KR20170009276A - Use of Radotinib for treatment of viral respiratory disease - Google Patents

Abstract

The present invention relates to a novel use of radotinib for treating viral respiratory diseases. According to the present invention, radotinib represented by chemical formula 1 or a pharmaceutically acceptable salt thereof can be used for preventing, alleviating and treating coronavirus infection. Also, radotinib can be used as an anti-virus agent useful for preventing and treating diseases induced by infection of Middle East respiratory syndrome coronavirus (MERS-CoV).

Description

Use of Radotinib for the Treatment of Respiratory Viral Diseases [

The present invention relates to the use of Radotinib for the prophylaxis and treatment of respiratory diseases caused by MERS-CoV, especially those with antiviral activity against coronavirus.

The coronavirus is a single-stranded positive RNA virus in the envelope and the genome size is 25-32 kb, which is a relatively large virus among known RNA viruses. It is said that the virus name comes from the Latin Corona, which has a specific structure of a flame or crown with spike protein, which is stick-shaped protuberance, on the outer skin.

The coronaviruses found in birds, animals, birds, cats, dogs, cows, pigs, and rats were first divided into four groups (Alpha-, Beta-, Gamma-, and Deltacoronavirus) Loses. The alpha and beta coronavirus groups are predominantly infected in mammals, while the gamma and delta coronavirus groups are found in birds. Coronavirus is known to cause a variety of diseases in animals such as gastrointestinal and respiratory diseases.

HCoV-229E and HCoV-OC43 found in the 1960s, HCoV-NL63 (2004) and HCoV-HKU1 (2005) found after the SARS pandemic, and these are generally associated with upper respiratory tract infections It is known, but it can lead to severe lung disease in immunodeficiency patients. It has been reported that the rate of coronavirus infection is increased mainly in winter and early spring, and coronavirus is the causative agent among adult cold patients. In 2003, SARS-CoV was first reported to cause Severe Acute Respiratory Syndrome (SARS). According to the World Health Organization (WHO) report, 8,273 patients worldwide There were 775 deaths (about 10% mortality), and by 2004, additional patient outbreaks and deaths were reported.

In September 2012, there was a serious respiratory disease patient with respiratory symptoms such as high fever, cough, and shortness of breath similar to SARS, and the causative agent was HCoV-EMC, which is different from the known virus.

The new coronavirus was classified under the name "Middle East respiratory syndrome-coronavirus (MERS-CoV)" in the coronavirus study group of the International Committee on Taxonomy of Viruses in May 2013. The virus is presumed to be the most potent source of bats, since genetic sequencing is similar to Pipistrellus Bat CoV-HKU5 and HKU4 found in bats. However, a recent study, published in The Lancet Infectious Diseases journals, found that all of the 50 blood samplings of Oman's dromedary camel were found, indicating that MERS-CoV antibody was found. In the Canary Islands, however, 15 out of 105 were positive for antibodies.

Although the virus itself was not found, the study showed that these camels were infected with mers or similar virus at some point and showed that the virus host is very likely to be a camel.

After the first case of infection in Saudi Arabia in September 2012, MERS-CoV coronavirus (MERS-CoV) infection was officially reported to WHO until June 2018, with 808 worldwide and 313 deaths (34.5%).

Early diagnosis of MERS-CoV coronavirus (MERS-CoV) The patient's clinical symptoms were significantly similar to SARS, and there was concern that SARS might have a significant association with SARS. However, It has been found that it is a new virus that is significantly different from the SARS virus.

First, the Middle East Respiratory Syndrome coronavirus (MERS-CoV) showed about 55% lower nucleotide homology with the SARS virus. In the phylogenetic analysis, the two viruses showed different evolutionary differences, The middle respiratory syndrome coronavirus (MERS-CoV) in the B group of beta-coronavirus belonged to the C group of beta-coronaviruses. Those belonging to the beta coronavirus group are known to be the SAR coronaviruses and human coronaviruses HCoV-HKU1 and HCoV-OC43.

In addition, the pathway for SARS and the respiratory syndrome coronavirus (MERS-CoV) is also different. The SARS virus is the ACE2 receptor distributed in human airway epithelium, the MERS-CoV DDP4 (also called CD26) in the parental cells is used. Both MERS-CoV and SARS viruses have high mortality rates, but they must be spread through large droplets after progressing to pneumonia. Therefore, influenza A Suggests that infectivity may be reduced by as much as compared to diseases that spread to aerosols, such as measles cases. However, in the case of SARS, the risk of infectious disease is still high because it can be significantly increased in the case of close exposure to SARS or aerosolization of respiratory droplets.

