JP6363765B1 - Novel compound and pharmaceutical composition for treating tuberculosis containing the same - Google Patents

Abstract

Disclosed is a pharmaceutical composition for the prevention or treatment of tuberculosis. A compound represented by Formula 2, a pharmaceutically acceptable salt thereof, and a composition containing them as an active ingredient. [Selection] Figure 1

Description

  The present invention relates to a novel compound and a composition for preventing or treating tuberculosis comprising the novel compound.

  Tuberculosis, which has a long history, is currently infected by 1/3 of the world population, and infected people can become TB patients whenever their immune function is reduced by infection with a disease such as AIDS. Currently, 16 million patients are estimated worldwide and 8 million new tuberculosis patients occur each year. Of those, 2 million die each year.

  In developed countries, the tuberculosis infection rate is 0.1% or less and the disease incidence is 25% or less. In the case of the Republic of Korea, the prevalence of active pulmonary tuberculosis (prevalence by radiography) decreased from 5.1% in 1965 to 1.0% in 1995, and the positive prevalence for bacteria from 0.94% The annual tuberculosis infection risk rate decreased from 5.3% to 0.5%. The death rate from tuberculosis decreased from 10.4% in 1991 to 6.3 in 2001 per 100,000 population.

  However, tuberculosis is still one of the top 10 causes of death, and as a principle for treatment, initial treatment is based on short-term therapy for 6 months. In general, four anti-tuberculosis drugs rifampin (RIF), isoniazid (INH), pyrazinamide (PZA), ethambutol (EMB) are used in combination, and two months later, RIF, INH And 3 treatments of EMB for 4 months. Streptomycin (STR) can be used instead of EMB for 2 months of initial intensive care. In this case, maintenance treatment is administered RIF and INH. Retreatment for relapsed patients remains largely sensitive to the original treatment drug, so the principle is to use the original drug again, extending it further by 3 months beyond the prescribed initial treatment period. To treat. For re-treatment of patients who have failed the first treatment, a drug sensitivity test is performed to exclude all drugs used for the first treatment, and at least 3 new sensitive drugs that have not been used in the past, if possible 4 , Or more, and in principle, 18 months or more.

  For the treatment of pulmonary tuberculosis, four drugs are used in combination in order to prevent outpatient treatment and acquisition of resistance. In the anti-tuberculosis-resistant bacteria, mutant strains appear at a frequency of about every 106 cell divisions. Therefore, the principle of multi-drug therapy using at least two anti-tuberculosis drugs together must be observed.

  The biggest problem with tuberculosis treatment is that there is no special treatment if resistance to primary anti-tuberculosis drugs is achieved. In fact, the development and systematic management of effective anti-tuberculosis drugs has drastically reduced the incidence and death of tuberculosis, but recently the occurrence and acquired nature of multi-drug resistant tuberculosis (MDR-TB) Tuberculosis is increasing due to the immunodeficiency syndrome (AIDS) epidemic. MDR-TB refers to the occurrence of simultaneous resistance to at least two drugs including rifampin (RIF) and isoniazid (INH). The criterion for determination is that the number of M. tuberculosis strains grown on a medium containing a drug exceeds 1% of the number of strains grown on a medium not containing the drug.

  Recently, resistant tuberculosis, including multidrug resistant tuberculosis, has emerged as a serious health problem. In recent years, the incidence of tuberculosis in South Korea has not decreased any more, which is closely related to the fact that the number of resistant tuberculosis patients has not decreased. Also, 50% of resistant tuberculosis patients are known to die. Therefore, many causes of death from tuberculosis are due to resistant tuberculosis.

  In 1995, the resistance rate to INH was 9.2% in a survey of tuberculosis in the Republic of Korea. Resistance to one or more drugs was 9.9% and resistance to two or more drugs including INH and RIF was 5.3%. These are the roots of persistent diffusion of drug-resistant bacteria to the community.

  Currently, several types of secondary anti-tuberculosis drugs have been developed, but there is no drug that can replace the primary drug in terms of efficacy. Therefore, the development of new anti-tuberculosis drugs that have different mechanisms of action from existing drugs is a very urgent issue.

Korean Published Patent Publication No. 10-2015-0096041 (2015.08.24)

  An object of the present invention is to provide a compound represented by Formula 2 or a pharmaceutically acceptable salt thereof.

  An object of the present invention is to provide a pharmaceutical composition for the prevention or treatment of tuberculosis comprising the compound represented by Formula 2 or a pharmaceutically acceptable salt thereof as an active ingredient.

  An object of the present invention is to provide a pharmaceutical composition for preventing or treating resistant tuberculosis comprising a compound represented by Formula 2 or a pharmaceutically acceptable salt thereof as an active ingredient.

  An object of the present invention is to provide a food composition for preventing or improving tuberculosis comprising a compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof as an active ingredient.

  An object of the present invention is to provide a food composition for preventing or ameliorating resistant tuberculosis, comprising as an active ingredient a compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof.

