KR20120053395A - Pharmaceutical composition for treating asthma comprising 4-phenylbutyric acid or pharmaceutically acceptable salts thereof - Google Patents

Abstract

PURPOSE: A pharmaceutical composition containing phenyl butyric acid is provided to reduce inflammatory cell and cytokine expression, airway inflammation, and airway hyperresponsiveness. CONSTITUTION: A pharmaceutical composition for preventing or treating bronchial inflammation and asthma contains 4-phenylbutyric acid of chemical formula 1 or pharmaceutically acceptable salt thereof as an active ingredient. The asthma is acute asthma caused by bacterial infection. The pharmaceutical composition additionally contains pharmaceutically acceptable carrier, excipient, or diluents. A health functional food contains the compound of chemical formula 1 as an active ingredient. The health functional food is manufactured in the form of a tablet, capsule, pill, or liquid.

Description

Pharmaceutical composition for treating asthma comprising phenyl butyric acid (４-ｐｈｅｎｙｌ thereofｕｔｙｒｉｃａｃｉｄ) or a pharmaceutically acceptable salt thereof as an active ingredient.

The present invention relates to a pharmaceutical composition for preventing or treating bronchial inflammation and asthma, comprising phenylbutyl acid (4-phenylbutyric acid) or a pharmaceutically acceptable salt thereof as an active ingredient. More specifically, the present invention provides phenylbutyric acid or a pharmaceutically acceptable salt thereof having an activity of reducing the number of inflammatory cells in lung tissue, inhibiting expression of inflammatory cytokines, airway inflammation, and airway hyperresponsiveness as an active ingredient. It relates to a pharmaceutical composition for the prevention or treatment of bronchial inflammation and asthma comprising.

Asthma is a disease characterized by airway hypersensitivity to various stimuli. Clinical symptoms such as wheezing, dyspnea, and cough caused by extensive airway narrowing are reversibly improved naturally or by treatment. Can be. Most asthma are allergic, characterized by chronic airway inflammation and bronchial hyperresponsiveness (Minoguchi K and Adachi M. Pathophysiology of asthma.In: Cherniack NS, Altose MD, Homma I, editors Rehabilitation of the patient with respiratory disease.New York: McGraw-Hill, 1999, pp97-104).

Allergic asthma patients account for about 10% of the world's population, and 17 million people in the United States alone suffer from asthma. In 2000, the US market for allergic asthma drugs was about $ 6.4 billion, and the Korean market was reported to be about $ 1 billion, accounting for 20% of the total Korean drug market.

Asthma can be divided into exogenous and endogenous asthma depending on the cause. In exogenous asthma, it is asthma that causes symptoms when exposed to the causative antigen. The most causative antigens are house dust and mites. Pollen, animal epithelium, and fungi act as causative antigens. In the case of endogenous asthma, upper respiratory tract infections, exercise, emotional instability, cold climate and humidity changes cause or worsen asthma, which is common in adult-type asthma. Other medications include asthma, exercise-induced asthma and occupational asthma.

In terms of pathophysiology, asthma is recognized as an inflammatory disease caused by the proliferation, differentiation, and activation of inflammatory cells by cytokines produced by Th2 immune cells, which migrate and invade airways and surrounding airways (Elias JA, Lee CG, Zheng T, Ma B, Homer RJ, Zhu Z. New insights into the pathogenesis of Asthma., J. Clin. Invest., 111, pp 291-297, 2003). In this case, inflammatory cells such as activated eosinophils, mast cells, and alveolar macrophages secrete various inflammatory mediators to induce bronchial contraction and increase eosinophils. It is an important factor exacerbating asthma (Barnes PJ, Chung KF, Page CP. Inflammatory mediators of asthma: An update., Pharmacol Rev., 50, pp 515-596, 1998).

Therefore, for the treatment of asthma, airway inflammation is inhibited by inhibiting the production of Th2 cytokines such as interleukin-4, interleukin-5, and interleukin-13, which are involved in the production and activation of inflammatory cells (lymphocytes, eosinophils, neutrophils) in lung tissue. It is necessary to induce reaction inhibition. In particular, in the case of acute asthma exacerbation by bacterial infection, it is important to reduce airway hyperresponsiveness with the airway inflammatory response involved in typical pathophysiology. However, many researches on the treatment of asthma, such as developing drugs that can treat asthma by inhibiting the production of inflammatory mediators, but the treatment of acute asthma exacerbation and severe asthma is still very short.

