KR102258912B1 - Agent for inhibiting weight loss and body temperature loss caused by influenza virus comprising alarin as main ingredient - Google Patents

Abstract

The present invention relates to a use of alarin, which has an amino acid sequence similar to galanin derived from a splice variant of RNA of galanin-like peptide (GALP), and more particularly, to a use of an agent using alarin as a main ingredient for suppressing acute inflammation and immunity loss caused by influenza virus, further improving immunity and treating inflammation, in which the above-mentioned therapeutic agent of the present invention for acute inflammation and immunity loss caused by influenza virus contains alarin as a main ingredient. As described above, an inhibitor of weight loss and body temperature drop caused by influenza virus, which has alarin as a main component, according to the present invention, contains alarin, a kind of peptide that is present in vivo and can be simply synthesized, as a main ingredient, and thus the ingredient can be easily obtained, thereby effectively inhibiting the proliferation and survival of the influenza virus at a specific concentration in a subject infected with the influenza virus, so as to provide therapeutic efficacy without causing a specific side effect in the human body. Accordingly, the inhibitor can be safely used with an excellent effect of providing an agent for inhibiting and treating the induced acute inflammation and immunosuppression caused by the influenza virus.

Description

Agent for inhibiting weight loss and body temperature loss caused by influenza virus comprising alarin as main ingredient

The present invention relates to an inhibitor of weight loss and body temperature drop caused by influenza virus containing allarin as a main component, and more particularly, it is derived from a splice variant of galanin-like peptide (GALP) RNA and has an amino acid sequence similar to galanin. A new use of Alarin having Alarin as a main component, it relates to a weight loss caused by influenza virus and its use as an inhibitor of body temperature drop.

Despite the continuous and remarkable development of medical technology, an effective treatment method has not yet been provided for a number of diseases. In particular, diseases caused by viral infections of many continuously mutating and evolving species always pose new problems. Among these viruses, influenza virus in most cases recovers after several days of infection, but it can cause fatal complications such as pneumonia in the elderly or chronically ill, so an effective treatment is required. Influenza viruses are classified into A, B, and C types, and the annual influenza A virus subtypes are determined by the surface antigens hemagglutinin (H1-H16) and neuraminidase (N1-N9).

Influenza viruses generally cause epidemics through antigenic mutations, and typical seasonal influenza is seasonally prevalent every year through antigenic mutations. H1N1, a new influenza virus that has been prevalent around the world in recent years, is a new influenza virus that has undergone antigenic mutation through genetic recombination. This influenza A (H1N1) virus spread from birds to humans and pigs and caused a pandemic, and the human influenza A (H1N1) virus disappeared and another type of human influenza A (H2N2) appeared, and again human influenza A (H1N1) appeared and became the main causative virus of seasonal influenza together with human influenza A (H3N2), which appeared later. Such influenza treatment or inhibitors include M2 inhibitors amantadine and rimantadine, and neuraminidase (NA) inhibitors zanamivir and oseltamivir. Since M2 inhibitors act on the M2 protein present only in influenza A virus, amantadine and rimantadine act specifically only in influenza A virus. On the other hand, NA inhibitors have an inhibitory effect on both type A and type B influenza viruses. However, recently, resistance to these influenza therapeutics has been reported, and as a swine influenza virus that is resistant to Tamiflu, which is the most widely used effectively for treating swine influenza A (H1N1) virus, has been discovered, the demand for new therapeutic agents continues. In addition, since the influenza virus is spread by droplet infection, the infection spreads rapidly and is easy to cause an influenza pandemic, so an effective treatment or inhibitor is continuously required.