である場合に比べて抗結核活性が２〜４倍優れており、

又は−ＣＨ_２ＣＨ_２ＣＨ_２ＳＯ_３Ｈに比べても抗結核活性が１２８倍以上優れていることを確認し、化学式１におけるＲ_１の置換基がＢｕｔｙｌである化学式２の化合物を用いて抗結核治療剤を提供しようとする。 In the phenanthrenyl compound represented by Chemical Formula 1, the anti-tuberculosis activity against H37Ra (Mycobacterium tuberculosis H37Ra) was evaluated by changing the substituent of R ₁ . At this time, when the substituent of R ₁ is Butyl having 4 carbon atoms, the antituberculous activity is at least 2 to 4 times or more superior to -CH ₃ , -Ethyl, and -Proyl having a small number of carbon atoms. cage, more number of carbon atoms -n-Pentyl, -Hexyl, to confirm that the anti-tuberculosis activity than -Heptyl is excellent 16-32 times, also, the substituents _{R 1}

Antituberculosis activity is 2 to 4 times better than

Alternatively, it is confirmed that the antituberculosis activity is 128 times or more superior to that of —CH ₂ CH ₂ CH ₂ SO ₃ H, and the compound of the chemical formula 2 in which the substituent of R ₁ in the chemical formula 1 is Butyl is used. Try to provide TB treatment.

Wherein R ₁ in Chemical Formula 1 is butyl, R ₂ is a compound of Formula 2 is methyl, is as following Formula 2.

  In order to achieve the above object, the present invention provides a compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof.

  The term “pharmaceutically acceptable salt” as used in the present invention retains the desired biological and / or physiological activity of the compound and exhibits minimal unwanted toxicological effects. Means any salt. It means a salt produced by a usual method in the art, and such a production method is known to those skilled in the art. Specifically, the pharmaceutically acceptable salt includes, but is not limited to, salts derived from pharmacologically or physiologically acceptable inorganic acids, organic acids and bases.

  For example, pharmaceutically acceptable base addition salts can be prepared from inorganic and organic bases. Salts derived from inorganic bases can include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, and magnesium salts. Salts derived from organic bases include, but are not limited to, primary, secondary and tertiary amines; substituted amines including naturally occurring substituted amines; and isopropylamine, trimethylamine, diethylamine , Triethylamine, tripropylamine, ethanolamine, 2-dimethylaminoethanol, tromethamine, lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline, betaine, Cyclic containing ethylenediamine, glucosamine, N-alkylglucamine, theobromine, purine, piperazine, piperidine, and / or N-ethylpiperidine It can include a salt of Min. Also, carboxylic acid amides including other carboxylic acid derivatives such as carboxamides, lower alkyl carboxamides, di (lower alkyl) carboxamides, and the like may be included.

  For example, pharmaceutically acceptable acid addition salts can be prepared from inorganic and organic acids. Salts derived from inorganic acids include hydrochloric acid, bromic acid, sulfuric acid, nitric acid, phosphoric acid and the like. Salts derived from organic acids include acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, malic acid, malonic acid, succinic acid, maleic acid, fumaric acid, tartaric acid, citric acid , Benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, and / or salicylic acid, and the like.

  In another aspect, the present invention provides a pharmaceutical composition for preventing or treating tuberculosis, comprising as an active ingredient a compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof.

  In the present invention, the active ingredient of the pharmaceutical composition for prevention or treatment of tuberculosis may be (S) -form or (R) -form of the compound represented by Formula 2.

  The (S) -form may have a structure represented by the following chemical formula 3: (S) -methyl 1-((6-butoxy-2,3-dimethoxyphenanthrene-10-yl) methyl) -5-oxopyrrolidine -2-carboxylate ((S) -methyl 1-((6-butyoxy-2,3-dimethylphenanthren-10-yl) methyl) -5-oxopyrrolidine-2-carboxylate), where (R) -form is And (R) -methyl 1-((6-butoxy-2,3-dimethoxyphenanthren-10-yl) methyl) -5-oxopyrrolidine-2-carboxylate (( R) -methyl 1-((6-butyoxy-2,3-dimox phenanthren-10-yl) methyl) can be named -5-oxopyrrolidine-2-carboxylate).

  In the present invention, the tuberculosis is caused by infection by tuberculosis bacteria, and the tuberculosis includes resistant tuberculosis, pulmonary tuberculosis, hemorrhoid tuberculosis, bone tuberculosis, throat tuberculosis, lymphoid tuberculosis, breast tuberculosis, spinal tuberculosis and the like. The Mycobacterium tuberculosis (Mycobacterium tuberculosis) and Tubercula bacillus (Tubecle bacillus) may be used as the tuberculosis causative agent of the tuberculosis, and more specifically, Mycobacterium tuberculosis ( Mycobacterium tuberculosis).

  In one embodiment of the present invention, the compounds of Formula 3 and Formula 4 corresponding to (S) -form and (R) -form of the compound of Formula 2 are prepared against Mycobacterium tuberculosis. It was confirmed to have antitubercular activity. In addition, it was confirmed that the compound of Chemical Formula 3 corresponding to (S) -form has an antituberculosis activity two times or more superior to the compound of Chemical Formula 4 corresponding to (R) -form.