On the other hand, phenyl butyric acid (hereinafter referred to as PBA) is a chemical chaperone (chaperone) is known to inhibit the ER stress, and recently, one of the genetic liver disease alpha 1-antitrypsin deficiency (alpha1- In animal models of antitrypsin deficiency, alpha 1-antitrypsin levels have been reported to increase with PBA treatment.

However, there has been no teaching or disclosure of phenylbutyl acid to exhibit anti-inflammatory and anti-asthmatic activity.

Accordingly, the present inventors have made diligent efforts to develop effective therapeutic agents for suppressing airway inflammatory response and airway hypersensitivity, which play an important role in typical pathophysiology in asthma. As a result, phenylbutyl acid reduces the number of inflammatory cells in lung tissue and expresses inflammatory cytokines. Since it has activities such as inhibition, reduction of airway inflammation and airway hypersensitivity, it has been confirmed that it can be used as a useful asthma treatment, and completed the present invention.

It is an object of the present invention to provide a pharmaceutical composition for the prevention or treatment of bronchial inflammation and asthma, comprising phenylbutyl acid (4-phenylbutyric acid) or a pharmaceutically acceptable salt thereof as an active ingredient.

Another object of the present invention is to provide a health functional food containing petyl butyric acid as an active ingredient.

The present invention to achieve the above object, the present invention comprises a phenyl butyric acid (4-phenylbutyric acid) represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient, prevention of bronchial inflammation and asthma Or a pharmaceutical composition for treatment.

In the present invention, pharmaceutically acceptable salts of phenylbutyl acid represented by Formula 1 include salts of acidic or basic groups which may be present in the compound of Formula 1, unless otherwise indicated. For example, pharmaceutically acceptable salts include sodium, calcium and potassium salts of the hydroxy group, and other pharmaceutically acceptable salts of the amino group include hydrobromide, sulfate, hydrogen sulphate, phosphate, hydrogen phosphate, dihydrogen Phosphate, acetate, succinate, citrate, tartrate, lactate, mandelate, methanesulfonate (mesylate), and p-toluenesulfonate (tosylate) salts, and methods or preparations for preparing salts known in the art It can be prepared through the process.

As used herein, the term "bronchial inflammation" is one of the defense responses of bronchial tissue to a certain stimulus, and complex lesions involving three types of tissue degeneration, circulatory disorder and exudation, and tissue proliferation. Say. The causes may include mechanical injury, physical factors such as temperature and radiation, chemical factors such as poisons, and parasites such as bacterial infection. In the present invention, the cause of bronchial inflammation may be infection by bacteria.

In the present invention, the asthma is distinguished from chronic bronchitis, which is a chronic obstructive obstructive pulmonary disease, and chronic obstructive pulmonary disease (COPD). Significant airway obstruction is present and many of these irreversible factors are irreversible, while asthma shows airway hyperresponsiveness and inflammatory response to various stimuli, causing airway obstruction but not classified as chronic obstructive pulmonary disease And, even if airway obstruction appears, there is a reversible characteristic that is lost by treatment or naturally.

In the present invention, the asthma may be acute asthma. Acute asthma is the sudden onset of asthma. Acute asthma of the present invention may have a variety of causes, but preferably may be acute asthma caused by bacterial infection.

Phenylbutyric acid of the present invention has the activity of reducing the number of inflammatory cells in lung tissue, inhibiting the expression of inflammatory cytokines, reducing airway inflammation and airway hypersensitivity, and thus can be used as a useful asthma treatment.

In one embodiment of the present invention, a mouse model of acute exacerbation of asthma caused by bacterial infection via OVA inhalation according to a defined protocol following simultaneous sensitization of laboratory mice with lipopolysaaccharide (LPS) and egg yolk albumin (ovalbumin, OVA) After PBA treatment, the levels of inflammatory cells (lymphocytes, eosinophils, neutrophils) and lung tissues, inflammation, airway resistance, and Th2 cytokine levels in lung tissues after PBA treatment were measured. Investigating how the airway inflammatory response and airway hyperresponsiveness, which play an important role, changes with intraperitoneal administration of PBA, the intraperitoneal administration of PBA results in interleukin-4 (IL-4) and interleukin-5 (IL-5). ) And effectively inhibited increased expression of inflammatory cytokines such as interleukin-13 (IL-13) and reduced airway inflammation and airway hypersensitivity in bronchial tissues. It was confirmed that this is effective to excellent and improved (Figs. 15).

