Novel compound and composition containing the same as an active ingredient for preventing, improving or treating bacterial inflammation and inflammasome-mediated diseases

Abstract

The present application relates to a compound of Chemical Formula 1, its isomer, or a pharma- ceutically acceptable salt thereof, and its use in the prevention, amelioration, and / or treatment of inflammasome-mediated diseases, and / or diseases caused by or infected with Gram-negative bacteria, which is safe without causing cytotoxicity, can effectively inhibit inflammasomes, and has excellent preventive, ameliorative, and / or therapeutic effects against the above diseases.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority based on Korean Patent Application No. 10-2022-0022320 dated February 21, 2022, Korean Patent Application No. 10-2022-0140639 dated October 27, 2022, Korean Patent Application No. 10-2022-0146145 dated November 4, 2022, and Korean Patent Application No. 10-2023-0022804 dated February 21, 2023, and all contents disclosed in the documents of the Korean Patent Applications are incorporated herein by reference. [Background technology]

[0002] Gram-negative bacteria are known to cause periodontal disease, urinary tract infection, cystitis, pyelonephritis, abdominal cavity infection, pneumonia, etc., and are highly virulent in the human body, often leading to secondary infections such as hospital infections and opportunistic infections. If bacteria that infect localized areas like this cannot be properly controlled in the early stages, the virulence of the bacteria increases, eventually circulating in the bloodstream and causing a systemic inflammatory response (sepsis), which can lead to organ failure and even death from shock.

[0003] Gram-negative bacteria have an outer membrane that Gram-positive bacteria do not have, so it is difficult for drugs to penetrate, and because of the bacterial membrane developed by quorum sensing in Gram-negative bacteria, high concentrations of antibiotics are required, and the effective period of the drug after administration is short, which can lead to problems of antibiotic overuse and resistance. However, by using a quorum sensing inhibitor to inhibit the formation of the bacterial membrane of Gram-negative bacteria and control the virulence of the bacteria, it is possible to fundamentally prevent the progression to related diseases (see Figures 9 and 10).

[0004] In particular, when a quorum sensing inhibitor is administered together with an antibiotic, it can counteract the bacterial membrane effect and increase the effectiveness of the antibiotic, thereby lowering the effective concentration and prolonging the effective period, thereby increasing the antibiotic effect and minimizing the induction of drug resistance.

[0005] Currently, there is a need to develop new therapeutic agents that target gram-negative bacteria (E. coli, P. aeruginosa, Acinetobacter baumannii, Porphyromonas gingivalis, etc.) that cause various types of inflammation, including sepsis, a systemic inflammatory response syndrome, periodontitis, osteomyelitis, urethral infection, cystitis, abdominal infection, and pneumonia.

[0006] Meanwhile, the inflammasome inflammatory complex can be activated by gram-negative bacterial infection and other factors, inducing an immune response, and excessively activated NLRP3 inflammasome is known to cause a variety of inflammasome-mediated diseases, such as neurodegenerative diseases, autoinflammatory diseases, and cancer. Therefore, there is a need to develop therapeutic agents that can effectively treat the above diseases by suppressing the NLRP3 inflammasome. Summary of the Invention

[0007] Problem to be solved by the invention One example of the present application provides a novel compound of formula 1, an isomer thereof, or a pharma- ceutically acceptable salt thereof.

[0008] [ka]

[0009] Another example provides a composition (such as a pharmaceutical composition, food composition, or feed composition) for preventing, ameliorating, and / or treating inflammasome-mediated diseases and / or infections with or diseases induced by Gram-negative bacteria, protecting nerve cells, and / or regenerating (or generating) nerve cells, comprising one or more compounds selected from the group consisting of the compound of Chemical Formula 1, its isomers, and pharma- ceutical acceptable salts thereof as an active ingredient.

[0010] Other examples provide uses of one or more compounds selected from the group consisting of compounds of formula 1, their isomers, and pharma- ceutically acceptable salts thereof for the prevention, amelioration, and / or treatment of inflammasome-mediated diseases, and / or infections with or diseases induced by Gram-negative bacteria, for protecting nerve cells, and / or for regenerating (or generating) nerve cells.

[0011] Another example provides the use of one or more compounds selected from the group consisting of compounds of formula 1, their isomers, and pharma- ceutically acceptable salts thereof for the manufacture of a composition (such as a pharmaceutical composition, food composition, or feed composition) for preventing, ameliorating, and / or treating an inflammasome-mediated disease, and / or an infection with or a disease induced by a gram-negative bacterium, protecting nerve cells, and / or regenerating (or generating) nerve cells.

[0012] Another example provides a method for preventing, ameliorating, and / or treating an inflammasome-mediated disease, and / or infection with or a disease induced by a gram-negative bacterium, a method for protecting nerve cells, and / or a method for regenerating or generating nerve cells, comprising the step of administering an effective amount of one or more selected from the group consisting of a compound of chemical formula 1, an isomer thereof, and a pharmaceutically acceptable salt thereof to a subject (individual) in need of the prevention, amelioration, and / or treatment of an inflammasome-mediated disease, and / or infection with or a disease induced by a gram-negative bacterium, protection of nerve cells, and / or regeneration or generation of nerve cells.

[0013] Another example provides an antibiotic comprising a compound of formula 1. The antibiotic may exhibit antibiotic activity against gram-negative bacteria.

[0014] The antibiotic (first antibiotic) provided herein has the advantages of increasing the antibiotic susceptibility of bacteria, increasing the bactericidal effect of the second antibiotic due to the bacteriostatic effect of the first antibiotic itself, and enabling the second antibiotic to exert an excellent antibiotic effect even at a low concentration. This can reduce side effects such as toxicity (e.g., nephrotoxicity, hepatotoxicity, etc.) caused by the use of high concentrations of antibiotics. In addition, the emergence of antibiotic-resistant bacteria caused by the combined use of antibiotics with different mechanisms can be suppressed.

[0015] For this reason, an example is (a) a compound, and (b) Second antibiotic The present invention provides a combination antibiotic comprising:

[0016] Another example provides a feed additive that includes an antibiotic or a combination of antibiotics as an active ingredient.

[0017] Another example provides a disinfectant that contains an antibiotic or combination of antibiotics as an active ingredient.

[0018] Another example provides a method of disinfection comprising applying an antibiotic or a combination of antibiotics to an object in need of disinfection.

[0019] Another example provides a cleaning agent that includes an antibiotic or combination of antibiotics as an active ingredient.

[0020] Another example provides a method of cleansing that includes applying an antibiotic or a combination of antibiotics to an object in need of cleansing.

[0021] Means for solving the problem One example of the present application provides a compound of the following formula 1, an isomer thereof, or a pharma- ceutically acceptable salt thereof:

[0022] [ka] [Brief description of the drawings]

[0023] [Figure 1] 1 and 2 are graphs showing the degree of biofilm formation (O.D590nm) of P. gingivalis, which was confirmed by treating the compounds of Examples 1-1 to 1-3 at different concentrations (2μM, 0.2μM, 0.02μM, 0.002μM) in a medium in which P. gingivalis was cultured. "NT" is a group that was not treated with anything, "DPD" is a group in which the compound DPD was treated in the culture medium and P. gingivalis was cultured, "CT" is a group in which the culture medium was treated with PBS instead of DPD and P. gingivalis was cultured, and DPD + Example 1-1, DPD + Example 1-2, and DPD + Example 1-3 are groups that were treated with DPD and the compound of the example together. [Diagram 2] Same as above [Figure 3A] Figures 3A to 3E show graphs of the degree of biofilm formation (O.D.590nm) of P. gingivalis when the compounds of Examples 1-2 and the compounds of the comparative examples were treated at different concentrations (2 μM, 0.2 μM, 0.02 μM, 0.002 μM) in a medium in which P. gingivalis was cultured. [Figure 3B] Same as above [Figure 3C] Same as above [Figure 3D] Same as above [Figure 3E] Same as above [Figure 4A] 4A to 4D are graphs showing the stability in human plasma and rat plasma of the compound of Example 1-2 and the compound of the comparative example. [Figure 4B] Same as above [Figure 4C] Same as above [Figure 4D] Same as above [Figure 5A]5A to 5C are graphs showing the expression levels of three gingipain proteins (lysine-gingipain (Kgp), arginine-gingipain A (RgpA), and arginine-gingipain B (RgpB)), which are major virulence factors and proteolytic enzymes produced by P. gingivalis, when the medium in which P. gingivalis was cultured was treated with the compounds of Examples 1-1 to 1-3, respectively. The 16s rRNA gene was used to normalize the expression levels of the gingipain genes. "Control" is a group that was not treated with anything, "DPD" is a group in which P. gingivalis was cultured in a culture medium treated with the compound DPD, and DPD + Example 1-1 and DPD + Example 1-2 are groups that were treated with DPD and the compounds of the examples together. [Figure 5B] Same as above [Figure 5C] Same as above [Figure 6] 6 is a graph showing the presence or absence of cytotoxicity when mouse bone marrow-derived macrophages were treated with the compound of Example 1-2 at various concentrations (0.5 μM to 16 μM). "Con" is a control group that was not treated with anything. [Figure 7A] 7A to 7D show that treatment of Gram-negative bacteria with the compounds of the examples significantly inhibited E. coli biofilms compared to a positive control group (CT) that was not treated with the compounds of the examples. (Figures 7A and 7B: Results for E. coli, Figure 7C: Results for P. aeruginosa, Figure 7D: Results for A. baumannii) [Figure 7B] Same as above [Figure 7C] Same as above [Figure 7D] Same as above [Figure 8] FIG. 8 shows that when Escherichia coli was treated with a compound of the example in combination with an antibiotic, the minimum growth inhibitory concentration of the antibiotic was significantly reduced compared to treatment with the antibiotic alone. [Figure 9] 9 and 10 show schematic diagrams of how the compounds of the present application inhibit biofilms of Gram-negative bacteria. [Figure 10] Same as above [Figure 11] FIG. 11 shows that inflammation was induced in nerve cells by the infection stimulus P. gingivalis (P. gingivalis), resulting in cell damage, but the nerve cells were protected by treatment with the compounds of the Examples. [Figure 12] 12 and 13 are graphs showing that gene expression of inflammatory cytokines was increased in neurons due to the infection stimulus of P. gingivalis, but the neuroinflammation was alleviated by treatment with the compound of the example. [Figure 13] Same as above [Figure 14] FIG. 14 is a graph showing that the compounds of the examples do not exhibit toxicity to nerve cells. [Figure 15] FIG. 15 is a graph showing plasma concentration as a function of time when an example compound was intravenously injected into male rats and blood samples were taken. [Figure 16] FIG. 16 is a graph showing the brain concentration over time when the compound of the embodiment is intravenously injected into male rats and brain samples are taken. This shows that the compound of the embodiment travels through the blood and then passes through the blood-brain barrier to reach the brain, thereby acting on the lesion in the target brain site and treating neurodegenerative diseases. [Figure 17] FIG. 17 is a graph showing that the inflammatory response induced in macrophages is suppressed by the compound of the example, as determined by treating mouse bone marrow-derived macrophages with the compound of the example and measuring the change in the production of inflammatory cytokines (IL-1β) in the macrophages. [Figure 18] FIG. 18 is a graph showing that the compounds of the examples do not exhibit cytotoxicity in mouse bone marrow-derived macrophages. [Figure 19] FIG. 19 is a graph showing that the compounds of the examples do not exhibit cytotoxicity in human monocytic cell lines. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0024] Definition of Terms The terms used in this specification are briefly explained below.

[0025] The term "pharmaceutically acceptable salt" as used herein means a salt form of a compound that does not induce serious irritation to an organism to which the compound is administered and does not impair the biological activity and physical properties of the compound, and in the present invention, may collectively refer to any salt that equally retains the biological effectiveness and properties of the compound of Chemical Formula 1 and is preferable in terms of pharmaceutical, biological or other properties. The pharmaceutically acceptable salt may be an acid addition salt formed with an acid that forms a non-toxic acid addition salt containing a pharmaceutically acceptable anion, for example, an inorganic acid such as hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, hydrobromic acid, hydroiodic acid, etc.; an organic carboxylic acid such as tartaric acid, formic acid, citric acid, acetic acid, trichloroacetic acid, trifluoroacetic acid, gluconic acid, benzoic acid, lactic acid, fumaric acid, maleic acid, salicylic acid, etc.; or a sulfonic acid such as methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, etc. As a specific example, a compound in free base form can be reacted with a stoichiometric amount of an appropriate acid to obtain an acid addition salt of the compound of one embodiment. In this case, the reaction can proceed in water, an organic solvent, or a mixture thereof, specifically, in a non-aqueous medium such as ether, ethyl acetate, ethanol, isopropanol, or acetonitrile. In addition, depending on the form of the pharmaceutically acceptable salt, each form of salt can be obtained by a normal reaction obvious to a person skilled in the art. In addition, the pharmaceutically acceptable salt may be an alkali metal salt or an alkaline earth metal salt formed by lithium, sodium, potassium, calcium, magnesium, etc.; an amino acid salt such as lysine, arginine, or guanidine; or an organic salt such as dicyclohexylamine, N-methyl-D-glucamine, tris(hydroxymethyl)methylamine, diethanolamine, choline, or triethylamine.

[0026] As used herein, the term "isomer" refers to compounds or salts thereof that have the same chemical or molecular formula but are optically or sterically different. Such isomers, salts thereof, and racemic mixtures of isomers are also included within the scope of the present invention.

[0027] As used herein, the term "aryl" refers to a carbocyclic group having a shared pi electron system and having at least one ring (e.g., a phenyl group). The term includes monocyclic or fused polycyclic (i.e., rings which share adjacent pairs of carbon atoms) groups.

[0028] The term "heteroaryl" as used herein means a heterocyclic aryl group having a shared pi electron system and having at least one ring, including, but not limited to, furan, thiophene, pyrrole, imidazole, oxazole, isoxazole, oxadiazole, tetrazole, thiazole, imidazole, pyrazole, isothiazole, triazole, thiadiazole, pyridine, pyridazine, pyrimidine, pyrazine, triazine, and the like.

[0029] The aryl and / or heteroaryl are 5-10 or may be substituted or unsubstituted, or may be an aryl and / or heteroaryl having 5 to 10 carbon atoms (C5-C10), 5 to 9 carbon atoms (C5-C9), 5 to 8 carbon atoms (C5-C8), 5 to 7 carbon atoms (C5-C7), 5 to 6 carbon atoms (C5-C6), 6 to 10 carbon atoms (C6-C10), 6 to 9 carbon atoms (C6-C9), 6 to 8 carbon atoms (C6-C8), 6 to 7 carbon atoms (C6-C7), 5 carbon atoms, 6 carbon atoms, 7 carbon atoms, 8 carbon atoms, 9 carbon atoms, or 10 carbon atoms.

[0030] The term "alkyl" as used herein refers to an aliphatic hydrocarbon group. The alkyl moiety may be a "saturated alkyl" group that does not contain any alkene or alkyne moieties, or an "unsaturated alkyl" group that contains at least one alkene or alkyne moiety. An "alkene" moiety refers to a group that consists of at least one carbon-carbon double bond, and an "alkyne" moiety refers to a group that consists of at least one carbon-carbon triple bond. For example, an "alkyl" may be a saturated or unsaturated alkyl having a linear, branched, or cyclic structure. Typical alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl, hexyl, ethenyl, propenyl, butenyl, and the like. For example, C1-C4-alkyl has 1 to 4 carbon atoms in the alkyl chain and is selected from the group consisting of methyl, ethyl, propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl, and t-butyl.