It is not yet clear what the initial source of the MERS-CoV coronavirus (MERS-CoV) was and how it spread to humans and caused the disease. However, two cases of confirmed new coronavirus cases were found to be on farm and animal contact. This shows the possibility of common infection of the new coronavirus.

Major clinical symptoms are fever (87%), cough (89%), dyspnea, and some patients have vomiting and diarrhea (35%). Renal failure also occurs in patients with impaired immune function, with a mortality rate of 34.5%. (Saudi Arabia, Qatar, etc.), and it is estimated that these areas are infected areas, but they can not accurately identify the exact route of infection. However, there has been no evidence of widespread infection among people, but it has been confirmed that close contact is made with family members or medical staff. The incubation period is estimated from 9 days to 12 days. In the MEJM (2008.02) report, it was 1.9 days to 14.7 days (average 5.2 days).

There is no developed drug or preventive vaccine for which the therapeutic effect of the MERS-CoV virus has been confirmed yet in the Middle East respiratory syndrome coronavirus (MERS-CoV). Until now, there has been no special treatment or prevention method except to avoid contact with suspected viral infection patients and thorough personal hygiene. Currently, antibiotics, ribavirin, interferon, and patient's serum have been used to treat infections of the Middle East Respiratory Syndrome Coronavirus (MERS-CoV), but these treatments have not shown any specific effect.

Therefore, it is very urgent to develop alternative medicines for the treatment and prevention of diseases of the respiratory syndrome coronavirus (MERS-CoV) in the Middle East.

The present invention provides an antiviral agent exhibiting strong antiviral activity against coronavirus.

The present invention provides a therapeutic composition comprising radotinib (Formula 1) or a pharmaceutically acceptable salt thereof as an active ingredient for the prevention and treatment of respiratory diseases caused by coronavirus.

≪ Formula 1 >

Hereinafter, the present invention will be described in detail.

The composition of the present invention contains radotinib or a pharmaceutically acceptable salt thereof as an active ingredient and can be used for the purpose of prevention and treatment of coronavirus diseases.

The present invention provides receptor tyrosine kinase inhibitors for the treatment or prevention of viral respiratory disease in a patient, and in particular, the use of radtunib or a pharmaceutically acceptable salt thereof. In one non-limiting embodiment of the invention, there is provided a use of radotinib for the inhibition of both replication, transmission or replication and transmission of coronavirus. The present invention provides the use of receptor tyrosine kinase inhibitors such as radothinib for the treatment of viral respiratory disease in animal patients, preferably mammalian patients, and more preferably human patients.

The term " radothinib "of the present invention refers to a compound represented by the above formula (1), such as 4-methyl-N- [3- (4-methylimidazol- - [4- (3-methylpyrazin-2-yl) -pyrimidin-2- ylamino) benzamide.

Compounds of formula 1 are known to inhibit one or more tyrosine kinases such as c-Abl, Bcr-Abl, PDGFR, Flt3, VEGF-R, EGF-R and c-Kit as receptor tyrosine kinases, It has excellent anticancer effect against various cancers such as stomach cancer, colon cancer, pancreatic cancer, liver cancer, prostate cancer, breast cancer, chronic or acute leukemia, blood cancer, brain tumor, bladder cancer, rectal cancer, cervical cancer, lymphoma, 10-0674813. However, they are effective against respiratory virus diseases such as respiratory viruses such as Coronavirus, Adenovirus, Influenza virus, parainfluenza virus, and respiratory syncytial virus. There is no report.

The compounds of the present invention may be used as such or in the form of a pharmaceutically acceptable salt thereof. The term " pharmaceutically acceptable " as used herein means physiologically acceptable and does not generally cause an allergic reaction or a similar reaction when administered to humans, and the salt includes a pharmaceutically acceptable free acid, Lt; / RTI > is preferred. Acid addition salts are prepared by reacting a compound of the present invention with a compound of the formula I with hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, methanesulfonic acid, toluenesulfonic acid, benzenesulfonic acid, acetic acid, propionic acid, ascorbic acid, citric acid, malonic acid, fumaric acid, maleic acid, lactic acid, ≪ / RTI > or tartaric acid. These salts can be prepared by known methods, for example, by treating the compound with a suitable acid in the presence of a suitable solvent.

