Novel compound sensitive to nitrogen monoxide and pharmaceutical composition comprising same

Abstract

The present invention relates to a compound in which a nitric oxide scavenger is conjugated to a low-molecular-weight compound having pharmaceutical activity, and a nitric oxide-sensitive pharmaceutical composition comprising same. The compound is designed such that a nitrogen monoxide scavenger is separated while removing nitrogen monoxide only in a diseased region such as an inflammatory region having a high concentration of nitrogen monoxide, with the concomitant impartment of pharmaceutical activity to the low-molecular-weight compound in an inactivated state. The compound is activated only at the disease site, thereby minimizing side effects of existing medicines and simultaneously securing the pharmaceutical activity effect of the low molecular weight compound and the effect of removing nitrogen monoxide, and thus can be used for preventing or treating diseases accompanied by nitrogen monoxide generation.

Description

Novel nitric oxide-sensitive compound and pharmaceutical composition containing the same

[0001] This application is based upon and claims priority to Korean Patent Application No. 10-2023-0086949, filed with the Korean Intellectual Property Office on July 5, 2023, the entire contents of which are incorporated herein by reference.

[0002] The present invention relates to a nitric oxide-sensitive pharmaceutical composition, wherein the pharmaceutical composition comprises a nitric oxide scavenger moiety that detects and removes nitric oxide and a low-molecular-weight compound having a separate pharmaceutical activity, so that in normal tissues where the concentration of nitric oxide is low, the activity of the low-molecular-weight compound having the pharmaceutical activity is minimized, and only in areas of inflammatory diseases where the concentration of nitric oxide is high, etc., while removing nitric oxide, the nitric oxide scavenger is separated from the low-molecular-weight compound, thereby locally and selectively exerting the pharmaceutical activity of the low-molecular-weight compound only in the target area, thereby minimizing side effects and maximizing the efficacy, and can be usefully used for the prevention or treatment of all diseases related to nitric oxide.

[0003] Nitric oxide is a highly reactive radical molecule with a short half-life of less than 5 seconds, and acts as a major mediator of cell signaling, and is associated with various diseases such as inflammatory diseases, cancer growth, and metastasis.

[0004] Specifically, inflammatory diseases include rheumatism, osteoarthritis, asthma, inflammatory bowel disease (UC), Crohn's disease, inflammatory diseases caused by bacterial infection, inflammatory diseases caused by viral infection, inflammatory diseases caused by parasitic infection, and septic shock. Neurological diseases such as Alzheimer's disease, amyotrophic sclerosis, Huntington's disease, Parkinson's disease, and stroke are linked to nitric oxide. In addition, cardiovascular diseases such as hypertension and heart failure, as well as cancer, obesity, and hepatic encephalopathy are also known to be associated with nitric oxide.

[0005] The purpose of the present invention is to provide a pharmaceutical composition for the prevention or treatment of all diseases related to nitric oxide by minimizing side effects and toxicity of the low-molecular-weight compound and maximizing the efficacy by locally and selectively removing nitric oxide only from a diseased area with a high concentration of nitric oxide and releasing a low-molecular-weight compound having pharmaceutical activity.

[0006] However, the problems to be solved by the present invention are not limited to the problems mentioned above, and other problems not mentioned will be clearly understood by those skilled in the art from the description below.

[0007] All combinations of the various elements disclosed herein fall within the scope of the present invention. Furthermore, the scope of the present invention is not limited by the specific description below.

[0008] In order to achieve the above purpose, the present invention provides a pharmaceutical composition for preventing or treating all diseases related to nitrogen monoxide, which comprises a low molecular weight compound combined with a nitrogen monoxide scavenger as an active ingredient.

[0009] The compound according to the present invention is a substance that combines a nitrogen monoxide scavenger and a low molecular weight compound having pharmaceutical activity, and can combine an amine group or hydroxyl group present in the low molecular weight compound with the nitrogen monoxide scavenger using an organic synthesis reaction.

[0010] One embodiment of the present invention provides a compound represented by the following chemical formula 1, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof:

[0011] [Chemical Formula 1]

[0012]

[0013] In the above chemical formula 1, X is a low molecular weight compound that contains an amine group or a hydroxyl group in the molecular structure of the compound and has pharmaceutical activity, and may include, for example, a kinase inhibitor, particularly a Janus kinase inhibitor (JAK inhibitor), and other anticancer agents, anti-inflammatory agents, etc.

[0014] The above kinase inhibitors may include Netarsudil (ROCK1 / 2, glaucoma treatment), Fostamatinib (Syk, thrombocytopenia treatment), and Belumosudil (ROCK2, graft-versus-host disease treatment).

[0015] The above Janus kinase inhibitor may be selected from the group consisting of Tofacitinib, Upadacitinib, Baricitinib, Filgotinib, Abrocitinib, Delgocitinib, Oclacitinib, Peficitinib, and Ruxolitinib.

[0016] The above anticancer drugs include Doxorubicin, Cyclophosphamide, Cisplatin, Oxaliplatin, 5-Fu (5-Fluorouracil), Gemcitabine, Paclitaxel, Docetaxel, Irinotecan, MMAE (Monomethyl auristatin E), Crizotinib, Osimertinib, Sorafenib, Ibrutinib, Ruxolitinib, Vemurafenib, Ceritinib, Alectinib, Brigatinib, Lorlatinib, Capmatinib, and Gefitinib. Gefitinib, Erlotinib, Lapatinib, Icotinib, Afatinib, Neratinib, Dacomitinib, Almonertinib, Tucatinib, Midostaurin, Gilteritinib, Quizartinib, Pexidartinib, Sunitinib, Pazopanib, Vandetanib, Axitinib, Cabozantinib, Regorafenib, Apatinib, Lenvatinib, Tivozanib, Fruquintinib (Fruquintinib), Nintedanib, Anlotinib, Erdafitinib, Pemigatinib, Avapritinib,Ripretinib, Pralsetinib, Larotrectinib, Entrectinib, Imatinib, Dasatinib, Nilotinib, Bosutinib, Radotinib, Ponatinib, Acalabrutinib, Zanubrutinib, Fedratinib, Dabrafenib, Encorafenib, Trametinib, Cobimetinib, Binimetinib, Selumetinib, Palbociclib, Ribociclib Abemaciclib, Idelalisib, Copanlisib, Duvelisib, Alpelisib, Tazemetostat, Vorinostat, Belinostat, Tucidinostat, Panobinostat, Enasidenib, Ivosidenib, Venetoclax, Vismodegib, Sonidegib, Glasdegib, Bortezomib, Carfilzomib, Ixazomib, Olaparib, Rucaparib, Niraparib Talazoparib, Umbralisib, Trilaciclib, Infigratinib, Mobocertinib, Asciminib,It may be selected from the group consisting of futibatinib, pacritinib, and everolimus.