Therefore, many studies have been conducted to recognize the above situation and to solve it, for example, Korean Patent Publication No. 2014-0104652 (Patent Document 1) in Korea Patent Publication No. 2014-0104652 (Patent Document 1), as an active ingredient, Under the name of a composition for anti-influenza comprising It can be usefully used as an influenza treatment because it has an excellent effect in reducing and inhibiting the proliferation of influenza viruses such as seasonal influenza virus (A/Wisconsin/15/09 H3N2) or seasonal influenza virus (B/Brisbane/60/2008)" In the Korean Patent Application Laid-Open No. 2015-0013527 (Patent Document 2), as the name of an enhanced anti-influenza agent conjugated with anti-inflammatory activity, "inhibition of influenza virus neuraminidase and inhibition of inflammation-inducing cytokines simultaneously caffeic acid (CA)-bearing zanamivir (ZA) conjugates ZA-7-CA (1), ZA-7-CA-amide (7) and ZA-7-Nap (43) for The bifunctional ZA conjugates act synergistically on the protection of mice lethally infected with H1N1 or H5N1 influenza viruses, and the efficacy of ZA-7-CA, ZA-7-CA-amide and ZA-7-Nap conjugates is similar to that of anti-inflammatory and ZA much greater than combination therapy.”

In addition, a number of pharmaceutical compositions or methods for inhibiting influenza have been disclosed in addition to the contents disclosed in the above patent documents, but the influenza virus is still mutated and a drug having an inhibitory efficacy of the influenza virus is sought in a situation where new species are appearing, and this The demand for practical application is still there.

On the other hand, Alarin derived from a splice variant of galanin-like peptide (GALP) RNA was biosynthesized in vivo and synthesized genetically, which is a peptide having the amino acid sequence of APAHRSSTFPKWVTKTERGRQPLRS. . Since allarin shares only 5 amino acids with GALP, it cannot activate GALP-specific receptors, and neuropeptides have been reported to be expressed in ganglion cells of human neuroblastic tumors and may be a potential tumor marker. For more detailed information on allarin, reference may be made to International Patent Publication No. WO/2006/094973 (Patent Document 3), the entire contents of which are incorporated herein by reference. However, in Patent Document 3, only allarin's use as a therapeutic agent for keratitis nervosa, retinol disease, acute and chronic inflammatory and autoimmune diseases, obesity or growth deficiency treatment, wound healing, uterine fibroids, endometriosis, and anorexia may be suggested. only Accordingly, the present applicant studied other efficacy of allarin having the above-mentioned properties. As a result, the applicant showed an excellent therapeutic effect for uremia, while anemia, renal osteodystrophy, epicarditis, alumina osteosis, amyloidosis, crystalline arthritis, polyps. It has been found that it can be used as an excellent treatment for uremia that can be used for a long time because it does not cause side effects such as cystic kidney and diarrhea, and it does not cause a decrease in quality in the social life of the patient and the occurrence of high treatment cost according to the treatment. It has been registered as 1827245 (Patent Document 4).

Therefore, the present inventors recognized the above-mentioned conventional needs in the art against the background of the prior art as described above, and as a result of intensive research to solve the above-mentioned prior art, allarin, a peptide that can be simply provided, has a specific dose When used as a medicine, it exerts an excellent effect in suppressing the acute inflammation and weakening of immunity caused by influenza virus, and does not cause any specific side effects to the human body, so it can be used safely, suppressing acute inflammation and lowering of immunity caused by influenza, and further improving immunity The present invention was completed by obtaining experimental results that it can be used as an agent to treat inflammation.

Patent Document 1: Korean Patent Publication No. 2014-0104652 Patent Document 2: Korean Patent Publication No. 2015-0013527 Patent Document 3: International Patent Publication No. WO/2006/094973 Patent Document 4: Korean Patent Registration No. 1827245

Accordingly, an object of the present invention is to meet the requirements in the prior art based on the various problems in the conventional influenza virus therapeutics or inhibitors as described above and the complex and diverse technical background in these technical fields. Its main purpose is to suppress acute inflammation and immune decline caused by influenza virus, which is satisfactory as a substance that can be simply provided, and further enhances immunity and provides the effect of treating inflammation, while providing effects to the human body. This is to provide a therapeutic agent for influenza virus that can be safely used without causing any specific side effects.

