Novel febuxostat derivative and use thereof

Abstract

The present invention relates to a novel febuxostat derivative compound and a composition for the prevention or treatment of tuberculosis, comprising same as an active ingredient. The compound of the present invention can directly kill Mycobacterium tuberculosis, which is difficult to treat due to multidrug resistance (MDR) and extensive drug resistance (XDR), or can efficiently inhibit infected Mycobacterium tuberculosis through host-directed therapy mediated by macrophages of a host. Therefore, the compound of the present invention can be used alone as a therapeutic composition, and can also be administered in combination with a conventional anti-tuberculosis agent, thereby exhibiting a synergistic effect, and thus can be effectively used to improve a therapeutic effect while minimizing the dose and administration period of the anti-tuberculosis agent.

Description

Novel Febuxostat derivatives and uses thereof

[0001] The present invention relates to a novel Febuxostat derivative compound and a composition for preventing or treating tuberculosis comprising the same as an active ingredient.

[0002]

[0003] Tuberculosis (TB) is one of the three major infectious diseases threatening human health designated by the World Health Organization (WHO). It is estimated that one-third of the world's population is infected with the tuberculosis bacteria, and approximately 90% of those infected with the bacteria have a latent infection that does not show any symptoms until active tuberculosis appears, and only 10% of these patients will progress to tuberculosis during their lifetime [1]. According to the World Health Organization's 2022 Global TB Report, an estimated 10.6 million new cases of tuberculosis occurred worldwide in 2021, and approximately 1.6 million people (including 200,000 people infected with HIV) with existing tuberculosis will die from tuberculosis. Thus, the incidence and mortality rates due to tuberculosis and co-infection with HIV and tuberculosis are increasing every year.

[0004] Due to its unique characteristics, tuberculosis bacteria are resistant to most antibiotics, and their treatment methods differ from those of other bacterial infections. To date, more than a dozen anti-tuberculosis drugs have been developed and are in use. Early-stage tuberculosis treatment often results in a complete cure when these drugs are combined and taken consistently for a prescribed period. However, failure of initial treatment can lead to increased resistance, which can lead to multidrug-resistant tuberculosis (MDR-TB), making treatment difficult and reducing success rates. Tuberculosis treatment requires long-term use of multiple anti-tuberculosis drugs for six months or more. Patients are prone to irregular or premature discontinuation of these drugs, and long-term side effects and the development of MDR-TB following discontinuation pose a significant challenge. Therefore, the development of new, effective and rapid-acting treatments is urgently needed.

[0005] Currently, the first-line antituberculosis drugs recommended by the WHO, such as isoniazid (INH), rifampin (RIF), ethambutol (EMB), and pyrazinamide (PZA), are recommended for daily treatment for 2 to 4 months in patients with tuberculosis. However, patients infected with M. tuberculosis bacteria with multidrug resistance and extensive resistance to first-line antituberculosis drugs are emerging. Furthermore, although the treatment efficacy of first-line antituberculosis drugs is high, the use of first-line antituberculosis drugs such as EMB and PZA can cause side effects such as hyperuricemia and acute gouty arthritis [2, 3].

[0006] Meanwhile, research is actively underway on host-directed therapy (HDT), a treatment method that induces an increase in the immune response of a host infected with tuberculosis bacteria through metabolites that can regulate the immune / metabolic response of host cells.

[0007] The present inventors have previously elucidated the anti-tuberculosis effect of febuxostat, a representative treatment for hyperuricemia and gouty arthritis, through previous research [4], and in the present invention, they sought to discover various febuxostat derivatives that can directly kill tuberculosis bacteria or activate the immune response within host cells to kill tuberculosis bacteria.

[0008]

[0009] Numerous papers and patents are referenced and cited throughout this specification. The disclosures of these cited papers and patents are incorporated herein by reference in their entirety to provide a clearer understanding of the state of the art and the scope of the present invention.

[0010]

[0011] The present inventors have diligently researched and developed a small-molecule therapeutic agent that can effectively inhibit Mycobacterium strains, specifically Mycobacterium tuberculosis, which have poor therapeutic response to conventional antibiotics and a high possibility of developing multidrug resistance. As a result, the present invention was completed by discovering that a Febuxostat derivative compound represented by the following chemical formula 1 can effectively induce direct death of Mycobacterium tuberculosis or significantly inhibit the growth of infected Mycobacterium tuberculosis through host-directed therapy.

[0012] Accordingly, the purpose of the present invention is to provide a novel derivative compound of Febuxostat and a composition for preventing or treating tuberculosis containing the same as an active ingredient.

[0013]

[0014] Other objects and advantages of the present invention will become more apparent from the detailed description, claims and drawings below.

[0015]

[0016] According to one aspect of the present invention, the present invention provides a compound represented by the following chemical formula 1:

[0017] Chemical Formula 1

[0018]

[0019] In the above chemical formula, R1 is C1-C5 alkoxy or heterocycloalkyl of a 6-membered ring; L1 is a direct bond or -NH-; A is heteroaryl of a 5-membered or 9-membered ring which is unsubstituted or substituted with one or more substituents selected from the group consisting of C1-C5 alkyl, C1-C5 hydroxyalkyl, -COOR2 (R2 is hydrogen or C1-C3 alkyl) and NR3R4 (R3 and R4 are each independently hydrogen or C1-C7 alkyl).

[0020] The present inventors have conducted extensive research to develop a small-molecule therapeutic agent that can effectively inhibit Mycobacterium tuberculosis, a disease that has a low therapeutic response to conventional antibiotics and a high risk of developing multidrug resistance. As a result, we discovered that the compound represented by Chemical Formula 1 can effectively induce direct killing of Mycobacterium tuberculosis or significantly inhibit the growth of infected Mycobacterium tuberculosis by activating an immune response mediated by host macrophages.

[0021] The term “alkyl” as used herein means a straight-chain or branched saturated hydrocarbon group, and includes, for example, methyl, ethyl, propyl, isopropyl, etc. C1-C5 alkyl means an alkyl group having an alkyl unit having 1 to 5 carbon atoms, and when C1-C5 alkyl is substituted, the carbon number of the substituent is not included.

[0022] The term “alkoxy” in this specification means a radical formed by the removal of hydrogen from an alcohol, for example, C1-C5 alkoxy means a radical formed by the removal of hydrogen from an alcohol having 1 to 5 carbon atoms.

[0023] As used herein, the term “hydroxyalkyl” refers to a straight-chain or branched saturated hydrocarbon group substituted with a hydroxy group. C1-C5 hydroxyalkyl refers to an alkyl group in which any one hydrogen of the alkyl unit having 1 to 5 carbon atoms is replaced with a hydroxy group.

[0024] As used herein, the term “heterocycloalkyl” refers to a saturated carbon ring containing oxygen, sulfur, or nitrogen as a heteroatom within the ring. Specifically, the heteroatom is nitrogen or oxygen, and more specifically, nitrogen. The number of heteroatoms is 1-2, and specifically, 1. “Heterocycloalkyl of a six-membered ring” refers to a ring in which the sum of the numbers of carbons and heteroatoms forming the ring is 6.

[0025] According to a specific embodiment of the present invention, the heterocycloalkyl of the hexagonal ring is piperidine.

[0026] As used herein, the term “heteroaryl” refers to a heterocyclic aromatic group containing oxygen, sulfur, or nitrogen as a heteroatom within the ring. The number of heteroatoms contained within the ring is 1-3, and specifically 1-2. The term “penta- to non-penta-membered ring heteroaryl” refers to a heteroaryl having 5 to 9 ring atoms including both carbon and heteroatoms, and the ring may be a monocycle or a bicycle.

[0027] According to a specific embodiment of the present invention, A is a 5-membered-9-membered heteroaryl ring which is unsubstituted or substituted with one or two substituents selected from the group consisting of C1-C4 alkyl, C1-C3 hydroxyalkyl, -COOR2 (R2 is hydrogen or C1-C2 alkyl) and NR3R4 (R3 is hydrogen and R4 is C4-C7 alkyl).

[0028] According to a specific embodiment of the present invention, the 5-membered-9-membered heteroaryl is selected from the group consisting of thiophene, thiazole, benzothiazole, indole, benzoxazole, benzimidazole, and imidazopyridine.