[0017] The above anti-inflammatory agent may be selected from the group consisting of dexamethasone, methotrexate, cyclosporine, acetaminophen, etodolac, piroxicam, aceclofenac, etc.

[0018] In the above chemical formula 1, Y is hydrogen, C1-C6 alkyl group, C1-C6 alkoxy group, hydroxy group, nitro group, amino group, halogen, thiol group, cyano group and Can be selected from among.

[0019] The above n is an integer from 0 to 4.

[0020] The above R1 is a C1-C6 alkyl group, a C1-C6 alkoxy group, a hydroxy group, a nitro group, an amino group, a halogen, a thiol group, a cyano group, a C3-C 14 Aryl group, C3-C 14 Heteroaryl group, -NR 11 R 12 , -NR 12 C(=O)R 12 , -C(=O)R 11 and -C(=O)OR 11 and when there are two or more R1, they are the same or different from each other, and two adjacent R1 can be arbitrarily combined to form a fused ring. Here, the aryl group or heteroaryl group is unsubstituted or has at least one substituent independently selected from the group consisting of halogen, hydroxyl group, thiol group, cyano group, C1-C6 alkyl group and C1-C6 alkoxy group, and R 11 and R 12 are each independently hydrogen, halogen, CF3, C1-C6 alkyl group, C3-C14 Aryl group and C3-C 14 It can be selected from among heteroaryl groups.

[0021] In the above chemical formula 1, Z is , , and Can be selected from among.

[0022] In the above chemical formula 1, m is an integer from 0 to 4.

[0023] The above R2 is a C1-C6 alkyl group, a C1-C6 alkoxy group, a hydroxy group, a nitro group, an amino group, a halogen, a thiol group, a cyano group, a C3-C 14 Aryl group, C3-C 14 Heteroaryl group, -NR 21 R 22 , -NR 22 C(=O)R 22 , -C(=O)R 21 and -C(=O)OR 21 and when there are two or more R2, they are the same or different from each other, and two adjacent R2 can be arbitrarily combined to form a fused ring. Here, the aryl group or heteroaryl group is unsubstituted or has at least one substituent independently selected from the group consisting of halogen, hydroxyl group, thiol group, cyano group, C1-C6 alkyl group and C1-C6 alkoxy group, and R 21 and R 22 are each independently hydrogen, halogen, CF3, C1-C6 alkyl group, C3-C 14 Aryl group and C3-C 14 It can be selected from among heteroaryl groups.

[0024] The above R2 may be the same as the definition of the above R1. The above R 21 and the above R 22 are respectively the above R 11 and the above R 12 It may be identical to the definition of .

[0025] The above R3 and R4 are each independently selected from hydrogen, halogen, hydroxyl group, thiol group, cyano group, C1-C6 alkyl group and C1-C6 alkoxy group, or R3 and R4 may be combined to form a fused ring.

[0026] Another embodiment of the present invention provides a compound represented by the following chemical formula 2, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof:

[0027] [Chemical Formula 2]

[0028]

[0029] In the above chemical formula 2, R2 and m have the same definitions as in the above chemical formula 1.

[0030] The compound represented by the above chemical formula 2 may be a compound described in Table 1 below, or a solvate, hydrate, stereoisomer, or a pharmaceutically acceptable salt thereof, wherein X may be tofacitinib.

[0031] Compound (1)Compound (2) Compound (3)Compound (4) Compound (5)Compound (6) Compound (7)Compound (8) Compound (9)Compound (10)

[0032]

[0033] The compound represented by the above chemical formula 2 may be a compound described in Table 2 below, or a solvate, hydrate, stereoisomer, or a pharmaceutically acceptable salt thereof, wherein X may be Upadacitinib.

[0034] Compound (29)Compound (30) compound (31)

[0035]

[0036] The compound represented by the above chemical formula 2 may be a compound described in Table 3 below, or a solvate, hydrate, stereoisomer, or a pharmaceutically acceptable salt thereof, wherein X may be dexamethasone.

[0037] Compound (33)Compound (34)

[0038]

[0039] The compound represented by the above chemical formula 2 may be a compound described in Table 4 below, or a solvate, hydrate, stereoisomer, or a pharmaceutically acceptable salt thereof, wherein X may be cyclosporine.

[0040] Compound (35)

[0041]

[0042] Another embodiment of the present invention provides a compound represented by the following chemical formula 3, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof:

[0043] [Chemical Formula 3]

[0044]

[0045] In the above chemical formula 3, R1, R2, n and m have the same definitions as in the above chemical formula 1.

[0046] The compound represented by the above chemical formula 3 may be a compound described in Table 5 below, or a solvate, hydrate, stereoisomer, or a pharmaceutically acceptable salt thereof, wherein X may be tofacitinib.

[0047] Compound (11)Compound (12) compound (13)

[0048]

[0049] Another embodiment of the present invention provides a compound represented by the following chemical formula 4, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof:

[0050] [Chemical Formula 4]

[0051]

[0052] In the above chemical formula 4, R2 and m have the same definitions as in the above chemical formula 1.

[0053] The compound represented by the above chemical formula 4 may be a compound described in Table 6 below, or a solvate, hydrate, stereoisomer, or a pharmaceutically acceptable salt thereof, wherein X may be tofacitinib.

[0054] Compound (14)Compound (15) Compound (16)Compound (17) Compound (18)Compound (19) Compound (20)Compound (21) Compound (22)

[0055]

[0056] Another embodiment of the present invention provides a compound represented by the following chemical formula 5, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof:

[0057] [Chemical Formula 5]

[0058]

[0059] In the above chemical formula 5, R2, R3, R4 and m are the same as defined in the above chemical formula 1.

[0060] The compound represented by the above chemical formula 5 may be a compound described in Table 7 below, or a solvate, hydrate, stereoisomer, or a pharmaceutically acceptable salt thereof, wherein X may be tofacitinib.

[0061] Compound (23)Compound (24)

[0062]

[0063] Another embodiment of the present invention provides a compound represented by the following chemical formula 6, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof:

[0064] [Chemical Formula 6]

[0065]

[0066] In the above chemical formula 6, R2, R3, R4 and m have the same definitions as in the above chemical formula 1.

[0067] The compound represented by the above chemical formula 6 may be a compound described in Table 8 below, or a solvate, hydrate, stereoisomer, or a pharmaceutically acceptable salt thereof, wherein X may be tofacitinib.

[0068] Compound (25)Compound (26) Compound (27)Compound (28)

[0069]

[0070] The compound represented by the above chemical formula 6 may be a compound described in Table 9 below, or a solvate, hydrate, stereoisomer, or a pharmaceutically acceptable salt thereof, wherein X may be Upadacitinib.