[0031] As used herein, an alkyl group is defined as a C 1-10 The alkoxy group may be a straight or branched chain alkyl group of the formula 1-10 It may be a straight or branched chain alkoxy group of the formula C. 1-10 or a straight or branched chain alkyl group of C 1-10each independently represents a straight or branched alkoxy group having 1 to 10 carbon atoms (C1-C10), 1 to 9 carbon atoms (C1-C9), 1 to 8 carbon atoms (C1-C8), 1 to 7 carbon atoms (C1-C7), 1 to 6 carbon atoms (C1-C6), 1 to 5 carbon atoms (C1-C5), 1 to 4 carbon atoms (C1-C4), 1 to 3 carbon atoms (C1-C3), 1 to 2 carbon atoms (C1-C2), 1 carbon atom (C1), 2 to 10 carbon atoms (C2-C10), 2 to 9 carbon atoms (C2-C9), 2-8 carbons (C2-C8), 2-7 carbons (C2-C7), 2-6 carbons (C2-C6), 2-5 carbons (C2-C5), 2-4 carbons (C2-C4), 2-3 carbons (C2-C3), 2 carbons (C2), carbon Number of 3 to 10 carbons (C3-C10), 3 to 9 carbons (C3-C9), 3 to 8 carbons (C3-C8), 3 to 7 carbons (C3-C7), 3 to 6 carbons (C3-C6), 3 to 5 carbons (C3-C5), 3 to 4 carbons (C3-C4) ), 3 carbons (C3), 4-10 carbons (C4-C10), 4-9 carbons (C4-C9), 4-8 carbons (C4-C8), 4-7 carbons (C4-C7), 4-6 carbons (C4-C6), 4-5 carbons (C4-C5) ), 4 carbons (C4), 5-10 carbons (C5-C10), 5-9 carbons (C5-C9), 5-8 carbons (C5-C8), 5-7 carbons (C5-C7), 5-6 carbons (C5-C6), 5 carbons (C5), 6 carbons The alkyl group may be 6 to 10 carbon atoms (C6-C10), 6 to 9 carbon atoms (C6-C9), 6 to 8 carbon atoms (C6-C8), 6 to 7 carbon atoms (C6-C7), 6 carbon atoms (C6), 7 to 10 carbon atoms (C7-C10), 7 to 9 carbon atoms (C7-C9), 7 to 8 carbon atoms (C7-C8), 7 carbon atoms (C7), 8 to 10 carbon atoms (C8-10), 8 to 9 carbon atoms (C8-C9), 9 carbon atoms (C9), 9 to 10 carbon atoms (C9-C10), or 10 carbon atoms (C10).

[0032] The term "halo" or "halogen" as used herein may refer to a fluoro group (-F), a chloro group (-Cl), a bromo group (-Br) or an iodo group (-I).

[0033] As used herein, the term "heterocycle" refers to a group in which a ring carbon is replaced by oxygen, nitrogen, sulfur, etc., and may optionally contain a double bond. Examples of heterocycles include, but are not limited to, pyrroline, pyrrolidine, tetrahydrofuran, imidazoline, imidazolidine, pyrazoline, pyrazolidine, pyran, piperidine, piperazine, morpholine, thiomorpholine, and the like.

[0034] "Each independently substituted" means that when the number of replaceable hydrogens is two or more, each hydrogen may be substituted with the same or different substituents.

[0035] As used herein, "prevention" refers to any action that inhibits or delays the onset of a disease (disorder) by administering a composition according to an embodiment, "treatment" refers to any action that results in the improvement or favorable change of symptoms in a suspected or diseased individual by administering a composition according to an embodiment, and "amelioration" can refer to any action that at least reduces a parameter associated with the condition in which the disease is treated, e.g., the severity of symptoms, by administering a composition according to an embodiment. The disease may be an inflammasome-mediated disease.

[0036] As used herein, the term "pharmaceutical effective amount" refers to the amount of an active ingredient that provides a desired pharmacological effect, and can, in some cases, refer to the concentration or dosage of the active ingredient in a pharmaceutical composition to achieve the desired pharmacological effect.

[0037] Terms other than those mentioned above can be interpreted as having the meanings commonly understood by those skilled in the art in the field to which the present invention belongs.

[0038] Compound of Formula 1 One example of the present application provides a compound of the following formula 1, an isomer thereof, or a pharma- ceutically acceptable salt thereof:

[0039] [ka]

[0040] In chemical formula 1, [ka] is a substituted or unsubstituted C 3-7 cycloalkyl, substituted or unsubstituted C 3-7 cycloalkenyl, substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S; 3-7 heterocycloalkyl, substituted or unsubstituted C containing one or more double bonds and containing one or more heteroatoms selected from the group consisting of N, O, and S; 3-7 heterocycloalkenyl, substituted or unsubstituted C 6-10 or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. 5-10 In one embodiment, the heteroaryl may be [ka] is a substituted or unsubstituted C 6-10 It may also be an aryl of the formula:

[0041] Substituted cycloalkyl, substituted cycloalkenyl, substituted heterocycloalkyl, substituted heterocycloalkenyl, substituted aryl, or substituted heteroaryl are any of the following: hydroxy (-OH), halogen (-F, -Br, -Cl, or -I), cyano (-CN), nitro (-NO2), amino (-NH), substituted or unsubstituted C 1-10 Linear or branched alkyl of, and substituted or unsubstituted C 1-10 and straight-chain or branched-chain alkoxy. In this specification, "substituted with one or more" includes substitution with two or more of the same substituents, for example, substitution with two OH.

[0042] Substituted alkyl or substituted alkoxy can be any of the following: hydroxy (-OH), halogen, cyano (-CN), nitro (-NO2), amino (-NH), C 1-5 Linear or branched alkyl of C 1-5 and n is 0, 1 or 2. The aryl group may be substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy.

[0043] R3 is a substituted or unsubstituted C 1-10 Straight or branched chain alkyl, substituted or unsubstituted C 1-10 linear or branched chain alkoxy, substituted or unsubstituted C 3-7 cycloalkyl, substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S; 3-7 Heterocycloalkyl, substituted or unsubstituted C 6-10 or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. 5-10 In one embodiment, R3 is a substituted or unsubstituted C 6-10 It may also be an aryl of the formula:

[0044] Substituted alkyl is hydroxy (-OH), halogen, cyano (-CN), nitro (-NO2), amino (-NH), C 1-5 Linear or branched alkyl of C 1-5 or substituted or unsubstituted C 6-10 or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. 5-10 may be substituted with heteroaryl of the formula:

[0045] Substituted alkoxy, substituted cycloalkyl, substituted heterocycloalkyl, substituted aryl, or substituted heteroaryl are hydroxy (-OH), halogen, cyano (-CN), nitro (-NO2), amino (-NH), C 1-5Linear or branched alkyl of C 1-5 and n is 0, 1 or 2. The aryl group may be substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy.

[0046] R1 and R2 are each independently (or together) hydrogen (H), halogen (-F, -Br, -Cl, or -I), substituted or unsubstituted C 1-10 Straight or branched chain alkyl, substituted or unsubstituted C 1-10 linear or branched chain alkoxy, substituted or unsubstituted C 6-10 or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. 5-10 In one embodiment, R and R are each independently a substituted or unsubstituted C 6-10 or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. 5-10 In one embodiment, the heteroaryl may be a substituted C 6-10 Aryl or substituted C 5-10 The heteroaryl may be substituted with one or more substituents selected from the group consisting of a methyl group, an ethyl group, a methoxy group, and an ethoxy group.

[0047] Substituted alkyl is hydroxy (-OH), halogen, cyano (-CN), nitro (-NO2), amino (-NH), C 1-5 Linear or branched alkyl of C 1-5 or substituted or unsubstituted C 6-10 or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. 5-10 may be substituted with heteroaryl of the formula:

[0048] The substituted alkoxy, substituted aryl, or substituted heteroaryl are hydroxy (-OH), halogen, cyano (-CN), nitro (-NO2), amino (-NH), C 1-5 Linear or branched alkyl of C 1-5 and n is 0, 1 or 2. The aryl group may be substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy.

[0049] In one example, the compound of formula 1 may be one or more selected from the group consisting of the following compounds: 1) 3-(diphenylmethylene)-2-methylisoindolin-1-one, 2) 3-(di-p-tolylmethylene)-2-methylisoindolin-1-one, and 3) 3-(bis(4-methoxyphenyl)methylene)-2-methylisoindolin-1-one Another embodiment of the present application provides a compound of formula 1A, an isomer thereof, or a pharma- ceutically acceptable salt thereof:

[0050] [ka]

[0051] In the compound of formula 1A, [ka] R1, R2, and R3 are as described above, except that the compound of formula 1A may not include the following compounds: 3-(diphenylmethylene)-2-methylisoindolin-1-one (CAS No. 92172-54-8) In one example, the compound of formula 1A can be one or more selected from the group consisting of the following compounds: 1) 3-(di-p-tolylmethylene)-2-methylisoindolin-1-one, and 2) 3-(bis(4-methoxyphenyl)methylene)-2-methylisoindolin-1-one.

[0052] An isomer of a compound of Chemical Formula 1 or Chemical Formula 1A (hereinafter, in this specification, a compound of Chemical Formula 1 or Chemical Formula 1A will be uniformly described as a compound of Chemical Formula 1 unless otherwise specified) may be an optical isomer, a stereoisomer, or a mixture of isomers (racemic mixture) of a compound of Chemical Formula 1. In a compound of Chemical Formula 1 according to an embodiment, it is understood that each substituent can be attached to a chiral center of a carbon atom. And, any asymmetric carbon atom on a compound of an embodiment can be present in any form of (R)-, (S)- or (R, S)-configuration, and preferably in (R)- or (S)-configuration, which are each separated form. Also, a compound of an embodiment can be present in any form of any possible isomer or mixture thereof, for example, in any form of a pure geometric isomer, partial stereoisomer, optical isomer, racemate, or mixture thereof. In addition, when a compound of an embodiment has a double bond, each substituent attached to the double bond may be in an E or Z configuration.

[0053] Inflammasome-Mediated Diseases As used herein, the term "inflammasome" refers to a caspase-1-activating multiprotein complex that can include 1) a sensor protein, NLR (nucleotide-binding oligomerization domain and leucine-rich repeat-containing receptor) protein, 2) an adaptor protein, ASC (adaptor protein apoptosis-associated spec-like protein containing a caspase-recruitment domain), and 3) an effector protein, pro-caspase-1. IL-1β, a proinflammatory cytokine, is formed by a multiprotein polymer called inflammasome and can mediate pyroptosis, a type of inflammatory cell death. When Toll-like receptors (TLRs) are activated by pathogen-associated molecular pattern molecules (PAMPs), which are substances possessed by pathogens, or damage-associated molecular pattern molecules (DAMPs), which are produced by dead cells, the expression of IL-1β precursor (pro-IL-1β) increases, and then, a protein complex called the inflammasome activates caspase-1, which breaks down the IL-1β precursor, thereby producing activated IL-1β.

[0054] The inflammasome can be selected from the group consisting of, but is not limited to, NLRP3 (NOD-like receptor family, pyrin domain-containing 3) inflammasome, AIM2 (Absent in melanoma 2) inflammasome, and NLRC4 (NLR family CARD domain-containing protein 4) inflammasome. NLRP3, AIM2, or NLRC4 inflammasome includes, but is not limited to, nucleic acids, polynucleotides, oligonucleotides, sense and anti-sense polynucleotide strands, complementary sequences, peptides, polypeptides, proteins, homologous and / or heterologous molecules, isoforms, precursors, mutants, variants, derivatives, splice variants, alleles, distinct species, and active fragments thereof.

[0055] When inflammasome is excessively activated, inflammasome-mediated diseases may occur. Inflammasome activation may be induced by gram-negative bacteria infection or gram-negative bacteria, but is not limited thereto. In one example, the compound of the present application can effectively regulate the secretion amount of inflammatory cytokine IL-1β by suppressing inflammasome activation (e.g., suppressing the production of inflammatory cytokine IL-1β), and can show excellent effects in preventing, improving or treating inflammasome-mediated diseases.

[0056] Inflammasome-mediated diseases can refer to any type of disease that occurs when inflammasomes are abnormally and excessively activated.

[0057] In one example, the inflammasome-mediated disease can be periodontal disease.

[0058] In one example, the inflammasome-mediated disease can be a neurodegenerative disease.

[0059] In one example, the inflammasome-mediated disease may be an inflammatory disease. The inflammatory disease may be an inflammasome-mediated inflammatory disease.

[0060] In one example, the inflammasome-mediated disease may be cancer.

[0061] In one example, the inflammasome-mediated disease can be a gram-negative bacterial infection or a disease induced by a gram-negative bacteria.

[0062] In one example, the inflammasome-mediated disease may be, but is not limited to, one or more selected from the group consisting of periodontal disease, neurodegenerative disease, inflammatory disease (e.g., inflammasome-mediated inflammatory disease), cancer, gram-negative bacterial infection or disease induced by gram-negative bacteria, metabolic disease, sepsis, septic shock, inflammatory bowel disease, osteoarthritis, hyperimmunoglobulin D syndrome, and cryopyrin-associated periodic syndromes.

[0063] The compounds of the present application may exhibit antibiotic effects against gram-negative bacteria, specifically, inhibitory activity against the virulence of Porphyromonas bacteria, as described below, and may have activity for preventing, improving, and / or treating periodontal disease.

[0064] The activity of suppressing the virulence of Gram-negative bacteria may be achieved by, but is not limited to, suppressing quorum sensing of Gram-negative bacteria and / or suppressing the expression (e.g., suppression of the mRNA transcription process or the protein translation process) of virulence factors of Gram-negative bacteria (e.g., biofilm of Gram-negative bacteria, or gingipain in the case of Porphyromonas bacteria). The gingipain may be one or more selected from the group consisting of lysine-gingipain (Kgp), arginine-gingipain A (RgpA), arginine-gingipain B (RgpB), etc.

[0065] By suppressing the virulence of Gram-negative bacteria, it is possible to prevent, improve, and / or treat periodontal disease. That is, the prevention, improvement, and / or treatment of periodontal disease may be achieved by suppressing the virulence of Gram-negative bacteria, for example, by suppressing quorum sensing of Porphyromonas bacteria, and / or by suppressing the expression of virulence factors of Porphyromonas bacteria (for example, gingipain, biofilm, etc.) (for example, suppression of mRNA transcription process, or suppression of protein translation process, etc.), but is not limited thereto.

[0066] The periodontal disease may be one or more diseases selected from periodontitis, gingivitis, and the like.

[0067] The compounds of the present application are characterized by having activity of suppressing the expression of inflammatory proteins (e.g., inflammatory cytokines, etc.), inflammation and death of nerve cells (e.g., neuroinflammation), and / or nerve cell death and / or inflammation caused by Porphyromonas gingivalis in patients with neurodegenerative diseases.

[0068] As used herein, the term "neurodegenerative disease" may be selected from all diseases that exhibit abnormalities in motor control ability, cognitive function, sensory function, and autonomic nerve function due to death, reduction, functional decline, or functional loss of nerve cells, etc. For example, the neurodegenerative disease may be one or more selected from the group consisting of dementia (e.g., Alzheimer's disease (AD), vascular dementia, mixed dementia, Lewy body dementia, frontotemporal dementia, etc.), Parkinson's disease (PD), Huntington's disease, amyotrophic lateral sclerosis (Lou Gehrig's disease), tremor, Tarant's disease, multiple sclerosis, motor neuron disease, spinal muscular atrophy, Creutzfeldt-Jakob disease, Pick's disease, prion disease, spinocerebellar degeneration, etc., but is not limited thereto.