[0029] According to a specific embodiment of the present invention, when A is a 5-membered-9-membered heteroaryl substituted with two substituents, A is thiazole, benzimidazole or imidazopyridine substituted with two substituents selected from the group consisting of C1-C4 alkyl, C1-C3 hydroxyalkyl, -COOR2 (R2 is hydrogen or C1-C2 alkyl) and NR3R4 (R3 is hydrogen and R4 is C4-C7 alkyl). More specifically, A is thiazole substituted with C1-C4 alkyl and C1-C3 hydroxyalkyl; imidazopyridine substituted with C1-C3 hydroxyalkyl or -COOH and NR3R4 (R3 is hydrogen and R4 is C4-C7 alkyl); or benzimidazole substituted with C1-C4 alkyl and -COOR2 (R2 is C1-C2 alkyl).

[0030]

[0031] According to a specific embodiment of the present invention, the compound represented by the chemical formula 1 is selected from the group consisting of compounds represented by the following chemical formulas 2 to 21:

[0032] Chemical Formula 2 Chemical Formula 3

[0033]

[0034] Chemical Formula 4 Chemical Formula 5

[0035]

[0036] Chemical Formula 6 Chemical Formula 7

[0037]

[0038] Chemical Formula 8 Chemical Formula 9

[0039]

[0040] Chemical Formula 10 Chemical Formula 11

[0041]

[0042] Chemical Formula 12 Chemical Formula 13

[0043]

[0044] Chemical Formula 14 Chemical Formula 15

[0045]

[0046] Chemical Formula 16 Chemical Formula 17

[0047]

[0048] Chemical Formula 18 Chemical Formula 19

[0049]

[0050] Chemical Formula 20 Chemical Formula 21

[0051] .

[0052]

[0053] According to another aspect of the present invention, the present invention provides a composition for preventing or treating a Mycobacterium infection disease, comprising a compound represented by the chemical formula 1 of the present invention as described above or a pharmaceutically acceptable salt thereof as an active ingredient.

[0054] According to another aspect of the present invention, the present invention provides a method for preventing or treating a Mycobacterium infection disease, comprising administering to a subject a composition comprising a compound represented by the chemical formula 1 of the present invention or a pharmaceutically acceptable salt thereof as an active ingredient.

[0055] The term “Mycobacterium infection disease” as used herein encompasses all clinical symptoms caused by infection with pathogens of the genus Mycobacterium, including nontuberculous or tuberculous mycobacteria, including, but not limited to, pulmonary disease, lymphadenitis, skin, soft tissue, and bone infections, and disseminated diseases, and includes all pathological conditions caused directly or indirectly by infection with pathogens of the genus Mycobacterium.

[0056] The above mycobacterium includes, but is not limited to, Mycobacterium tuberculosis (tuberculosis), Mycobacterium bovis, M. bovis BCG, M. africanum, M. canetti, M. caprae, M. microti, and M. tuberculosis K strain.

[0057] According to a specific embodiment of the present invention, the mycobacterium infection disease that can be prevented or treated with the composition of the present invention is tuberculosis (M. tuberculosis), and more specifically, active tuberculosis.

[0058] The term “prevention” as used herein means inhibiting the occurrence of a disease or condition in a subject who has not been diagnosed as having the disease or condition but is susceptible to such disease or condition.

[0059] As used herein, the term “treatment” means (a) suppressing the development of a disease, condition, or symptom; (b) alleviating the disease, condition, or symptom; or (c) eliminating the disease, condition, or symptom. When the composition of the present invention is administered to a subject, it acts to suppress the development of symptoms caused by tuberculosis infection, eliminate them, or alleviate them by directly killing tuberculosis bacteria or activating an immune response by macrophages in the infected subject. Therefore, the composition of the present invention can be a composition for treating these diseases on its own, or can be used as an adjuvant treatment for tuberculosis by being administered together with other anti-tuberculosis agents. Accordingly, the terms “treatment” or “therapeutic agent” as used herein include the meaning of “adjuvant treatment” or “adjuvant treatment agent.” Antituberculosis agents that can be administered together with the composition of the present invention include, for example, rifamficin, rifapentine, isoniazid, pyrazinamide, ethambutol, streptomycin, fluoroquinolone, kanamycin, cycloserine, prothionamide, levofloxacin, moxifloxacin, ofloxacin, rifabutin, capeomycin, amikacin, ciprofloxacin, protionamide, ethionamide, cycloserine, These include, but are not limited to, thioacetazone, clofazimine, amoxicillin / clavulanate, clarithromycin, azithromycin, and linezolid.

[0060] As used herein, the term “administration” or “administer” refers to directly administering a therapeutically effective amount of the composition of the present invention to a subject so that the same amount is formed in the body of the subject.

[0061] In the present invention, the term “therapeutically effective amount” means the content of a composition in which the pharmacological ingredient in the composition is contained in an amount sufficient to provide a therapeutic or preventive effect to a subject to whom the pharmaceutical composition of the present invention is to be administered, and includes a “prophylactically effective amount”.

[0062] The term “subject” as used herein includes, without limitation, a human, mouse, rat, guinea pig, dog, cat, horse, cow, pig, monkey, chimpanzee, baboon, or rhesus macaque. Specifically, the subject of the present invention is a human.

[0063] As used herein, the term “pharmaceutically acceptable salt” includes salts derived from pharmaceutically acceptable inorganic acids, organic acids, or bases. Examples of suitable acids include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, perchloric acid, fumaric acid, maleic acid, phosphoric acid, glycolic acid, lactic acid, salicylic acid, succinic acid, toluene-p-sulfonic acid, tartaric acid, acetic acid, trifluoroacetic acid, citric acid, methanesulfonic acid, formic acid, benzoic acid, malonic acid, naphthalene-2-sulfonic acid, benzenesulfonic acid, and the like. Salts derived from suitable bases may include alkali metals such as sodium, alkaline earth metals such as magnesium, and ammonium.

[0064] When the composition of the present invention is prepared as a pharmaceutical composition, the pharmaceutical composition of the present invention includes a pharmaceutically acceptable carrier.

[0065] Pharmaceutically acceptable carriers included in the pharmaceutical composition of the present invention are those commonly used in formulations, and include, but are not limited to, lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia gum, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. In addition to the above components, the pharmaceutical composition of the present invention may further include lubricants, wetting agents, sweetening agents, flavoring agents, emulsifiers, suspending agents, preservatives, and the like. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington's Pharmaceutical Sciences (19th ed., 1995).

[0066] The pharmaceutical composition of the present invention can be administered orally or parenterally, and specifically can be administered orally, intravenously, or intramuscularly.

[0067] The appropriate dosage of the pharmaceutical composition of the present invention may be prescribed in various ways depending on factors such as the formulation method, administration method, patient age, weight, sex, pathological condition, food, administration time, administration route, excretion rate, and response sensitivity. The preferred dosage of the pharmaceutical composition of the present invention is within the range of 0.001-100 mg / kg for adults.

[0068] The pharmaceutical composition of the present invention can be manufactured in a unit dose form or can be manufactured by inserting it into a multi-dose container by formulating it using a pharmaceutically acceptable carrier and / or excipient according to a method that can be easily performed by a person having ordinary skill in the art to which the present invention pertains, and the like. In this case, the formulation may be in the form of a solution, suspension, syrup or emulsion in an oil or aqueous medium, or in the form of an extract, powder, granule, tablet or capsule, and may additionally include a dispersing agent or stabilizer.

[0069]

[0070] According to another aspect of the present invention, the present invention provides a functional food composition for improving or preventing Mycobacterium infection disease, comprising a compound represented by the chemical formula 1 of the present invention as described above or a food-related acceptable salt thereof as an active ingredient.

[0071] The chemical formula 1 compound used in the present invention and the mycobacterium infection disease that can be improved or prevented by using the compound have already been described above, so description thereof is omitted to avoid excessive duplication.

[0072] In this specification, the term “food-acceptable salt” means a salt in a form that can be used in a food composition among salts in which cations and anions are bonded by electrostatic attraction, and specific examples thereof include the examples of “pharmaceutically acceptable salts” described above.

[0073] When the composition of the present invention is manufactured as a food composition, it may contain not only the compound of the present invention as an active ingredient, but also carbohydrates, seasonings, and flavoring agents that are commonly added during food manufacturing. Examples of carbohydrates include, but are not limited to, 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 [thaumatin, stevia extracts (e.g., rebaudioside A, glycyrrhizin, etc.]) and synthetic flavoring agents (saccharin, aspartame, etc.) can be used. For example, when the food composition of the present invention is manufactured as a drink, in addition to the pine bark extract, which is an active ingredient of the present invention, citric acid, liquid fructose, sugar, glucose, acetic acid, malic acid, fruit juice, Eucommia ulmoides extract, jujube extract, licorice extract, etc. can be additionally included.