Another object of the present invention is to provide a method that can more easily provide an agent for suppressing acute inflammation and immunosuppression caused by influenza virus, and further improving immunity and treating inflammation, which has the above characteristics. .

The present invention may also have as its object to achieve these objects and other objects that can be easily derived by a person skilled in the art from the general description of the present specification in addition to the above clear objects.

As described above, the present inventors recognized the situation in which a substance having conventional influenza virus suppression and therapeutic efficacy is continuously required, and as a result of intensive research to respond to this, the neuropeptide Alla, which shares only 5 amino acids with GALP When Lin is used in a specific dose, it has an excellent effect in inhibiting and treating influenza virus and does not cause any specific side effects to the human body, so it can be safely used. The object of the present invention can be achieved by finding out that it is possible to suppress acute inflammation and decrease in immunity and further improve immunity and provide a formulation for treating inflammation.

In order to achieve the above object, an inhibitor of weight loss and body temperature drop caused by influenza virus of the present invention;

It is characterized in that it contains Alarin as a main component.

According to another configuration of the present invention, the allarin is characterized in that the human-derived allarine acetate salt having the following amino acid sequence:

H-Ala-Pro-Ala-His-Arg-Ser-Ser-Thr-Phe-Pro-Lys-Trp-Val-Thr-Lys-Thr-Glu-Arg-Gly-Arg-Gln-Pro-Leu-Arg- Ser-OH-acetate salt

According to another configuration of the present invention, the alarin has a molecular weight of 2894.29 and is characterized in that it has the following molecular formula:

C ₁₂₇ H ₂₀₆ N ₄₃ O ₃₈

According to another configuration of the present invention, the therapeutic agent for acute inflammation and immunosuppression caused by the influenza virus of the present invention is formulated to be administered in an adult 60 kg standard of 1.2 mg to 1.4 mg of Alarin acetate. characterized.

According to another configuration of the present invention, the formulation is characterized in that it is an injection.

According to another configuration of the present invention, the formulation is characterized in that it is injected by subcutaneous intramuscular injection.

The weight loss and body temperature lowering inhibitor caused by influenza virus containing allarin as a main component of the present invention, which is configured as described above, contains allarin, a type of peptide that exists in vivo and can be simply synthesized, as a main component. Body weight caused by influenza virus that can be obtained simply and can be safely used because it does not cause specific side effects in the human body while providing the efficacy of effectively inhibiting the proliferation and survival of influenza virus at a specific concentration in a subject infected with the influenza virus It has an excellent effect of reducing and providing an inhibitor of body temperature drop.

1 shows gene splice variants of Galanin-like peptides (GALPs) derived from Murine.
2 shows the results of comparing amino acid sequences obtained by obtaining allarin according to a preferred embodiment of the present invention from murine, rat, macaque and human.
3 to 6 show hematoxylin & eosin (H&E) staining in the G1, G2, G3 and G4 groups, respectively, performed according to an embodiment of the present invention, and an optical microscope (Olympus BX53, Japan) It is a micrograph of the histopathological examination result using

Hereinafter, preferred embodiments of the present invention will be described in more detail with reference to the accompanying drawings, but it goes without saying that the present invention is not limited to the following embodiments and can be implemented with various modifications within the scope of the gist. .

The following embodiments are provided to complete the disclosure of the present invention, and to fully inform those of ordinary skill in the art to which the present invention belongs, the scope of the invention, the scope of the present invention being defined only by the claims will only be Accordingly, in some embodiments, well-known components, well-known operations, and well-known techniques have not been described in detail in order to avoid obscuring interpretation of the present invention.

The terms used in this specification have been selected as currently widely used general terms as possible while considering the functions in the present invention, which may vary depending on the intention of those skilled in the art to which the present invention belongs or the advent of new technology . In addition, in a specific case, there is also a term arbitrarily selected by the applicant, and in this case, the meaning will be described in detail in the corresponding description of the present invention. It should be defined based on the meaning and content throughout this specification.