[0069] The neurodegenerative disease may be caused by infection with a gram-negative bacterium, such as, but not limited to, a bacterium of the genus Porphyromonas, or may be induced by a bacterium of the genus Porphyromonas. The bacterium of the genus Porphyromonas may be one or more selected from the group consisting of Porphyromonas gingivalis, Porphyromonas endodontalis, and the like.

[0070] The neuronal cells may be central neuronal cells, for example brain (neuronal) cells.

[0071] The inflammatory protein (eg, an inflammatory cytokine) may be, but is not limited to, IL-6, IL-8, IL-18, TNF-α, IL-1, and the like.

[0072] The inflammatory disease may refer to an abnormal inflammatory disease that occurs when inflammasomes are abnormally and excessively activated, and may refer to a neuroinflammatory disease, an autoinflammatory disease, an autoimmune disease, etc.

[0073] The neuroinflammatory disease may refer to a disease caused by damage to nerve tissue due to an inflammatory response, and may be one or more selected from the group consisting of Alzheimer's disease, Parkinson's disease, Huntington's disease, Lou Gehrig's disease, Creutzfeldt-Jakob disease, multiple sclerosis, amyotrophic lateral sclerosis, inflammatory spinal cord injury, diffuse Lewy body disease, leukoencephalitis, temporal lobe epilepsy, and inflammatory spinal cord injury, but is not limited thereto.

[0074] The auto-inflammatory disease may be one or more selected from the group consisting of, but is not limited to, Muckle-Wells syndrome (MWS), latent adult autoimmune diabetes (LADA), familial cold autoinflammatory syndrome (FCAS), cryopyrin-associated periodic syndrome (CAPS), neonatal-onset multisystem inflammatory syndrome (NOMID), chronic infantile neurological cutaneous articular (CINCA) syndrome, familial Mediterranean fever (FMF), pediatric arthritis (e.g., systemic onset pediatric idiopathic arthritis (SJIA)), pediatric rheumatoid arthritis (e.g., systemic onset pediatric idiopathic rheumatoid arthritis), and gout.

[0075] The autoimmune disease may be, but is not limited to, one or more selected from the group consisting of rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), atopic dermatitis (AD) and psoriasis.

[0076] Inflammasome activation is also known to be involved in the pathogenesis of many cancers, including colorectal cancer, acute myeloid leukemia, adrenal cortical carcinoma, Kaposi's sarcoma, lymphoma, anal cancer, appendix cancer, teratoid rhabdoid tumor, basal cell carcinoma, bile duct cancer, bladder cancer, bone cancer, brain cancer, breast cancer, bronchial tumor, carcinoid tumor, cardiac tumor, cervical cancer, chondroma, chronic lymphocytic leukemia, chronic myeloproliferative neoplasm, colon cancer, colorectal cancer, craniopharyngioma, endometrial cancer, ependymoma, esophageal cancer, esthesioneuroblastoma, Ewing's sarcoma, eye cancer, and ovariectomy. The cancer may be, but is not limited to, one or more selected from the group consisting of ductal cancer, gallbladder cancer, gastrointestinal carcinoid tumor, gastrointestinal stromal tumor, germinoma, hairy cell leukemia, head and neck cancer, heart cancer, liver cancer, hypopharyngeal cancer, pancreatic cancer, renal cancer, laryngeal cancer, chronic myeloid leukemia, lip cancer, oral cancer, lung cancer, melanoma, Merkel cell carcinoma, mesothelioma, oral cancer, oral cancer, osteosarcoma, ovarian cancer, penile cancer, pharyngeal cancer, prostate cancer, rectal cancer, salivary gland cancer, skin cancer, small intestine cancer, soft tissue sarcoma, testicular cancer, throat cancer, thyroid cancer, urethral cancer, uterine cancer, vaginal cancer, and vulvar cancer.

[0077] The compounds of the present application can inhibit inflammasomes and effectively prevent, ameliorate or treat cancer.

[0078] Metabolic disease is a general term for diseases that occur due to disorders of the metabolism of substances in the body, and may be one or more selected from the group consisting of obesity, hyperlipidemia, hypercholesterolemia, arteriosclerosis, type 2 diabetes, non-alcoholic fatty liver disease (NAFLD), and non-alcoholic steatohepatitis (NASH), but is not limited to these.

[0079] Antibiotics against gram-negative bacteria and gram-negative infections or diseases caused by gram-negative bacteria Another example provides an antibiotic containing compound.

[0080] Other examples provide uses for the compounds as antibiotics against gram-negative bacteria (e.g., inhibiting (or suppressing) the growth (or development) of gram-negative bacteria, bacteriostasis of gram-negative bacteria, bactericidal or pest control of gram-negative bacteria, etc.).

[0081] Another example provides a method for killing (or bacteriostatic, control, or growth-inhibiting) gram-negative bacteria, comprising the step of applying (or administering) an effective amount of a compound to a subject in need of antibiotic treatment against gram-negative bacteria (or bacteriostatic, bactericidal, control, etc. of gram-negative bacteria).

[0082] Another example provides the use of the compounds for the manufacture of antibiotics.

[0083] Bacteriostatic effect can mean that an antibiotic binds reversibly to bacteria and inhibits bacterial growth without killing the bacteria. Bactericidal effect can mean that an antibiotic binds irreversibly to bacteria and kills the bacteria beyond inhibiting bacterial growth. The distinction between bacteriostatic effect and bactericidal effect is not absolute, and antibiotics can function as a mixture of bacteriostatic and bactericidal agents depending on the concentration of the antibiotic administered.

[0084] The compounds of formula 1 may have (i) virulence inhibitory activity against gram-negative bacteria, (ii) antibiotic effect (e.g., bacteriostatic and / or bactericidal effect) against gram-negative bacteria, and / or (iii) activity to prevent, ameliorate, and / or treat infections or diseases induced by gram-negative bacteria.

[0085] The virulence inhibitory activity against Gram-negative bacteria, or the antibiotic effect against Gram-negative bacteria, may be achieved by, but is not limited to, inhibiting quorum sensing of Gram-negative bacteria and / or inhibiting the expression of virulence factors of Gram-negative bacteria (e.g., biofilms, extracellular polysaccharides (EPSs), toxins, etc.) (e.g., inhibiting the mRNA transcription process or the protein translation process, etc.).

[0086] The prevention, amelioration, and / or treatment effect of infection with gram-negative bacteria or diseases induced by gram-negative bacteria can be achieved by suppressing the virulence of gram-negative bacteria and by exerting an antibiotic effect on gram-negative bacteria. That is, the prevention, amelioration, and / or treatment of infection with gram-negative bacteria or diseases induced by gram-negative bacteria can be achieved by suppressing the virulence of gram-negative bacteria or by exerting an antibiotic effect on gram-negative bacteria, for example, by suppressing quorum sensing of gram-negative bacteria, and / or by suppressing the production and / or development of virulence factors of gram-negative bacteria (e.g., biofilm, extracellular polysaccharides (EPSs), toxins, etc.) (e.g., suppression of the mRNA transcription process, or suppression of the protein translation process, etc.), but is not limited thereto.

[0087] The antibiotic may have an antibiotic effect against gram-negative bacteria. The gram-negative bacteria may be one or more (e.g., one, two, three, four, or five) species selected from the group consisting of bacteria of the genus Escherichia, bacteria of the genus Pseudomonas, bacteria of the genus Acinetobacter, bacteria of the genus Enterobacter, bacteria of the genus Klebsiella, bacteria of the genus Porphyromonas, bacteria of the genus Fusobacteria, bacteria of the genus Tannerella, and the like. For example, the Escherichia bacterium may be Escherichia coli, the Pseudomonas bacterium may be Pseudomonas aeruginosa, the Acinetobacter baumannii, the Enterobacter bacterium may be Enterobacter aerogenes (also known as Klebsiella aerogenes) or Enterobacter cloacae, the Klebsiella bacterium may be Klebsiella pneumoniae, and the Porphyromonas bacterium may be Porphyromonas gingivalis, Porphyromonas endodontalis, or Porphyromonas keloides. The Fusobacteria bacterium may be Fusobacterium nucleatum, and the Tannerella bacterium may be, but is not limited to, Tannerella forsythia.

[0088] The gram-negative bacterial infection or the disease induced by gram-negative bacteria can be selected from all diseases caused by infection with gram-negative bacteria, and may be, for example, one or more selected from the group consisting of enteritis, Crohn's disease, ulcerative colitis, bacterial dysentery, urinary tract infection, skin infection, bacteremia, sepsis, skin infection, bedsore, pneumonia, endocarditis, meningitis, otitis externa, otitis media, keratitis, osteomyelitis, peritonitis, cystic fibrosis, and periodontal disease (e.g., gingivitis, periodontitis, etc.), but is not limited thereto.

[0089] The gram-negative bacterial infection or disease induced by gram-negative bacteria may be selected from the group consisting of, but is not limited to, diseases induced by Escherichia bacteria, such as enteritis, Crohn's disease, ulcerative colitis, bacterial dysentery, urinary tract infection, skin infection, bacteremia, sepsis, etc.; diseases induced by Pseudomonas bacteria, such as skin infection, bedsore, pneumonia, bacteremia, sepsis, endocarditis, meningitis, otitis externa, otitis media, keratitis, osteomyelitis, enteritis, peritonitis, cystic fibrosis, etc.; diseases induced by Acinetobacter bacteria, such as skin infection, pneumonia, bacteremia, sepsis, etc.; diseases induced by Porphyromonas bacteria, Fusobacterium bacteria, or Tannerella bacteria, such as gingivitis, periodontitis, etc.

[0090] The antibiotics provided herein can be used in combination with other antibiotics (second antibiotics) in addition to the compounds described above.

[0091] Thus, another embodiment provides a combination antibiotic comprising an antibiotic and a second antibiotic.

[0092] The concomitant antibiotic may have an antibiotic effect against gram-negative bacteria. The gram-negative bacteria are as described above.

[0093] The second antibiotic may be one or more selected from commonly used antibiotics, for example, antibiotics having antibiotic activity against gram-negative bacteria. The second antibiotic may be one or more selected from the group consisting of beta-lactam antibiotics, bacterial cell membrane permeability inhibitors, bacterial ribosome inhibitors, bacterial nucleic acid synthesis inhibitors, and bacterial folate synthesis inhibitors.

[0094] In one embodiment, the second antibiotic is a beta-lactam antibiotic, one or more bacterial cell membrane permeability inhibitors selected from the group consisting of polymicin, amphotericin B, ketoconazole, fluconazole, and itraconazole; one or more bacterial ribosome inhibitors selected from the group consisting of aminoglycosides, tetracyclines, macrolides, lincosamides, chloramphenicol, etc.; One or more bacterial nucleic acid synthesis inhibitors selected from the group consisting of quinolone, fluoroquinolone, rifampicin, and the like; and one or more bacterial folate synthesis inhibitors selected from the group consisting of sulfonamides, trimethoprim, etc.; The compound may be one or more selected from the group consisting of, but is not limited to, the compound.

[0095] The beta-lactam antibiotic may be one or more penicillin antibiotics selected from the group consisting of penicillin G, methicillin, oxacillin, flucloxacillin, amoxicillin, ampicillin, ampicillin / sulbactam, piperacillin, piperacillin / tazobactam, ticarcillin, azlocillin, carbenicillin, and amoxicillin / clavulanic acid; one or more cephalosporin antibiotics selected from the group consisting of cefazolin, cephalexin, cephalothin, ceftezol, cefazedone, cefaclor, cefamandole, cefmetazole, cefotiam, cefuroxime, cefotaxime, ceftriaxone, ceftazidime, and cefepime, Monobactam antibiotics, such as aztreonam, or The antibiotic may be one or more carbapenem antibiotics selected from the group consisting of imipenem, imipenem / cilastatin, faropenem, meropenem, doripenem, and artapenem, but is not limited thereto.

[0096] The second antibiotic may be a beta-lactamase inhibitor such as clavulanic acid, sulbactam, or tazobactam, and may have enhanced antibiotic activity when used in combination with cilastatin.

[0097] In one example, the second antibiotic may be used in combination with substances such as clavulanic acid, sulbactam, tazobactam, cilastatin, etc., such as amoxicillin / clavulanic acid, ampicillin / sulbactam, piperacillin / tazobactam, imipenem / cilastatin, etc. In this case, the second antibiotic may be used in combination with clavulanic acid, sulbactam, tazobactam, cilastatin, etc. acid, sulbactam, tazobactam, cilastatin, etc., on a weight basis, 1:10~0.1, 1:9~0.1, 1:8~0.1, 1:7~0.1, 1:6~0.1, 1:5~0.1, 1:4~0.1, 1:3~0.1, 1:2~0.1, 1:10~0.2, 1:9~0.2, 1:8~0.2, 1:7~0.2, 1:6~0.2, 1:5~0.2, 1:4~0.2, It can be used in various ratios such as 1:3~0.2, 1:2~0.2, 1:10~0.5, 1:9~0.5, 1:8~0.5, 1:7~0.5, 1:6~0.5, 1:5~0.5, 1:4~0.5, 1:3~0.5, 1:2~0.5, 1:10~0.75, 1:9~0.75, 1:8~0.75, 1:7~0.75, 1:6~0.75, 1:5~0.75, 1:4~0.75, 1:3~0.75, 1:2~0.75, 1:2, 1:0.5.

[0098] In addition, the second antibiotic may be used in combination of two or more types. When two or more types of second antibiotics are used in combination, the combination ratio of each second antibiotic is, on a weight basis, 1:10 to 0.1, 1:9 to 0.1, 1:8 to 0.1, 1:7 to 0.1, 1:6 to 0.1, 1:5 to 0.1, 1:4 to 0.1, 1:3 to 0.1, 1:2 to 0.1, 1:10 to 0.2, 1:9 to 0.2, 1:8 to 0.2, 1:7 to 0.2, 1:6 to 0.2, 1:5 to 0.2, 1:4 It can be used in various ratios such as 1:10~0.75, 1:9~0.75, 1:8~0.75, 1:7~0.75, 1:6~0.5, 1:5~0.5, 1:4~0.5, 1:3~0.5, 1:2~0.5, 1:10~0.75, 1:9~0.75, 1:8~0.75, 1:7~0.75, 1:6~0.75, 1:5~0.75, 1:4~0.75, 1:3~0.75, 1:2~0.75, 1:2, 1:0.5.

[0099] In this way, by using an antibiotic (first antibiotic) in combination with another antibiotic (second antibiotic), there are advantages in that the antibiotic sensitivity of bacteria is increased, the bactericidal effect of the second antibiotic is increased due to the bacteriostatic action of the first antibiotic itself, and the second antibiotic can exert an excellent antibiotic effect even at a low concentration. Alternatively, there is an advantage in that the second antibiotic, which has an antibiotic effect against gram-negative bacteria, can exert an excellent antibiotic effect even at a low concentration when the first antibiotic and the second antibiotic are used in combination, due to the quorum sensing inhibition and / or biofilm formation inhibition and / or virulence weakening effect of the first antibiotic. This makes it possible to reduce side effects such as toxicity (e.g., nephrotoxicity, hepatotoxicity, etc.) caused by the use of high concentrations of antibiotics. In addition, it is possible to suppress the emergence of antibiotic-resistant bacteria caused by the use of antibiotics with different mechanisms in combination.

[0100] As used herein, the term "antibiotic" encompasses all forms of preparations that have the ability to inhibit the growth of (e.g., inhibit quorum sensing, inhibit biofilm formation, and / or attenuate virulence) and / or kill Gram-negative bacteria, and unless otherwise specified, can be used interchangeably with antibacterial agents, preservatives, disinfectants, etc.