[0074]

[0075] The features and advantages of the present invention are summarized as follows:

[0076] (a) The present invention provides a novel derivative compound of Febuxostat and a composition for preventing or treating tuberculosis containing the same as an active ingredient.

[0077] (b) The compound of the present invention can directly kill tuberculosis bacteria that are difficult to treat due to multidrug resistance (MDR) and extensive drug resistance (XDR) or can efficiently suppress tuberculosis bacteria through host-directed therapy mediated by host macrophages.

[0078] (c) Accordingly, the compound of the present invention can be used as a single therapeutic composition by itself, and can be usefully used to improve the therapeutic effect while minimizing the dosage and administration period of the anti-tuberculosis agent by co-administering with a conventional anti-tuberculosis agent to exhibit a synergistic effect.

[0079]

[0080] Figure 1 is a diagram showing the results of analyzing the drug sensitivity of tuberculosis bacteria to FBX-based derivatives discovered in the present invention using a liquid dilution method. PC: positive control; NC: negative control.

[0081] Figure 2 is a drawing showing the results of measuring the growth inhibition effect of Mycobacterium tuberculosis by the FBX derivative of the present invention in macrophages.

[0082] FIG. 3 is a drawing showing the results of confirming the synergistic effect of combined administration of the FBX derivative of the present invention and a first-line anti-tuberculosis agent in macrophages, showing a decrease in tuberculosis CFU in macrophages when rifampicin (RIF) and FBX (FIG. 3a) and FBX derivative (FT009) (FIG. 3b) were combined, respectively.

[0083] Figure 4 is a drawing showing the synergistic effect of low-concentration co-administration of PZA (Pyrazinamide), a first-line anti-tuberculosis treatment drug, and FT009, an FBX derivative of the present invention.

[0084] Figure 5 is a drawing showing the results of measuring the growth inhibition effect of seven FBX derivatives of the present invention on Mycobacterium tuberculosis in macrophages.

[0085] Figure 6 is a drawing showing the results of measuring the killing effect of FBX derivatives FT009 and FT010 of the present invention against nontuberculous mycobacteria (MAC). PC: positive control; NC: negative control.

[0086] Figure 7 is a drawing showing the results of analyzing the killing effect of FBX derivatives FT017 and FT060 of the present invention against nontuberculous mycobacteria (MAC) using the liquid dilution method (Figure 7a) and colony forming unit (CFU) (Figure 7b) measurement. PC: positive control; NC: negative control.

[0087]

[0088] Hereinafter, the present invention will be described in more detail through examples. These examples are intended solely to illustrate the present invention more specifically, and it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples, in accordance with the gist of the present invention.

[0089] Example

[0090]

[0091] Synthesis of compound FT009

[0092] To a solution of Compound A270 mg (0.78 mmol) in THF / MeOH (4 mL / 0.4 mL), 51.2 mg (2.35 mmol) of LiBH4 was added, and the mixture was stirred at room temperature for 16 hours. 10 mL of sat. NH4Cl(aq) was added dropwise to the reaction mixture, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 3:1 volume ratio) to obtain FT009170 mg (0.56 mmol, yield 72%).

[0093] 1 H NMR (600 MHz, Methanol-d4)δ8.12(d, J=2.3Hz, 1H), 8.09(dd, J=8.8, 2.3Hz, 1H), 7.25(d, J=8.9Hz, 1H), 4.75(s, 2H), 3.97(d, J=6.3Hz, 2H), 2.41(s, 3H), 2.21-2.12(m, 1H), 1.10(s, 3H), 1.09(s, 3H).

[0094]

[0095]

[0096] Synthesis of compound FT010

[0097] (i) Synthesis of compound D

[0098] Compound B 155 mg (0.51 mmol) and compound C 100 mg (0.37 mmol) were dissolved in 4 mL of 1,4-dioxane solvent, and then Pd(PPh3) 443 mg (0.037 mmol) and Na2CO3 78 mg (0.73 mmol, 2 M aqueous solution) were added and heated and stirred at 90°C for 16 hours. After cooling the reaction mixture to room temperature, 10 mL of sat. NH4Cl(aq) was added dropwise and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 6:1 volume ratio) to obtain compound D 74 mg (0.20 mmol, yield 55%).

[0099] (ⅱ) Synthesis of compound FT010

[0100] Dissolve 60 mg (0.16 mmol) of compound D in EtOH: THF: H2O (1 mL, 0.5 mL, 0.5 mL) solvent and add LiOH . 27 mg (0.65 mmol) of H2O was added and stirred at 50°C for 4 hours. After cooling the reaction mixture to room temperature, an acidic ion exchange resin was added to adjust the pH to 3–4, filtered, and the ion exchange resin was washed with EtOAc. The entire filtrate was collected and distilled under reduced pressure to obtain FT01039 mg (0.11 mmol, yield 67%).

[0101] 1H NMR (600 MHz, Chloroform-d) δ 8.56 (d, J = 1.8 Hz, 1H), 8.23 ​​(d, J = 2.3 Hz, 1H), 8.18 (dd, J = 8.9, 2.3 Hz, 1H), 8.10 (dd, J = 8.5, 1.6 Hz, 1H), 8.00 - 7.96 (m, 1H), 7.04 (d, J = 8.9 Hz, 1H), 3.87 (d, J = 6.5 Hz, 2H), 2.18 - 2.10 (m, 1H), 1.03 (s, 3H), 1.02 (s, 3H).

[0102]

[0103]

[0104] Synthesis of compound FT016

[0105] To a solution of compound D55 mg (0.15 mmol) in THF / MeOH (2 mL / 0.2 mL), 9.8 mg (0.45 mmol) of LiBH4 was added, and the mixture was stirred at room temperature for 16 hours. 10 mL of sat. NH4Cl(aq) was added dropwise to the reaction mixture, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, and the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 3:1 volume ratio z) to obtain FT01644 mg (0.13 mmol, yield 86%).

[0106] 1H NMR (400 MHz, Chloroform-d) δ 8.26 (d, J = 2.3 Hz, 1H), 8.22 (dd, J = 8.8, 2.3 Hz, 1H), 8.01 (d, J = 8.3 Hz, 1H), 7.92 (s, 1H), 7.47(dd, J = 8.5, 1.7 Hz, 1H), 7.05 (d, J = 8.8 Hz, 1H), 4.85 (s, 2H), 3.91 (d, J = 6.5 Hz, 2H), 2.28-2.14 (m, 1H), 1.98 (brs, 1H), 1.11 (s, 3H), 1.09 (s, 3H).

[0107]

[0108]

[0109] Synthesis of compound FT017

[0110] Compound E185 mg (0.91 mmol) and methyl 3,4-diaminobenzoate 182 mg (1.10 mmol) were dissolved in 10 mL of DMF, and then NaHSO3 133 mg (1.27 mmol) was added. The reaction mixture was heated and stirred at 100 °C for 16 hours. After cooling the reaction mixture to room temperature, 10 mL of water was added dropwise, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, and the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 5:1 volume ratio) to obtain FT017 230 mg (0.66 mmol, yield 72%).

[0111] 1H NMR (400MHz, DMSO-d6) δ13.27(brs, 1H), 8.49-8.43(m, 2H), 8.18(s, 1H), 7.85(dd, J=8.4, 1.6Hz, 1H), 7.68(d, J=8.5Hz, 1H), 7.48(d, J=8.9Hz, 1H), 4.03 (d, J=6.5Hz, 2H), 3.88(s, 3H), 2.16-2.06(m, 1H), 1.05(s, 3H), 1.03(s, 3H).

[0112]

[0113]

[0114] Synthesis of compound FT024

[0115] (i) Synthesis of compound H

[0116] Compound F40 mg (0.17 mmol) and compound G57 mg (0.21 mmol) were dissolved in 2 mL of DMSO, and then CuBr 0.2 mg (0.002 mmol) and TBABr 16 mg (0.17 mmol) were added. The reaction mixture was heated and stirred at 40 °C for 6 hours. After cooling the reaction mixture to room temperature, 10 mL of water was added dropwise, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 3:1 volume ratio) to obtain compound H35 mg (0.092 mmol, yield 53%).