[0071] compound (32)

[0072]

[0073] Another embodiment of the present invention provides a pharmaceutical composition for the prevention or treatment of a disease involving nitrogen monoxide generation, comprising a compound represented by the above chemical formula 1, or a solvate, hydrate, stereoisomer of the compound, or a pharmaceutically acceptable salt thereof. Specifically, the pharmaceutical composition for the prevention or treatment of a disease involving nitrogen monoxide generation may comprise at least one of the compounds represented by the above chemical formulas 1 to 6, or a solvate, hydrate, stereoisomer thereof, or a pharmaceutically acceptable salt thereof. The pharmaceutical composition may be a nitric oxide-sensitive pharmaceutical composition. The nitric oxide-sensitive pharmaceutical composition may remove at least a portion of the nitrogen monoxide from a site of a disease involving nitrogen monoxide generation, and release a compound having pharmaceutical activity to the site of the disease.

[0074] The disease involving the generation of the above-mentioned nitric oxide may be selected from the group consisting of inflammatory diseases, nervous system diseases, cardiovascular diseases, autoimmune diseases, allergic diseases, cancer, obesity, myelofibrosis, and hepatic encephalopathy.

[0075] The above inflammatory diseases may be selected from the group consisting of all inflammatory diseases caused by increased concentrations of nitric oxide, such as inflammatory diseases caused by infection (inflammatory diseases caused by viral infection, inflammatory diseases caused by bacterial infection, sepsis, etc.), degenerative inflammatory diseases (osteoarthritis), inflammatory diseases caused by injury, autoimmune diseases (rheumatoid arthritis, ulcerative bowel disease, Crohn's disease, systemic lupus erythematosus, psoriasis, multiple sclerosis, juvenile idiopathic arthritis), allergic diseases (asthma, rhinitis, atopic dermatitis), cancer (lung cancer, breast cancer, colon cancer, stomach cancer, liver cancer, brain cancer, pancreatic cancer, thyroid cancer, skin cancer, bone marrow cancer, lymphoma, uterine cancer, cervical cancer, ovarian cancer, kidney cancer, melanoma, etc.), and other diseases caused by abnormal overactivation of immune cells, such as myelofibrosis.

[0076] The above neurological disease may be selected from the group consisting of Alzheimer's disease, amyotrophic sclerosis, Huntington's disease, Parkinson's disease, and stroke.

[0077] The above cardiovascular disease may be hypertension or heart failure.

[0078] The pharmaceutical composition may be administered orally or parenterally. When administered parenterally, the pharmaceutical composition may be administered intra-articularly, intravenously, subcutaneously, intramuscularly, intraperitoneally, intranasally, intrapulmonary, or rectally.

[0079] The pharmaceutical composition may be administered as a pharmaceutical composition by any conventional route, orally (e.g., in the form of tablets or capsules), parenterally (e.g., in the form of injectable solutions or suspensions), topically (e.g., in the form of lotions, gels, ointments, or creams), or in the form of nasal drops or suppositories. A pharmaceutical composition comprising the compound of the present invention in free form or in the form of a pharmaceutically acceptable salt together with at least one pharmaceutically acceptable carrier or diluent may be prepared by a conventional method, such as mixing, granulating, or coating. For example, the oral composition may be a tablet, granule, or capsule containing an excipient, a disintegrant, a binder, a lubricant, and the like, and an active ingredient, etc. In addition, the injectable composition may be a solution or suspension, may be sterilized, and may further contain a preservative, a stabilizer, a buffer, etc.

[0080] In order to use the pharmaceutical composition above for actual treatment and obtain the desired therapeutic effect, the dosage of the compound represented by the chemical formula 1 of the present invention, or a solvate, hydrate, stereoisomer thereof, or a pharmaceutically acceptable salt thereof, which is the active ingredient, may be determined according to the patient's age, sex, body weight, the type of disease accompanied by the generation of nitric oxide, the extent of treatment, etc. For example, in the case of oral administration, the dosage may be generally in the range of 0.001 to 3,000 mg / kg per day for an adult (assuming a body weight of 60 kg), administered once or in several divided doses, and may be administered orally or parenterally once or in several doses every two days, once a week, or once a month. It will be understood that the dosage may be determined depending on the type of the low molecular weight compound represented by X. It should be understood that the dosage for a specific individual or patient should be determined in light of various related factors such as the patient's weight, age, sex, health condition, diet, administration time, administration method, and severity of the disease, and may be appropriately increased or decreased by a specialist, and the above dosage is not intended to limit the scope of the present invention in any way.

[0081] The compounds of the present invention may be used in combination with other therapeutic agents. The compounds may be administered in combination with a therapeutic agent selected from the group consisting of cytotoxic drugs, radiotherapy, and immunotherapy.

[0082] Another embodiment of the present invention provides a method for treating a disease involving the generation of nitric oxide, comprising administering to a subject a pharmaceutical composition comprising a compound represented by the above chemical formula 1, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof of the compound.

[0083] The pharmaceutical composition for preventing or treating a disease involving the generation of nitrogen monoxide of the present invention can be used as a preventive or therapeutic agent for a disease related to nitrogen monoxide by selectively releasing the drug only to a diseased area with a high concentration of nitrogen monoxide.

[0084] Figure 1 is a conceptual diagram showing the operating principle of a nitric oxide-sensitive pharmaceutical composition according to one embodiment of the present invention.

[0085] Figure 2 shows the HPLC results of compound (4) prepared in Example 4 according to the presence or absence of nitrogen monoxide. Figure 2a shows the HPLC results of compound (4) in an environment without nitrogen monoxide. Figure 2b shows the HPLC results of compound (4) reacting in an environment with an excess of nitrogen monoxide. Figure 2c shows the HPLC results for tofacitinib.

[0086] Figure 3 is NMR data of compound (35) prepared in Example 35.

[0087] Figure 4 is a graph evaluating the efficacy of compound (1) prepared in Example 1 for inflammatory bowel disease.

[0088] Figure 5 is a graph evaluating the efficacy of compound (4) prepared in Example 4 for inflammatory bowel disease.

[0089] Figure 6 is a graph evaluating the efficacy of compound (1) prepared in Example 1 for rheumatoid arthritis.

[0090] Figure 7 is a graph showing the efficacy evaluation of compound (11) prepared in Example 11 for rheumatoid arthritis.

[0091] The present invention can be more readily understood by reference to the following detailed description of preferred embodiments and examples included herein. It should be understood that the terminology used herein is for the purpose of describing specific embodiments only and is not intended to be limiting. Furthermore, unless specifically defined herein, terms used herein should be understood to have their conventional meanings as known in the relevant art.

[0092] As used herein, the singular forms include the plural forms unless otherwise indicated. For example, a substituent includes one or more substituents.

[0093] As used herein, the term “halo” or “halogen” refers to fluorine (F), chlorine (Cl), bromine (Br), or iodine (I), unless otherwise stated.