[0101] Pharmaceutical Compositions Another aspect provides a pharmaceutical composition for preventing or treating inflammasome-mediated diseases and / or infections with or diseases induced by Gram-negative bacteria, comprising the compound, its isomer, or a pharma- ceutically acceptable salt thereof as an active ingredient.

[0102] Another embodiment provides a use of one or more compounds selected from the group consisting of compounds of formula 1, their isomers, and pharma- ceutically acceptable salts thereof for the prevention, amelioration, or treatment of inflammasome-mediated diseases and / or infections with or diseases induced by Gram-negative bacteria.

[0103] Another embodiment provides a composition for preventing, ameliorating, or treating an inflammasome-mediated disease, and / or an infection with or a disease induced by a gram-negative bacterium, comprising one or more selected from the group consisting of a compound of Chemical Formula 1, an isomer thereof, and a pharma-ceutically acceptable salt thereof as an active ingredient.

[0104] Another embodiment provides a method for preventing, ameliorating, or treating an inflammasome-mediated disease, and / or infection with or a disease induced by a gram-negative bacterium, comprising administering to a subject in need of the prevention, amelioration, or treatment of an inflammasome-mediated disease, and / or infection with or a disease induced by a gram-negative bacterium, an effective amount of one or more selected from the group consisting of a compound of formula 1, an isomer thereof, and a pharma- ceutically acceptable salt thereof. The method may additionally comprise a step of identifying a subject in need of the prevention, amelioration, and / or treatment of an inflammasome-mediated disease, and / or infection with or a disease induced by a gram-negative bacterium, prior to the administering step.

[0105] Another embodiment provides a use of one or more compounds selected from the group consisting of compounds of formula 1, their isomers, and pharma- ceutically acceptable salts thereof, for the manufacture of a composition for preventing, ameliorating, or treating an inflammasome-mediated disease, and / or an infection with or a disease induced by a gram-negative bacterium.

[0106] Other examples provide pharmaceutical compositions for protecting nerve cells and / or for regenerating or generating nerve cells that include the compounds.

[0107] Other examples provide methods for protecting and / or regenerating or generating neural cells, comprising administering a compound to a subject in need of such protection and / or regeneration (or generation) of neural cells.

[0108] Another aspect provides the use of a compound for the manufacture of a pharmaceutical composition for protecting neuronal cells and / or for regenerating (or generating) neuronal cells.

[0109] Hereinafter, unless otherwise specified, the compound of Chemical Formula 1 as an active ingredient of a pharmaceutical composition includes all isomers of the compound and pharma- ceutically acceptable salts of the compound or isomers, and all of these compounds, isomers, and salts should be construed as being included in the scope of the present application. However, for convenience, in this specification, unless otherwise specified, one or more selected from the group consisting of the compound of Chemical Formula 1, its isomers, and pharma- ceutically acceptable salts thereof as an active ingredient of a pharmaceutical composition will be simply referred to as the "compound of Chemical Formula 1".

[0110] Inflammasome-mediated diseases have been described above.

[0111] The subject to which the pharmaceutical composition is administered may be one or more selected from mammals including humans, primates such as monkeys, rodents such as mice and rats, livestock such as dogs, cats, pigs, cows, horses, sheep and goats, and poultry such as chickens, ducks, geese, pheasants, quails and turkeys, or cells, tissues, or cultures thereof derived therefrom. For example, the subject to which the pharmaceutical composition is administered may be a subject in need of prevention, amelioration, and / or treatment of inflammasome-mediated diseases and / or infections with or diseases induced by gram-negative bacteria, and may be, for example, a human.

[0112] In one example, the subject of administration can be selected from among mammals, including humans, primates such as monkeys, and rodents such as rats and mice, whose patients suffer from neurodegenerative diseases, or cells (brain cells) or tissues (brain tissues) isolated from mammals, or cultures thereof, for example, from humans suffering from neurodegenerative diseases, or brain cells, brain tissues, or cultures thereof isolated therefrom.

[0113] A pharmaceutical composition containing one or more selected from the group consisting of the compound of formula 1, its isomers, and pharma- ceutically acceptable salts thereof as an active ingredient can be formulated into a conventional pharmaceutical preparation and used. For example, the pharmaceutical preparation may be prepared into various preparations for oral or parenteral administration, and the preparation form can be determined in various ways depending on the method of use, the method of administration, the purpose of administration, etc. When prepared into various preparations for oral or parenteral administration, it can be formulated using one or more selected from the group consisting of commonly used fillers, extenders, binders, wetting agents, disintegrants, diluents such as surfactants, excipients, etc.

[0114] When a pharmaceutical composition is orally administered, since proteins or peptides are digested, oral compositions must be formulated to coat the active agent or protect it from decomposition in the stomach. Solid preparations for oral administration may include tablets, pills, powders, granules, capsules, etc., and such solid preparations can be prepared by mixing an active ingredient with at least one or more excipients, such as one or more selected from the group consisting of starch, calcium carbonate, sucrose, lactose, gelatin, etc. In addition to simple excipients, lubricants such as magnesium stylate talc can also be used. Liquid preparations for oral administration may include suspensions, oral liquids, emulsions, syrups, etc. When formulating into liquid preparations, water and / or liquid paraffin, which are commonly used simple diluents, can be used, and optionally, one or more other excipients selected from the group consisting of wetting agents, sweeteners, flavorings, preservatives, etc. may be additionally included. Additionally, the compositions may be administered by any device capable of delivering the active agent to target cells.

[0115] When the pharmaceutical composition is administered parenterally, the parenteral administration may be by any convenient route, such as, but not limited to, vascular administration (e.g., intraarterial administration, intravenous administration, etc.), injection or insertion into a patient's affected site (e.g., gums, etc.), intramuscular administration, subcutaneous administration, intraperitoneal administration, intranasal administration, transdermal administration, intradermal administration, topical administration, intrapulmonary administration, intrarectal administration, intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, etc. Preparations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized preparations, suppositories, etc. As non-aqueous solvents for preparing non-aqueous solutions, or suspension solvents for preparing suspensions, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, injectable esters such as ethyl oleate, etc. can be used as bases for suppositories. witepsol, macrogol, Tween 61, cacao butter, lauric butter, glycerogelatin, etc. can be used. For intranasal administration, the pharmaceutical composition may be diluted and administered via a nasal spray, which is absorbed into the nasal cavity using a sprayer or spray system. For nasal spray, a nasal spray or respiratory dosage form such as an aerosol may be used.

[0116] The content of the active ingredient contained in the pharmaceutical composition is, based on the weight of the entire pharmaceutical composition, 0.01% by weight to 99.9% by weight, 0.01% by weight to 90% by weight, 0.01% by weight to 80% by weight, 0.01% by weight to 70% by weight, 0.01% by weight to 60% by weight, 0.01% by weight to 50% by weight, 0.01% by weight to 40% by weight, 0.01% by weight to 30% by weight, 1% by weight to 99.9% by weight, 1% by weight to 90% by weight, 1% by weight to 80% by weight, 1% by weight to 70% by weight, 1% by weight to 60% by weight, 1% by weight to 50% by weight, 1% by weight to 40% by weight, 1% by weight to 30% by weight, The amount may be 5% to 99.9% by weight, 5% to 90% by weight, 5% to 80% by weight, 5% to 70% by weight, 5% to 60% by weight, 5% to 50% by weight, 5% to 40% by weight, 5% to 30% by weight, 10% to 99.9% by weight, 10% to 90% by weight, 10% to 80% by weight, 10% to 70% by weight, 10% to 60% by weight, 10% to 50% by weight, 10% to 40% by weight, or 10% to 30% by weight, but is not limited thereto and can be appropriately adjusted depending on the form of the formulation, the administration method, the purpose of administration, and the like. In addition, the pharmaceutical composition may additionally contain a pharma- ceutical acceptable carrier in addition to the active ingredient. The pharma- ceutical acceptable carrier is one that is usually used in the formulation of drugs including proteins, nucleic acids, or cells, and may mean a carrier that does not stimulate the organism and does not inhibit the biological activity and / or properties of the active ingredient. In one example, the carrier may be, but is not limited to, one or more selected from the group consisting of lactose, dextrose, sucrose, sorbitol, mannitol, starch, gum acacia, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, mineral oil, etc. The pharmaceutical composition may additionally contain one or more selected from the group consisting of diluents, excipients, lubricants, wetting agents, sweeteners, flavorings, emulsifiers, suspending agents, preservatives, etc. that are commonly used in the manufacture of pharmaceutical compositions.

[0117] The pharma- ceutical effective amount of the pharmaceutical composition can be variously formulated depending on factors such as the formulation method, administration method, age, weight, sex, pathological condition, food, administration time, administration interval, administration route, excretion rate, and reaction sensitivity, and can be administered or taken once or several times a day, but is not limited thereto, and can be administered in various dosages and methods. In one example, the dosage of the pharmaceutical composition per dose is 0.0001 to 5000 mg / kg, 0.0001 to 4000 mg / kg, 0.0001 to 3000 mg / kg, 0.0001 to 2500 mg / kg, 0.0001 to 2000 mg / kg, 0.0001 to 1500 mg / kg, 0.0001 to 1000 mg / kg, 0.0001 to 500 mg / kg, 0.0001 to 100 mg / kg, 0.0001 to 50 mg / kg, 0.0001 to 50 mg / kg, 0.0001 to 10 mg / kg, 0.0001 to 5 mg / kg, 0.0001-1mg / kg, 0.001-5000mg / kg, 0.001-4000mg / kg, 0.001-3000mg / kg, 0.001-2500mg / kg, 0.001-2000mg / kg, 0.001-1500mg / kg, 0.001-1000mg / kg, 0.001-500mg / kg, 0.001-100 / kg, 0.001-50mg / kg, 0.001-10mg / kg, 0.001-5mg / kg, 0.001-1mg / kg, 0.01~5000mg / kg, 0.01~4000mg / kg, 0.01~3000mg / kg, 0.01~2500mg / kg, 0.01~2000mg / kg, 0.01~1500mg / kg, 0.01~1000mg / kg, 0 .01~500mg / kg, 0.01~100mg / kg, 0.01~50mg / kg, 0.01~10mg / kg, 0.01~5mg / kg, 0.01~1mg / kg, 0.1~5000mg / kg, 0.1~4000mg / kg, 0 .1~3000mg / kg, 0.1~2500mg / kg, 0.1~2000mg / kg, 0.1~1500mg / kg, 0.1~1000mg / kg, 0.1~500mg / kg, 0.1~100mg / kg, 0.1~50mg / kg , 0.1~10mg / kg, 0.1~5mg / kg, 0.1~1mg / kg, 1~5000mg / kg, 1~4000mg / kg, 1~3000mg / kg, 1~2500mg / kg, 1~2000mg / kg, 1~1500mg / kg,1~1000mg / kg、1~500mg / kg、1~100mg / kg、1~50mg / kg、1~10mg / kg、1~5mg / kg、5~5000mg / kg、5~4000mg / kg、5~3000mg / kg、5~2500mg / kg、5~2000mg / kg、5~1500mg / kg、5~1000mg / kg、5~500mg / kg、5~100mg / kg、5~50mg / kg、5~10mg / kg、10~5000mg / kg、10~4000mg / kg、10~3000mg / kg、10~2500mg / kg、10~2000mg / kg、10~1500mg / kg、10~1000mg / kg、10~500mg / kg、10~100mg / kg、10~50mg / kg、20~5000mg / kg、20~4000mg / kg、20~3000mg / kg、20~2500mg / kg、20~2000mg / kg、20~1500mg / kg、20~1000mg / kg、20~500mg / kg、20~100mg / kg、20~50mg / kg、50~5000mg / kg、50~4000mg / kg、50~3000mg / kg、50~2500mg / kg、50~2000mg / kg、50~1500mg / kg、50~1000mg / kg、50~500mg / kg、50~100mg / kg、100~5000mg / kg、100~4000mg / kg、100~3000mg / kg、100~2500mg / kg、100~2000mg / kg、100~1500mg / kg、100~1000mg / kg、100~500mg / kg、200~5000mg / kg、200~4000mg / kg、200~3000mg / kg、200~2500mg / kg、200~2000mg / kg、200~1500mg / kg、200~1000mg / kg、200~500mg / kg、500~5000mg / kg、500~4000mg / kg、500~3000mg / kg、500~2500mg / kg、500~2000mg / kg、500~1500mg / kg、500~1000mg / kg、1500~5000mg / kg、1500~4000mg / kg、1500~3000mg / kg、1500~2500mg / kg、1500~2000mg / kg、2000~5000mg / kg、2000~4000mg / kg、2000~3000mg / kg、Alternatively, the amount may be 2000 to 2500 mg / kg, but is not limited thereto.

[0118] Food Composition Another example provides a food composition for preventing and / or improving an inflammasome-mediated disease, and / or an infection with or a disease induced by a gram-negative bacterium, for protecting nerve cells, and / or for regenerating (or generating) nerve cells, comprising the compound. The food composition may be a health functional food.

[0119] Inflammasome-mediated diseases and / or Gram-negative bacterial infections or diseases induced by Gram-negative bacteria are as described above.

[0120] Examples of food compositions include meat, sausages, bread, chocolate, candies, snacks, confectionery, pizza, ramen, other noodles, gums, dairy products including ice cream, various soups, beverages, tea, energy drinks, alcoholic beverages, vitamin complexes, health functional foods and health foods, etc., and include all foods in the usual sense.

[0121] Functional food is the same term as food for special health use (FoSHU), and refers to food with high medical and medical effects that is processed to efficiently exhibit bioregulatory functions in addition to nutritional supply. Here, "function" means obtaining useful effects for health purposes such as regulating nutrients to the structure and function of the human body and physiological effects. The food of the present application can be manufactured by a method commonly used in the industry, and can be manufactured by adding raw materials and ingredients commonly added in the industry during manufacturing. In addition, the dosage form of the food can be manufactured without any restrictions as long as it is a dosage form recognized as a food. The food composition of the present application can be manufactured into various dosage forms, and unlike general medicines, it has the advantage of being free of side effects that may occur when taking medicines for a long period of time because it is made from food as a raw material, and is highly portable, and the food of the present application can be taken as an adjuvant to promote the effect of preventing or improving inflammasome-mediated inflammatory diseases.

[0122] Health foods refer to foods that have more active health maintenance and promotion effects than general foods, while health supplement foods refer to foods intended for supplemental health purposes. In some cases, the terms functional health foods, health foods, and health supplements can be used interchangeably.

[0123] Specifically, health functional foods are foods in which the composition according to the example is added to food ingredients such as beverages, teas, spices, gum, and sweets, or is produced as a capsule, powder, suspension, etc., and when ingested, they bring about specific health benefits. However, unlike general medicines, they have the advantage that they are made from food ingredients and do not have the side effects that can occur when taking medicines for a long period of time.

[0124] The food composition according to the example can be ingested on a daily basis, and is therefore expected to have excellent effects in preventing or improving inflammasome-mediated inflammatory diseases, making it highly useful.

[0125] The food composition may further include a physiologically acceptable carrier, but the type of carrier is not particularly limited, and any carrier commonly used in the art may be used. The food composition may also include additional ingredients commonly used in food compositions to improve odor, taste, vision, etc. For example, vitamins A, C, D, E, B1, B2, B6, B12, niacin, biotin, folate, panthotenic acid, etc. may also be included. Minerals such as zinc (Zn), iron (Fe), calcium (Ca), chromium (Cr), magnesium (Mg), manganese (Mn), copper (Cu), and chromium (Cr) may also be included. Amino acids such as lysine, tryptophan, cysteine, and valine may also be included.