  The explanation for the term “pharmaceutically acceptable salt” used in the present invention is as described above.

  The term “prevention” as used in the present invention may mean any act of administering the composition according to the present invention to an individual to suppress or delay the onset of tuberculosis. The term “treatment” as used in the present invention may mean any act that causes the composition of the present invention to improve or improve the symptoms of tuberculosis.

  A pharmaceutical composition comprising the compound represented by Formula 2 of the present invention or a pharmaceutically acceptable salt thereof as an active ingredient is an appropriate carrier, excipient or dilution generally used for the production of a pharmaceutical composition. An agent can further be included. At this time, the compound represented by Formula 2 or a pharmaceutically acceptable salt thereof contained in the composition is not particularly limited, but is 0.001% by weight to 99% by weight based on the total weight of the composition. %, Preferably 0.01% to 50% by weight.

  The pharmaceutical composition comprises tablets, pills, powders, granules, capsules, suspensions, liquids for internal use, oils, syrups, sterilized aqueous solutions, non-aqueous solvents, suspensions, oils, lyophilization agents. And any one dosage form selected from the group consisting of suppositories, and may be various oral or parenteral dosage forms. In the case of formulating, it is prepared using diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, surfactants and the like that are usually used.

  Solid dosage forms for oral administration include tablets, pills, powders, granules, capsules and the like, such solid dosage forms comprising at least one or more excipients such as starch, calcium carbonate, sucrose. (Sucrose) or lactose, gelatin or the like may be mixed. In addition to simple excipients, lubricants such as magnesium stearate and talc can also be used. Liquid preparations for oral administration include suspensions, liquids for internal use, oils, syrups, etc., and various excipients such as water and liquid paraffin, which are commonly used simple diluents. , Wetting agents, sweeteners, fragrances, preservatives and the like. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, oils, lyophilized formulations, suppositories. As the non-aqueous solvent and suspension solvent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like may be used. As a suppository base, witepsol, macrogol, tween 61, cacao butter, lauric fat, glycerogelatin and the like may be used.

  The pharmaceutical composition of the present invention can be administered in a pharmaceutically effective amount. In the present invention, the term “pharmaceutically effective amount” means an amount sufficient to treat a disease at a reasonable belief / risk ratio applicable to medical treatment, the level of effective dose. Is the type and severity of the individual, age, sex, type of illness, drug activity, drug sensitivity, administration time, route of administration and excretion rate, duration of treatment, factors including the drugs used simultaneously, and other It may be determined by factors well known in the medical field. The compositions of the present invention can be administered as individual therapeutic agents or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents. Single or multiple doses may be administered. Taking all of the above factors into account, it is important to administer an amount that gives the maximum effect in a minimum amount without side effects and can be readily determined by one skilled in the art.

  The pharmaceutical composition of the present invention is not particularly limited as long as it is an individual for the purpose of treatment or prevention of tuberculosis, and any one is applicable. For example, the pharmaceutical composition can be used for any of non-human animals such as monkeys, dogs, cats, rabbits, guinea pigs, rats, mice, cows, sheep, pigs, goats, etc., humans, birds and fishes. The product can be administered parenterally, subcutaneously, intraperitoneally, intrapulmonary and intranasally, and can be administered for local treatment by any suitable method, including intralesional administration, if necessary. The preferred dosage of the pharmaceutical composition of the present invention varies depending on the individual's condition, body weight, degree of illness, drug form, administration route and period, but can be appropriately selected by those skilled in the art. For example, it may be administered by oral, rectal, intravenous, intramuscular, subcutaneous, intrauterine dura or cerebrovascular injection, but is not limited thereto.

  A suitable total daily dose may be determined by the attending physician within the scope of good medical judgment and is generally in the range of 0.001 to 1000 mg / kg, preferably 0.05 to 200 mg / kg, More preferably, an amount of 0.1 to 100 mg / kg can be administered once to several times a day.

  In another aspect, the present invention provides a pharmaceutical composition for preventing or treating resistant tuberculosis, which comprises a compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof as an active ingredient.

  In the present invention, the resistant tuberculosis may be super tuberculosis (Extensible drug-resistant tuberculosis, XDR-TB), isoniazid-resistant tuberculosis, INHr-TB, rifampin-resistant tubule-resistant tuberculosis. Or one or more selected from the group consisting of pyrazinamide-resistant tuberculosis (PZAr-TB) and streptomycin-resistant tuberculosis (STRr-TB).

  In one embodiment of the present invention, the compound represented by Chemical Formula 3 and Chemical Formula 4 corresponding to (S) -form and (R) -form of the compound represented by Chemical Formula 2 is super tuberculosis ( Extensively drug-resistant tuberculosis, XDR-TB), isoniazid resistant TB (isoniazid-resistant tuberculosis, iNHr-TB), rifampin-resistant tuberculosis (rifampin-resistant tuberculosis, RIFr-TB), pyrazinamide-resistant tuberculosis (pyrazinamide-resistant tuberculosis, PZAr -TB), and streptomycin-resistant tube Rculosis, were confirmed to have the Strr-TB) to anti-tuberculosis activity.