[0117] (ⅱ) Synthesis of compound FT024

[0118] Compound H25 mg (0.066 mmol) was dissolved in 1 mL of THF solvent, the temperature was lowered to 0 °C, and 164 μL (0.164 mmol) of DIBAL-H 1.0 M in THF was added dropwise. The reaction mixture was stirred at room temperature for 4 hours, and 10 mL of sat. NH4Cl(aq) was added dropwise to the reaction mixture, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, and the solid was removed by filtering, followed by distillation under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 2:1 volume ratio) to obtain FT02410 mg (0.028 mmol, yield 43%).

[0119] 1 H NMR (600 MHz, Methanol-d4) δ8.12(d, J=2.8Hz, 1H), 7.83(dd, J=9.1, 2.8Hz, 1H), 7.69(s, 1H), 7.58(d, J=8.2Hz, 1H), 7.32(dd, J=8.3, 1.7Hz, 1H), 7.16(d, J=9.2Hz, 1H), 4.65(s, 2H), 3.90(d, J=6.4Hz, 2H), 2.16-2.10(m, 1H), 1.10(s, 3H), 1.08(s, 3H).

[0120]

[0121]

[0122] Synthesis of compound FT029

[0123] (i) Synthesis of compound K

[0124] Add 100 mg (0.49 mmol) of compound I and 99 mg (0.59 mmol) of compound J to 5 mL of toluene and give Cu(OAc) 2×After adding 1.3 mg (0.074 mmol) of H2O, the reaction mixture was heated and stirred at 110°C for 16 hours. After cooling the reaction mixture to room temperature, 10 mL of water was added dropwise, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 3:1 volume ratio) to obtain compound K88 mg (0.25 mmol, yield 51%).

[0125] (ii) Synthesis of compound FT029

[0126] Compound K30 mg (0.086 mmol) was dissolved in THF:MeOH:H2O (1 mL, 2 mL, 1 mL) solvent, NaOH 41 mg (1.03 mmol) was added, and the mixture was heated and stirred at 60°C for 12 hours. After cooling the reaction mixture to room temperature, acidic ion exchange resin was added to adjust the pH to 3-4, filtered, and the ion exchange resin was washed with EtOAc. The entire filtrate was collected and distilled under reduced pressure to obtain FT02920 mg (0.059 mmol, yield 69%).

[0127] 1 H NMR (600 MHz, Chloroform-d) δ 8.48 (d, J=2.2 Hz, 1H), 8.43 (dd, J=8.8, 2.2Hz, 1H), 8.33 (d, J=1.3 Hz, 1H), 8.17 (dd, J=8.3, 1.4 Hz, 1H), 7.80 (d, J=8.4 Hz, 1H), 7.12 (d, J=8.9 Hz, 1H), 3.94 (d, J=6.5 Hz, 2H), 2.26-2.20 (m, 1H), 1.11 (s, 3H), 1.10 (s, 3H).

[0128]

[0129]

[0130] Synthesis of compound FT033

[0131] (i) Synthesis of compound M

[0132] Compound B 245 mg (0.81 mmol), Compound L 120 mg (0.54 mmol), Pd(PPh3) 463 mg (0.054 mmol), and Na2CO3 115 mg (1.09 mmol, 2 M aqueous solution) were dissolved in 5 mL of 1,4-dioxane solvent, and stirred and heated at 90°C for 6 hours. After cooling the reaction mixture to room temperature, 10 mL of sat. NH4Cl(aq) was added dropwise, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, and the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 8:1 volume ratio) to obtain Compound M 120 mg (0.38 mmol, yield 70%).

[0133] (ⅱ) Synthesis of compound FT033

[0134] Compound M50 mg (0.16 mmol) was dissolved in THF:MeOH:H2O (1 mL, 2 mL, 1 mL), NaOH 76 mg (1.90 mmol) was added, and the mixture was heated and stirred at 60°C for 10 hours. After cooling the reaction mixture to room temperature, acidic ion exchange resin was added to adjust the pH to 3-4, filtered, and the ion exchange resin was washed with EtOAc. The entire filtrate was collected and distilled under reduced pressure to obtain FT03340 mg (0.13 mmol, yield 84%).

[0135] 1 H NMR (600 MHz, Methanol-d4) δ7.91(d, J=5.3Hz, 1H), 7.86(tt, J=7.7, 2.9 Hz, 1H), 7.69(q, J=4.0, 3.5Hz, 1H), 7.36(q, J=4.6, 3.9Hz, 1H), 7.18(t, J=8.2Hz, 1H), 3.91(d, J=6.3Hz, 2H), 2.18-2.08(m,1H), 1.07(s, 3H), 1.06(s, 3H).

[0136]

[0137]

[0138] Synthesis of compound FT034

[0139] To a solution of Compound M30 mg (0.095 mmol) in 1.5 mL of THF, 10.4 mg (0.48 mmol) of LiBH4 was added, and the mixture was heated and stirred at 55°C for 8 hours. After cooling the reaction mixture to room temperature, 10 mL of sat. NH4Cl(aq) was added dropwise, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex: EtOAc = 3:1 volume ratio) to obtain FT03415 mg (0.052 mmol, yield 55%).

[0140] 1 H NMR (600 MHz, Chloroform-d) δ 7.71 (s, 1H), 7.66 (d, J=8.9 Hz, 1H), 7.06(d, J=3.7 Hz, 1H), 6.99-6.90(m, 2H), 4.81(s, 2H), 3.84(d, J=6.4 Hz, 2H), 2.22-2.10 (m, 1H), 1.90 (brs, 1H), 1.07 (s, 3H), 1.06 (s, 3H).

[0141]

[0142]

[0143] Synthesis of compound FT041

[0144] (i) Synthesis of compound O

[0145] Compound B 358 mg (1.19 mmol), Compound N 300 mg (0.849 mmol), Pd(PPh3) 498 mg (0.085 mmol), and Na2CO3 180 mg (1.70 mmol, 2 M aqueous solution) were dissolved in 8 mL of DMF solvent and stirred at 95°C for 16 hours. After cooling the reaction mixture to room temperature, 10 mL of sat. NH4Cl(aq) was added dropwise and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, the solid was removed by filtering, and the filtrate was distilled under reduced pressure.

[0146] The obtained mixture was dissolved in 5 mL of CH2Cl2 solvent, and 1 mL of trifluoroacetic acid was added dropwise at 0°C. The reaction mixture was stirred at room temperature for 2 hours, then 10 mL of sat. NaHCO3(aq) was added dropwise, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, and the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex: EtOAc = 1:1 volume ratio) to obtain 31 mg (0.38 mmol, yield 44%) of compound O1.

[0147] (ⅱ) Synthesis of compound FT041

[0148] Dissolve 30 mg (0.086 mmol) of compound O in THF:H2O (1 mL, 1 mL) solvent and add LiOH . 15 mg (0.34 mmol) of H2O was added and stirred at 50°C for 5 hours. After cooling the reaction mixture to room temperature, an acidic ion exchange resin was added to adjust the pH to 3–4, filtered, and the ion exchange resin was washed with EtOAc. The entire filtrate was collected and distilled under reduced pressure to obtain FT04122 mg (0.066 mmol, yield 76%).

[0149] 1H NMR (600 MHz, Chloroform-d / Methanol-d4(4:1))δ8.49(d, J=1.5Hz, 1H), 7.85(dd, J=8.6, 1.7Hz, 1H), 7.76(dd, J=8.6, 2.4Hz, 1H), 7.72(d, J=2.4Hz, 1H), 7.39(d, J=8.4Hz, 1H),7.33(s, 1H), 6.99(d, J=8.7Hz, 1H), 3.82(d, J=6.4Hz, 2H), 2.18-2.07(m, 1H), 1.02(s, 3H), 1.01(s, 3H).

[0150]

[0151]

[0152] Synthesis of compound FT043

[0153] (i) Synthesis of compound Q

[0154] Compound P600 mg (2.26 mmol), 4,4,4',4',5,5,5',5'-octamethyl-2,2'-bi(1,3,2-dioxaborolane) 632 mg (2.49 mmol), and potassium acetate 444 mg (4.53 mmol) were added to 20 mL of 1,4-dioxane solvent, and 111 mg (0.1357 mmol) of Pd(dppf)Cl2 were added, and the gas inside the vessel was replaced with nitrogen for 10 minutes. The reaction mixture was heated and stirred at 100°C for 12 hours. After cooling the reaction mixture to room temperature, 20 mL of sat. NH4Cl(aq) was added dropwise, and extracted twice with 20 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, solids were removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 5:1 volume ratio) to obtain compound Q706 mg (2.26 mmol, yield 92%).