[0126] The food composition may also contain food additives such as preservatives (potassium sorbate, sodium benzoate, salicylic acid, sodium dehydroacetate, etc.), bactericides (bleaching powder and highly bleached powder, sodium hypochlorite, etc.), antioxidants (butyl hydroxyanisole (BHA), butyl hydroxytoluene (BHT), etc.), colorants (tar dyes, etc.), color formers (sodium nitrite, sodium acetate, etc.), bleaching agents (sodium sulfite), seasonings (MSG monosodium glutamate, etc.), sweeteners (dulcin, cyclamate, saccharin, sodium, etc.), flavors (vanillin, lactones, etc.), leavening agents (alum, potassium D-hydrogen tartrate, etc.), strengtheners, emulsifiers, thickeners (thickening agents), coating agents, gum base agents, foam inhibitors, solvents, and improvers. The additives are selected according to the type of food and can be used in appropriate amounts.

[0127] The composition according to the example can be added as it is or used together with other foods or food ingredients, and can be used appropriately by a conventional method. The amount of the active ingredient mixed can be suitably determined depending on the purpose of use (prevention, health or therapeutic treatment). In general, when producing a food or beverage, the food composition of the present application can be added to the food or beverage in an amount of 50 parts by weight or less, specifically 20 parts by weight or less. However, when ingested for a long period of time for health and hygiene purposes, the content can be below the range, and there is no problem in terms of safety, so the active ingredient can be used in an amount above the range.

[0128] An example of the food composition can be a health drink composition, which can contain various flavorings or natural carbohydrates as additional ingredients, like ordinary drinks. The natural carbohydrates mentioned above can be 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. The sweetener can be a natural sweetener such as thaumatin or stevia extract; or an artificial sweetener such as saccharin or aspartame.

[0129] In addition, the health drink composition may contain various nutrients, vitamins, electrolytes, flavors, colorants, pectic acid, salts of pectic acid, alginic acid, salts of alginic acid, organic acids, protective colloid thickeners, pH regulators, stabilizers, preservatives, glycerin, alcohol, or carbonation agents. In addition, fruit pulp for the production of natural fruit juice, fruit juice beverages, or vegetable beverages may be contained. Such ingredients may be used independently or in mixtures. The ratio of such additives is not particularly important, but is generally selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the health drink composition of the present application.

[0130] An example food composition may contain various weight percentages if it can show an effect of preventing or improving inflammasome-mediated inflammatory diseases.

[0131] Feed composition, etc. Another example provides a feed additive (feed additive composition) for preventing and / or ameliorating inflammasome-mediated diseases, and / or infections with or diseases induced by gram-negative bacteria, for protecting nerve cells, and / or for regenerating (or generating) nerve cells, comprising a compound, an antibiotic or a combination of antibiotics as an active ingredient.

[0132] Inflammasome-mediated diseases and / or Gram-negative bacterial infections or diseases induced by Gram-negative bacteria are as described above.

[0133] Another example provides a feed comprising the feed additive.

[0134] The feed may be prepared by mixing the antibiotic or the combination antibiotic with the feed, or by adding the antibiotic or the combination antibiotic to the feed, separately prepared as a feed additive, during the preparation of the feed.

[0135] The compound, antibiotic or combined antibiotic in the feed may be in a liquid or dry state, for example, in a dried powder form. The antibiotic may be included in an amount of 0.005-10% by weight, 0.05-10% by weight, 0.1-10% by weight, 0.005-5% by weight, 0.05-5% by weight, 0.1-5% by weight, 0.005-2% by weight, 0.05-2% by weight, or 0.1-2% by weight of the total weight of the feed, but is not limited thereto. In addition, the feed may additionally include a conventional additive that enhances the shelf life of the feed, in addition to the compound, antibiotic or combined antibiotic.

[0136] In the present specification, the feed to which the compound, antibiotic or combination antibiotic can be added can be selected from the group consisting of commercially available feed, grains, roots and fruits, food processing by-products, algae, fibers, pharmaceutical by-products, dairy fats, starches, melons, grain by-products, proteins, inorganic substances, oils and fats, minerals, single cell proteins, zooplankton, remaining food and drink, etc., but is not limited thereto.

[0137] Another example provides a food additive or drinking water additive containing the compound, antibiotic, or combination of antibiotics as an active ingredient. By mixing the compound, antibiotic, or combination of antibiotics with drinking water and supplying it, the number of gram-negative bacteria in the drinking water can be reduced. The gram-negative bacteria are as described above.

[0138] Another example provides a disinfectant containing a compound, an antibiotic, or a combination of antibiotics as an active ingredient. Another example provides a disinfection method including a step of applying a compound, an antibiotic, or a combination of antibiotics to an object requiring disinfection. Disinfectants are a general term for preparations for preventing pathogen infection, and can be used as disinfectants for general use, disinfectants for food and cooking areas and facilities, and disinfectants for buildings such as poultry farms and livestock sheds, livestock bodies, drinking water, straw, egg beds, transport vehicles, tableware, and other various growing items.

[0139] Another example provides a cleaning agent containing the compound, antibiotic, or combination antibiotic as an active ingredient. Another example provides a cleaning method including a step of applying the compound, antibiotic, or combination antibiotic to an object in need of cleaning. Since the antibiotic has an antibiotic effect against gram-negative bacteria, it is applicable to cleaning the skin surface or various parts of the body of an individual that are exposed or may be exposed to gram-negative bacteria. The gram-negative bacteria are as described above.

[0140] Manufacturing method In another embodiment, a method for preparing a compound of Formula 1 is provided.

[0141] In one example, the preparation method may include a step of introducing phenylboronic acid or a derivative thereof (e.g., phenylboronic acid (Cas No. 98-80-6), 4-methylphenylboronic acid (4-Methylphenylboronic Acid or p-Tolylboronic acid (Cas No. 5720-05-8), or (4-methoxyphenyl)boronic acid (Cas No. 5720-07-0)) into a compound of the following chemical formula 4 to prepare a compound of the following chemical formula 1.

[0142] [ka]

[0143] In the compound of formula 1, [ka] When R is an unsubstituted C6 aryl and R is a C1 alkyl (methyl group), the step of preparing a compound of formula 1 may be carried out by reacting a compound of formula 4 with phenylboronic acid (Cas No. 98-80-6), 4-methylphenylboronic acid (or p-Tolylboronic acid, Cass No. 5720-05-8), or (4-methoxyphenyl)boronic acid (Cass No. 5720-07-0).

[0144] A person skilled in the art to which the present invention pertains can prepare a compound by various methods based on the structure of Chemical Formula 1, and all such methods should be interpreted as being within the scope of the present invention. In other words, a compound of Chemical Formula 1 can be prepared within the scope of the present invention by any combination of various synthetic methods described in this specification or the prior art. Therefore, the preparation method according to the present invention is not limited to only those presented below.

[0145] Effect of the Invention The present application relates to a compound of Chemical Formula 1, its isomer, or a pharma- ceutically acceptable salt thereof, and its use for preventing, improving, and / or treating inflammasome-mediated diseases, and / or diseases caused by infection with or induced by Gram-negative bacteria, which is safe without causing cytotoxicity, can effectively inhibit inflammasomes, and has excellent preventive, improving, and / or therapeutic effects against diseases. EXAMPLES

[0146] The present invention will now be described in more detail with reference to the following examples, which are merely for the purpose of illustrating the present invention and are not intended to limit the scope of the present invention.

[0147] Example 1. Preparation of Compounds Example 1-1. Preparation of 3-(diphenylmethylene)-2-methylisoindolin-1-one The compound of Example 1-1 (3-(diphenylmethylene)-2-methylisoindolin-1-one (CAS No. 92172-54-8)) was produced by the following steps.

[0148] Step 1: Preparation of Compound A [ka]

[0149] Triisopropylphosphite (P(i-PrO)3, Triisopropylphosphite, Cas No. 116-17-6, sigma aldrich) (2.7 ml, 11.00 mmol) in 10 mL of THF (Cas No. 109-99-9, sigma aldrich) containing Zn dust (Cas No. 7440-66-6, sigma aldrich) (490 mg, 7.50 mmol) was cooled to 0 °C under Ar (Cas No. 7440-37-1, Songkwan Specialty Gas). A solution of CBr4 (Cas No. 558-13-4, TCI) (2.49 g, 7.50 mmol) in THF was slowly added to the solution of P(i-PrO)3 ​​and the reaction was stirred until the color changed to yellow. A solution of N-Methylphthalimide anhydride (Cas No. 550-44-7, sigma aldrich) (806 mg, 5.00 mmol) in THF was then added. The reaction was stirred at room temperature for 12 hours. After quenching by adding sat.aq NaHCO3 solution (Cas No. 144-55-8, Samchun Chemical) (2.5 mL), the phases were then separated and the crude product was extracted from the aqueous layer with ether. The residue was dissolved in ether and filtered through a Celite pad. The resulting filtrate was concentrated by rotary evaporation and purified by column chromatography (50% DCM (dichloromethane) in hexane) to give compound A.

[0150] Step 2: Preparation of the compound of Example 1-1 Under an argon atmosphere, compound A (158 mg, 0.5 mmol) prepared in step 1, Na2CO3 (212 mg, 2 mmol), Pd(OAC)2 (Cas No. 3375-31-3, TCI) (22 mg, 0.1 mmol), PPh3 (Cas No. 603-35-0, TCI) (52 mg, 0.2 mmol), H2O (2 ml) and THF (8 ml) were added to a Schlenk tube. Phenylboronic acid (Cas No. 98-80-6, TCI) (1.2 mmol) was added to the stirred mixture, and the mixture was stirred at room temperature for 24 hours. The reaction mixture was extracted with DCM (Cas No. 75-09-2, sigma aldrich) (10 mL), washed with brine, dried over MgSO4 (Cas No. 10034-99-8, Samchun Chemical) and filtered. The crude product was purified by flash chromatography on silica gel to obtain the compound of Example 1-1.

[0151] [ka]

[0152] Example 1-2. Preparation of 3-(di-p-tolylmethylene)-2-methylisoindolin-1-one The compound of Example 1-2 (3-(di-p-tolylmethylene)-2-methylisoindolin-1-one) was produced by the following steps.

[0153] Step 1: Preparation of Compound A Compound A was produced in the same manner as in step 1 of Example 1-1.

[0154] Step 2: Preparation of the compound of Example 1-2 Under argon atmosphere, compound A (158 mg, 0.5 mmol) prepared in step 1, Na2CO3 (212 mg, 2 mmol), Pd(OAC)2 (22 mg, 0.1 mmol), PPh3 (52 mg, 0.2 mmol), H2O (2 ml) and THF (8 ml) were added to a Schlenk tube. 4-Methylphenylboronic Acid (or p-Tolylboronic acid, Cas No. 5720-05-8, TCI) (1.2 mmol) was added to the stirred mixture, and the mixture was stirred at room temperature for 24 hours. The reaction mixture was extracted with DCM (10 mL), washed with brine, dried with MgSO4 and filtered. The crude product was purified by flash chromatography on silica gel to obtain the compound of Example 1-2.

[0155] [ka]

[0156] Example 1-3. Preparation of 3-(bis(4-methoxyphenyl)methylene)-2-methylisoindolin-1-one The compound of Example 1-3 (3-(bis(4-methoxyphenyl)methylene)-2-methylisoindolin-1-one) was produced by carrying out the following steps.

[0157] Step 1: Preparation of Compound A Compound A was produced in the same manner as in step 1 of Example 1-1.

[0158] Step 2: Preparation of Compounds of Examples 1-3 Under argon atmosphere, compound A (158 mg, 0.5 mmol) prepared in step 1, Na2CO3 (212 mg, 2 mmol), Pd(OAC)2 (22 mg, 0.1 mmol), PPh3 (52 mg, 0.2 mmol), H2O (2 ml) and THF (8 ml) were added to a Schlenk tube. 4-Methoxyphenylboronic acid (Cas No. 5720-07-0, TCI) (1.2 mmol) was added to the stirred mixture, and the mixture was stirred at room temperature for 24 hours. The reaction mixture was extracted with DCM (10 mL), washed with brine, dried over MgSO4 and filtered. The crude product was purified by flash chromatography on silica gel to give the compounds of Examples 1-3.

[0159] [ka]

[0160] Preparation of Comparative Compound DPD The compound DPD (4,5-dihydroxy-2,3-pentanedione, CAS NO. 142937-55-1) used in the following experimental examples was prepared by carrying out the following steps 1 to 5 (see Molecules. 2018 Oct., 23(10):2545).

[0161] Step 1: 1-propynylmagnesium bromide (0.5M in THF, 1.3eq) was slowly added to anhydrous (t-butyldimethylsilyloxy)acetaldehyde (1.0eq) in THF at 0°C. The mixture was stirred at room temperature for 3 hours. The reaction solution was concentrated by rotary evaporation and then quenched by adding a cold saturated solution of NH4Cl. The reaction mixture was then extracted with Et2O, washed with brine, dried over MgSO4, and filtered. A yellow oily substance 1 was obtained.

[0162] Step 2: The obtained substance 1 (1.0 eq) was dissolved in MeOH, and Dowex50WX8 100-200 mesh (100 mg / 1 mL) was added thereto, and the mixture was stirred at room temperature for 12 hours. The product was filtered to obtain substance 2.

[0163] Step 3: The obtained material 2 (1.0 eq) was added with cyclohexanone dimethyl ketal (3.0 eq) and catalyst p-TSA and stirred at room temperature for 12 hours. The reaction solution was concentrated by rotary evaporation, dissolved in Et2O again, washed with NaHCO3, dried over MgSO4, and filtered to obtain material 3.

[0164] Step 4: The obtained material 3 was placed in a 1:1:1 CHCl3 / ACN / H2O solution, NaIO4 (4.4eq) and RuO2_H2O (2.5%mol) were added, and stirred at room temperature for 12 hours. The reaction solution was concentrated by rotary evaporation, extracted with DCM, and dried over MgSO4 to obtain material 4.

[0165] Step 5: The obtained material 4 was taken up in MeOH, Dowex 50WX8 resin was added, and the mixture was stirred at room temperature for 12 hours. After filtration, the compound DPD was obtained.

[0166] Example 2. Inhibitory effect of P. gingivalis on biofilm formation The inhibitory effect of the compounds of Examples 1-1 to 1-3 on the formation of a biofilm caused by Porphyromonas gingivalis was confirmed.

[0167] A medium in which P. gingivalis (provided by Korea Oral Microorganism Resource Center (KCOM)) was cultured was treated with each of the compounds of Examples 1-1 to 1-3, and the degree of biofilm formation by P. gingivalis was confirmed.

[0168] Sterilized cover glass slips (round, 12 mm radius) were placed one by one on the bottom of each well of a cell culture plate (24-well plate) using tweezers, and DPD (4,5-dihydroxy-2,3-pentanedione, CAS NO. 142937-55-1), a precursor of the interbacterial signal substance AI-2 (Autoinducer-2), was added to the medium (BHI medium + supplement) for culturing P. gingivalis at 10 μM (BHI, a component of the medium, is Brain Heart Infusion Medium, and hemin (10 μg / ml) and vitamin K (0.2 μg / ml) were used as supplements). It is known that bacteria use DPD for quorum sensing to increase bacterial biofilm formation. The compounds of the examples (compounds of Examples 1-1 to 1-3) were each diluted in a medium containing DPD to prepare final compound concentrations of 0.002 μM, 0.02 μM, 0.2 μM, and 2 μM, respectively.