The term "airway hypersensitivity" used in the present invention means that the airway is normal and responds sensitively to weak stimuli that will not respond. Such hypersensitivity includes allergens caused by antigen-antibody responses to specific stimuli, as well as those present in anaphylaxis and specific constitutions not dependent on antigen-antibody responses.

Therefore, since the phenyl butyric acid of the present invention has anti-inflammatory and anti-asthmatic activity, a composition comprising phenyl butyric acid or a pharmaceutically acceptable salt thereof may be used as a pharmaceutical composition for the prevention or treatment of bronchial inflammation and asthma. Can be.

As used herein, the term "treatment of bronchial inflammation and asthma" refers to the improvement of bronchial inflammation and asthma symptoms by administering to the body a composition comprising the phenylbutyl acid of the present invention or a pharmaceutically acceptable salt thereof as an active ingredient. Means any action that could be or beneficially altered.

In addition, the pharmaceutical composition comprising phenylbutyl acid of Formula 1 according to the present invention may further comprise a suitable carrier, excipient or diluent according to a conventional method. Carriers, excipients and diluents that may be included in the compositions of the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, Methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.

The pharmaceutical compositions according to the invention can be used in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like oral formulations, external preparations, suppositories or sterile injectable solutions according to conventional methods. have. Specifically, when formulated, it may be formulated using diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrants, surfactants and the like that are commonly used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and such solid preparations include at least one excipient in the compound, for example, starch, calcium carbonate, sucrose. ), Lactose, gelatin and the like can be mixed. In addition to simple excipients, lubricants such as magnesium stearate and talc may also be used. Liquid preparations for oral use include suspensions, solvents, emulsions, and syrups.In addition to the commonly used simple diluents, water and liquid paraffin, various excipients such as wetting agents, sweeteners, fragrances, and preservatives may be included. have. Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations and suppositories. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate and the like can be used. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

The preferred dosage of the pharmaceutical composition according to the present invention depends on the condition and weight of the patient, the extent of the disease, the form of the drug, the route of administration and the duration, and may be appropriately selected by those skilled in the art. However, for the desired effect, the phenylbutyl acid of the present invention may be administered in an amount of 0.001 to 100 mg / kg divided once to several times daily. More preferably, the phenyl butyric acid may be administered at a rate of 80 mg / kg each time.

Pharmaceutical dosage forms of the compounds according to the invention can also be used in the form of their pharmaceutically acceptable salts, or can be used alone or in combination with other pharmaceutically active compounds as well as in a suitable collection.

The pharmaceutical composition of the present invention can be administered to mammals such as rats, mice, livestock, humans, and the like by various routes. All modes of administration can be expected, for example, by oral, intraperitoneal, rectal or intravenous, intramuscular, subcutaneous, intrauterine dural or intracerebroventricular injection. Preferably the administration of the pharmaceutical composition may be administered intraperitoneally.

As another aspect, the present invention provides a health functional food containing petylbutyl acid represented by the formula (1) as an active ingredient.

The phenylbutyl acid of the present invention can be added to food or beverages for the purpose of preventing anti-inflammatory, anti-allergic and asthmatic diseases. At this time, the amount of the compound in the food or beverage may be added at 0.01 to 15% by weight of the total food weight, the health beverage composition may be added in a ratio of 0.02 to 5 g, preferably 0.3 to 1g based on 100 ml. However, this can be easily determined by those skilled in the art to suit the product.

In addition to the compound, the health functional food may further include food supplements that are food acceptable, and may be prepared in the form of tablets, capsules, pills, liquids, and the like.

More specifically, the beverage composition of the health functional food of the present invention is not particularly limited to other ingredients except for containing the compound as an essential ingredient in the indicated ratio and additional ingredients such as various flavors or natural carbohydrates, such as ordinary drinks It may contain as. Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And conventional sugars such as polysaccharides such as dextrin, cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those described above, natural flavoring agents (tauumatin, stevia extract (e.g., Rebaudioside A, glycyrrhizin, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. The proportion of said natural carbohydrates is generally about 1-20 g, preferably about 5-12 g per 100 ml of the composition of the present invention.