The compounds of the present invention may be in crystalline form or in the form of a solvate (e.g., hydrate) and both forms are contemplated to be within the scope of the present invention. The term "solvate" is a complex of various stoichiometries formed by a solute (a compound of the present invention in the present invention) and a solvent. These solvents should not interfere with the biological activity of the solute. For example, the solvent may be water, ethanol, methanol, ethyl acetate or acetone. Solventing methods are generally known in the art.

Pharmaceutically acceptable salts of the compounds of formula (1) include both unsolvated and solvated forms.

The present invention provides a method for preventing the infection of a mammalian coronavirus comprising administering to said mammal a therapeutically effective amount of a compound of formula 1 or a pharmaceutically acceptable derivative thereof.

Although the present invention describes the treatment of coronaviruses, in particular human coronaviruses, the present invention relates to the use of coronaviridae, more particularly Betacoronavirus, more specifically the MERS-CoV virus, For the treatment of animal and human strains.

The composition of the present invention may also be provided in the form of a pharmaceutical composition wherein the pharmaceutical composition comprises a pharmaceutically effective amount of a compound of the present invention or a pharmaceutically acceptable salt thereof alone or in combination with one or more A pharmaceutically acceptable carrier may further be included. Preferably, the carrier may be a pharmaceutically acceptable inert carrier. As used herein, the term "pharmaceutically effective amount" refers to an amount that exhibits a further reaction than the negative control, and preferably refers to an amount sufficient to treat or prevent a viral respiratory disease. The therapeutic dose when the patient is a human is generally from 50 mg to 2000 mg / day, more preferably from 100 mg to 1000 mg / day, depending on the seriousness of the condition and whether the radtinib is administered alone or in combination with other drugs . May be administered via the oral or parenteral route once or divided once a day. However, the pharmaceutically effective amount may be appropriately changed depending on various factors such as the disease and its severity, the patient's age, body weight, health condition, sex, administration route and treatment period.

The term "pharmaceutically acceptable" as used herein means a non-toxic composition which is physiologically acceptable and which, when administered to humans, does not inhibit the action of the active ingredient and does not normally cause an allergic reaction such as gastrointestinal disorder, dizziness, . The composition of the present invention can be variously formulated according to the route of administration by a method known in the art together with the pharmaceutically acceptable carrier. Routes of administration include, but are not limited to, oral or parenteral administration. Parenteral routes of administration include, for example, transdermal, intranasal, intraperitoneal, intramuscular, subcutaneous, intravenous, and other routes.

When the pharmaceutical composition of the present invention is orally administered, the pharmaceutical composition of the present invention may be formulated into a powder, a granule, a tablet, a pill, a sugar, a tablet, a capsule, a liquid, a gel , Syrups, suspensions, wafers, and the like. Examples of suitable carriers include saccharides including lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol and maltitol, and starches including corn starch, wheat starch, rice starch and potato starch, Cellulose such as methylcellulose, sodium carboxymethylcellulose and hydroxypropylmethyl-cellulose and the like, fillers such as gelatin, polyvinylpyrrolidone and the like. In addition, crosslinked polyvinylpyrrolidone, agar, alginic acid, or sodium alginate may optionally be added as a disintegrant. Further, the pharmaceutical composition may further comprise an anti-coagulant, a lubricant, a wetting agent, a flavoring agent, an emulsifying agent and an antiseptic agent.

In addition, when administered parenterally, the pharmaceutical composition of the present invention can be formulated together with a suitable parenteral carrier according to methods known in the art in the form of injection, transdermal drug delivery, and nasal inhalation. In the case of such injections, they must be sterilized and protected against contamination of microorganisms such as bacteria and fungi. Examples of suitable carriers for injectables include, but are not limited to, solvents or dispersion media containing water, ethanol, polyols (e.g., glycerol, propylene glycol and liquid polyethylene glycol, etc.), mixtures thereof and / or vegetable oils . More preferably, suitable carriers include isotonic solutions such as Hanks' solution, Ringer's solution, phosphate buffered saline (PBS) containing triethanolamine, or sterile water for injection, 10% ethanol, 40% propylene glycol and 5% dextrose Etc. may be used. In order to protect the injection from microbial contamination, various antibacterial and antifungal agents such as parabens, chlorobutanol, phenol, sorbic acid, thimerosal and the like may be further included. In addition, the injections may in most cases additionally include isotonic agents, such as sugars or sodium chloride.