In this specification, the singular form also includes the plural form unless specifically stated in the phrase. In addition, components and operations referred to as'comprising (or having)' do not exclude the presence or addition of one or more other components and operations.

As used herein, the terms “having”, “having” or any other variation thereof are intended to cover non-exclusive inclusions. For example, a method, article, or device having a list of features is not necessarily limited to those features, but may have other features not explicitly listed or inherent in such a method, article, or device. Also, unless expressly stated to the contrary, "or" is intended to be in an inclusive sense and not an exclusive one.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

The inhibitor of weight loss and body temperature drop caused by influenza virus according to a preferred embodiment of the present invention comprises Alarin as a main component.

According to another preferred embodiment of the present invention, the allarin used as a therapeutic agent for acute inflammation and immunosuppression caused by influenza virus of the present invention may be human-derived allarin having the following amino acid sequence:

Ala-Pro-Ala-His-Arg-Ser-Ser-Thr-Phe-Pro-Lys-Trp-Val-Thr-Lys-Thr-Glu-Arg-Gly-Arg-Gln-Pro-Leu-Arg-Ser- OH

According to another preferred embodiment of the present invention, allarin used as an inhibitor of weight loss and body temperature drop caused by influenza virus of the present invention may be a pharmaceutically acceptable salt of human-derived allarin having the above amino acid sequence.

The pharmaceutically acceptable salt refers to a salt that retains the biological effects of the free acids and bases of a particular compound and is not biologically or otherwise harmful. The compounds of the present invention may have functional groups that are either sufficiently acidic, sufficiently basic, or both, and thus may be reacted with any number of inorganic or organic bases, and inorganic and organic acids, to form pharmaceutically acceptable salts. Exemplary pharmaceutically acceptable salts include those prepared by reaction of a compound of the present invention with an inorganic or organic acid or inorganic base, for example, salts comprising: acetate, trifluoroacetate, sulfate , pyrosulfate, bisulfate, sulfite, bisulfite, phosphate, monohydrogen phosphate, dihydrogen phosphate, metaphosphate, pyrophosphate, chloride, bromide, iodide, acetate, propionate, decanoate, Caprylate, acrylate, formate, isobutyrate, caproate, heptanoate, propiolate, oxalate, malonate, succinate, suberate, sebacate, fumarate, maleate, butine-1 ,4-dioleate, hexain-1,6-dioleate, benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, hydroxybenzoate, methoxybenzoate, phthalate, sulfonate, xyl Rensulfonate, phenylacetate, phenylpropionate, phenylbutyrate, citrate, lactate, γ-hydroxybutyrate, glycolate, tartrate, methane-sulfonate, propanesulfonate, naphthalene-1 -sulfonates, naphthalene-2-sulfonates and mandelates.

According to another preferred embodiment of the present invention, allarin used as an inhibitor of weight loss and body temperature decrease caused by influenza virus of the present invention may be a human-derived trifluoroacetate salt having the following amino acid sequence. :

H-Ala-Pro-Ala-His-Arg-Ser-Ser-Thr-Phe-Pro-Lys-Trp-Val-Thr-Lys-Thr-Glu-Arg-Gly-Arg-Gln-Pro-Leu-Arg- Ser-OH-acetate salt

According to another preferred embodiment of the present invention, said allarine has a molecular weight of 2894.29 and has the following molecular formula:

C ₁₂₇ H ₂₀₆ N ₄₃ O ₃₈

According to another preferred embodiment of the present invention, the alarin can be stored stably, preferably at -20 ± 5 °C.