[0094] As used herein, the term "alkyl" refers to a saturated monovalent aliphatic hydrocarbon radical, including straight and branched chains, having a specific number of carbon atoms, unless otherwise stated. An alkyl group typically has from 1 to 20 carbon atoms ("C1-C 20 alkyl"), preferably having 1 to 12 carbon atoms ("C1-C 12 Alkyl"), more preferably containing 1 to 8 carbon atoms ("C1-C8 alkyl"), or 1 to 6 carbon atoms ("C1-C6 alkyl"), or 1 to 4 carbon atoms ("C-1-C4 alkyl"). Examples of alkyl groups include methyl, ethyl, n-propyl, isopropyl, n-butyl, iso-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl, n-heptyl, n-octyl, and the like.

[0095] As used herein, the term "alkoxy", unless otherwise stated, refers to a monovalent -O-alkyl group wherein the alkyl moiety has the specified number of carbon atoms. Alkoxy groups typically have 1 to 8 carbon atoms ("C1-C8 alkoxy"), or 1 to 6 carbon atoms ("C1-C6 alkoxy"), or 1 to 4 carbon atoms ("C1-C4 alkoxy"). For example, C1-C4 alkoxy includes methoxy (-OCH3), ethoxy (-OCH2CH3), isopropoxy (-OCH(CH3)2), tert-butyloxy (-OC(CH3)3), and the like. The alkoxy group is optionally substituted on the alkyl moiety by the same groups described herein as appropriate for alkyl. In particular, the alkoxy group may be optionally substituted with one or more halo atoms, particularly one or more fluoro atoms, up to the total number of hydrogen atoms present on the alkyl moiety. Such groups are referred to as "haloalkoxy" groups having a specific number of carbon atoms and substituted with one or more halo substituents, e.g., when fluorinated, more specifically as "fluoroalkoxy" groups, typically containing from 1 to 6 carbon atoms, preferably from 1 to 4 carbon atoms, often 1 or 2 carbon atoms, and 1, 2 or 3 halo atoms (i.e., "C1-C6 haloalkoxy", "C1-C4 haloalkoxy" or "C1-C2 haloalkoxy"). More specifically, fluorinated alkyl groups can be specifically referred to as fluoroalkoxy groups, typically substituted with 1, 2 or 3 fluoro atoms, e.g., C1-C6, C1-C4 or C1-C2 fluoroalkoxy groups. Therefore, C1-C4 fluoroalkoxy includes trifluoromethyloxy (-OCF3), difluoromethyloxy (-OCF2H), fluoromethyloxy (-OCFH2), difluoroethyloxy (-OCH2CF2H), etc.

[0096] As used herein, the term "aryl" or "aromatic", unless otherwise stated, refers to an optionally substituted or unsubstituted monocyclic or fused bicyclic or polycyclic ring system having the well-known characteristic of aromaticity, wherein at least one ring contains a fully conjugated pi-electron system. Typically, an aryl group has from 3 to 20 carbon atoms ("C3-C") as ring members. 20 aryl"), preferably having 6 to 14 carbon atoms ("C6-C 14 aryl"), more preferably 6 to 12 carbon atoms ("C6-C 12 A fused aryl group may comprise an aryl ring (e.g., a phenyl ring) fused to another aryl or heteroaryl ring, or to a saturated or partially unsaturated carbocyclic or heterocyclic ring, wherein the point of attachment to the base molecule on such a fused ring system is an atom of the aromatic portion of the ring system. For example, an aryl group includes phenyl, biphenyl, naphthyl, toluyl, anthracenyl, phenanthrenyl, indanyl, indenyl, or tetrahydronaphthyl. The aryl group is optionally substituted as further described herein.

[0097] As used herein, the term "heteroaryl" or "heteroaromatic" refers to a monocyclic or fused bicyclic or polycyclic ring system having the well-known characteristic of aromaticity, containing a specified number of ring atoms as ring members within the aromatic ring and including one or more heteroatoms selected from B, N, O, S, P and Si. The inclusion of heteroatoms allows aromaticity in 5-membered and 6-membered rings. Typically, heteroaryl groups have from 5 to 20 ring atoms ("5- to 20-membered heteroaryl"), preferably from 5 to 14 ring atoms ("5- to 14-membered heteroaryl"), and more preferably from 5 to 12 ring atoms ("5- to 12-membered heteroaryl"). The heteroaryl ring is attached to the base molecule through a ring atom of the heteroaromatic ring such that aromaticity is maintained. Thus, a six-membered heteroaryl ring can be attached to the base molecule via a ring C atom, whereas a five-membered heteroaryl ring can be attached to the base molecule via a ring C or N atom. Heteroaryl groups can also be fused to other aryl or heteroaryl rings, or to saturated or partially unsaturated carbocyclic or heterocyclic rings, provided that the point of attachment to the base molecule on such fused ring systems is an atom of the heteroaromatic portion of the ring system. For example, unsubstituted heteroaryl groups include pyrrole, furan, thiophene, pyrazole, imidazole, isoxazole, oxazole, isothiazole, thiazole, triazole, oxadiazole, thiadiazole, tetrazole, pyridine, pyridazine, pyrimidine, pyriazine, benzofuran, benzothiophene, indole, benzimidazole, indazole, quinoline, isoquinoline, purine, triazine, naphthyridine, or carbazole. In various embodiments, the 5 or 6 membered heteroaryl group is selected from the group consisting of pyrrolyl, furanyl, thiophenyl, pyrazolyl, imidazolyl, isoxazolyl, oxazolyl, isothiazolyl, thiazolyl, triazolyl, pyridinyl, and pyrimidinyl, pyrazinyl, or pyridazinyl rings.The heteroaryl group is substituted or unsubstituted as further described herein.

[0098]

[0099] Hereinafter, the present invention will be described in detail through examples and experimental examples. However, the following examples and experimental examples are merely illustrative of the present invention and the scope of the present invention is not limited thereto.

[0100]

[0101] Example 1: Preparation of compound (1) (N-(2-aminophenyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0102] [Reaction Formula 1]

[0103]

[0104] Step (1): Preparation of compound (1-1)

[0105] 300 mg of tofacitinib (0.96 mmol, 1 equiv) and 394 mg of bis(4-nitrophenyl) carbonate (1.30 mmol, 1.35 equiv) were added to 8 mL of acetonitrile, and the mixture was cooled to 0–5°C. 229 μL of DIPEA (1.34 mmol, 1.4 equiv) was added to the mixture, and the mixture was stirred at room temperature for 4 hours. After the mixture was cooled to 0–5°C, 214 mg of tert-butyl (2-aminophenyl)carbamate (0.96 mmol, 1 equiv) dissolved in 6 mL of dichloromethane was added dropwise, and the mixture was stirred at room temperature for 16 hours. After the reaction was completed, the reaction solution was extracted by adding water and dichloromethane. After separating the organic layer, drying it with magnesium sulfate and filtering it, and concentrating the remaining filtrate under reduced pressure. The concentrate was purified by silica gel column chromatography (ethyl acetate: hexane = 1:1) to obtain the target compound (1-1). (Yield 19%)

[0106] Step (2): Preparation of compound (1)