[0169] The culture medium was dispensed onto a cell culture plate covered with a glass slip, and 2 × 10 bacteria were added. 8 The plates were inoculated to give a total of 1000 cells / ml. For the positive control group (DPD) not treated with the compound of the embodiment, DPD was added to the medium (BHI medium + supplement) and then the same number of bacteria were inoculated. For the negative control group (CT) not treated with either the compound of the embodiment or DPD, the same amount of PBS (phosphate buffered saline) was added to the culture medium (BHI medium + supplement) instead of DPD. The completed cell culture plate was maintained at 37°C under anaerobic conditions (10% H2, 10% CO2 and 80% N2) for 48 hours to culture the bacteria.

[0170] The biofilms formed on the glass slips by the bacteria cultured in each group were stained with 1% crystal violet solution for 15 minutes, washed three times with PBS (phosphate-buffered saline) solution, and decolorized with acetone-alcohol (20:80, vol / vol). 200 μl of the decolorizing solution containing crystal violet was added to each microplate for each group, and the absorbance (OD) was measured using an absorbance microplate reader (Epoch2, Bio-Tek, USA). 590nm The results are shown in Figures 1 and 2.

[0171] As shown in Figures 1 and 2, no biofilm formation was observed in the medium (NT) in which P. gingivalis was not cultured. Compared with the medium (CT) in which P. gingivalis was cultured in PBS, the medium (DPD) in which P. gingivalis was cultured in DPD significantly increased biofilm formation by P. gingivalis due to the quorum sensing effect. In the medium in which DPD and the compound of the example were added together to the medium and P. gingivalis was cultured (DPD + Example 1-1, DPD + Example 1-2, DPD + Example 1-3), it was confirmed that the biofilm of P. gingivalis increased by DPD was suppressed in a concentration-dependent manner by the compound of the example.

[0172] Comparative experiment In order to prove that the compounds of Examples 1-1 to 1-3 have a superior effect to the compound of the Comparative Example, the following experiment was carried out. The compound of the Comparative Example was the compound ((Z)-3-(bromo(phenyl)methylene)isobenzofuran-1(3H)-one) of <Example 1> of Korean Patent Publication (KR10-2019-0130983A).

[0173] 1. Inhibitory effect of P. gingivalis on biofilm formation The inhibitory effect of the compounds of Examples 1-2 and Comparative Example on the formation of a biofilm by Porphyromonas gingivalis was confirmed. The experimental process was carried out in the same manner as in Experimental Example 1, and the experimental results are shown in Figures 3A to 3E.

[0174] As can be seen from FIG. 3A to FIG. 3E, no biofilm formation was observed in the medium (NT) in which P. gingivalis was not cultured. Compared with the medium (CT) in which P. gingivalis was cultured in PBS, the biofilm formation of P. gingivalis was significantly increased in the medium (DPD) in which P. gingivalis was cultured in DPD due to the quorum sensing effect. In the medium in which DPD and a compound were added together to the medium and P. gingivalis was cultured (DPD + Comparative Example, DPD + Example 1-2), it was confirmed that the biofilm of P. gingivalis increased by DPD was inhibited in a compound concentration-dependent manner, and in a comparison of the biofilm formation inhibitory ability of P. gingivalis at the same concentration, it was confirmed that the biofilm formation inhibitory ability of the compound of Example 1-2 was better than that of the compound of the Comparative Example.

[0175] 2.Plasma stability The plasma stability in human plasma and rat plasma of the compounds of Examples 1-2 and Comparative Examples was compared.

[0176] 1) Sample preparation The compounds of Examples 1-2 and Comparative Examples, and the substances used as positive controls (eucatropine, loterpresnol) were dissolved in DMSO to prepare stock solutions at a concentration of 10 mM. The stock solutions of each test substance were then diluted to prepare working solutions of 0.2 mM and 1 mM, which were then used in the test (70% acetonitrile was used as the solvent for the compounds of Comparative Examples, and 50% acetonitrile was used as the solvent for the compounds of Examples 1-2).

[0177] 2) Test method The positive control group and the standard solution of the test substance were placed in tubes containing two types of plasma (human and rat) at concentrations of 1 μM and 5 μM, respectively, and then incubated at 37° C. for a certain period of time (0, 30, 60, 120, 240 minutes) while being mixed by shaking. After each period, 100 μl of the mixed solution sample was dispensed into a cell culture plate (96-well plate) containing 400 μl of quench reagent to terminate the test, and after centrifugation (5000×g, 10 minutes), 100 μl of the supernatant was removed and mixed with 200 μl of ultrapure water, and the solution was injected into an LC-MS / MS system to analyze the drug at each time point, thereby evaluating the plasma stability of the compounds of Comparative Example and Example 1-2.

[0178] 3) Test results The results showing stability in human plasma are shown in Figure 4A (compound of Example 1-2) and Figure 4B (compound of comparative example), and the results showing stability in rat plasma are shown in Figure 4C (compound of Example 1-2) and Figure 4D (compound of comparative example).

[0179] As can be seen from Figures 4A to 4D, the amount of the compound remaining in human plasma and rat plasma decreased over time in the comparative example (Figures 4B and 4D), whereas the compound of Example 1-2 was confirmed to remain stably in plasma (Figures 4A and 4C).

[0180] Example 3. Inhibitory effect of gingipain expression in P. gingivalis The medium in which P. gingivalis was cultured was treated with each of the compounds of the examples, and the level of expression of three gingipain proteins (gingipain lysine-gingipain (Kgp), arginine-gingipain A (RgpA), and arginine-gingipain B (RgpB)), which are major virulence factors produced by P. gingivalis and are also proteolytic enzymes, was confirmed.

[0181] Culture medium containing 10 μM of DPD (4,5-dihydroxy-2,3-pentanedione), a precursor of the bacterial signaling substance (autoinducer-2), and 2 μM of the compound of the example, or containing no compound of the example, was dispensed into a cell culture plate (48-well plate), and then P. gingivalis (2 × 10 8 cells / ml) were inoculated into each group, and the inoculated cultures were incubated at 37°C under anaerobic conditions (10% H2, 10% CO2, and 80% N2) for 48 h.

[0182] After 48 hours, each culture was centrifuged at 13,000 rpm for 1 minute to remove the supernatant, and total RNA was extracted using easy-BLUETM Total RNA Extraction kit (iNtRON Biotechnology, Seongnam, Korea). cDNA was synthesized, and the expression of gingipain gene was confirmed by real-time PCR. cDNA was synthesized using AccuPower® CycleScript RT PreMix dN6 (Bioneer, Daejeon, Korea) and PCR was performed using Power SYBR TM Real-time (quantitative) reverse transcription PCR (RT-qPCR) was performed using Green PCR Master Mix (Applied Biosystems, Foster City, CA, USA).

[0183] The primer sequences used for amplification are shown in Table 1 below, and the RT-qPCR amplification conditions were enzyme activation at 95°C for 3 minutes, followed by denaturation at 95°C for 10 seconds and primer annealing at 60°C for 30 seconds, which were repeated 40 times, and analyzed using a Bio-Rad CFX Connect Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA). The expression level of the gingipain gene was normalized using the 16s rRNA gene, and the gene expression amount was calculated using the Comparative Ct Method (ΔΔCt), which is shown in Figures 5A to 5C.

[0184] [Table 1]

[0185] As shown in Figures 5A to 5C, in the group treated with DPD, a precursor of AI-2 (DPD), P. gingivalis recognized quorum sensing signal molecules and the expression levels of all three gingipain genes (rgpA, rgpB, kgp) increased compared to the control group, as compared to the control group, as compared to the control group. In the groups treated with DPD and the compounds of the examples (DPD + Example 1-1, DPD + Example 1-2), it was confirmed that the expression levels of the three gingipain genes increased by DPD decreased.

[0186] Example 4. Cytotoxicity evaluation The compounds of the examples were evaluated for their cytotoxicity in mouse bone marrow derived macrophages.

[0187] Bone marrow was isolated from the femur and tibia of 8-12 week old C57BL / 6J mice (obtained from Daehan Biolink) using a 26g syringe. The isolated bone marrow was differentiated into macrophages for 6 days in a 150π cell culture plate in a medium containing IMDM medium + 30% L929 cell supernatant + 10% fetal bovine serum + 1% penicillin / streptomycin + 1% non-essential amino acids + 1% sodium pyruvate, and then used for the experiment.

[0188] Differentiated macrophages were 1 × 10 6 200 μl of each were seeded onto a 48-well plate at a concentration of 100 cells / ml, and stabilized for 12 hours at 37° C. and 5% CO2 conditions. After removing the supernatant of the cultured cells, 200 μl of each of the compounds of Example 2 diluted to concentrations of 0.5, 1, 2, 4, 8, and 16 μM were added to serum-free medium and cultured for 24 hours at 37° C. and 5% CO2 conditions.

[0189] After that, the supernatant of the cultured cells was removed, and 200 μl of 400 μg / ml MTT solution was added, and the cells were incubated at 7°C and 5% CO2 for 4 hours. After that, the reaction solution was removed, 200 μl of DMSO was added, and the absorbance at 570 nm was measured. At this time, the cell viability of the untreated cells was set as 100%, and the cell viability of the cells treated with the samples was calculated and shown in Figure 6.

[0190] As can be seen from FIG. 6, it was confirmed that the compound of Example 1-2 did not exhibit cytotoxicity in mouse bone marrow-derived macrophages at a concentration of 16 μM or less.

[0191] Example 5. Inhibitory effect on biofilm formation of E. coli, P. aeruginosa, and A. baumannii 1) Inhibitory effect on E. coli biofilm formation The inhibitory effect of the compound of Example 1-2 on biofilm formation in Escherichia coli (E. coli) (KCTC2571 (ATCC8739), obtained from the Korea Oral Microorganism Resource Center (KCOM)) was confirmed.

[0192] Sterilized cover glass slips (round, 12 mm radius) were placed on the bottom of each well of a cell culture plate (24-well plate) using tweezers, and the compound of Example 1-2 was diluted in the culture medium for bacteria (LB medium) to final concentrations of 0.006 μg / ml, 0.06 μg / ml, 0.6 μg / ml, 1.2 μg / ml, 2.4 μg / ml, and 4.8 μg / ml.

[0193] The culture medium was dispensed onto a cell culture plate covered with a glass slip, and 2 × 10 E. coli were cultured. 7 The plates were inoculated at 1000 cells / ml. For the positive control group (CT) not treated with the compound, the same number of bacteria was inoculated into a medium (LB) containing no compound. For the negative control group (NT) not treated with any compound or bacteria, only the medium (LB medium) was used. The completed cell culture plates were maintained at 37°C under anaerobic conditions (10% H2, 10% CO2 and 80% N2) for 48 hours to culture the bacteria.

[0194] The biofilms formed on the glass slips by the bacteria cultured in each group were stained with 1% crystal violet solution for 15 minutes, washed three times with phosphate-buffered saline (PBS) solution, and decolorized with acetone-alcohol (20:80, vol / vol). 200 μl of the decolorizing solution containing crystal violet was added to each well of a 96-well microplate, and the absorbance (OD) was measured using an absorbance microplate reader (Epoch2, Bio-Tek, USA). 590nm ) were measured to compare the degree of biofilm formation, and the results are shown in Figures 7A and 7B.

[0195] 2) Inhibitory effect on biofilm formation by Pseudomonas aeruginosa and Acinetobacter The inhibitory effect of the compound of Example 1-2 on biofilm formation in Pseudomonas aeruginosa (obtained from Korea Oral Microorganism Resource Center (KCOM)) and A. baumannii (obtained from Korea Conservation Center for Microorganisms (KCCM)) was confirmed.

[0196] Sterilized cover glass slips (round, 12 mm radius) were placed one by one on the bottom of each well of a cell culture plate (24-well plate) using tweezers, and the compound of Example 1-2 was diluted in the bacterial culture medium (Pseudomonas aeruginosa: LB medium, Acinetobacter: NB medium) to final concentrations of 0.006 μg / ml, 0.06 μg / ml, and 0.6 μg / ml.

[0197] The culture medium was dispensed onto a cell culture plate with a glass slip, and 2 × 10 7 The cells were inoculated at 1000 cells / ml. For the positive control group (CT) not treated with the compound, the same number of bacteria was inoculated into the medium (LB or NB) without the compound. For the negative control group (NT) not treated with either compound or bacteria, only the medium (LB or NB) was used. The completed cell culture plate was maintained at 37°C under anaerobic conditions (10% H2, 10% CO2 and 80% N2) for 48 hours to culture the bacteria.

[0198] The biofilms formed on the glass slips by the bacteria cultured in each group were stained with 1% crystal violet solution for 15 minutes, washed three times with phosphate-buffered saline (PBS) solution, and decolorized with acetone-alcohol (20:80, vol / vol). 200 μl of the decolorizing solution containing crystal violet was added to each well of a 96-well microplate, and the absorbance (OD) was measured using an absorbance microplate reader (Epoch2, Bio-Tek, USA). 590nm ) to compare the degree of biofilm formation, and the results are shown in Figures 7C and 7D.

[0199] As can be seen from Figures 7A to 7D, after 48 hours of culture, biofilms of each bacteria (Escherichia coli, Pseudomonas aeruginosa, and Acinetobacter) increased in the control group (CT), but in the group treated with the compound of Example 1-2, it was confirmed that the biofilms were significantly suppressed in a concentration-dependent manner.

[0200] Example 6. Effect of combined use of antibiotics on E. coli 100 μl of E. coli culture medium (LB) was dispensed into cell culture plates (96-well plates), and three antibiotics (Imipenem (IPM, Sigma-Aldrich), Meropenem (MEM, Sigma-Aldrich), and Ceftriaxone (CRO, Sigma-Aldrich)) were added as a control group to final concentrations of 0.0312 μg / ml to 32 μg / ml, respectively, and serially diluted. In order to confirm whether the efficacy of the antibiotics increases due to treatment with the compound of Example 1-2, the compound of Example 1-2 was mixed in each antibiotic dilution solution to contain 1 μg / ml in the experimental groups. The final volume of the control group and the experimental group was 200 μl, and 1 × 10 E. coli was added to each well. 5 After inoculation at 1000 cells / ml, the plates were cultured at 37°C for 24-48 hours. After the culture was completed, the optical density (OD) of each well was measured using an absorbance microplate reader (Epoch2, Bio-Tek, USA). 600nm ) was measured and the minimum growth inhibitory concentration was compared between the control group and the experimental group, and the results are shown in FIG.

[0201] As can be seen from Figure 8, the minimum growth inhibitory concentrations of three antibiotics (imipenem (IPM), meropenem (MEM), and ceftriaxone (CRO)) against E. coli were reduced by 20% to 50% or more by the compounds of the Examples. This confirmed that the efficacy of existing antibiotics was improved when the compounds of the Examples of the present invention were used in combination, compared to when the antibiotics were used alone.

[0202] Example 7. Confirmation of neuronal inflammation suppression and protective effects Human neuroblastoma (SH-SY5Y cells) (obtained from Korea Cell Line Bank) was cultured in a cell culture plate (6-well plate) at 3 × 10 6 After treatment with cells / well, the cells were cultured in a 5% CO2 incubator at 37°C for 24 hours. The negative control group was treated with 3ml of culture medium alone on the neurons attached to the cell culture plate, and the positive control group was inoculated with P. gingivalis (provided by the Korea Oral Microorganism Resource Center (KCOM)) at an infection ratio of 500 (MOI 500). To confirm the effect of the compound of the examples on alleviating inflammation in neurons, the experimental group was inoculated with the same number of P. gingivalis and then treated with the compound of Example 1-2 diluted in medium to 1 μg / ml (2.95 μM).