In addition to the above, the health functional food of the present invention includes various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic and natural flavors, coloring and neutralizing agents (such as cheese and chocolate), pectic acid and salts thereof, alginic acid And salts thereof, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated drinks, and the like. In addition, the health functional food of the present invention may contain natural fruit juice and fruit flesh for producing fruit juice beverages and vegetable beverages. These components can be used independently or in combination. The proportion of such additives is not so critical but is usually selected in the range of 0 to about 20 parts by weight per 100 parts by weight of the compound of the present invention.

As described above, the pharmaceutical composition for treating asthma comprising phenyl butyric acid or a pharmaceutically acceptable salt thereof having anti-inflammatory and anti-asthmatic activity of the present invention as an active ingredient is an inflammation in lung tissue. It can be used as a therapeutic agent for inflammation and asthma because it has a decrease in cell number, expression of inflammatory cytokines, airway inflammation, and airway hypersensitivity.

Figure 1 shows acute asthma exacerbation caused by bacterial infection made by LPS treatment in the OVA-induced asthma mouse model, after PBA treatment, the total number of inflammatory cells and fractions of lymphocytes, neutrophils, eosinophils confirmed in bronchoalveolar lavage fluid. .
Figure 2 shows the results of the observation of the effect of PBA on the Peribronchial and perivascular inflammation score using optical microscopy in acute asthma exacerbation model caused by bacterial infection (A: control, B: acute asthma exacerbation group, C: PBA Treated group).
Figure 3 shows the effect of reducing airway hypersensitivity of PBA in acute asthma exacerbation model by bacterial infection.
Figure 4 shows the change by PBA administration of Th2 cytokine, which is important for acute asthma exacerbation by bacterial infection.
Figure 5 shows the results of measuring the expression in the lung tissue of the mRNA for the CHOP protein reflecting the ER stress using the RT-PCR method.

Hereinafter, the present invention will be described in more detail with reference to examples. However, these examples are for illustrative purposes only, and the scope of the present invention is not limited to these examples.

Materials and Experiments

Hereinafter, 8 week old female C57BL / 6 mice used in the Examples were purchased from Orientbio Inc. (Seongnam, Korea), and PBA was used by Calbiochem (San Diego, CA, USA). In addition, in the following experimental step, PBA was administered 80 mg per kg of mice twice intraperitoneally 1 hour before the first OVA inhalation and 6 hours after the last OVA inhalation.

Example  1: In a model of exacerbation of acute asthma caused by bacterial infection PBA  Total inflammatory cell count and lymphocytes after treatment, Neutral sphere Fractional effect of eosinophils

This example investigated the effect of PBA on the reduction of total inflammatory cell numbers and fractions of lymphocytes, neutrophils, and eosinophils in bronchial alveolar lavage fluid of acute asthma exacerbation model caused by bacterial infections treated with OVA and LPS.

After inhaling saline, the drug vehicle-treated group was (SAL + VEH), OVA and LPS-sensitized group, and the OVA-inhaled model was treated with drug vehicle (OVA + LPS + VEH), OVA and After sensitization with LPS, a group treated with PBA was inoculated (OVA + LPS + PBA) in a model in which OVA was inhaled, and bronchial alveolar lavage fluid was analyzed.

As a result, it was confirmed that the PBA-treated group had an effect of reducing the total number of inflammatory cells and fractions of lymphocytes, neutrophils, and eosinophils (see FIG. 1).

Example  2: In acute asthma exacerbation model due to bacterial infection Peribronchial and  perivascular inflammation score For PBA Effect

To confirm histologically the airway inflammatory response, the mice were euthanized 48 hours after the last OVA inhalation, fixed fluid was injected into the trachea and lungs, the lungs were separated from the mice, and fixed again with 10% neutral formalin. After dehydration of the tissue, a block was made using paraffin, and then cut using a micro cutter to a 4-μm thickness, and the paraffin was removed on a glass to prepare a sample slide, and H & E staining was performed. The results were obtained by observing the tissue at 20 magnification using an optical microscope.