  In the present invention, explanations for the terms “compound represented by Formula 2”, “pharmaceutically acceptable salt”, “prevention”, and “treatment” are as described above.

  In another aspect, the present invention provides a food composition for preventing or ameliorating tuberculosis, comprising as an active ingredient a compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof.

  In the present invention, the explanation for “compound represented by Formula 2”, “tuberculosis”, and “prevention” is as described above.

  In the present invention, the term “improving” means that a tuberculosis symptom is improved when a food composition containing a compound represented by Formula 2 or a pharmaceutically acceptable salt thereof as an active ingredient is administered to an individual. Can mean any act that makes you better.

  Specifically, the compound represented by Chemical Formula 2 of the present invention or a pharmaceutically acceptable salt thereof can be added to a food composition for the purpose of preventing or improving tuberculosis.

  The food composition of the present invention can include pills, powders, granules, soaking agents, tablets, capsules, liquids, and the like, and the compound represented by Chemical Formula 2 of the present invention or a foodically acceptable product thereof. There is no restriction | limiting in particular in the kind of foodstuff which can add salt, For example, there exist various drinks, gum, tea, a vitamin complex, health supplement foods, etc.

  In addition to the compound represented by Chemical Formula 2 or a pharmaceutically acceptable salt thereof, other ingredients can be added to the food composition, and the type thereof is not particularly limited. For example, as with normal foods, various herbal medicine extracts, food-acceptable food supplements, natural carbohydrates, and the like can be contained as additional components, but are not limited thereto.

  The food-acceptable salt of the present invention may be an acid addition salt formed by a food-acceptable free acid or a metal salt formed by a base. An inorganic acid and an organic acid can be used as the free acid. As the inorganic acid, hydrochloric acid, sulfuric acid, bromic acid, sulfurous acid or phosphoric acid can be used, and as the organic acid, citric acid, acetic acid, maleic acid, fumaric acid, gluconic acid, methanesulfonic acid, etc. are used. be able to. As the metal salt, an alkali metal salt, an alkaline earth metal salt, a sodium, potassium or calcium salt can be used. However, it is not necessarily limited to this.

  In the present invention, the term “food supplement additive” means a component that can be supplementarily added to food, and is added to produce a health functional food of each dosage form. It can be selected and used appropriately. Examples of food supplements include various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic and natural flavors, colorants, fillers, pectinic acid and salts thereof, alginic acid and salts thereof, Examples include organic acids, protective colloid thickeners, pH regulators, stabilizers, preservatives, glycerin, alcohol, carbonates used in carbonated beverages, etc. The type of is not limited.

  Examples of the natural carbohydrate include monosaccharides such as glucose and fructose; disaccharides such as maltose and sucrose; polysaccharides such as dextrin and cyclodextrin; and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents, natural flavoring agents (such as thaumatin), stevia extracts (such as rebaudioside A, glycyrrhizin) and synthetic flavoring agents (such as saccharin, aspartame) can be advantageously used.

  The food composition of the present invention may contain a health functional food. The term “health functional food” used in the present invention is manufactured and processed in the form of tablets, capsules, powders, granules, liquids, and rounds using ingredients and components having functionalities useful for the human body. Food. Here, the term “functionality” means that nutritional effects are adjusted on the structure and function of the human body, and useful effects are obtained for health uses such as physiological effects. The health functional food of the present invention can be produced by a method usually used in the art, and at the time of production, it can be produced by adding raw materials and ingredients usually added in the art. Further, unlike general medicines, using food as a raw material has the advantage that there are no side effects that may occur during long-term use of the medicine, and is excellent in portability.

  The mixing amount of the active ingredient can be suitably determined depending on the purpose of use (prevention, health or therapeutic treatment). Generally, the compound represented by the chemical formula 2 of the present invention or a pharmaceutically acceptable salt thereof is 1 to 50% by weight, preferably 5 to 10% by weight, of the raw material composition during the production of food. It may be added in an amount, but is not limited thereto. However, in the case of long-term intake for the purpose of health and hygiene or for the purpose of health regulation, the amount may be used within the above range.

  There is no restriction | limiting in particular in the kind of said foodstuff. Examples of foods to which the substance can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gums, dairy products including ice creams, various soups, There are drinking water, tea, drinks, alcoholic beverages, vitamin composites, etc., and all include health functional foods in a general sense.

  As another aspect, the present invention provides a food composition for preventing or ameliorating resistant tuberculosis comprising, as an active ingredient, a compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof.

  In the present invention, explanations for the terms “compound represented by Formula 2”, “foodically acceptable”, “resistant tuberculosis”, “prevention”, and “improvement” are as described above.

  The present invention relates to a compound represented by Chemical Formula 2 and uses of the compound represented by Chemical Formula 2 for prevention or treatment of tuberculosis. The compound represented by Chemical Formula 2 having antituberculous activity has characteristics that it is excellent in activity even in tuberculosis strains and resistant tuberculosis strains and has low cytotoxicity, and can be used as a pharmaceutical composition for prevention or treatment of tuberculosis. it can.