According to another preferred embodiment of the present invention, the weight loss and body temperature lowering inhibitor caused by the influenza virus of the present invention is 60 kg body weight to provide an effect of suppressing and treating acute inflammation and immunosuppression caused by the influenza virus. 1 mg to 2 mg of alarin salt, preferably alarin acetate, may be administered based on adults of Preferably, the therapeutic agent for acute inflammation and reduced immunity caused by influenza virus of the present invention is based on an adult weighing 60 kg to provide an effect of suppressing and treating acute inflammation and reduced immunity caused by influenza virus. 1.2 mg to 1.6 mg of lean acetate may be administered. More preferably, the therapeutic agent for acute inflammation and reduced immunity caused by influenza virus of the present invention is based on an adult weighing 60 kg to provide an effect of suppressing and treating acute inflammation and reduced immunity caused by influenza virus. Alarin acetate 1.2 mg to 1.4 mg may be administered. Even more preferably, the therapeutic agent for acute inflammation and reduced immunity caused by influenza virus of the present invention is based on an adult weighing 60 kg to provide an effect of suppressing and treating acute inflammation and reduced immunity caused by influenza virus. 1.2mg to 1.3mg of alarin acetate can be administered. Most preferably, the therapeutic agent for acute inflammation and reduced immunity caused by influenza virus of the present invention is based on an adult weighing 60 kg to provide an effect of suppressing and treating acute inflammation and reduced immunity caused by influenza virus. Alarin acetate 1.2 mg may be administered.

According to another preferred embodiment of the present invention, the weight loss and body temperature lowering inhibitor caused by the influenza virus of the present invention is used to suppress and treat acute inflammation and immunosuppression caused by the influenza virus. Based on an adult weighing 60 kg, alarin 1.0 mg to 2.0 mg, preferably 1.1 mg to 1.8 mg, more preferably 1.2 mg to 1.6 mg, more preferably 1.2 mg to 1.4 mg, and most preferably may be formulated in a dosage form such that 1.2 mg is administered.

According to another preferred embodiment of the present invention, the formulation, although not particularly limited thereto, may preferably be an injection.

According to another preferred embodiment of the present invention, the formulation, although not particularly limited thereto, may preferably be administered by subcutaneous intramuscular injection.

According to another preferred embodiment of the present invention, when the formulation for suppressing weight loss and body temperature decrease caused by influenza virus of the present invention is an injection, a solvent, solubilizer or emulsifier is used as a carrier component, such as water , ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butyl glycol oil, glycerol aliphatic ester, polyethylene glycol or fatty acid ester of sorbitan may be used.

Hereinafter, the present invention will be described in more detail with reference to preferred embodiments, but it is needless to say that the present invention is not limited to these examples.

Example

Influenza A virus (H1N1, PR8) infected specific pathogen-free (SPF) C57BL/6 mice were evaluated for the effect of the test substance to inhibit and treat acute inflammation and immunosuppression caused by influenza virus. The specific pathogen-free (SPF) C57BL/6 mouse is an experimental animal widely used in the efficacy test of a therapeutic agent (ie, antiviral) against influenza virus, and a number of basic data for comparing and analyzing the results have been reported. Therefore, it was determined that it was suitable as an experimental animal and was selected. After receiving the experimental animal, it went through a 7-day quarantine and acclimatization period. During the acclimatization period, general symptoms were observed to confirm the health status, and a healthy animal was used for the test.

In this example, the inhibitory efficacy test is performed at a temperature of 23±3° C., a relative humidity of 55±15%, the number of ventilation 10 to 20 times/hr, a lighting time of 12 hours (lights at 8 am and lights out at 8 pm) and illuminance 150 to 300 Lux. It was conducted in a rodent breeding area set as In addition, environmental conditions such as temperature/humidity, ventilation frequency, and illuminance of the animal room were measured regularly during the breeding period.

Feed was supplied with feed for laboratory animals produced by Samtaco Co., Ltd. and freely consumed, and purified water was freely ingested using a polycarbonate drinking bottle. For the rug, beta chips were supplied from Orient Bio Co., Ltd. (located at 322, Galmachi-ro, Jungwon-gu, Seongnam-si, Gyeonggi-do) and used.

During the acclimatization and experiment period, no more than 5 animals were bred per breeding box in a polycarbonate breeding box for rodents (W 170 x L 235 x H 125 mm), and breeding boxes, rugs and water bottles were exchanged at least once a week.