[0107] 34 mg of compound (1-1) (0.06 mmol, 1 equiv) was dissolved in 1 mL of dioxane, and the mixture was cooled to 0-5°C. 130 μL of hydrochloric acid (4 M dioxane solution, 0.44 mmol, 7.25 equiv) was added to the mixture, and the mixture was stirred at room temperature for 16 hours. After completion of the reaction, the reaction solution was cooled to 0-5°C, and 5% NaHCO3 aqueous solution was added dropwise to neutralize. Water and ethyl acetate were added to the mixture, and extraction was performed. The organic layer was separated, dried over magnesium sulfate, and filtered, and the remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (ethyl acetate: hexane = 1: 1) to obtain the target compound (1). (Yield 99%)

[0108] 1 H-NMR (300 MHz, DMSO-d6) δ 11.55 (s, 1H), 8.42 (d, 1H, J=3 Hz), 8.12 (d, 1H, J=9 Hz), 7.77 (d, 1H, J=3 Hz), 7.61 (d, 1H, J=6 Hz), 7.29 - 7.18 (m, 3H), 6.96 - 6.91 (m, 2H), 4.21 - 3.62 (m, 4H), 3.49 - 3.38 (m, 2H), 3.33 (s, 3H), 2.41 (m, 1H), 1.98 - 1.60 (m, 3H), 1.02 (d, 3H, J=6 Hz).

[0109]

[0110] Example 2: Preparation of compound (2) (N-(2-amino-5-methylphenyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0111] [Reaction Formula 2]

[0112]

[0113] Step (1): Preparation of tert-Butyl (4-methyl-2-nitrophenyl)carbamate

[0114] 1 g of 4-methyl-2-nitroaniline (6.57 mmol, 1 equiv) was dissolved in 18 mL of anhydrous THF, and the mixture was cooled to 0-5°C. 578 mg of NaH (60% content) (14.46 mmol, 2.2 equiv) was added, and the mixture was stirred for 10 minutes. The mixture was warmed to room temperature and stirred for 30 minutes. 1.58 g of (Boc)2O (7.23 mmol, 1.1 equiv) dissolved in 1.8 mL of anhydrous THF was added to the mixture, and the mixture was stirred for 16 hours. After the reaction was completed, water and ethyl acetate were added to the reaction solution and extracted. The organic layer was separated, dried over magnesium sulfate, filtered, and the remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (ethyl acetate: hexane = 1:100) to obtain the target compound, tert-butyl (4-methyl-2-nitrophenyl)carbamate (yield 92%).

[0115] Step (2): Preparation of compound (2-1)

[0116] 465 mg of tert-butyl (4-methyl-2-nitrophenyl)carbamate (1.84 mmol, 1 equiv) was dissolved in 32 mL of methanol. 47 mg of 10% Pd / C was added to the mixture at room temperature and stirred for 2 hours under hydrogen gas at atmospheric pressure. After completion of the reaction, the mixture was purified through a Celite filter and washed with ethyl acetate. The filtrate was concentrated under reduced pressure to obtain the target compound (2-1). (Yield: 99%)

[0117] Step (3): Preparation of compound (2-2)

[0118] In step (1) of the above Example 1, compound (2-1) was used instead of tert-butyl (2-aminophenyl)carbamate, and the same procedure was repeated to obtain the target compound, compound (2-2). (Yield: 74%)

[0119] Step (4): Preparation of compound (2)

[0120] In step (2) of the above Example 1, compound (2-2) was used instead of compound (1-1) and the same procedure was repeated to obtain the target compound, compound (2). (Yield: 42%)

[0121] 1 H-NMR (300 MHz, DMSO-d6) δ 11.21 (s, 1H), 8.38 (d, 1H, J=3 Hz), 7.74 (d, 1H, J=3 Hz), 7.29 (s, 1H), 6.91 - 6.72 (m, 3H), 4.79 (s, 2H), 4.13 - 3.42 (m, 6H), 3.32 (s, 3H), 2.49 (m, 1H), 2.19 (s, 3H), 1.89 - 1.59 (m, 3H), 1.02 (d, 3H, J=6 Hz).

[0122]

[0123] Example 3: Preparation of compound (3) (N-(2-amino-4-methylphenyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0124] [Reaction Formula 3]

[0125]

[0126] Step (1): Preparation of tert-Butyl (5-methyl-2-nitrophenyl)carbamate

[0127] In step (1) of the above Example 2, 5-methyl-2-nitroaniline was used instead of 4-methyl-2-nitroaniline and the same procedure was repeated to obtain the target compound, tert-butyl (5-methyl-2-nitrophenyl)carbamate. (Yield 56%)

[0128] Step (2): Preparation of compound (3-1)

[0129] In step (2) of the above Example 2, tert-butyl (5-methyl-2-nitrophenyl)carbamate was used instead of tert-butyl (4-methyl-2-nitrophenyl)carbamate, and the same procedure was repeated to obtain the target compound (3-1). (Yield 97%)

[0130] Step (3): Preparation of compound (3-2)

[0131] In step (1) of the above Example 1, compound (3-1) was used instead of tert-butyl (2-aminophenyl)carbamate, and the same procedure was repeated to obtain the target compound, compound (3-2). (Yield: 64%)

[0132] Step (4): Preparation of compound (3)

[0133] In step (2) of the above Example 1, compound (3-2) was used instead of compound (1) and the same procedure was repeated to obtain the target compound, compound (3). (Yield: 67%)

[0134] 1H-NMR (300 MHz, DMSO-d6) δ 11.09 (s, 1H), 8.38 (d, 1H, J=3 Hz), 7.74 (d, 1H, J=3 Hz), 7.28 - 7.25 (m, 1H), 6.89 (d, 1H, J=6 Hz), 6.64 (s, 1H), 6.46 (d, 1H, J=9 Hz), 4.94 (s, 2H), 4.13 - 3.42 (m, 6H), 3.32 (s, 3H), 2.41 - 2.39 (m, 1H), 2.20 (s, 3H), 1.86 - 1.59 (m, 3H), 1.01 (d, 3H, J=6 Hz)

[0135]

[0136] Example 4: Preparation of compound (4) (N-(2-amino-4-(dimethylamino)phenyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0137] [Reaction Formula 4]

[0138]

[0139] Step (1): Preparation of tert-Butyl (5-(dimethylamino)-2-nitrophenyl)carbamate

[0140] In step (1) of the above Example 2, N1,N1-dimethyl-4-nitrobenzene-1,3-diamine was used instead of 4-methyl-2-nitroaniline, and the same procedure was repeated to obtain the target compound tert-butyl (5-(dimethylamino)-2-nitrophenyl)carbamate (yield 52%).