Is a graph showing the results of NMR ¹ H compounds of SCH1702 of the present invention. It is a graph showing the ^{NMR 13} results in C of the compound of SCH1702 of the present invention. It is a graph which shows the analysis result of S-form and R-form using the polarizer of the compound of SCH1702 of this invention. The cytotoxicity result of SCH1702 (S-form) corresponding to the compound of Chemical formula 3 in THP-1, Raw264.7, L929, and HEK-293 cells is shown. The experiment was repeated 3 times and n = 3. Values indicate mean ± standard deviation.

  Hereinafter, the present invention will be described in more detail by way of examples. However, the following examples are merely illustrative of the present invention, and the content of the present invention is not limited to the following examples.

  Production Example 1: Synthesis of Compound of Formula 2 (SCH1702)

  A compound of formula 2 (SCH1702) was synthesized by the method shown in the following reaction scheme 1.

In the above reaction formula 1, homoveratric acid represented by the chemical formula (I) and 4-butoxybenzaldehyde represented by the chemical formula (B) are converted into triethylamine, Et ₃ N, The mixture was reacted with acetic anhydride (Acetic anhydride, Ac ₂ O) under N ₂ for 10 hours, cooled, filtered through a filter, and purified with EtOAc to give a yellow compound (Step 1-1).

After thionyl chloride (SOCl ₂ ) was added to the compound represented by the chemical formula (c) and reacted for 40 minutes and concentrated in vacuo to remove the solvent, pyridine and methanol were added again, and then again. Reaction was performed in an ice bath for 40 minutes to obtain a compound represented by the chemical formula (2) (step: 1-2).

CH ₂ Cl ₂ and anhydrous FeCl ₃ are added to the compound represented by the chemical formula (2) in an ice bath, followed by reaction at room temperature for 12 hours. After that, NaHCO ₃ is added and the organic layer is fractionated and dried. The yellow compound represented by the chemical formula (e) was obtained (step: 1-3).

LiAlH ₄ (lithium aluminum hydride) and THF (tetrahydrofluoric acid) are added to the compound represented by the chemical formula (e) at 0 ° C. and reacted for 50 minutes, and then the reaction is stopped by adding THF and H ₂ O. To obtain a white compound represented by the chemical formula (f) (step: 1-4).

After the compound represented by the formula (to) reacting alcohol with NaI, 10 min by addition of acetonitrile (Acetonitril) and TMSCl (chlorotrimethylsilane), diisopropyl glutamate (L-diisopropyl glutamate or D-diisopropyl glutamate) and _K 2 CO ₃ was added and reacted for 5 hours, followed by fractionation with water and CH ₂ Cl ₂ . The organic layer was concentrated under reduced pressure, acetic acid and methanol were added, and the mixture was reacted at 50 ° C. for 5 hours. After concentration under reduced pressure, 1,4-dioxane, methanol and KOH were added, reacted at room temperature for 1 hour, and fractionated with water and CH ₂ Cl ₂ . The aqueous layer was acidified to pH 4.0 by adding H ₃ PO ₄ . After filtration through a filter, recrystallization from methanol gave white chemical formula SCH1702 (Chemical Formula 2) (Step 1-5). In the reaction of Step 1-5, the compound [(S) -methyl 1-((6-butoxy-2,3-dimethoxyphenanthren-10-yl) methyl) -5-oxopyrrolidine when L-diisopropylpropylate is applied] 2-carboxylate ((S) -methyl 1-((6-butyoxy-2,3-dimethylphenanthrene-10-yl) methyl) -5-oxopyrrolidene-2-carboxylate); when D-disopropyl glutamate is applied ( R) -methyl 1-((6-butoxy-2,3-dimethoxyphenanthrene-10-yl) methyl) -5-oxopyrrolidine-2-carboxylate ((R) -methyl 1-((6-butoxy-2 , 3- imethoxyphenanthren-10-yl) methyl) -5-oxopyrrolidine-2-carboxylate)] was obtained (step: 1-5).

  The compound represented by SCH1702 is the compound represented by Chemical Formula 2 of the present invention, and the NMR analysis results (1H, 13C) are shown in FIGS. 1 and 2, respectively. Moreover, the analysis result of S-form and R-form of SCH1702 using a polarizer is shown in FIG.

  Comparative Example 1: Compound of formula 1 (SCH1701)

  The compound of formula 5 (SCH1701) was used as a comparative example for comparison of activity with the SCH1702 compound produced in Production Example 1.

In the chemical formula 1, the SCH 1701 compound has a substituent of R ₁ of methyl (—CH ₃ ), and the SCH 1702 compound has a substituent of R ₁ of butyl (—Butyl).

  Example 1: Evaluation of antituberculous efficacy

  Anti-tuberculosis efficacy was double confirmed using REMA (Rezurin microtiter assay) and MGIT assay.