During the acclimatization period, the weights of animals determined to be healthy were measured and randomly distributed according to the ranked weights so that the average weight of each group was distributed as evenly as possible.

As shown in Table 1 below, in this example, group G1 is a group that does not induce influenza, G2 is a control group that induces influenza and does not administer drugs, and G3 is 20 μg/kg of test substance administered to experimental animals. One group, and G4 is the group in which 40 μg/kg of the test substance was administered to the experimental animals. Also, G5 to G7 correspond to G2 to G4, respectively, provided that G2 to G4 used an challenge _{titer of 1 LD 50} , whereas G5 to G7 used an _{challenge titer of 10 LD 50} .

Here, the test substance is Alarin (human) acetate salt of the present invention.

group gender number of animals
(Marie) animal number Attack inoculation titer Whether to inoculate dosing substance Dosage
(μg/kg) dose amount
(mL/kg) G1 F 10 1-10 - N - One G2 F 10 11-20 1 LD ₅₀ Y - One G3 F 10 21-30 1 LD ₅₀ Y Allarine Acetate 20 One G4 F 10 31-40 1 LD ₅₀ Y Allarine Acetate 40 One G5 F 10 41-50 10 LD ₅₀ Y - One G6 F 10 51-60 10 LD ₅₀ Y Allarine Acetate 20 One G7 F 10 61-70 10 LD ₅₀ Y Allarine Acetate 40 One

The influenza virus was H1N1 influenza A virus (A/Puerto Rico/08/1934, H1N1), _{and the challenge inoculation titers were 1 LD 50} and 10 LD ₅₀ , and the challenge inoculation route was intranasal. In addition, the test substances in each group were administered once through intravenous (IV), 7 days after virus inoculation.

During the period of acclimatization, administration, and observation, the type of general symptoms including death, the date of onset, and the severity of symptoms were observed once a day and recorded for each individual.

Through the primary survival rate evaluation experiment, animals were anesthetized by inhalation with isoflurane on the 2nd day (9th day after infection) after drug administration, and when anesthesia was confirmed, an open chest was opened to expose the lungs and the tracheal bronchus was exposed and the right bronchus was ligated using 3-0 silk. A syringe connected with a catheter was installed in the left bronchus, and then, about 1 mL each was injected twice through the connected syringe, and then BALF (bronchoalveolar lavage fluid) was collected. After that, whole lungs were collected, divided into left and right lobes, and the left lobe was frozen and stored, and the right lobe was sufficiently immersed in 10% NBF and fixed.

After blood was collected from the posterior vena cava using a syringe, a portion of the collected blood was placed in an EDTA-3K tube and stored for later analysis. The remaining blood was injected into a vacuum collection vessel containing a clot activator, allowed to stand at room temperature for about 15-20 minutes to coagulate, and then centrifuged at 3,000 rpm for 10 minutes to separate serum.

After confirming the number of cells through total cell count to differentiate each cell present in BALF, eosinophils, neutrophils, macrophages, and leukocytes ( Lymphocyte) was checked and the white blood cell percentage was calculated (differential cell counting), and the results are shown in Table 2 below.

After homogenization using the left lobe of the collected lung tissue, proteins were extracted and cytokines such as TNF-alpha, IL-6, and IL-10 were analyzed using an ELISA kit. In the isolated serum, cytokines such as TNF-alpha, IL-6, and IL-10 were analyzed using an ELISA kit to test for inflammatory cytokines, and the results are shown in Tables 3 to 6 below.

In addition, tissues fixed in 10% NBF undergo general tissue processing such as trimming, dehydration, paraffin embedding, and sectioning to prepare samples for histopathological examination, and then stain with Hematoxylin & Eosin (H&E). was performed, and histopathological changes were observed using an optical microscope (Olympus BX53, Japan) to perform histopathological examination, and the results are shown in FIGS. 3 to 6 .