[0141] Step (2): Preparation of compound (4-1)

[0142] In step (2) of the above Example 2, tert-butyl (5-(dimethylamino)-2-nitrophenyl)carbamate was used instead of tert-butyl (4-methyl-2-nitrophenyl)carbamate, and the same procedure was repeated to obtain the target compound (4-1). (Yield 91%)

[0143] Step (3): Preparation of compound (4-2)

[0144] In step (1) of the above Example 1, compound (4-1) was used instead of tert-butyl (2-aminophenyl)carbamate, and the same procedure was repeated to obtain the target compound, compound (4-2). (Yield 59%)

[0145] Step (4): Preparation of compound (4)

[0146] In step (2) of the above Example 1, compound (4-2) was used instead of compound (1) and the same procedure was repeated to obtain the target compound, compound (4). (Yield: 77%)

[0147] 1 H-NMR (300 MHz, DMSO-d6) δ 10.87 (s, 1H), 8.37 (d, 1H, J=3 Hz), 7.73 (d, 1H, J=3 Hz), 7.10 (d, 1H, J=9 Hz), 6.88 (d, 1H, J=3 Hz), 6.19 (d, 1H, J=3 Hz), 6.06 (dd, 1H, 12, 3 Hz), 4.84 (s, 2H), 4.20 - 3.41 (m, 6H), 3.32 (s, 3H), 2.84 (s, 6H), 2.41 (m, 1H), 2.20 (s, 3H), 1.86 - 1.59 (m, 3H), 1.02 (d, 3H, J=9 Hz).

[0148]

[0149] Example 5: Preparation of compound (5) (N-(2-amino-5-(dimethylamino)phenyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0150] [Reaction Formula 5]

[0151]

[0152] Step (1): Preparation of compound (5-2)

[0153] To a toluene (10 mL) solution containing 1 g of compound (5-1) (3.15 mmol, 1 equiv) were added Me2NH-HCl (1.29 g, 15.7 mmol, 5.00 eq), XPhos (150 mg, 315 μmol, 0.1 eq), t-BuONa (909 mg, 9.46 mmol, 3.00 eq), and Pd2(dba)3 (144 mg, 157 μmol, 0.05 eq). The mixture was stirred at 90°C for 12 hours. After completion of the reaction, the reaction solution was extracted with water and ethyl acetate. The organic layer was separated, dried over magnesium sulfate, filtered, and the remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (ethyl acetate: petroleum ether = 1:100 to 1:0) to obtain the target compound (5-2). (Yield 29%)

[0154] Step (2): Preparation of compound (5-3)

[0155] 390 mg of compound (5-2) (1.39 mmol, 1 eq) was dissolved in 3.25 mL of ethanol and 0.65 mL of water. Fe (387 mg, 6.93 mmol, 5.00 eq) and NH4Cl (370 mg, 6.93 mmol, 5.00 eq) were added to the mixture, and the mixture was stirred at 80°C for 12 hours. After the reaction was completed, water and ethyl acetate were added to the reaction solution and extracted. The organic layer was separated, dried over magnesium sulfate, and filtered. The remaining filtrate was concentrated under reduced pressure to obtain the target compound (5-3). (Yield: 57%)

[0156] Step (3): Preparation of compound (5-4)

[0157] In step (1) of the above Example 1, compound (5-3) was used instead of tert-butyl (2-aminophenyl)carbamate, and the same procedure was repeated to obtain the target compound (5-4). (Yield: 71%)

[0158] Step (4): Preparation of compound (5)

[0159] In step (2) of the above Example 1, compound (5-4) was used instead of compound (1) and the same procedure was repeated to obtain the target compound, compound (5). (Yield: 96%)

[0160] 1H-NMR (400 MHz, DMSO-d6) δ 11.37 - 11.33 (m, 1H), 8.42 - 8.36 (m, 1H), 7.76 (d, 1H, J = 4.2 Hz), 7.16 - 7.13 (m, 1H), 6.94 - 6.90 (m, 1H), 6.80 - 6.75 (m, 1H), 6.56 - 6.51 (m, 1H), 5.01 - 4.85 (m, 1H), 4.50 - 4.02 (m, 5H), 3.98 (br s, 5H), 3.43 (br d, 2H, J = 4.4 Hz), 2.77 (s, 6H), 1.97 - 1.53 (m, 2H), 1.03 (br d, 3H, J = 7.0 Hz).

[0161]

[0162] Example 6: Preparation of compound (6) (N-(2-amino-4-methoxyphenyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0163] [Reaction Formula 6]

[0164]

[0165] Step (1): Preparation of compound (6-2)

[0166] In step (1) of the above Example 1, compound (6-1) was used instead of tert-butyl (2-aminophenyl)carbamate, and the same procedure was repeated to obtain the target compound, compound (6-2). (Yield: 92%)

[0167] Step (2): Preparation of compound (6)

[0168] In step (2) of the above Example 1, compound (6-2) was used instead of compound (1) and the same procedure was repeated to obtain the target compound, compound (6). (Yield: 6%)

[0169] 1 H-NMR (400 MHz, DMSO-d6) δ 11.07 - 10.86 (m, 1H), 8.49 - 8.01 (m, 1H), 7.74 (d, 1H, J = 4.0 Hz), 7.31 - 7.14 (m, 1H), 6.96 - 6.79 (m, 1H), 6.60 - 6.38 (m, 1H), 6.22 (dd, 1H, J = 2.4, 8.8 Hz), 5.10 (br s, 2H), 5.00 - 4.82 (m, 1H), 4.21 - 4.01 (m, 2H), 3.99 - 3.74 (m, 2H), 3.71 (m, 4H), 3.43 (br dd, 1H, J = 3.2, 5.4 Hz), 3.33 - 3.32 (m, 2H), 3.32 - 3.26 (m, 1H), 2.46 - 2.37 (m, 1H), 1.93 - 1.69 (m, 1H), 1.68 - 1.53 (m, 1H), 1.03 (d, 3H, J = 7.2 Hz).

[0170]

[0171] Example 7: Preparation of compound (7) (N-(2-amino-5-methoxyphenyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0172] [Reaction Formula 7]

[0173]

[0174] Step (1): Preparation of compound (7-2)

[0175] In step (1) of the above Example 1, compound (7-1) was used instead of tert-butyl (2-aminophenyl)carbamate, and the same procedure was repeated to obtain the target compound (7-2).

[0176] Step (2): Preparation of compound (7)

[0177] In step (2) of the above Example 1, compound (7-2) was used instead of compound (1) and the same procedure was repeated to obtain the target compound, compound (7). (Yield: 15%)

[0178] 1 H-NMR (400 MHz, DMSO-d6) δ 11.50 - 11.31 (m, 1H), 8.43 - 8.35 (m, 1H), 7.76 (d, 1H, J = 4.0 Hz), 7.35 - 7.23 (m, 1H), 6.91 (br s, 1H), 6.81 (br d, 1H, J = 8.6 Hz), 6.66 - 6.49 (m, 1H), 4.91 (br d, 1H, J = 3.2 Hz), 4.56 (br s, 2H), 4.23 - 4.05 (m, 2H), 3.96 (br dd, 1H, J = 3.6, 12.4 Hz), 3.88 - 3.58 (m, 5H), 3.45 - 3.40 (m, 1H), 3.33 (br s, 2H), 3.28 (br d, 1H, J = 4.0 Hz), 2.40 (br dd, 1H, J = 5.6, 12.0 Hz), 1.89 - 1.52 (m, 2H), 1.03 (br d, 3H, J = 7.2 Hz).

