) production efficiency, slowing down aging and aging of stem cells, delaying aging and aging of adipose stem cells and cardiac stem cells.

In addition, it should be understood that various modifications and changes may be made to the present invention, and the equivalents of the scope of the invention.

 DRAWINGS

 Figure 1 shows that SR59230A increases the expression of p53 in MCF-7 cells;

 Figure 2 shows that SR59230A reduces SIRT1 expression in MCF-7 cells;

 Figure 3 shows BRL37344 reduces myocardial apoptosis in rats with heart failure;

 Figure 4 shows that BRL37344 reduces myocardial p53 and increases SIRT1;

 Figure 5 shows BRL37344 improving heart function in rats with heart failure;

 Figure 6 shows β-galactosidase staining in adipose stem cells;

 Figure 7 shows Rictor quantitative PCR ;

 Figure 8 shows mTOR quantitative PCR;

 Figure 9 shows SIRT1, 3, 4 quantitative PCR.

 detailed description

 The invention is further explained by the following examples, but the examples are not intended to limit the invention in any way.

 Example 1

 Blocking the β 3 receptor reduces SIRT1 in MCF-7 cells and increases p53 expression. Different concentrations of SR59230A MCF-7 cells were treated and cells were lysed 24 h later to extract total protein. The protein concentration was determined by BCA method, 10 ug protein was taken, and 10% SDS-PAGE was separated to transfer the protein to PVDF. On the membrane, TBST containing 4% skim milk (10 mmol/L Tris HC1, pH 7.5, 150 mmol/L NaC1, 0.1 % Tween) 20) Blocking membrane 1 h , p53 antibody ( 1 : 1000 ), SIRT1 antibody ( 1 : 1000 ), incubate at 4 °C overnight, wash the membrane and incubate with secondary antibody for 1 h , ECL color development, the experiment was repeated 3 times. The Fluorchem 8900 software analyzes the gray value of the protein band and calculates the ratio of the target band to the internal reference band (GAPDH). Results are shown in Figure 1. 2, as the concentration of SR59230A increases, p53 expression increases, while SIRT1 gradually decreases.

Example 2

 In a rat model of myocardial infarction, excitatory beta 3 receptors increase myocardial cell SIRT1 and reduce p53 Expression, improve heart function and exert anti-apoptotic effects. The left anterior descending coronary artery was ligated in rats, resulting in a myocardial infarction model. Intraperitoneal injection of BRL37344 1mg/kg/day, heart 4 weeks later Super. Heart tissue was taken for apoptosis (tinel method) and SIRT1, p53 expression (Western blot). Take the heart and measure the ratio of left ventricular weight to total heart weight (LVW/BW ). The results are shown in Figures 3 to 5 . BRL37344 reduces myocardial p53 and increases SIRT1 , reduces cardiomyocyte apoptosis and reduces heart failure in rats LVW/BW , reverse cardiac hypertrophy and improve heart function.

 Example 3

Blocking the β 3 receptor increases β-galactosidase in adipose stem cells and promotes stem cell aging. Rat adipose-derived stem cells were treated with BRL37344 and SR59230A for 10 ^-7 M each. After 24 h, paraformaldehyde was fixed and stained and photographed according to the β-galactosidase staining kit. The results are shown in Figure 6. Compared to SR59230A, BRL37344 reduced β-galactosidase in adipose-derived stem cells, suggesting that the β 3 adrenergic receptors that agonize adipose-derived stem cells can play an anti-aging role.

 Example 4

Exciting beta 3 receptors reduces myocardial cell mTOR-Rictor complexes. Rat cardiomyocytes were treated with BRL37344 and SR59230A for 10 ^-7 M each. After 8 h, total RNA was extracted and reverse transcribed. Quantitative PCR was performed to detect mTOR and Rictor gene expression. The results are shown in Figures 7 and 8: BRL37344 reduced the mTOR-Rictor complex compared to the control group.

 Example 5

 Excited beta 3 receptor increases SIRT1, SIRT3 and SIRT4 in the heart, liver, brain, kidney and aorta of aging rats Expression. Twenty male Sprague-Dawley rats, 12 months old, were randomly divided into 2 groups, BRL37344 100nmol/only, intraperitoneal injection, qd, continuous use 2 Week. The control group was given the same volume of solvent. After the rats were sacrificed, the heart, liver, brain, kidney and aorta were taken, total RNA was extracted, and after reverse transcription, quantitative PCR was performed to detect SIRT1 and SIRT3. SIRT4 gene expression. The results are shown in Figure 9: BRL37344 increases SIRT1, SIRT3 and SIRT4 in the heart, liver, brain, kidney and aorta compared to the control group. Expression.

 Example 6

Excitatory β 3 receptor increases mitochondrial content in rat cardiomyocytes. Rat rat neonatal cardiomyocytes were treated with BRL37344 10 ^-7 M, MitoTracker mitochondrial probe was used to detect mitochondria number, JC1 mitochondrial membrane potential, fluorescent probe DCFH-DA was used to detect intracellular reactive oxygen species (ROS) content, ATP The intracellular ATP level was detected by the kit, and the MDC staining was detected by flow cytometry. The rat cardiomyocytes were transfected into GFP-LC3 eukaryotic expression vector. It was found that BRL37344 increased intracellular mitochondrial content, maintained cell mitochondrial membrane potential, decreased intracellular ROS content, and increased intracellular ATP levels. SR59230A increases MDC staining, reduces mitochondria, damages mitochondrial membrane potential, and promotes the formation of multiple bright green fluorescent spots.

 Summary: The present invention discloses a signaling pathway for the β 3 adrenergic receptor, ie, the β 3 receptor regulates mTOR , SIRT1 , The expression and activity of p53 and other proteins are important for the occurrence and development of tumors, metabolic diseases, immune system diseases and aging-related diseases. β 3 receptor can be used as a therapeutic target by regulating β 3 Receptors, treating the above diseases.

Claims (10)

1. Preparation of SIRT1, SIRT3 and SIRT4 by β 3 adrenergic receptor agonists Application in agonists.
2. The use of β 3 adrenergic receptor agonists in the preparation of mTOR inhibitors.
3. The use of β 3 adrenergic receptor agonists in the preparation of p53 inhibitors.
4. 3 3 adrenergic receptor agonists in the preparation of therapeutic SIRT1 , mTOR and p53 Applications in drugs for signaling pathway-related diseases.
5. β 3 The use of adrenergic receptor agonists for the preparation of a medicament for the treatment of cardiac hypertrophy, myocardial fibrosis, pulmonary hypertension, cirrhosis, renal failure, Alzheimer's disease and malignancies.
6. The use of β 3 adrenergic receptor agonists in the preparation of anti-atherosclerosis, anti-coronary stent restenosis or coronary drug stent coating drugs.
7. The use of β 3 adrenergic receptor agonists in the preparation of cardiac arrest protection fluids.
8. The use of β 3 adrenergic receptor agonists in the preparation of anti-aging drugs.
9. The use of a β 3 adrenergic receptor agonist for the preparation of a medicament for increasing the efficiency of induced pluripotent stem cells.
10. The use of β 3 adrenergic receptor agonists in the preparation of anti-stem cell aging drugs.

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