As a result of calculating the percentage of white blood cells, the levels of eosinophils, neutrophils, macrophages and leukocytes of all H1N1 influenza A virus challenge inoculation groups (G2-G4) were statistically significantly higher than those of the normal control group (G1) (p<0.001). ), macrophage and eosinophil levels in the allarin acetate 20 μg/kg administration group (G3) were significantly lower than in the induced control group (G2) (p<0.05).

As a result of ELISA analysis, the lung TNF-α and IL-6 levels of all H1N1 influenza A virus challenge inoculation groups were significantly higher than those of G1 (p<0.001), and the IL-6 levels of G3 were significantly higher than those of G2. was very low (p<0.05). The IL-10 level of all test substance administration groups (G3 and G4) was significantly higher than that of G1 (p<0.05 and p<0.01), and the TNF-α/IL-10 ratio level of G2 and G4 was G1. was significantly higher (p<0.01 and p<0.05).

After homogenizing the lung tissue, the protein was extracted and ELISA analysis was performed. As a result, TNF-α, IL-6 and IL-10 levels of all H1N1 influenza A virus challenge groups were significantly higher than those of G1 (p<0.001). , p<0.01 or p<0.05).

As a result of collecting BALF and performing ELISA analysis, the TNF-α, IL-6, IL-10 and IL-6/IL-10 ratio levels of all H1N1 influenza A virus challenge groups were significantly higher than those of G1. (p<0.001, p<0.01, or p<0.05), and the TNF-α/IL-10 ratio of G2 was significantly higher than that of G1 (p<0.001). The IL-6 level of G3 was significantly lower than that of G2 (p<0.05), and the TNF-α/IL-10 and IL-6/IL-10 ratio levels of all test substance-administered groups were significantly lower than that of G2. low (p<0.05).

As a result of performing ELISA analysis by separating the serum, the TNF-α, IL-6, TNF-α/IL-10 and IL-6/IL-10 ratio levels of all H1N1 influenza A virus challenge groups were significantly higher than those of G1. was found to be high (p<0.001), the IL-6 level of G3 was significantly lower than that of G2 (p<0.05), and the IL-6/IL-10 ratio level of all test substance-administered groups was significantly lower than that of G2. was found (p<0.05).

As a result of histopathological examination, no abnormal findings were observed in the normal control group (G1). In the induced group, macrophages and neutrophils were mainly infiltrated with diffuse intraalveolar space, and the levels were found to be moderate or severe, and some individuals were accompanied by severe hyperemia. In the case of G3 and G4, the test substance administration group, weak and mainly lymphocyte infiltration was observed, and local neutrophil infiltration was also observed.

As a result of calculating the percentage of leukocytes in this example, the levels of eosinophils, neutrophils, macrophages and leukocytes of all test substance-administered groups were significantly higher than those of the normal control group. and the levels of macrophages and eosinophils showed a significant difference.

In addition, as a result of ELISA analysis, the TNF-α and IL-6 levels of the test substance administration group measured in all tissues were lower than those of the induced control group G2, and IL-6 measured in the lung, BALF and serum of the G3 group. The level was significantly lower than that of the induced control group. In addition, as a result of comparing the TNF-α/IL-10 and IL-6/IL-10 ratios, which are indicators indicating the severity of injury, there was no significant difference in the lungs, but it showed a low level compared to the induced control group, TNF-α/IL-10 and IL-6/IL-10 ratio levels measured in BALF and serum were significantly lower than those of the challenge control group, G2. As a result of histopathological examination, infiltration of inflammatory cells was found to be weak or weak in all subjects in the test substance administration group, and the lesion was also localized, indicating that the induced control group G2 was moderate or severe. showed an improved trend compared to (see FIGS. 3 to 6).

In addition, in this Example, each individual body weight was measured using an electronic scale before and after administration of the test substance once/day for 2 weeks, and body temperature was also measured once/day for 2 weeks using an electronic thermometer, and the results were obtained. It is shown in Tables 7 to 9 below. In Table 7, data are expressed as mean±standard deviation.