[0203] The completed cell culture plates for each group were cultured in a 37℃, 5% CO2 incubator for 24 hours. After the culture was completed, the morphological changes of the nerve cells due to the inflammatory response were confirmed by optical microscopy for each group, and the results are shown in Figure 11.

[0204] As can be seen from FIG. 11, in nerve cells in which inflammation was induced by the infection stimulus of P. gingivalis, cell damage such as deformation of cell morphology was observed, but it was confirmed that the nerve cells were protected by treatment with the compound of the embodiment.

[0205] Real-time PCR analysis After the optical microscopy analysis, the supernatant was removed from the cell culture plate, and the remaining cells were analyzed for the inflammatory cytokine levels in the neurons for each group by real-time PCR analysis. For each group, neurons were lysed using 1 ml of easy-BLUETM Total RNA Extraction kit (iNtRON Biotechnology, Seongnam, Korea), total RNA was extracted from the lysate, cDNA was synthesized, and the expression of inflammatory cytokine genes was confirmed using real-time PCR. cDNA was synthesized using AccuPower® CycleScript RT PreMix dT20 (Bioneer, Daejeon, Korea) and amplified using Power SYBR TM Real-time (quantitative) reverse transcription PCR (RT-qPCR) was carried out using Green PCR Master Mix (Applied Biosystems, Foster City, CA, USA).

[0206] The primer sequences used are shown in Table 2 below, and the amplification conditions were enzyme activation at 95°C for 10 minutes, followed by denaturation at 95°C for 10 seconds and annealing at 60°C for 30 seconds, repeated 40 times, using a Bio-Rad CFX Connect Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA). The relative expression levels of inflammatory cytokine genes were calculated using the Comparative Ct Method (ΔΔCt) based on the expression level of the housekeeping gene GAPDH (glyceraldehyde-3-phosphate dehydrogenase), and the results are shown in Figure 12.

[0207] [Table 2]

[0208] ELISA analysis The amount of inflammatory cytokines secreted from neurons was measured by ELISA in the supernatant samples separated from the cell culture plate. The capture antibody was coated on a 96-well plate at an appropriate concentration, and then the plate was incubated at 25°C for 24 hours, and then washed three times with wash buffer. Blocking was carried out at 25°C for 1 hour using 300 μl of Reagent Diluent buffer, and the plate was washed three times with wash buffer. Then, 100 μl of test sample and standard solution were treated and incubated for 2 hours. All experimental groups were treated identically in two wells (duplicates) per group, and washed three times with wash buffer during treatment. 100 μl of detection antibody was treated at an appropriate concentration, incubated for 2 hours, and then washed three times with wash buffer. After treatment with 100 μl of streptavidin-HRP, the plate was protected from light for 20 minutes, incubated at 25°C, and then treated with 50 μl of stop solution after incubation, and the absorbance (OD 450nm ) was measured. The result was calculated as the average of the measured values, and the calculated amounts of inflammatory cytokines are shown in FIG.

[0209] As can be seen from Figures 12 and 13, it was confirmed that the gene expression and production of major inflammatory cytokines (IL-6, IL-8, TNF-α) in nerve cells was suppressed by treatment with the compound of the embodiment, thereby alleviating neuroinflammation.

[0210] Example 8. Cytotoxicity evaluation In order to confirm the cellular safety of the compounds of the examples, the cytotoxicity against the nerve cells used in the experiment was confirmed.

[0211] To evaluate whether the compounds of the examples are toxic to nerve cells, human neuroblastoma (SH-SY5Y, 3 × 10 5After dispensing 1000 cells / well, the cells were treated with the compound of Example 1-2 at concentrations of 2, 4, 8, 16, and 32 μM and incubated at 37°C in a 5% CO2 incubator for 24 hours. After 24 hours, 50 μl of CCK-8 (cell counting kit-8, Dojindo Molecular Technologies, Inc.) solution was added to each well, and the cells were incubated again at 37°C in a 5% CO2 incubator for 1-3 hours. The absorbance (OD 450nm ) to evaluate cell activity, and the cell viability of each experimental group was calculated based on the cell viability of the control group not treated with the compound of the embodiment as 100%. The results are shown in FIG. 8.

[0212] As can be seen from FIG. 14, the compounds of the examples did not show cytotoxicity in nerve cells at concentrations of 16 μM or less.

[0213] Example 9. Blood-brain barrier permeation test results The compound of Example 1-2 was diluted to a concentration of 0.4mg / ml in a vehicle of 10%DMSO+65%PEG400+25% physiological saline, and then injected into male SD rats (Zhejiang Vital River Laboratory Animal Technology Co., Ltd.) at 5ml per kg by intravenous injection (IV), and then 60μl blood samples were collected at each time (0.25h, 0.5h, 1h, 5h, 10h). The collected blood samples were centrifuged at 8000rpm for 6 minutes at 4℃, the supernatant was removed, and plasma samples were collected at each time, and stored at -50℃ before analysis, and then used for analysis.

[0214] To analyze the plasma concentrations of substances, 20 μl of each time point sample, standard sample, and QC (quality control) sample were mixed with 60 μl of acetonitrile (ACN) (Cas No. 75-05-8) containing internal standards (tolbutamide (Cas No. 64-77-7) 200 ng / ml, propranolol (Cas No. 318-98-9) 50 ng / ml, and Dic (N,N'-diisopropylcarbodiimide) (Cas No. 693-13-0) 500 ng / ml). The mixture was vortexed for 1 min, centrifuged (13000 rpm, 4°C, 10 min), and the supernatant (50 μl) was separated and transferred to a 96-well plate containing purified water (150 μl) and mixed by shaking for 10 min. The final mixture (3 μl) was injected into an LC-MS / MS system to analyze the concentrations of the compounds in the plasma samples at each time point, and the results are shown in FIG.

[0215] For brain samples, in order to obtain only samples from the brain tissue, the brain tissue was isolated after heart perfusion, and three times (w / v) saline was added to the isolated brain tissue, which was then ground together to prepare a homogenized sample solution. 20 μl of each brain sample, standard sample, and QC sample were mixed with 60 μl of acetonitrile (ACN) (Cas No. 75-05-8) containing internal standards (tolbutamide (Cas No. 64-77-7) 200 ng / ml, propranolol (Cas No. 318-98-9) 50 ng / ml, and Dic (N,N'-diisopropylcarbodiimide) (Cas No. 693-13-0) 500 ng / ml), and the mixture was vortexed for 1 min, centrifuged (13000 rpm, 4 °C, 10 min), and the supernatant (50 μl) was separated and transferred to a 96-well plate containing purified water (150 μl), and then shaken and mixed for 10 min. The final mixture (3 μl) was injected into the LC-MS / MS system to analyze the concentration of compounds in the brain samples at each time point. The compound concentration in the brain tissue was analyzed by converting the calculated concentration (ng / ml) of the sample solution into the concentration in the brain tissue (ng / g) reflecting the dilution factor, and the results are shown in FIG.

[0216] As can be seen from Figures 15 and 16, the compound of the embodiment was administered IV to a male SD rat at a dose of 2mg / kg, and it was confirmed that the compound of the embodiment traveled through the blood, passed through the blood-brain barrier, and reached the brain, and remained in the brain for up to 10 hours after administration. This confirmed that the compound of the embodiment can act on the lesion in the brain at the target site to treat neurodegenerative diseases.

[0217] Example 10. Confirmation of anti-inflammatory effect in macrophages In order to confirm the inflammation-reducing effect of the compounds of the Examples, mouse bone marrow-derived macrophages were differentiated and the change in the production of inflammatory cytokines (IL-1β) by macrophages due to treatment with the compounds was measured.

[0218] Bone marrow was isolated from the femur and tibia of 8-12 week old C57BL / 6J mice (obtained from Daehan Biolink) using a 26g syringe. The isolated bone marrow was differentiated into macrophages for 6 days in a medium composed of IMDM medium + 30% L929 cell supernatant + 10% fetal bovine serum + 1% penicillin / streptomycin + 1% non-essential amino acids + 1% sodium pyruvate in a 150pi cell culture plate, and then used for the experiment.

[0219] Differentiated macrophages were 1 × 10 6 The cells were dispensed in 200 μl portions into a 48-well plate at a concentration of 100 cells / ml, and stabilized at 37°C and 5% CO2 for 12 hours. After removing the supernatant of the cultured cells, serum-free medium was treated with 180 μl of LPS at a concentration of 100 ng / ml for 5 hours for priming. The compound of Example 1-2 was diluted to concentrations of 0.78125, 1.5625, 3.125, 6.25, 12.5, and 25 mg / ml and added in 20 μl portions (working concentrations: 0.078125, 0.15625, 0.3125, 0.625, 1.25, and 2.5 mg / ml), and after 1 hour, ATP (20 μl added at a concentration of 10 mM. 30 minutes of incubation) was added according to the conditions. After collecting samples of the cultured cell supernatant, the amount of IL-1β, a representative inflammatory cytokine released by NLRP3 inflammasome activity, was measured by ELISA and shown in Figure 17.

[0220] As can be seen from Figure 17, it was confirmed that the release of IL-1β, which was increased by a stimulant in macrophages, was inhibited in a concentration-dependent manner by the compound of Example 1-2, thereby confirming that the inflammatory response induced in macrophages was inhibited by the compound of the Example.

[0221] Example 11. Cytotoxicity evaluation To confirm the cellular safety of the compounds of the examples, their cytotoxicity was evaluated using mouse bone marrow-derived macrophages and human mononuclear cell line (THP-1) (obtained from Korea Cell Line Bank (KCLB)).

[0222] For macrophages, 1 × 10 differentiated macrophages 6 200 μl of each were seeded onto a 48-well plate at a concentration of 100 cells / ml, and stabilized for 12 hours at 37° C. and 5% CO2. After removing the supernatant of the cultured cells, 200 μl of each of the compounds of Example 1-2 diluted to concentrations of 0.5, 1, 2, 4, 8, and 16 μM were added to serum-free medium, and the cells were cultured for 24 hours at 37° C. and 5% CO2.

[0223] After that, the supernatant of the cultured cells was removed, and 200 μl of MTT solution with a concentration of 400 μg / ml was added, and the cells were reacted for 4 hours at 7°C and 5% CO2. Then, the reaction solution was removed, 200 μl of DMSO was added, and the absorbance at 570 nm was measured. At this time, the cell viability of the cells that were not treated was set as 100% as the standard, and the cell viability of the cells that were treated with the samples was calculated and shown in Figure 18.

[0224] Human mononuclear cell line (THP-1) was cultured in RPMI1640 medium (Hyclone, Waltham, MA, USA) supplemented with 10% fetal bovine serum (Hyclone, Waltham, MA, USA), 2.05 mM L-glutamic acid, and antibiotics (100 units / ml penicillin, 100 μg / ml streptomycin), and then cultured at a concentration of 1 × 10 5The cells were seeded in a 96-well microtiter plate at a concentration of 1000 cells / well, and the compound of Example 1-2 was diluted to a final concentration of 0.002 μM, 0.02 μM, 0.2 μM, and 2 μM, and the compound was treated in each well and cultured for 24 hours. After that, the cultured cells were treated with Cell Counting Kit-8 (CCK-8, Kumamoto Prefecture, Japan, DOJINDO) solution at 10 μl per well, and the absorbance at 450 nm was measured at each time (1 h, 2 h, 3 h), and the results were derived based on the absorbance data at the last time, 3 h.

[0225] At this time, the cell viability of untreated cells was set as 100% as a standard, and the cell viability of the cells treated with the samples was calculated and shown in FIG.

[0226] As can be seen from Figures 18 and 19, the compound of Example 1-2 did not show cytotoxicity at concentrations of 16 μM or less in mouse bone marrow-derived macrophages, and did not show cytotoxicity at any concentration in the human monocytic cell line (THP-1).

[0227] Example 12. Drug safety evaluation using rat micronucleus test In order to confirm the safety of the compounds of the examples in experimental rats (purchased from ORIENTBIO INC., Republic of Korea), the substances were administered to bone marrow cells of Sprague-Dawley rats, and the general symptoms, changes in body weight, and the presence or absence of micronucleus induction (genotoxicity) were evaluated.

[0228] The administration site (subcutaneous) of the SD rat was shaved, and the compound of the embodiment was administered subcutaneously to the left and right sides of the neck of the test rat using a disposable syringe at a low dose (500mg / kg / day), a medium dose (1000mg / kg / day), and a high dose (2000mg / kg / day) in separate doses (10ml / kg / day each). The general symptoms such as appearance, behavior, and excretion of each individual were observed twice a day (just before and immediately after administration) during the administration period and once a day during the other periods, and the results are shown in Table 3 below.

[0229] In addition, body weight was measured on the day administration began and on the day bone marrow was harvested to confirm whether there were any changes due to administration of the substance. The results are shown in Table 4 below.

[0230] 24 hours after the second administration of the test substance, femurs were removed from each group to collect bone marrow cells, and the bone marrow cell suspension was fixed with 0.5 mL of 10% neutral formalin for 5 minutes, and then centrifuged for 5 minutes (4°C, 1,000 rpm, Micro17TR, Hanil Science Industrial, Korea) to remove the supernatant. The precipitated bone marrow cells were then suspended in 0.3 mL of 10% neutral formalin, filtered through a cell strainer, and transferred to a storage tube. The fixed bone marrow cell suspension was dropped onto a cover glass, and covered with a slide glass coated with 20 μL of 0.05% acridine orange to prepare an observation slide, and the observation slide was observed under a microscope (600x magnification, BX51, Olympus, Japan). 4,000 polychromatic erythrocytes (PCE) were observed per individual, and the occurrence rate of micronucleated polychromatic erythrocytes (MNPCE) to polychromatic erythrocytes (MNPCE / PCE) was calculated for each individual, and the results are shown in Table 3 below.

[0231] From the observation results, it was confirmed whether the ratio of polychromatic erythrocytes to the total number of erythrocytes in the test substance group (treated with the compound of Example 1-2) was 20% or more of that in the negative control group, and the ratio of polychromatic erythrocytes to the total number of erythrocytes (PCE / (PCE+NCE)) was calculated by observing 500 total erythrocytes per individual as an index of inhibition of proliferation of bone marrow cells, and the results are shown in the following Table 5. Cyclophosphamide (Cas No: 50-18-0) was administered at a dose of 20 mg / kg / day to the positive control group.

[0232] [Table 3]

[0233] [Table 4]

[0234] [Table 5]

[0235] As can be seen from the experimental results in Table 3, no abnormal phenomena were observed in the clinical symptoms of rats in the test substance group (group administered with the compound of Example 1-2) at any administered dose of the compound of Example 1-2 during the observation period.

[0236] Furthermore, as can be seen from Table 4, at all administered doses of the compound of Example 1-2, the test substance group did not show any statistically significant change in body weight compared to the negative control group.

[0237] Furthermore, as can be seen from Table 5, in all doses of the compound of Example 1-2, the test substance group did not show a statistically significant difference in the frequency of micronucleated polychromatic erythrocytes in polychromatic erythrocytes (MNPCE / PCE) compared with the negative control group, and the ratio of polychromatic erythrocytes in total erythrocytes [PCE / (PCE+NCE)] also did not show a statistically significant difference compared with the negative control group. In contrast, in the positive control group, the ratio of polychromatic erythrocytes did not show a statistically significant difference compared with the negative control group, but the frequency of micronucleated polychromatic erythrocytes in polychromatic erythrocytes was confirmed to be statistically significantly increased compared with the negative control group (p<0.01).

[0238] Through the results, it was confirmed that the compounds of the examples were safe and did not induce micronuclei in rat bone marrow cells.