These formulations are described in Remington's Pharmaceutical Science, 15th Edition, 1975, Mack Publishing Company, Easton, Pennsylvania, which is a commonly known formulary in pharmaceutical chemistry.

In the case of an inhalation dosage form, the compounds used according to the invention can be formulated into a pressurized pack or a pressurized pack using a suitable propellant, for example dichlorofluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gases. It can be conveniently delivered in the form of an aerosol spray from a nebulizer. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve that delivers a metered amount. For example, capsules and cartridges for use in an inhaler or insufflator may be formulated to contain a powder mix of a suitable powder base.

Other pharmaceutically acceptable carriers can be found in Remington's Pharmaceutical Sciences, 19th ed., Mack Publishing Company, Easton, Pa., 1995).

The composition of the present invention provides the use of a compound of formula (I) or a pharmaceutically acceptable salt thereof for the inhibition of replication or transmission of an RNA virus of Coronavirus.

The composition may be administered in combination with one or more additional drugs having antiviral efficacy to prevent and treat respiratory viral infectious diseases.

The composition can be administered with one or more drugs of these drugs to treat respiratory viral infectious disease prevention and symptoms, along with antiviral drugs such as antibiotics, interferon and ribavirin, etc. .

In the examples of the present invention, the efficacy of the antiviral agent was evaluated through Example 1 and Example 2 in order to confirm whether the compound of Chemical Formula 1 has antiviral activity against respiratory diseases caused by coronavirus. There was no virus-inhibiting effect on the concentration of ribavirin treated with the comparative drug. However, as shown in FIGS. 1 and 2, it was confirmed that the radoninib showed a decrease in the RNA copy number compared with the control group at most of the concentration-treated conditions, and thus it has an antiviral effect against MERS-CoV.

Accordingly, the present invention provides a composition for preventing and treating respiratory diseases caused by coronavirus, comprising radodinib or a pharmaceutically acceptable salt thereof as an active ingredient. The present invention also relates to a process for the preparation of 4-methyl-N- [3- (4-methylimidazol-1-yl) -5-trifluoromethyl- Methylpyrazin-2-yl) -pyrimidin-2-ylamino) benzamide or a pharmaceutically acceptable salt thereof as an active ingredient for preventing and treating respiratory viral diseases.

More preferably, the present invention provides a composition for the prevention and treatment of respiratory diseases caused by MERS-CoV (Middle East Respiratory Syndrome) coronavirus.

The above description does not limit the claimed invention in any way, and the combination of the features discussed further is not absolutely necessary for the solution of the present invention.

The present invention provides a composition for preventing and treating respiratory viral diseases, which comprises radotinib or a pharmaceutically acceptable salt thereof as an active ingredient. The composition of the present invention can be effectively used for prevention and treatment by effectively inhibiting respiratory diseases caused by MERS-CoV (Middle Respiratory Tract Syndrome) coronavirus (MERS-CoV).

FIG. 1 shows the results of viral gene activity analysis at 12 and 24 hours after infection of cells with MERS-CoV and the compounds of the present invention.
FIG. 2 shows the viral gene activity analysis results at 8 hours and 24 hours after the cells were first infected with MERS-CoV, and then the cell culture solution containing the compound of the present invention was added at different concentrations.

The present invention will be explained in more detail through the following examples. However, the embodiments are for illustrative purposes only and should not limit the scope of the invention in any way.

<Examples>

Example  One

In Example 1, the virus was directly treated with radotinib before the cells were infected with MERS-CoV (Korean isolate, P4), and the drug was directly applied to the envelope protein to evaluate the effect of inhibiting the virus infection Respectively.

1) virus

    - MERS-CoV Korea separator (P4)

    - Virus titers: 1 x 105 PFU / ml

    - Virus Inoculation Titer: MOI 0.01 (3 x 103 PFU / ml)

2) Cell line - Vero cells (24-well plate)

3) Test method

Radotinib was diluted with 200 μM, 50 μM, 12.5 μM, 3.13 μM, and 0.0 μM (control group without radotinib treatment) and reacted with 104 PFU of MERS-CoV in a 37 ° C 5% CO ₂ incubator for 1 hour . After completion of the reaction, Vero cell line is infected for 1 hour, washed with PBS, and then cell supernatant is obtained at 12 and 24 hours after virus infection to extract RNA. The extracted RNAs were subjected to qRT-PCR to compare the RNA copy number to evaluate the antiviral efficacy.

As a result, RNA copy number was decreased in experimental group treated with 200 μM, 50 μM and 3.13 μ, except for 12.5 μM of radotinib, as shown in FIG.