[0155] (ⅱ) Synthesis of compound S

[0156] Compound Q442 mg (1.42 mmol), compound R289 mg (1.01 mmol), Pd(PPh3)4117 mg (0.10 mmol), and Na2CO3214 mg (2.02 mmol, 2 M aqueous solution) were dissolved in 10 mL of 1,4-dioxane solvent, and stirred and heated at 90°C for 15 hours. After cooling the reaction mixture to room temperature, 10 mL of sat. NH4Cl(aq) was added dropwise, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, and the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 8:1 volume ratio) to obtain compound S186 mg (0.48 mmol, yield 47%).

[0157] (ⅲ) Synthesis of compound FT043

[0158] Dissolve compound S45 mg (0.11 mmol) in THF:H2O (1 mL, 1 mL) solvent and add LiOH . 19 mg (0.46 mmol) of H2O was added and stirred and heated at 50°C for 12 hours. After cooling the reaction mixture to room temperature, an acidic ion exchange resin was added to adjust the pH to 3–4, filtered, and the ion exchange resin was washed with EtOAc. The entire filtrate was collected and distilled under reduced pressure to obtain FT04334 mg (0.093 mmol, yield 81%).

[0159] 1 H NMR (600 MHz, Chloroform-d / Methanol-d4(4:1))δ8.56(d, J=1.6Hz, 1H), 8.19(d, J=2.3Hz, 1H), 8.10(dd, J=8.5, 1.7Hz, 1H), 8.07(dd, J=8.8, 2.3Hz, 1H), 7.96(d, J=8.5Hz, 1H), 7.04(d, J=8.8Hz, 1H), 3.35-3.30(m, 4H), 1.77-1.69(m, 4H), 1.67-1.59(m, 2H).

[0160]

[0161]

[0162] Synthesis of compound FT046

[0163] (i) Synthesis of compound U

[0164] Compound I241 mg (1.18 mmol) and compound T150 mg (0.985 mmol) were dissolved in 5 mL of MeOH solvent, and then 90 mg (1.08 mmol) of pivalonitrile and 38 mg (0.197 mmol) of p-toluenesulfonic acid monohydrate were added. The reaction mixture was stirred at room temperature for 16 hours, 10 mL of sat. NaHCO3(aq) was added dropwise, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, and the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex: EtOAc = 2:1 volume ratio) to obtain compound U227 mg (0.54 mmol, yield 55%).

[0165] (ⅱ) Synthesis of compound FT046

[0166] Compound U30 mg (0.071 mmol) was dissolved in THF: MeOH: H2O (0.5 mL, 1 mL, 0.5 mL) solvent, NaOH 34 mg (0.86 mmol) was added, and the mixture was heated and stirred at 60°C for 10 hours. After cooling the reaction mixture to room temperature, acidic ion exchange resin was added to adjust the pH to 3–4, filtered, and the ion exchange resin was washed with EtOAc. The entire filtrate was collected and distilled under reduced pressure to obtain FT04622 mg (0.054 mmol, yield 76%).

[0167] 1H NMR (600 MHz, Chloroform-d / Methanol-d4(4:1))δ8.97-8.88(m, 1H), 8.21(d, J=2.3Hz, 1H), 8.12(dd, J=8.9, 2.2Hz, 1H), 7.73(d, J=9.3Hz, 1H), 7.47 (d, J=9.1Hz, 1H), 7.01(d, J=8.9Hz, 1H), 3.83(d, J=6.5Hz, 2H), 2.18-2.03(m, 1H), 1.03(s, 3H), 1.02(s, 3H), 1.01(s, 9H).

[0168]

[0169]

[0170] Synthesis of compound FT047

[0171] Compound U40 mg (0.095 mmol) was dissolved in 3 mL of THF solvent, the temperature was lowered to 0°C, and 285 μL (0.285 mmol) of DIBAL-H 1.0 M in THF was added dropwise. The reaction mixture was stirred at room temperature for 4 hours, and 10 mL of sat. NH4Cl(aq) was added dropwise to the reaction mixture, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex: EtOAc = 1:1 volume ratio) to obtain FT04719 mg (0.043 mmol, yield 52%).

[0172] 1H NMR (600 MHz, Chloroform-d) δ 8.25 (d, J = 2.2 Hz, 1H), 8.16 (dd, J = 8.8, 2.2Hz, 1H), 8.07-8.05 (m, 1H), 7.39 (dd, J = 9.1, 0.9 Hz, 1H), 7.10 (dd, J = 9.2, 1.7 Hz, 1H), 6.96 (d, J = 8.9 Hz, 1H), 4.65(s, 2H), 3.84(d, J = 6.5 Hz, 2H), 2.21-2.10(m, 1H), 1.08(s, 3H), 1.06(s, 3H), 1.02(s, 9H).

[0173]

[0174]

[0175] Synthesis of compound FT049

[0176] (i) Synthesis of compound V

[0177] Compound I 147 mg (0.723 mmol) and compound T 100 mg (0.657 mmol) were dissolved in 5 mL of MeOH solvent, and then 2-isocyano-2,3,3-trimethylbutane 107 mg (0.855 mmol) and p-toluenesulfonic acid monohydrate 25 mg (0.131 mmol) were added. The reaction mixture was stirred at room temperature for 16 hours, then 10 mL of sat. NaHCO3(aq) was added dropwise, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, and the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 2:1 volume ratio) to obtain compound V 201 mg (0.434 mmol, yield 66%).

[0178] (ⅱ) Synthesis of compound FT049

[0179] Dissolve 50 mg (0.11 mmol) of compound V in THF:H2O (1 mL, 1 mL) solvent and add LiOH .18 mg (0.43 mmol) of H2O was added and stirred and heated at 50°C for 12 hours. After cooling the reaction mixture to room temperature, an acidic ion exchange resin was added to adjust the pH to 3–4, filtered, and the ion exchange resin was washed with EtOAc. The entire filtrate was collected and distilled under reduced pressure to obtain FT04935 mg (0.078 mmol, yield 72%).

[0180] 1 H NMR (600 MHz, Chloroform-d / Methanol-d4(4:1))δ8.98(s, 1H), 8.14(d, J=2.1Hz, 1H), 8.05(dd, J=8.8, 2.2Hz, 1H), 7.77-7.69(m, 1H), 7.50-7.42(m, 1H), 7.04(d, J=8.9Hz, 1H), 3.85(d, J=6.4Hz, 2H), 2.19-2.08(m, 1H), 1.04(s, 3H), 1.03(s, 3H), 0.99(s, 9H), 0.93(s, 6H).

[0181]

[0182]

[0183] Synthesis of compound FT050

[0184] Compound V50 mg (0.11 mmol) was dissolved in 2 mL of THF solvent, the temperature was lowered to 0°C, and 324 μL (0.324 mmol) of DIBAL-H 1.0 M in THF was added dropwise. The reaction mixture was stirred at room temperature for 16 hours, and 10 mL of sat. NH4Cl(aq) was added dropwise to the reaction mixture, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex: EtOAc = 2:1 volume ratio) to obtain FT05015 mg (0.035 mmol, yield 32%).

[0185] 1H NMR (600 MHz, Chloroform-d) δ 8.19 (dd, J = 7.2, 1.8 Hz, 2H), 8.14 (dd, J = 8.7, 2.2 Hz, 1H), 7.49 (d, J = 9.1 Hz, 1H), 7.18 (dd, J = 9.1, 1.6 Hz, 1H), 7.01(d, J = 8.8 Hz, 1H), 4.72(s, 2H), 3.88(d, J = 6.5 Hz, 2H), 2.22-2.16 (m, 1H), 1.27-1.23 (m, 6H), 1.10(s, 3H), 1.08(s, 3H), 1.04(s, 9H).

[0186]

[0187]

[0188] Synthesis of compound FT051

[0189] Compound B 280 mg (0.930 mmol), compound W 179 mg (0.664 mmol), Pd(PPh3) 477 mg (0.066 mmol), and Na2CO3 141 mg (1.33 mmol, 2 M aqueous solution) were dissolved in 7 mL of 1,4-dioxane solvent, and stirred and heated at 90°C for 12 hours. After cooling the reaction mixture to room temperature, 10 mL of sat. NH4Cl(aq) was added dropwise, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, and the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 4:1 volume ratio) to obtain FT051123 mg (0.339 mmol, yield 51%).