1) Preparation of resazurin solution

  40 mg of sterilized water was charged with 4 mg of resazurin sodium salt (sigma-aldrich) to produce a 0.01% resazurin solution, which was then filtered through a 0.45 μm filter and resazurin. The solution was prepared and used after refrigerated storage.

2) Preparation of Mycobacterium tuberculosis and resistant M. tuberculosis

H37Ra (Mycobacterium tuberculosis H37Ra) and H37rv (Mycobacterium tuberculosis H37Rv), Super tuberculosis (Extensively drug-resistant tuberculosis, XDR-TB), isoniazid resistant TB (isoniazid-resistant tuberculosis, INHr-TB), rifampin-resistant tuberculosis (rifampin-resistant tuberculosis , RIFr-TB), pyrazinamide-resistant tuberculosis (PZAr-TB), and streptomycin-resistant tuberculosis (Streptomycin-resistant tuber (Culosis, STRr-TB) was diluted with 7H9 medium (7H9 media), and the number of bacteria was prepared at a final concentration of 5 × 10 ⁵ cells / ml.

3) Assay method for antituberculous activity

  REMA was performed by the following method.

After 96 μl of 7H9 media (7H9 media) was dispensed at 100 μL, 100 μl of the compounds of Formulas 3 and 4 were added, and then serial dilution (1.5 μg / ml to 50 μg / ml) was performed, followed by M. tuberculosis strain H37Ra (Mycobacterium tuberculosis H37Ra) and H37rv (Mycobacterium tuberculosis H37rv), Super tuberculosis (Extensively drug-resistant tuberculosis, XDR-TB), isoniazid resistant TB (isoniazid-resistant tuberculosis, iNHr-TB), rifampin-resistant tuberculosis (rifampin-resistant tuberculosis, RIFr TB), pyrazinamide-resistant tuberculosis (pyrazinamide-resistant tuberculosis, PZAr- TB), streptomycin resistant TB (Streptomycin-resistant tuberculosis, after adjusting the concentration of Strr-TB) to 5 × ¹⁰ 5 cells / ml, was added in 100μL . One week later, 30 μL of the prepared resazurin solution was added to each well to confirm antituberculosis activity.

  Isoniazid (isoniadid, INH), rifampin (Rifampin, RIF), streptomycin (STReptomycin, STR), pyrazinamide (pyrazinamide, PZA), ethambutol (EMB) as the negative control group, and negative control group. An amount of DMSO was used.

  MGIT assay was performed by the following method.

Using MGIT 960 System, the ability of the compounds of Chemical Formulas 3 and 4 to inhibit the growth of Mycobacterium tuberculosis was confirmed. MGIT tube was charged with 7 ml of 7H9 media (7H9 media), 740 to 820 μL of growth supplement was added, and then the compounds of Formulas 3 and 4 were added to adjust the concentration. Subsequently, tuberculosis strains H37Ra (Mycobacterium tuberculosis H37Ra) and H37rv (Mycobacterium tuberculosis H37rv), super tuberculosis (i-T tuberculosis), rifampin-resistant tuberculosis (RIFr-TB), pyrazinamide-resistant tuberculosis (PZAr-TB), streptomycin-resistant tuberculosis (Streptomycin-resista t tuberculosis, after adjusting the concentration of Strr-TB) to 5 × ¹⁰ 5 cells / ml, was placed a 100 [mu] L, was measured daily growth of M. tuberculosis in MGIT instrument, Mycobacterium tuberculosis growth inhibitory activity of each compound It was measured.

  Isoniazid (isoniadid, INH), rifampin (Rifampin, RIF), streptomycin (STReptomycin, STR), pyrazinamide (pyrazinamide, PZA), ethambutol (EMB) as the negative control group, and negative control group. An amount of DMSO was used.

4) Anti-tuberculosis activity test results

  The results of evaluation of antituberculosis efficacy by the above experimental method are shown in the table below organized by MIC (minimum inhibitory concentration, μL / ml).

A. Inhibitory effect against tuberculosis (Ra) bacteria

  Table 1 shows the antituberculous activity of SCH1702 (S-form and R-form) against H37Ra (Mycobacterium tuberculosis H37Ra).

  The drugs used in Table 1 below are SCH1702 (S-form and R-form), SCH1701 (S-form) is a comparative example, and isoniazid (INH), rifampin (RIF). , And streptomycin (STR) were used as positive control groups.

  As a result, referring to Table 1 below, the SCH1702 compound corresponding to Chemical Formula 1 of the present invention was shown to have excellent antitubercular activity against Mycobacterium tuberculosis H37Ra. Moreover, it was confirmed that the activity of S-form in the SCH1702 compound is superior to that of R-form, and that the antituberculosis activity is more than twice that of SCH1701 (S-form) of the comparative example.

B. Inhibitory effect against tuberculosis (Rv) bacteria

  Table 2 shows antituberculous activity of SCH1702 (S-form and R-form) against H37Rv (Mycobacterium tuberculosis H37Rv).

  The drugs used in Table 2 below are SCH1702 (S-form and R-form), SCH1701 (S-form) is a comparative example, isoniazid (isoniadid, INH), rifampin (Rifampin, RIF), Streptomycin (STR), pyrazinamide (PZA), ethambutol (EMB) was used, and the same amount of DMSO was used as a negative control group.