As a result, the degree of airway inflammation was significantly reduced in the PBA-treated group (FIG. 2C) compared to the acute asthma worsening group (FIG. 2B) (see FIG. 2).

Example  3: Effect of PBA on Airway Hypersensitivity in Models of Acute Asthma Exacerbation by Bacterial Infection

Measurement of airway hyperresponsiveness was assessed by measuring changes in airway function after administration of methacholine in aerosol form via the airways. In other words, after anesthetizing the mouse with pentobarbital, and after tracheotomy, the animal ventilator was connected, and the ventilation was directly performed through the ventilator while mechanically breathing with a single breathing volume of 10 ml / kg, respiratory rate of 150 breaths / min, and aerobic pressure of 2 cmH ₂ O. Methacholine was administered with a nebulizer, gradually increasing the dose from 5.0 mg / ml to 50 mg / ml, and the airway resistance (Rrs) was continuously measured. Airway responsiveness was assessed as a percentage of baseline when the saline control group was administered the maximum of Rrs measured at each methacholine concentration.

As a result, as shown in FIG. 3, the airway resistance dose response curve of the acute asthma exacerbation model caused by bacterial infection treated with OVA and LPS is shifted to the left side compared to the saline-induced group, and the concentration of methacholine 50 At mg / ml, airway resistance (Rrs) was significantly increased compared to the control group, but in the PBA-administered group, the dose-response curve of airway resistance shifted to the right as compared to LPS alone and Rrs also significantly decreased. It was found that PBA has an effect of reducing airway hyperresponsiveness caused by OVA and LPS.

Example  4: In a model of exacerbation of acute asthma caused by bacterial infection PBA of On cytokines  For effect

Quantification of Th2 cytokines was performed by ELISA for Western blottin concentrations of the corresponding proteins in lung tissue and the concentrations of each cytokine in lung lavage fluid. After extracting the protein from the lung tissue of homogenized mice, the sample was completed to maintain a constant concentration of protein, and then loaded onto an SDS-PAGE gel. For IL-4, anti-IL-4 antibody from Serotec (UK) was used. The anti-IL-5 antibody of SantaCruz Biotechnology (USA) for 5 and the anti-IL-13 antibody of R & D Systems (USA) for IL-13 were quantified. ELISA kits include IL-4,5; Invitrogen, IL-13; Bender Medsystems product was used.

Figure 4 shows the change by PBA administration of Th2 cytokine, which plays an important role in the pathophysiology of acute asthma exacerbation by bacterial infection. It was confirmed that the cytokine increased by bacterial infection is significantly inhibited by PBA treatment in lung tissue.

As shown above, PBA has the effect of inhibiting the expression of inflammatory cells, an increase in inflammatory cells in bronchoalveolar lavage fluid, and an increase in airway resistance, which play an important role in acute asthma exacerbation by bacterial infections. It was confirmed that acute exacerbation due to infection can be treated.

Example  5: In acute asthma exacerbation model due to bacterial infection PBA by ER stress  Inhibitory effect

In order to examine the relationship between ER stress and asthma, the expression level of mRNA for the CHOP protein reflecting the ER stress in the asthma exacerbation model was measured by RT-PCR method. Was administered to investigate whether ER stress is suppressed.

As a result, CHOP mRNA was increased in the asthma exacerbation model as shown in FIG. 5, and this phenomenon was markedly decreased when PBA was administered. Although there is no known data on the role of ER stress in conventional bronchial asthma, the present invention showed that bronchial asthma and RR stress are related.

Claims (7)

A pharmaceutical composition for the prevention or treatment of bronchial inflammation and asthma comprising 4-phenylbutyric acid represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient:
[Formula 1]

The composition of claim 1, wherein the asthma is acute asthma. The composition of claim 2, wherein the asthma is acute asthma by a bacterial infection. The composition of claim 1, wherein the phenylbutyl acid has an activity of reducing the number of inflammatory cells in lung tissue, inhibiting expression of inflammatory cytokines, airway inflammation, and airway hyperresponsiveness. The composition of claim 1, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable carrier, excipient or diluent. Health functional food containing a petyl butyric acid represented by the formula (1) as an active ingredient:
[Formula 1]

The dietary supplement of claim 6 which is a tablet, capsule, pill or liquid.

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