[0190] 1H NMR (600 MHz, Chloroform-d) δ 8.25 (s, 1H), 8.23 ​​(s, 1H), 7.75 (d, J = 9.3 Hz, 1H), 7.73 (d, J = 2.4 Hz, 1H), 7.68 (dd, J = 8.4, 2.1 Hz, 1H), 7.41 (dd, J = 9.4, 1.8 Hz, 1H), 7.05 (dd, J = 8.8, 2.1 Hz, 1H), 4.46 (q, J = 7.1 Hz, 2H), 3.88 (d, J = 6.5 Hz, 2H), 2.24 - 2.15 (m, 1H), 1.43 (t, J = 7.1 Hz, 3H), 1.08 (d, J = 6.7 Hz, 6H).

[0191]

[0192]

[0193] Synthesis of compound FT053

[0194] FT05145 mg (0.124 mmol) was dissolved in 2 mL of THF, and 8.1 mg (0.371 mmol) of LiBH4 was added at room temperature and stirred at 55°C for 8 hours. 10 mL of sat. NH4Cl(aq) was added dropwise to the reaction mixture, and extracted twice with 10 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (Hex:EtOAc = 3:1 volume ratio) to obtain FT05321 mg (0.067 mmol, yield 54%).

[0195] 1H NMR (600 MHz, Chloroform-d / Methanol-d4 (4:1)) δ 8.29(s, 1H), 7.72- 7.58(m, 3H), 7.58-7.49(m, 1H), 7.44-7.33(m, 1H), 7.06-6.97(m, 1H), 4.72(s, 2H), 3.82(d, J=11.7 Hz, 2H), 2.20-2.04(m, 1H), 1.02(s, 3H), 0.99(s, 3H).

[0196]

[0197]

[0198] Synthesis of compound FT060

[0199] FT017450 mg (1.32 mmol) was dissolved in 10 mL of DMF solvent, then the temperature was lowered to 0℃, and Cs2CO3629 mg (1.93 mmol) and iodoethane155 μL (0.193 mmol) were added dropwise. The reaction mixture was stirred at room temperature for 16 hours, and the solvent was removed by distillation under reduced pressure. 10 mL of water was added dropwise to the obtained mixture, and it was extracted twice with 10 mL of CH2Cl2. The obtained CH2Cl2 layer was dried over MgSO4, and the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (CHCl3: MeOH = 100:1 volume ratio) to obtain 140 mg (0.39 mmol, yield 30%) of compound X and 150 mg (0.40 mmol, yield 31%) of FT060.

[0200] 1H NMR (600 MHz, Chloroform-d) δ 8.46(s, 1H), 8.01(dd, J=8.5, 1.4 Hz, 1H), 7.89(s, 1H), 7.88(d, J=2.3 Hz, 1H), 7.40(d, J=8.5 Hz, 1H), 7.08(d, J=9.4 Hz, 1H), 4.26(q, J=7.3 Hz, 2H), 3.92(s, 3H), 3.89(d, J=6.5 Hz, 2H), 2.23-2.14 (m, 1H), 1.45(t, J=7.2 Hz, 3H), 1.08(s, 3H), 1.07 (s, 3H).

[0201]

[0202]

[0203] Synthesis of compounds FT064 and FT065

[0204] FT01740 mg (0.114 mmol) was dissolved in 2 mL of DMF solvent, the temperature was lowered to 0 °C, and 112 mg (0.34 mmol) of Cs2CO3, 19 mg (0.11 mmol) of potassium iodide, and 37 μL (0.34 mmol) of isobutyl bromide were added dropwise. The reaction mixture was heated and stirred at 80 °C for 16 hours, and the solvent was removed by distillation under reduced pressure. 10 mL of water was added dropwise to the obtained mixture, and it was extracted twice with 10 mL of CH2Cl2. The obtained CH2Cl2 layer was dried over MgSO4, the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (CHCl3: MeOH = 100:1 volume ratio) to obtain FT0648 mg (0.020 mmol, yield 17%) and FT06512 mg (0.030 mmol, yield 26%).

[0205] FT064: 1H NMR (600 MHz, Chloroform-d) δ 8.17(d, J = 1.5 Hz, 1H), 8.04 (dd, J = 8.5, 1.5 Hz, 1H), 8.00-7.95(m, 1H), 7.92(d, J = 2.2 Hz, 1H), 7.84 (d, J = 8.5 Hz, 1H), 7.13(d, J = 8.8 Hz, 1H), 4.14(d, J = 7.7 Hz, 2H), 3.97 (s, 3H), 3.93(d, J = 6.5 Hz, 2H), 2.27-2.19(m, 1H), 2.19-2.12(m, 1H), 1.11 (s, 3H), 1.10 (s, 3H), 0.78 (s, 3H), 0.76 (s, 3H).

[0206] FT065: 1 H NMR (600 MHz, Chloroform-d) δ 8.50 (d, J = 1.2 Hz, 1H), 8.05 (dd, J = 8.6, 1.5 Hz, 1H), 7.97-7.88 (m, 2H), 7.43 (d, J = 8.6 Hz, 1H), 7.11 (d, J = 8.7 Hz, 1H), 4.09 (d, J = 7.6 Hz, 2H), 3.95(s, 3H), 3.93 (d, J = 6.5 Hz, 2H), 2.26-2.17(m, 1H), 2.15-2.06(m, 1H), 1.11(s, 3H), 1.10(s, 3H), 0.77 (s, 3H), 0.76 (s, 3H).

[0207]

[0208]

[0209] Synthesis of compound FT073

[0210] (i) Synthesis of compound C

[0211] Compound A 300 mg (1.79 mmol) and compound B 303 mg (1.50 mmol) were dissolved in 15 mL of toluene solvent, and then 0.04 mg (0.22 mmol) of Cu(OAc)2·H2O was added and stirred while heating at 110°C in air for 16 hours. After the reaction mixture was cooled to room temperature, 15 mL of water was added dropwise and extracted twice with 15 mL of EtOAc. The obtained EtOAc layer was dried over MgSO4, the solid was removed by filtering, and the filtrate was distilled under reduced pressure. The obtained mixture was separated and purified by silica gel column chromatography (CH2Cl2 single solvent) to obtain compound C 150 mg (0.43 mmol, yield 29%).

[0212] (ii) Synthesis of compound FT073

[0213] Compound C30 mg (0.08 mmol) was dissolved in MeOH:THF (1 mL, 1.5 mL) solvent, 0.51 mL (1.03 mmol) of 2M NaOH aqueous solution was added, and the mixture was heated and stirred at 60°C for 4 hours. Acidic ion exchange resin was added to the reaction mixture to adjust the pH to 3-4, filtered, and the ion exchange resin was washed with EtOAc. The obtained mixture was separated and purified by silica gel column chromatography (CHCl3:MeOH = 15:1 volume ratio) to obtain FT07328 mg (0.08 mmol, yield 97%).

[0214] 1 H NMR (600 MHz, Methanol-d4)δ 8.48-8.39(m, 2H), 8.34(d, J=1.6 Hz, 1H), 8.11(dd, J=8.5, 1.7 Hz, 1H), 7.73(d, J = 8.5 Hz, 1H), 7.36(d, J=8.6) Hz, 1H), 4.02(d, J=6.4 Hz, 2H), 2.23-2.15(m, 1H), 1.11(d, J=6.7 Hz, 6H).

[0215] Tuberculosis direct killing compound derivative molecular weight structure FT009 (chemical formula 2) 5-(5-(hydroxymethyl)-4-methylthiazol-2-yl)-2-isobutoxybenzonitrile FT010 (chemical formula 3) 2-(3-cyano-4-isobutoxyphenyl)benzo[d]thiazole-6-carboxylic acid FT016 (chemical formula 4) 5-(6-(hydroxymethyl)benzo[d]thiazol-2-yl)-2-isobutoxybenzonitrile FT024 (formula 5) 5-((6-(hydroxymethyl)benzo[d]thiazol-2-yl)amino)-2-isobutoxybenzonitrileFT029 (chemical formula 6) 2-(3-cyano-4-isobutoxyphenyl)benzo[d]oxazole-6-carboxylic acid FT034 (chemical formula 7) 5-(5-(hydroxymethyl)thiophen-2-yl)-2-isobutoxybenzonitrile FT043 (formula 8) 2-(3-cyano-4-(piperidin-1-yl)phenyl)benzo[d]thiazole-6-carboxylic acid FT047 (formula 9) 5-(3-(tert-butylamino)-6-(hydroxymethyl)imidazo[1,2-a]pyridin-2-yl)-2-isobutoxybenzonitrileFT049 (Formula 10) 2-(3-cyano-4-isobutoxyphenyl)-3-((2,3,3-trimethylbutan-2-yl)amino)imidazo[1,2-a]pyridine-6-carboxylic acid FT050 (Formula 11) 5-(6-(hydroxymethyl)-3-((2,3,3-trimethylbutan-2-yl)amino)imidazo[1,2-a]pyridin-2-yl)-2-isobutoxybenzonitrileFT051 (Formula 12) Ethyl 6-(3-cyano-4-isobutoxyphenyl)imidazo[1,2-a]pyridine-2-carboxylate FT053 (Formula 13) 5-(2-(hydroxymethyl)imidazo[1,2-a]pyridin-6-yl)-2-isobutoxybenzonitrileFT073 (chemical formula 21) 2-(3-cyano-4-isobutoxyphenyl)benzo[d]oxazole-5-carboxylic acid