Example 1 is an experiment in which only the inhibitory effect of intracellular infection (entry) of virus was confirmed by reacting radotinib with the virus first, inoculating the cell, and not adding a drug into the cell culture fluid. In other words, since there is no radtinib in the cell culture solution, it is difficult to expect further sustained inhibition of viruses. In this case, the numerical difference of some viruses penetrating into the cell through the inhibitory effect of the drug may initially appear at different concentrations. However, even if a small amount of virus is introduced due to the inhibitory effect of the drug, it is predicted that the copy number of the gene will be similar to that of the control group because the replication of the gene will be explosive over time. Therefore, when the drug is not added to the culture solution after inhibiting intracellular penetration of the virus as in the experiment of Example 1, there is no difference in the RNA copy number of the virus according to the concentration of drug at 24 hours after the infection Respectively.

Example  2

Example 2 shows the effect of adding radothinib to a cell culture solution after first infecting cells with MERS-CoV (Korean isolate, P4) and comparing the efficacy of inhibiting the infection and replication of viruses released from the infected cells to ribavirin as a reference drug / The efficacy of the antiviral agent was evaluated through analysis.

1) virus

    - MERS-CoV Korea separator (P4)

    - Virus titers: 1 x 105 PFU / ml

    - Virus Inoculation Titer: MOI 0.01 (3 x 103 PFU / ml)

2) Cell line - Vero cells (24-well plate)

3) Test method

MERS-CoV is infected with Vero cell line at MOI of 0.01 for 1 hour at 37 ° C. After washing the cells with PBS, the cell culture medium containing each concentration (50 μM, 12.5 μM, 3.13 μM, and 0.0 μM) of the drug is added and incubated in a 5% CO ₂ incubator at 37 ° C. Cellular supernatants are obtained at predetermined times of 8 hours and 24 hours, and then RNA is extracted. QRT-PCR is performed using the extracted RNA to compare the RNA copy number to evaluate the antiviral efficacy.

As a result, in the case of the experimental group treated with radotinib as shown in FIG. 2, it was confirmed that the RNA copy number was reduced by 50% or more in the supernatant obtained at 8 and 24 hours after the virus infection. Particularly, in the case of the experimental group treated with radotinib, the RNA copy number obtained by acquiring the supernatant after 24 hours after the virus infection was at least 56% as compared with the control group without the radotinib Up to 71%, and up to 78% RNA copy number reduction on upE gene basis. On the contrary, in the treated group treated with ribavirin, there was no decrease in the RNA copy number at 8 hours after the viral infection, and at the 12.5 μM treated group after 24 hours, the RNA copy number of orf1a decreased by 29%.

Taken together, the results of this study showed that the virus inhibitory effect of each concentration was hardly observed in the treated condition of ribavirin. However, at most concentrations of radtunib treatment, the RNA copy number was reduced compared to the control group, indicating that it has antiviral efficacy against MERS-CoV.

&Lt; Industrial applicability >

The invention has been described as a preferred embodiment. It will, however, be evident to those skilled in the art that numerous modifications and variations can be made without departing from the scope of the invention disclosed herein. In addition, the present invention can provide an antiviral agent for the prevention and treatment of viral respiratory diseases by providing a composition containing radotinib or a pharmaceutically acceptable salt thereof as an active ingredient.

Claims (9)

A pharmaceutical composition for preventing or treating a viral respiratory infectious disease in a patient, which comprises radotinib or a pharmaceutically acceptable salt thereof as an active ingredient. The pharmaceutical composition according to claim 1, wherein the radotinib is a compound represented by the following formula (1) or a salt thereof.
&Lt; Formula 1 >

The pharmaceutical composition according to claim 1, wherein the viral respiratory infectious disease is a disease caused by infection with coronavirus. The pharmaceutical composition according to claim 1, wherein the viral respiratory infectious disease is a disease caused by an infection of a Middle Respiratory Syndrome Coronavirus (MERS-CoV). 2. The pharmaceutical composition according to claim 1, wherein the patient is an animal. 6. The pharmaceutical composition according to claim 5, wherein the animal is a mammal. The pharmaceutical composition according to claim 6, wherein the mammal is a human. The pharmaceutical composition according to claim 1, further comprising at least one other therapeutic agent selected from the group consisting of antibiotics, interferon, and ribavirin. The pharmaceutical composition according to claim 1, further comprising a pharmaceutically acceptable inert carrier.

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