[0179]

[0180] Example 8: Preparation of compound (8) (N-(2-amino-4-hydroxyphenyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0181] [Reaction Formula 8]

[0182]

[0183] Step (1): Preparation of compound (8)

[0184] BBr3 (169 mg, 0.676 mmol, 3.00 eq) was added to a solution of 130 mg of compound (6-2) (0.225 mmol, 1 equiv) in dichloromethane (1.3 mL). The mixture was stirred at 0°C for 4 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography to obtain the target compound (8). (Yield: 30%)

[0185] 1 H-NMR (400 MHz, DMSO-d6) δ 10.91 - 10.85 (m, 1H), 9.11 - 8.97 (m, 1H), 8.38 - 8.35 (m, 1H), 7.72 (d, 1H, J = 3.6 Hz), 7.04 (br d, 1H, J = 8.4 Hz), 6.88 (br d, 1H, J = 2.8 Hz), 6.28 - 6.21 (m, 1H), 6.04 (br d, 1H, J = 8.8 Hz), 5.01 - 4.92 (m, 2H), 4.21 - 4.05 (m, 2H), 4.02 - 3.92 (m, 1H), 3.85 - 3.64 (m, 3H), 3.51 - 3.47 (m, 2H), 3.24 - 3.20 (m, 1H), 2.38 (br d, 2H, J = 2.4 Hz), 1.90 - 1.68 (m, 1H), 1.66 - 1.54 (m, 1H), 1.16 - 0.98 (m, 3H).

[0186]

[0187] Example 9: Preparation of compound (9) (N-(2-amino-5-hydroxyphenyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0188] [Reaction Formula 9]

[0189]

[0190] Step (1): Preparation of compound (9)

[0191] In step (1) of the above Example 8, compound (7-2) was used instead of compound (6-2) and the same procedure was repeated to obtain the target compound (9). (Yield 12%)

[0192] 1 H-NMR (400 MHz, DMSO-d6) δ 11.39 (br d, 1H, J = 8.0 Hz), 8.71 (s, 1H), 8.40 - 8.36 (m, 1H), 7.76 (d, 1H, J = 4.0 Hz), 7.23 - 7.13 (m, 1H), 6.94 - 6.87 (m, 1H), 6.70 (d, 1H, J = 8.4 Hz), 6.43 (dd, 1H, J = 2.8, 8.4 Hz), 4.98 - 4.83 (m, 1H), 4.45 - 4.30 (m, 2H), 4.21 - 4.05 (m, 2H), 4.03 - 3.91 (m, 1H), 3.86 - 3.65 (m, 2H), 3.47 - 3.42 (m, 2H), 3.22 (br d, 1H, J = 7.2 Hz), 2.42 (br d, 2H, J = 8.2 Hz), 2.09 - 1.37 (m, 2H), 1.24 (s, 1H), 1.14 - 1.13 (m, 1H), 1.02 (br d, 3H, J = 6.8 Hz).

[0193]

[0194] Example 10: Compound (10) Preparation of (N-(6-amino-1,3-dimethyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-5-yl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0195] [Reaction Formula 10]

[0196]

[0197] Step (1): Preparation of tert-butyl (1,3-dimethyl-6-nitro-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-5-yl)carbamate

[0198] In step (1) of the above Example 2, 5-amino-1,3-dimethyl-6-nitro-1,3-dihydro-2H-benzo[d]imidazol-2-one was used instead of 4-methyl-2-nitroaniline, and the same procedure was repeated to obtain the target compound, tert-butyl (1,3-dimethyl-6-nitro-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-5-yl)carbamate.

[0199] Step (2): Preparation of compound (10-1)

[0200] In step (2) of the above Example 2, tert-butyl (1,3-dimethyl-6-nitro-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-5-yl)carbamate was used instead of tert-butyl (4-methyl-2-nitrophenyl)carbamate, and the same procedure was repeated to obtain the target compound (10-1). (Yield 35%)

[0201] Step (3): Preparation of compound (10-2)

[0202] In step (1) of the above Example 1, compound (10-1) was used instead of tert-butyl (2-aminophenyl)carbamate, and the same procedure was repeated to obtain the target compound, compound (10-2). (Yield: 42%)

[0203] Step (4): Preparation of compound (10)

[0204] In step (2) of the above Example 1, compound (10-2) was used instead of compound (1), and the same procedure was repeated to obtain the target compound, compound (10). (Yield: 61%)

[0205] 1 H-NMR (300 MHz, DMSO-d6) δ 11.19 - 11.17 (m, 1H), 8.39 - 8.37 (d, 1H, J = 6 Hz), 7.74 (d, 1H, J = 2.1 Hz), 7.24 - 7.24 (m, 1H), 6.90 (s, 1H), 6.62 - 6.61 (d, 1H, J = 2.1 Hz), 4.91 (s, 1H), 4.81 - 4.79 (m, 2H), 4.17 - 3.94 (m, 4H), 3.83 - 3.70 (m, 2H), 3.47 - 3.36 (m, 3H), 3.25 - 3.25 (m, 5H), 2.44 - 2.38 (m, 1H), 1.88 - 1.59 (m, 3H), 1.03 - 1.02 (d, 3H, J = 3.6 Hz).

[0206]

[0207] Example 11: Preparation of compound (11) (N-(2-aminophenyl)-N-((2-aminophenyl)carbamoyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0208] [Reaction Formula 11]

[0209]

[0210] Step (1): Preparation of compound (11-2)

[0211] Compound (11-1) (0.63 g, 3.84 mmol, 1.2 eq) and triethylamine (421 mg, 4.16 mmol, 1.3 eq) were added to a dioxane (30 mL) solution containing 1 g of Tofacitinib (3.20 mmol, 1 equiv). The mixture was stirred at 120°C for 2 days. After completion of the reaction, the reaction solution was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (methanol: dichloromethane = 1:10) to obtain the target compound, compound (11-2). (Yield 60%)

[0212] Step (2): Preparation of compound (11-3)

[0213] 2.9 g of compound (11-2) (6.09 mmol, 1 equiv) was dissolved in 50 mL of THF, and the mixture was cooled to 0°C. 219 mg of NaH (60% content) (7.31 mmol, 1.2 equiv) was added, and the mixture was stirred for 30 minutes. The mixture was warmed to room temperature and stirred for 18 hours. After the reaction was completed, water and ethyl acetate were added to the reaction solution and extracted. The organic layer was separated, dried over magnesium sulfate, and filtered, and the remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (methanol: dichloromethane = 1:10) to obtain the target compound (11-3). (Yield 51%)