  As a result, referring to Table 2 below, it was shown that the SCH1702 compound corresponding to the chemical formula 2 of the present invention is excellent in antituberculosis activity against Mycobacterium tuberculosis H37Rv. Moreover, it was confirmed that the activity of S-form in the SCH1702 compound is superior to that of R-form, and that the antituberculosis activity is more than twice that of SCH1701 (S-form) of the comparative example.

C. Inhibitory effect against tuberculosis (XDR) resistant bacteria

  Table 3 shows the anti-tuberculosis activity of SCH1702 (S-form and R-form) against supertubular drug-resistant tuberculosis (XDR-TB).

  The drug used in Table 3 below is SCH1702 (S-form and R-form), SCH1701 (S-form) is a comparative example, isoniazid (isoniazid, INH), rifampin (Rifampin, RIF), Streptomycin (STR), pyrazinamide (PZA), ethambutol (EMB) was used, and the same amount of DMSO was used as a negative control group.

  As a result, referring to Table 3 below, in the case of the SCH1702 compound corresponding to the chemical formula 2 of the present invention, it has excellent antitubercular activity against supertubular tuberculosis (Extensible drug-resistant tuberculosis, XDR-TB). It was shown that there is. Moreover, it was confirmed that the activity of S-form in the SCH1702 compound is superior to that of R-form, and that the antituberculosis activity is more than twice that of SCH1701 (S-form) of the comparative example.

two. Inhibitory effect against tuberculosis (INHr) resistant bacteria

  Table 4 shows the antitubercular activity of SCH1702 (S-form and R-form) against isoniazid-resistant tuberculosis (INHr-TB).

  The drugs used in Table 4 below are SCH1702 (S-form and R-form), SCH1701 (S-form) is a comparative example, isoniazid (isoniadid, INH), rifampin (RIFampin, RIF), And streptomycin (STR) were used as positive control groups.

  As a result, referring to Table 4 below, in the case of the SCH1702 compound corresponding to the chemical formula 2 of the present invention, it has excellent antitubercular activity against isoniazid-resistant tuberculosis (INHr-TB), which is a resistant tuberculosis bacterium. It was shown that there is. Moreover, it was confirmed that the activity of S-form in the SCH1702 compound is superior to that of R-form, and that the antituberculosis activity is more than twice that of SCH1701 (S-form) of the comparative example.

E. Inhibitory effect against tuberculosis (RIFr) resistant bacteria

  Table 5 shows the antituberculous activity of SCH1702 (S-form and R-form) against rifampin-resistant tuberculosis (RIFr-TB).

  The drugs used in Table 5 below are SCH1702 (S-form and R-form), SCH1701 (S-form) is a comparative example, isoniazid (isoniazid, INH), rifampin (RIFampin, RIF), And streptomycin (STR) were used as positive control groups.

  As a result, referring to Table 5 below, in the case of the SCH1702 compound corresponding to the chemical formula 2 of the present invention, the antituberculous activity against rifampin-resistant tuberculosis (RIFr-TB) which is a resistant tuberculosis bacterium is excellent. It was shown that there is. Moreover, it was confirmed that the activity of S-form in the SCH1702 compound is superior to that of R-form, and the antituberculosis activity is superior to that of SCH1701 (S-form) by two times or more.

What. Inhibitory effect against tuberculosis (PZAr) resistant bacteria

  Table 6 shows the antitubercular activity of SCH1702 (S-form and R-form) against pyrazinamide-resistant tuberculosis (PZAr-TB).

  The drugs used in Table 6 below are SCH1702 (S-form and R-form), SCH1701 (S-form) is a comparative example, isoniazid (isononiad, INH), rifampin (RIFampin, RIF), And streptomycin (STR) were used as a positive control group.

  As a result, referring to Table 6 below, in the case of the SCH1702 compound corresponding to the chemical formula 2 of the present invention, it is excellent in antituberculosis activity against pyrazinamide-resistant tuberculosis (PZAr-TB) which is a resistant tuberculosis bacterium. It was shown that Moreover, it was confirmed that the activity of S-form in the SCH1702 compound is superior to that of R-form, and that the antituberculosis activity is more than twice that of SCH1701 (S-form) of the comparative example.

G. Inhibitory effect against tuberculosis (STRr) resistant bacteria

  Table 7 shows the antitubercular activity of SCH1702 (S-form and R-form) against streptomycin-resistant tuberculosis (STRr-TB).

  The drugs used in Table 7 below are SCH1702 (S-form and R-form), SCH1701 (S-form) is a comparative example, isoniazid (isoniadid, INH), rifampin (Rifampin, RIF), And streptomycin (STR) were used as a positive control group.

  As a result, referring to Table 7 below, in the case of the SCH1702 compound corresponding to the chemical formula 2 of the present invention, it has excellent antitubercular activity against streptomycin-resistant tuberculosis (STRr-TB), which is a resistant tuberculosis bacterium. It was shown that there is. Moreover, it was confirmed that the activity of S-form in the SCH1702 compound is superior to that of R-form, and that the antituberculosis activity is more than twice that of SCH1701 (S-form) of the comparative example.