[0216]

[0217] Host-directed therapeutic compound derivative molecular structure FT017 (chemical formula 14) Methyl 2-(3-cyano-4-isobutoxyphenyl)-1H-benzo[d]imidazole-6-carboxylate FT033 (Formula 15) 5-(3-cyano-4-isobutoxyphenyl)thiophene-2-carboxylic acid FT041 (chemical formula 16) 3-(3-cyano-4-isobutoxyphenyl)-1H-indole-5-carboxylic acid FT046 (chemical formula 17) 3-(tert-butylamino)-2-(3-cyano-4-isobutoxyphenyl)imidazo[1,2-a]pyridine-6-carboxylic acid FT060 (chemical formula 18) Methyl 2-(3-cyano-4-isobutoxyphenyl)-1-ethyl-1H-benzo[d]imidazole-5-carboxylate FT064 (Formula 19) Methyl 2-(3-cyano-4-isobutoxyphenyl)-1-isobutyl-1H-benzo[d]imidazole-6-carboxylate FT065 (Formula 20) Methyl 2-(3-cyano-4-isobutoxyphenyl)-1-isobutyl-1H-benzo[d]imidazole-5-carboxylate

[0218]

[0219] Preparation Example 1: Culturing of tuberculosis bacteria

[0220] To cultivate the standard strain of tuberculosis (ATCC_27294), Middlebrook 7H9 liquid medium containing 10% OADC was used, and the medium was cultured at 37°C with stirring after inoculating the strain. The cultured tuberculosis bacteria were cultured until the absorbance at 600 nm reached 0.3 - 0.5, and the absorbance was 0.005 (approximately 5 x 10 6 A MIC (Minimal Inhibitory Concentration) test was performed to analyze the inhibition of microbial growth by diluting the solution to 10 CFU / ml.

[0221]

[0222] Preparation Example 2: Measurement of minimal inhibitory concentrations (MICs)

[0223] The tuberculosis strain produced in Preparation Example 1 was treated with rifampicin (RIF), febuxostat (FBX), and FBX-based derivatives discovered in the present invention based on the Clinical and Laboratory Standards Institute (CLSI) guidelines and the Resazurin microtiter assay (REMA), and the susceptibility of the tuberculosis strain to each drug was analyzed using the liquid dilution method using 7H9 liquid medium (Fig. 1). The group treated with rifampicin antibiotics at a concentration of 0.15 to 78 μM (0.125 to 64 μg / ml) was used as a positive control group. The susceptibility test was performed with FBX in the concentration range of 632 to 0.6 μM (0.39 to 200 μg / ml), and with FBX-based derivatives in the concentration range of about 600 to 1.2 μM.

[0224] 100 μl was dispensed per well in a 96-well plate, and the absorbance at 600 nm was 0.005 (approximately 5 x 10 6 The strains diluted to 10 CFU / ml were additionally inoculated into each well at 100 μl and cultured at 37°C for 2 weeks. At this time, a group cultured only with medium inoculated with neither tuberculosis bacilli nor drugs (Negative Control group; NC) was cultured as a negative control group, a group cultured only with tuberculosis bacilli (Positive Control group; PC) was cultured as a positive control group, and a group administered rifampicin to tuberculosis bacilli was cultured as a drug control group. After 2 weeks of culture, 22 μl of a color-developing solution (0.02% resazurin solution) was added to each well and cultured again at 37°C for 24 hours. The antibacterial activity of the drug treated 24 hours later was compared with the negative control group, positive control group, and drug control group.

[0225]

[0226] Preparation Example 3: Induction of differentiation of bone marrow-derived macrophages

[0227] After isolating bone marrow cells from the mouse bone marrow, differentiation medium was prepared by adding 10% L929 cell line culture medium to high-glucose DMEM (Biowest, France) culture medium containing 10% fetal bovine serum (Biowest, France) and 1% penicillin / streptomycin (Biowest, France). Then, 10 ml of the differentiation medium was placed in a 90 x 15 mm petri dish (SPL life science, Korea) and cultured for 3 days in an incubator under 5% CO2 and 37°C conditions. Then, macrophages were obtained by adding an additional 10 ml of the differentiation medium to the petri dish and further culturing for 6 to 7 days. In subsequent experiments, the macrophages were separated using trypsin-EDTA (Biowest, France) and then 2 x 10 cells were seeded into each well of a 48-well cell culture plate. 5 Cells were seeded and cultured for 24 hours to allow sufficient attachment of macrophages to the plate.

[0228]

[0229] Preparation Example 4: Preparation of 7H10 solid medium for culturing tuberculosis bacteria

[0230] 9.5g of Difco in a 1L Erlenmeyer flask TM Middlebrook 7H10 agar (BD bioscience, USA) powder was added, 450 mL of distilled water was added, and mixed well. After sterilization using an autoclave at 121℃ for 15 minutes, it was cooled sufficiently to 60℃, 50 mL of OADC (oleic acid + albumin + dextrose + catalase; BD bioscience, USA) was added, and 23 mL was added to each 90 x 15 mm petri dish (SPL life science, Korea), and left at room temperature for one day to solidify sufficiently, and then stored in the refrigerator until used in future experiments.

[0231]

[0232] Example 1: Drug susceptibility test (MIC) for Febuxostat-based derivatives

[0233] In the present invention, the susceptibility to Mycobacterium tuberculosis was evaluated using the febuxostat-based derivatives discovered in accordance with the method described in Preparation Example 2. As a result, as shown in Fig. 1, it was confirmed that a total of 13 derivatives (FT009, FT010, FT016, FT024, FT029, FT034, FT043, FT047, FT049, FT050, FT051 FT053, FT073) directly inhibited the growth of Mycobacterium tuberculosis (H37Rv ATCC 27294) at a lower concentration than febuxostat.

[0234]

[0235] Example 2: Confirmation of the inhibitory effect of five derivatives on the growth of Mycobacterium tuberculosis in macrophages.

[0236] In the above Manufacturing Example 3, Mycobacterium tuberculosis (H37Rv ATCC 27294) was added to each well of the macrophages attached to the 48-well plate at an MOI (Multiplicity of infection) of 1:1 and cultured for 4 hours, and then the culture medium containing the tuberculosis bacteria was removed. The differentiation medium of Manufacturing Example 3 was applied to each well of the plate containing 5 kinds of derivatives and cultured for 4 hours or 72 hours. Here, only the differentiation medium was used as the control group (pi_72h). After the culture was completed, each well of the plate was washed with 1 x PBS (phosphate buffered saline), and then 200 ㎕ of 0.05% Triton X-100 was added to each well and cultured for 10 minutes to sufficiently lyse the membrane of the macrophages. Thereafter, the lysate released from the macrophages was diluted by a factor of 1 / 100 or 1 / 1000, and 50 μl of the diluted lysate was dispensed into each petri dish of the 7H10 solid medium of Preparation Example 4, and cultured in a microbiological incubator for 2 weeks. After completion of culture, the number of bacteria (CFU) generated in the petri dish was measured.

[0237] As a result of evaluating cytotoxicity according to the concentration of each derivative, as shown in Fig. 2, among 10 derivatives (FT009_30uM, FT010_80uM, FT016_10uM, FT024_2.5uM, FT029_80uM, FT034_40uM, FT043_80uM, FT051_80uM, FT053_80uM), compared to the control group (pi_72h), 7 derivatives (FT009, FT016, FT024, FT034, FT043, FT051, FT053) inhibited the growth of tuberculosis bacilli, and in particular, it was confirmed that the growth of tuberculosis bacilli could be significantly reduced even when a lower concentration than febuxostat 316μM (100ug / ml) was used.

[0238] The above results show that the febuxostat derivatives of the present invention can very effectively inhibit the growth of strains infected with a host by increasing the antibacterial action using low concentrations in macrophages.