In addition, the clinical symptoms of mice after influenza virus infection were scored according to the following criteria according to the behavior of the mice, and the results are shown in Table 10 below:

1. slight trembling of the hair

2. Wavy fur, reduced mobility

3. Wavy fur, reduced mobility, rapid breathing

4. Fluctuating fur, reduced mobility, crouching appearance, rapid and/or labored breathing indicative of pneumonia (euthanize animals showing evidence of pneumonia or losing more than 20% of their original body weight)

5. Death.

As can be seen from Tables 7 to 8, the allarin acetate-administered group (20 μg/kg) according to the present invention showed no difference compared to the control group G5, but the allarin acetate-administered group (40 μg/kg) was infected On the 7th day after the drug administration, a tendency to significantly increase the weight gain was observed, and it was confirmed that the recovery progressed significantly faster.

In addition, as can be seen from the body temperature measurement results in Table 9, the difference in body temperature between the test groups was not large, but in the case of the allarin acetate administered group (40 μg/kg), it is generally determined that the body temperature change was not large.

As shown in the clinical score evaluation results in Table 10, in the case of G7 in the allarin acetate administration group (40 μg/kg), it was judged that the improvement of clinical symptoms was significantly faster than G5, the control group, 9 days after infection. do.

When the test substance was administered to C57BL/6 mice challenged with H1N1 influenza A virus under the test conditions as described in this Example, as described above, the induced control group in leukocyte percentage calculation, ELISA analysis and histopathological examination An effective improvement trend was observed compared to phosphorus G2, and a secondary anti-inflammatory evaluation test was conducted under the condition _{of 10 LD 50 among the 1 LD 50} virus and 10 LD ₅₀ virus-infected groups inoculated in the above experiment. As a result, improvement in clinical symptoms was confirmed after 9 days in G7, a group treated with 40 μg/kg of allarin acetate, compared with control group G5, a group treated with virus alone. As such, when the test substance is administered to C57BL/6 mice challenged with the H1N1 influenza A virus, it appears that the administration of the test substance can help suppress the acute inflammation and the decrease in immunity caused by the influenza virus. Therefore, it is judged that allarin has the effect of suppressing acute inflammation and weakening of immunity caused by influenza virus and effectively treating it.

In the above detailed description of the present invention, a preferred embodiment for the use of an agent for inhibiting and treating acute inflammation and immunosuppression caused by influenza virus using allarin as a main component has been described, but the content is described in the above embodiment Without departing from the gist of the present invention as claimed in the following claims, it is within the scope of the present invention to the extent that various modifications can be made by anyone with ordinary knowledge in the field to which the invention pertains. You will find that it is included.

Claims (6)

In the composition for inhibiting symptoms caused by influenza virus comprising Alarin as a main component,
The alarin is a human-derived allarin acetate salt (trifluoroacetate salt) having the following amino acid sequence,
Suppressing the symptoms caused by the influenza virus is characterized in that by suppressing the symptoms of weight loss and body temperature drop to maintain the body weight and body temperature in a state not infected with the influenza virus, weight loss caused by the influenza virus and a composition for inhibiting body temperature drop:
H-Ala-Pro-Ala-His-Arg-Ser-Ser-Thr-Phe-Pro-Lys-Trp-Val-Thr-Lys-Thr-Glu-Arg-Gly-Arg-Gln-Pro-Leu-Arg- Ser-OH-acetate salt. delete The composition according to claim 1, wherein the allarin has a molecular weight of 2894.29 and has the following molecular formula:
C ₁₂₇ H ₂₀₆ N ₄₃ O ₃₈ . According to claim 1, wherein the weight loss and body temperature lowering inhibitor caused by the influenza virus is an influenza virus, characterized in that it is formulated to be administered in an adult 60kg standard of 1.2mg to 1.4mg of alarin acetate (Alarin acetate) A composition for inhibiting induced weight loss and body temperature drop. 5. The composition of claim 4, wherein the formulation is an injection, which inhibits weight loss and body temperature drop caused by influenza virus. The composition according to claim 4, wherein the formulation is administered by subcutaneous intramuscular injection.

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