[0239] From the above description, those skilled in the art to which the present invention pertains will understand that the present invention can be implemented in other specific forms without changing its technical ideas or essential features. In this regard, it should be understood that the above-described embodiments are illustrative in all respects and are not limiting. The scope of the present invention should be interpreted as including all modifications or alterations derived from the meaning and scope of the claims below, rather than the above detailed description, and the equivalent concepts thereof.

Claims

1. A pharmaceutical composition for preventing or treating an inflammasome-mediated disease, comprising as an active ingredient one or more selected from the group consisting of a compound of the following formula 1, an isomer thereof, and a pharma- ceutically acceptable salt thereof: 【Chemistry 1】 In the above Chemical Formula 1, The above 【Chemistry 2】 is a substituted or unsubstituted C ３－７ cycloalkyl, substituted or unsubstituted C ３－７ cycloalkenyl, substituted or unsubstituted C ３－７ heterocycloalkyl, substituted or unsubstituted C containing one or more double bonds and containing one or more heteroatoms selected from the group consisting of N, O, and S; ３－７ heterocycloalkenyl of the formula (I), substituted or unsubstituted C ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ is a heteroaryl of the formula The substituted cycloalkyl, substituted cycloalkenyl, substituted heterocycloalkyl, substituted heterocycloalkenyl, substituted aryl, or substituted heteroaryl may be selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, substituted or unsubstituted C １－１０ and substituted or unsubstituted C １－１０ and optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy; The substituted alkyl or substituted alkoxy is selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ and optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy; The R ３ is a substituted or unsubstituted C １－１０ linear or branched alkyl, substituted or unsubstituted C １－１０ linear or branched chain alkoxy, substituted or unsubstituted C ３－７ cycloalkyl, substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S; ３－７ Heterocycloalkyl, substituted or unsubstituted Ｃ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ is a heteroaryl of the formula The substituted alkyl is hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ or substituted or unsubstituted C ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ and optionally substituted with a heteroaryl of the formula: The substituted alkoxy, substituted cycloalkyl, substituted heterocycloalkyl, substituted aryl, or substituted heteroaryl may be selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ and optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy; The R １ and R ２ are each independently hydrogen, halogen, or substituted or unsubstituted C １－１０ linear or branched alkyl, substituted or unsubstituted C １－１０ linear or branched chain alkoxy, substituted or unsubstituted C ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ is a heteroaryl of the formula The substituted alkyl is hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ or substituted or unsubstituted C ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ and optionally substituted with a heteroaryl of the formula: The substituted alkoxy, substituted aryl, or substituted heteroaryl may be selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ and n is 0, 1 or 2. The aryl group may be substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy.

2. The above 【Chemistry 3】 is a substituted or unsubstituted C ６－１０ is an aryl of The substituted aryl may be selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, substituted or unsubstituted C １－１０ and substituted or unsubstituted C １－１０ and optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy; The substituted alkyl or substituted alkoxy is selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ 2. The pharmaceutical composition according to claim 1, which is optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy.

3. The R ３ is a substituted or unsubstituted C １－１０ The pharmaceutical composition of claim 1, wherein the alkyl group is a straight or branched chain alkyl group.

4. The R １ and R ２ each independently represents a substituted or unsubstituted C ６－１０ is an aryl of The substituted aryl may be hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ 2. The pharmaceutical composition according to claim 1, which is optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy.

5. The pharmaceutical composition according to claim 1, wherein the compound of formula 1 is selected from the group consisting of the following compounds: 1) 3-(diphenylmethylene)-2-methylisoindolin-1-one, 2) 3-(di-p-tolylmethylene)-2-methylisoindolin-1-one, and 3) 3-(bis(4-methoxyphenyl)methylene)-2-methylisoindolin-1-one.

6. The pharmaceutical composition according to claim 1 , wherein the inflammasome-mediated disease is one or more selected from the group consisting of periodontal disease, neurodegenerative disease, and inflammatory disease.

7. The pharmaceutical composition according to claim 6, wherein the periodontal disease is one or more selected from the group consisting of periodontitis and gingivitis.

8. The pharmaceutical composition according to claim 6, wherein the neurodegenerative disease is one or more selected from the group consisting of dementia, Parkinson's disease, tremor, Tarant's disease, amyotrophic lateral sclerosis, multiple sclerosis, Huntington's disease, motor neuron disease, spinal muscular atrophy, Creutzfeldt-Jakob disease, Pick's disease, prion disease and spinocerebellar degeneration.

9. The inflammatory disease is a metabolic disease which is one or more selected from the group consisting of obesity, hyperlipidemia, hypercholesterolemia, arteriosclerosis, type 2 diabetes, nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH); autoinflammatory diseases, which are one or more selected from the group consisting of Muckle-Wells syndrome (MWS), latent adult autoimmune diabetes (LADA), familial cold autoinflammatory syndrome (FCAS), cryopyrin-associated periodic syndrome (CAPS), neonatal-onset multisystem inflammatory syndrome (NOMID), chronic infantile neurological cutaneous articular (CINCA) syndrome, familial Mediterranean fever (FMF), pediatric arthritis, pediatric rheumatoid arthritis, and gout; an autoimmune disease selected from the group consisting of rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), atopic dermatitis (AD) and psoriasis; Septic shock, inflammatory bowel disease, osteoarthritis, hyperimmunoglobulin D syndrome and cryopyrin-associated periodic syndromes The pharmaceutical composition according to claim 6, wherein the compound is one or more selected from the group consisting of:

10. A food composition for preventing or improving an inflammasome-mediated disease, comprising as an active ingredient one or more selected from the group consisting of the compound of formula 1 described in any one of claims 1 to 5, its isomer, and a pharma- ceutically acceptable salt thereof.

11. A feed composition for preventing or ameliorating an inflammasome-mediated disease, comprising as an active ingredient one or more selected from the group consisting of the compound of formula 1 described in any one of claims 1 to 5, its isomer, and a pharma-ceutically acceptable salt thereof.

12. A pharmaceutical composition for preventing or treating infection with gram-negative bacteria or a disease induced by gram-negative bacteria, comprising at least one active ingredient selected from the group consisting of a compound of the following formula 1, an isomer thereof, and a pharma- ceutical acceptable salt thereof: 【Chemistry 4】 In the above Chemical Formula 1, The above 【Chemistry 5】 is a substituted or unsubstituted C ３－７ cycloalkyl, substituted or unsubstituted C ３－７ cycloalkenyl, substituted or unsubstituted C ３－７ heterocycloalkyl, substituted or unsubstituted C containing one or more double bonds and containing one or more heteroatoms selected from the group consisting of N, O, and S; ３－７ heterocycloalkenyl of the formula (I), substituted or unsubstituted C ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ is a heteroaryl of the formula The substituted cycloalkyl, substituted cycloalkenyl, substituted heterocycloalkyl, substituted heterocycloalkenyl, substituted aryl, or substituted heteroaryl may be selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, substituted or unsubstituted C １－１０ and substituted or unsubstituted C １－１０ and optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy; The substituted alkyl or substituted alkoxy is selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ and optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy; The R ３ is a substituted or unsubstituted C １－１０ linear or branched alkyl, substituted or unsubstituted C １－１０ linear or branched chain alkoxy, substituted or unsubstituted C ３－７ cycloalkyl, substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S; ３－７ Heterocycloalkyl of the formula: ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ is a heteroaryl of the formula The substituted alkyl is hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ or substituted or unsubstituted C ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ and optionally substituted with a heteroaryl of the formula: The substituted alkoxy, substituted cycloalkyl, substituted heterocycloalkyl, substituted aryl, or substituted heteroaryl may be selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ and optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy; The R １ and R ２ are each independently hydrogen, halogen, or substituted or unsubstituted C １－１０ linear or branched alkyl, substituted or unsubstituted C １－１０ linear or branched chain alkoxy, substituted or unsubstituted C ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ is a heteroaryl of the formula The substituted alkyl is hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ or substituted or unsubstituted C ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ and optionally substituted with a heteroaryl of the formula: The substituted alkoxy, substituted aryl, or substituted heteroaryl may be selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ and n is 0, 1 or 2. The aryl group may be substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy.

13. The above 【Chemistry 6】 is a substituted or unsubstituted C ６－１０ is an aryl of The substituted aryl may be selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, substituted or unsubstituted C １－１０ and substituted or unsubstituted C １－１０ and optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy; The substituted alkyl or substituted alkoxy is selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ 13. The pharmaceutical composition according to claim 12, which is optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy.

14. The R ３ is a substituted or unsubstituted C １－１０ The pharmaceutical composition of claim 12, wherein the alkyl group is a straight or branched chain alkyl group.

15. The R １ and R ２ each independently represents a substituted or unsubstituted C ６－１０ is an aryl of The substituted aryl may be hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ 13. The pharmaceutical composition according to claim 12, which is optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy.

16. The pharmaceutical composition according to claim 12, wherein the compound of formula 1 is selected from the group consisting of the following compounds: 1) 3-(diphenylmethylene)-2-methylisoindolin-1-one, 2) 3-(di-p-tolylmethylene)-2-methylisoindolin-1-one, and 3) 3-(bis(4-methoxyphenyl)methylene)-2-methylisoindolin-1-one.

17. The pharmaceutical composition according to claim 12, wherein the gram-negative bacterial infection or disease induced by gram-negative bacteria is one or more selected from the group consisting of enteritis, Crohn's disease, ulcerative colitis, bacterial dysentery, urinary tract infection, skin infection, bacteremia, sepsis, skin infection, bedsore, pneumonia, endocarditis, meningitis, otitis externa, otitis media, periodontitis, gingivitis, keratitis, osteomyelitis, peritonitis, and cystic fibrosis.

18. A food composition for preventing or improving a gram-negative bacterial infection or a disease induced by a gram-negative bacteria, comprising at least one selected from the group consisting of the compound of formula 1 according to any one of claims 12 to 17, its isomer, and a pharma- ceutically acceptable salt thereof as an active ingredient.

19. A feed composition for preventing or ameliorating a gram-negative bacterial infection or a disease induced by a gram-negative bacteria, comprising at least one selected from the group consisting of the compound of formula 1 according to any one of claims 12 to 17, an isomer thereof, and a pharma- ceutically acceptable salt thereof as an active ingredient.

20. An antibiotic comprising at least one active ingredient selected from the group consisting of a compound of the following formula 1, an isomer thereof, and a pharma- ceutically acceptable salt thereof: 【Chemistry 7】 In the above Chemical Formula 1, The above 【Chemistry 8】 is a substituted or unsubstituted C ３－７ cycloalkyl, substituted or unsubstituted C ３－７ cycloalkenyl, substituted or unsubstituted C ３－７ heterocycloalkyl, substituted or unsubstituted C containing one or more double bonds and containing one or more heteroatoms selected from the group consisting of N, O, and S; ３－７ heterocycloalkenyl of the formula (I), substituted or unsubstituted C ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ is a heteroaryl of the formula The substituted cycloalkyl, substituted cycloalkenyl, substituted heterocycloalkyl, substituted heterocycloalkenyl, substituted aryl, or substituted heteroaryl may be selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, substituted or unsubstituted C １－１０ and substituted or unsubstituted C １－１０ and optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy; The substituted alkyl or substituted alkoxy is selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ and optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy; The R ３ is a substituted or unsubstituted C １－１０ linear or branched alkyl, substituted or unsubstituted C １－１０ linear or branched chain alkoxy, substituted or unsubstituted C ３－７ cycloalkyl, substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S; ３－７ Heterocycloalkyl of the formula: ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ is a heteroaryl of the formula The substituted alkyl is hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ or substituted or unsubstituted C ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ and optionally substituted with a heteroaryl of the formula: The substituted alkoxy, substituted cycloalkyl, substituted heterocycloalkyl, substituted aryl, or substituted heteroaryl may be selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ and optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy; The R １ and R ２ are each independently hydrogen, halogen, or substituted or unsubstituted C １－１０ linear or branched alkyl, substituted or unsubstituted C １－１０ linear or branched chain alkoxy, substituted or unsubstituted C ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ is a heteroaryl of the formula The substituted alkyl is hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ or substituted or unsubstituted C ６－１０ or substituted or unsubstituted C containing one or more heteroatoms selected from the group consisting of N, O, and S. ５－１０ and optionally substituted with a heteroaryl of the formula: The substituted alkoxy, substituted aryl, or substituted heteroaryl may be selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ and n is 0, 1 or 2. The aryl group may be substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy.

21. The above 【Chemistry 9】 is a substituted or unsubstituted C ６－１０ is an aryl of The substituted aryl may be selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, substituted or unsubstituted C １－１０ and substituted or unsubstituted C １－１０ and optionally substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy; The substituted alkyl or substituted alkoxy is selected from the group consisting of hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ The antibiotic according to claim 20, which may be substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy.

22. The R ３ is a substituted or unsubstituted C １－１０ The antibiotic of claim 20, wherein the alkyl group is a straight or branched chain alkyl group.

23. The R １ and R ２ each independently represents a substituted or unsubstituted C ６－１０ is an aryl of The substituted aryl may be hydroxy, halogen, cyano, nitro, amino, C １－５ and C １－５ The antibiotic according to claim 20, which may be substituted with one or more substituents selected from the group consisting of straight or branched chain alkoxy.

24. The antibiotic of claim 20, wherein the compound of formula 1 is selected from the group consisting of the following compounds: 1) 3-(diphenylmethylene)-2-methylisoindolin-1-one, 2) 3-(di-p-tolylmethylene)-2-methylisoindolin-1-one, and 3) 3-(bis(4-methoxyphenyl)methylene)-2-methylisoindolin-1-one.

25. The antibiotic according to claim 20, having antibiotic activity against gram-negative bacteria.

26. The antibiotic according to claim 20, wherein the gram-negative bacteria is one or more selected from the group consisting of bacteria of the genus Escherichia, bacteria of the genus Pseudomonas, bacteria of the genus Acinetobacter, bacteria of the genus Enterobacter, bacteria of the genus Klebsiella, bacteria of the genus Porphyromonas, bacteria of the genus Fusobacteria, and bacteria of the genus Tannerella.

27. An antibiotic according to claim 20; one or more second antibiotics selected from the group consisting of beta-lactam antibiotics, bacterial cell membrane permeability inhibitors, bacterial ribosome inhibitors, bacterial nucleic acid synthesis inhibitors, and bacterial folate synthesis inhibitors; Concomitant antibiotics, including:

28. A feed additive comprising the antibiotic according to claim 20.

29. A disinfectant comprising the antibiotic according to claim 20.

30. A cleaning agent comprising the antibiotic of claim 20.

31. A compound of the following formula 1A, an isomer thereof, or a pharma- ceutically acceptable salt thereof: 【Chemistry 10】 In the above Chemical Formula 1A, The above 【Chemistry 11】 R １ , R ２ , R ３ is as defined in claim 1, The compound of Formula 1A does not include the following compounds: 3-(diphenylmethylene)-2-methylisoindolin-1-one.

32. 32. The compound of claim 31, wherein the compound of formula 1A is selected from the group consisting of the following compounds, an isomer thereof, or a pharma- ceutically acceptable salt thereof: i) 3-(di-p-tolylmethylene)-2-methylisoindolin-1-one, and ii) 3-(bis(4-methoxyphenyl)methylene)-2-methylisoindolin-1-one.

33. A method for producing the compound according to claim 31 or 32, comprising the step of introducing phenylboronic acid (Cas No. 98-80-6), 4-methylphenylboronic acid (Cas No. 5720-05-8), or 4-methoxyphenylboronic acid ((4-methoxyphenyl)boronic acid (Cas No. 5720-07-0) into a compound of the following chemical formula 4. 【Chemistry 12】

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