[0239]

[0240] Example 3: Synergistic effect of combined administration of FBX derivative FT009 and first-line antituberculosis drugs in macrophages

[0241] In order to confirm the inhibitory effect of tuberculosis growth by co-administration of existing first-line tuberculosis treatment agents and one derivative of the present invention (FT009), the synergistic effect with febuxostat (FBX) or FT009 derivatives was confirmed by co-administering each antibiotic at different concentrations in macrophages infected with tuberculosis using the same method as in Example 2. As a result, in the case of FBX, there was no synergistic effect in any combination of existing tuberculosis treatment agents, and rather, the anti-tuberculosis effect was reduced when co-administered compared to monotherapy with rifampicin (RIF) (Fig. 3a). In contrast, in macrophages co-administered with FT009 and existing standard treatment agents, a significant decrease in tuberculosis CFU corresponding to a synergistic effect was observed in all antibiotic combinations (isoniazid; INH, Ethambutol; EMB, Pyrazinamide; PZA) except for RIF.

[0242] Furthermore, as shown in Fig. 4, it was observed that the CFU of tuberculosis bacteria was significantly reduced even when the FT009 derivative and existing standard therapeutic agents were administered together at low concentrations, thereby finally confirming the synergistic effect between the FBX derivative of the present invention and existing anti-tuberculosis drugs. Through these results, the FT009 derivative of the present invention was confirmed to have the potential as an innovative therapeutic agent that can significantly inhibit the growth of tuberculosis bacteria even when the currently used anti-tuberculosis drugs are administered at low doses, thereby reducing the amount of highly toxic anti-tuberculosis drugs used while further enhancing the anti-tuberculosis effect.

[0243]

[0244] Example 4: Inhibition of tuberculosis growth by five derivatives in macrophages

[0245] In the above Preparation Example 3, Mycobacterium tuberculosis was added to each well of the macrophages attached to the 48-well plate at an MOI of 1:1 and cultured for 4 hours, after which the culture medium containing the tuberculosis bacteria was removed. The differentiation medium of the above Preparation Example 3 of the plate containing 5 types of derivatives was applied to each well and cultured for 4 hours or 72 hours. Here, only the differentiation medium was used as the control group (pi_72h). After the culture was completed, each well of the plate was washed with 1 x PBS, 200 ㎕ of 0.05% Triton X-100 was added to each well, and the plate was cultured for 10 minutes to sufficiently dissolve the membrane of the macrophages, and the lysate released from the macrophages was diluted by a factor of 1 / 100 or 1 / 1000, and 50 ㎕ of the diluted lysate was dispensed into each petri dish of the 7H10 solid medium of Preparation Example 4, and cultured in a microbial incubator for 2 weeks. After the culture was completed, the number of bacteria (CFU) generated in the petri dish was measured. As a result, as shown in Fig. 5, compared to the control group (pi_72h), the seven derivatives (FT017_0.5μM, FT033_80μM, FT034_80μM, FT046_80μM, FT060_20μM, FT064_30μM, FT065_80μM) reduced the growth of tuberculosis bacilli, and in particular, it was confirmed that the growth of tuberculosis bacilli could be significantly reduced even at concentrations significantly lower than febuxostat 316uM (100ug / ml) (FT017: 632-fold, FT033, 041, 046, 065: 4-fold, FT060: 16-fold, FT064: 10-fold lower concentration).

[0246] Furthermore, two derivatives (FT009, FT010) having a direct tuberculosis killing effect and two derivatives (FT017, FT060) among host-directed therapeutic substances were examined to determine whether they exhibited direct killing effects and host-directed therapeutic effects not only on tuberculosis but also on two standard strains of non-tuberculous mycobacteria (Mycobacterium avium Complex, MAC) (M. avium ATCC 700898 and M. intracellulare ATCC 13950). As a result, it was confirmed that not only direct inhibition by the derivatives but also no effect as a host-directed therapeutic substance was observed, indicating that the bacterial killing effect by the derivatives of the present invention is specific to tuberculosis (Figs. 6 and 7).

[0247] Through the above results, it can be seen that the febuxostat-based derivatives FT017, FT033, FT034, FT046, FT060, FT064, and FT065 discovered in the present invention can very effectively inhibit the growth of tuberculosis bacteria infected in a host through antibacterial activity formed by regulating the immune response within macrophages.

[0248]

[0249] While specific aspects of the present invention have been described in detail above, it should be apparent to those skilled in the art that these specific descriptions are merely preferred embodiments and do not limit the scope of the present invention. Therefore, the substantial scope of the present invention is defined by the appended claims and their equivalents.

[0250]

[0251] References

[0252] 1. Wipperman, MF, NS Sampson, and ST Thomas, Crit Rev Biochem Mol Biol, 49(4): 269-93 (2014).

[0253] 2. Shapiro, M. and L. Hyde,Am J Med,23(4): 596-599(1957).

[0254] 3. Postlethwaite, A.E., A.G. Bartel, and W.N. Kelley,N Engl J Med,286(14):761-762(1972).

[0255] 4. Kim, L.H., et al.,Antimicrob Agents Chemother,2022. 66(9): p. e0076222.

Claims

1. A compound represented by the following chemical formula 1: Chemical Formula 1 In the above chemical formula, R1 is C1-C5 alkoxy or heterocycloalkyl of a 6-membered ring; L1 is a direct bond or -NH-; A is heteroaryl of a 5-membered or 9-membered ring which is unsubstituted or substituted with one or more substituents selected from the group consisting of C1-C5 alkyl, C1-C5 hydroxyalkyl, -COOR2 (R2 is hydrogen or C1-C3 alkyl) and NR3R4 (R3 and R4 are each independently hydrogen or C1-C7 alkyl).

2. A compound according to claim 1, characterized in that the heterocycloalkyl of the hexagonal ring is piperidine.

3. A compound characterized in that in paragraph 1, A is a heteroaryl of a 5-membered-9-membered ring, which is unsubstituted or substituted with one or two substituents selected from the group consisting of C1-C4 alkyl, C1-C3 hydroxyalkyl, -COOR2 (R2 is hydrogen or C1-C2 alkyl), and NR3R4 (R3 is hydrogen and R4 is C4-C7 alkyl).

4. A compound characterized in that the heteroaryl of the 5-membered-9-membered ring in the first paragraph is selected from the group consisting of thiophene, thiazole, benzothiazole, indole, benzoxazole, benzimidazole, and imidazopyridine.

5. In the third paragraph, when A is a 5-membered-9-membered heteroaryl substituted with two substituents, a compound characterized in that A is a thiazole, benzimidazole or imidazopyridine substituted with two substituents selected from the group consisting of C1-C4 alkyl, C1-C3 hydroxyalkyl, -COOR2 (R2 is hydrogen or C1-C2 alkyl) and NR3R4 (R3 is hydrogen and R4 is C4-C7 alkyl).

6. A compound characterized in that in the 5th paragraph, when A is a 5-membered-9-membered heteroaryl substituted with two substituents, A is a thiazole substituted with C1-C4 alkyl and C1-C3 hydroxyalkyl; an imidazopyridine substituted with C1-C3 hydroxyalkyl or -COOH and NR3R4 (R3 is hydrogen and R4 is C4-C7 alkyl); or a benzimidazole substituted with C1-C4 alkyl and -COOR2 (R2 is C1-C2 alkyl).

7. In the first paragraph, the compound represented by the chemical formula 1 is a compound characterized in that it is selected from the group consisting of compounds represented by the following chemical formulas 2 to 21: Chemical Formula 2 Chemical Formula 3 Chemical Formula 4 Chemical Formula 5 Chemical Formula 6 Chemical Formula 7 Chemical Formula 8 Chemical Formula 9 Chemical Formula 10 Chemical Formula 11 Chemical Formula 12 Chemical Formula 13 Chemical Formula 14 Chemical Formula 15 Chemical Formula 16 Chemical Formula 17 Chemical Formula 18 Chemical Formula 19 Chemical Formula 20 Chemical Formula 21 .

8. A composition for preventing or treating a Mycobacterium infection disease, comprising a compound of any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof as an active ingredient.

9. A composition according to claim 8, characterized in that the Mycobacterium infection disease is tuberculosis (M. tuberculosis).

10. A functional food composition for improving or preventing Mycobacterium infection disease, comprising a compound of any one of claims 1 to 7 or a food-related salt thereof as an active ingredient.

11. A composition according to claim 10, wherein the Mycobacterium infection disease is tuberculosis (M. tuberculosis).

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