  In conclusion, Table 8 summarizes the inhibitory effect of the SCH1702 (S-form) compound on tuberculosis and resistant tuberculosis strains. That is, SCH1702 (S-form) compounds include tuberculosis strains H37Ra (Mycobacterium tuberculosis H37Ra) and H37rv (Mycobacterium tuberculosis H37rv), resistant tuberculosis strains, and Super tuberculosis (Extensive tuberculosis HDR). isonazid-resistant tuberculosis (INHr-TB), rifampin-resistant tuberculosis (RIFr-TB), pyrazinamide-resistant tuberculosis (resintant-Tuberculossis, Resistant tuberculosis) Streptomycin resistant TB (Streptomycin-resistant tuberculosis, STRr-TB) is excellent in respect to the anti-tuberculosis activity.

  Example 2: Evaluation of cytotoxicity

Cytotoxicity was measured in HP-1, Raw 264.7, L929 and HEK-293 cells associated with tuberculosis infection, proliferation. 10 × 10 ⁴ cells / well were treated with SCH1702 (S-form) in DMEM medium and cultured for 24 hours and 48 hours. As a new medium, 500 μL of medium and 5 μL of MTT solution were added to a 24-well plate (well plate) and further reacted for 4 hours. After removing the medium, 200 μL of DMSO was dissolved in formazan, and the absorbance was measured at 540 nm using a microplate reader. The result was determined that the cytotoxicity was 50% toxic effect (CC ₅₀ , This is shown in FIG. 4 in terms of 50% Cytotoxic concentration). All the measured values are shown as the average value of three repeated experiments. Referring to FIG. 4, it was confirmed that the compound of SCH1702 (S-form) has almost no cytotoxicity.

  Example 3: Evaluation of antituberculous activity against tuberculosis strains by substituents

Table 9 shows the antituberculosis activity against H37Ra (Mycobacterium tuberculosis H37Ra) by changing the substituent of R ₁ in the S-form of the compound of Chemical Formula 1.

である場合に比べて、抗結核活性が２〜４倍優れており、

又は−ＣＨ_２ＣＨ_２ＣＨ_２ＳＯ_３Ｈである場合に比べても１２８倍以上抗結核活性が著しく優れていた。結論的に、化学式１において、Ｒ_１の置換基が、炭素数が４個であるＢｕｔｙｌである場合、抗結核活性が最も優れていることを確認した。 As a result, as shown in Table 9, when the substituent of R ₁ is Butyl having 4 carbon atoms, it is smaller than 1 to 3 carbon atoms compared to methyl, ethyl and propyl groups. The antituberculosis activity was at least 2 to 4 times better. Moreover, compared with -n-Pentyl, -Hexyl, and -Heptyl having 5 to 7 carbon atoms, the antituberculosis activity was 16 to 32 times better. In addition, the substituent is

Compared to the case, the antituberculosis activity is 2 to 4 times better,

Or _{-CH 2 CH 2 CH 2 SO 3} 128 times or more anti-tuberculosis activity as compared with the case where H is significantly better. In conclusion, in Formula 1, when the substituent of R ₁ is Butyl having 4 carbon atoms, it was confirmed that the antituberculosis activity was most excellent.

Tables 10 and 11 summarize the effects of R ₁ substituents on the antituberculosis activity and cytotoxicity in Chemical Formula 1. That is, the anti-tuberculosis activity of the SCH1702 compound was the most excellent, and the cytotoxicity was also excellent.

  The preferred embodiment of the present invention has been described in detail above, but the scope of the present invention is not limited to this, and the basic concept of the present invention defined in the following claims is used. Various modifications and improvements of those skilled in the art are also within the scope of the present invention.

Claims (8)

A compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof.

A pharmaceutical composition for preventing or treating tuberculosis, comprising as an active ingredient a compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof:

The pharmaceutical composition for prevention or treatment of tuberculosis according to claim 2, which is a compound represented by the following chemical formula 3 or chemical formula 4.

  The pharmaceutical composition for prevention or treatment of tuberculosis according to claim 2, wherein the causative agent of tuberculosis is Mycobacterium tuberculosis. A pharmaceutical composition for preventing or treating resistant tuberculosis comprising a compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof as an active ingredient.

  The resistant tuberculosis is super tuberculosis (Extensible drug-resistant tuberculosis, XDR-TB), isoniazid-resistant tuberculosis, INHr-TB, rifampin-resistant tuberculosis (IFrB). 6. Tuberculosis (pyrazinamide-resistant tuberculosis, PZAr-TB) and streptomycin-resistant tuberculosis (STRr-TB) selected from the group consisting of one or more prophylactic tuberculosis-resistant tuberculosis. A therapeutic pharmaceutical composition. A food composition for preventing or improving tuberculosis comprising a compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof as an active ingredient.

A food composition for preventing or ameliorating resistant tuberculosis comprising a compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof as an active ingredient.

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