[0214] Step (3): Preparation of compound (11)

[0215] 1.1 g of compound (11-3) (1.72 mmol, 1 equiv) was dissolved in a methanol (50 mL) solution, and 100 mg of 10% Pd / C (10 wt.) was added thereto. The mixture was stirred for 18 hours under hydrogen gas at atmospheric pressure. The mixture was warmed to room temperature and stirred for 18 hours. After completion of the reaction, the reaction solution was filtered through Celite and washed with 50 mL of methanol. The filtrate was concentrated under reduced pressure, and the concentrate was purified by silica gel column chromatography (methanol: dichloromethane = 1:10) to obtain the target compound (11). (Yield 6%)

[0216] 1 H-NMR (400 MHz, DMSO-d6) δ 11.24-11.19 (m, 1H), 8.40-8.34 (m, 1H), 7.73 (dd, 1H, J = 11.2, 4.0 Hz), 7.43 (dd, 1H, J = 8.0, 1.2 Hz), 7.08-6.81 (m, 5H), 6.75-6.69 (m, 1H), 6.65-6.62 (m, 1H), 6.58-6.51 (m, 1H), 5.58 (d, 1H, J = 8.4 Hz), 5.02 (br, 4H), 4.00-3.79 (m, 2H), 3.75-3.55 (m, 2H), 3.40-3.32 (m, 5H), 2.46-2.32 (m, 1H), 1.94-1.74 (m, 1H), 1.72-1.64 (m, 1H), 1.09-0.99 (m, 3H).

[0217]

[0218] Example 12: Preparation of compound (12) (N-(2-amino-4-methylphenyl)-N-((2-amino-4-methylphenyl)carbamoyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0219] [Reaction Formula 12]

[0220]

[0221] Step (1): Preparation of compound (12-1)

[0222] 500 mg of 4-methyl-2-nitroaniline (3.29 mmol, 1 equiv) was dissolved in 28 mL of 1, 2-dichloroethane, and the mixture was cooled to 0–5°C. 448 mg of triphosgene (1.64 mmol, 0.5 equiv) was dissolved in 66 mL of 1, 2-dichloroethane, added, and the temperature was raised to room temperature. 458 μL of trimethylamine (3.29 mmol, 1 equiv) was then added, and the mixture was stirred under reflux for 2–16 hours. After the reaction was completed, the temperature was lowered and the mixture was used in the next reaction without purification.

[0223] Step (2): Preparation of compound (12-2)

[0224] Compound (12-1) (3.29 mmol, 2 equiv) was added with 513 mg of tofacitinib (1.64 mmol, 1 equiv) and 265 μL of pyridine (3.29 mmol, 2 equiv), and the mixture was stirred under reflux for 1 to 2 hours. After completion of the reaction, the mixture was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (ethyl acetate: hexane = 1:1) to obtain the target compound, compound (12-2). (Yield 85%)

[0225] Step (3): Preparation of compound (12-3)

[0226] 100 mg of compound (12-2) (0.20 mmol, 1 equiv) was dissolved in 2.6 mL of anhydrous THF, and the mixture was cooled to 0–5°C. 410 μL of 1 M LiHMDS in THF (0.41 mmol, 2 equiv) was added, and the mixture was warmed to room temperature. Compound (12-1) was added, and the mixture was stirred for 1 hour. After the reaction was completed, the reaction solution was extracted with water and ethyl acetate. The organic layer was separated, dried over magnesium sulfate, and filtered, and the remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (dichloromethane: methanol = 10:1) to obtain the target compound (12-3). (Yield 40%)

[0227] Step (4): Preparation of compound (12)

[0228] 54 mg of compound (12-3) (0.08 mmol, 1 equiv) was dissolved in 5 mL of ethanol, 1.5 mL of THF, and 1.5 mL of water. 45 mg of Fe (0.81 mmol, 10 equiv) and 44 mg of NH4Cl (0.81 mmol, 10 equiv) were added to the mixture at room temperature, and the mixture was stirred under reflux for 1 hour. After the reaction was completed, ethyl acetate was added to the reaction solution and extracted. The organic layer was separated, dried over magnesium sulfate, and filtered. The remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (dichloromethane: methanol = 10:1) to obtain the target compound (12). (Yield 38%)

[0229] 1H-NMR (300 MHz, DMSO-d6) δ 11.56 - 11.53, 11.12 - 11.09 (m, 1H), 10.46 - 10.42 (m, 1H), 8.16 - 8.13 (d, 1H, J = 9 Hz), 7.28 - 7.25 (d, 1H, J = 9 Hz), 6.93 - 6.87 (m, 2H), 6.80 - 6.71 (m, 1H), 6.63 - 6.30 (m, 6H), 5.55 - 5.53 (m, 1H), 4.93 (s, 4H), 4.00 - 3.51 (m, 7H), 2.29 - 2.27 (m, 6H), 1.73 - 1.58 (m, 2H), 1.05 - 1.03 (m, 3H).

[0230]

[0231] Example 13: Compound (13) (N-(2-amino-4-methoxyphenyl)-N-((2-amino-4-methoxyphenyl)carbamoyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide) manufacturing

[0232] [Reaction Formula 13]

[0233]

[0234] Step (1): Preparation of compound (13-1)

[0235] In step (1) of the above Example 12, 4-methoxy-2-nitroaniline was used instead of 4-methyl-2-nitroaniline and the same procedure was repeated to obtain the target compound (13-1).

[0236] Step (2): Preparation of compound (13-2)

[0237] In step (2) of the above Example 12, compound (13-1) was used instead of compound (12-1) and the same procedure was repeated to obtain the target compound, compound (13-2). (Yield: 88%)

[0238] Step (3): Preparation of compound (13-3)

[0239] In step (3) of the above Example 12, compound (13-2) was used instead of compound (12-2) and the same procedure was repeated, but compound (13-3), which is the target compound, was obtained without purification.

[0240] Step (4): Preparation of compound (13)

[0241] In step (4) of the above Example 12, compound (13-3) was used instead of compound (12-3) and the same procedure was repeated to obtain the target compound, compound (13). (Yield 27%)

[0242] 1 H-NMR (300 MHz, DMSO-d6) δ 10.97 - 10.94 (m, 1H), 10.49 - 10.36 (m, 1H), 8.39 - 8.36 (m, 1H), 7.75 - 7.71 (m, 1H), 7.21 - 7.18 (d, 1H, J = 9 Hz), 6.93 - 6.85 (m, 1H), 6.81 - 6.78 (d, 1H, J = 9 Hz), 6.52 - 6.49 (m, 2H), 6.38 - 6.11 (m, 4H), 5.54 - 5.50 (m, 1H), 5.08 - 5.02 (m, 4H), 3.96 - 3.61 (m, 13H), 1.76 - 1.66 (m, 2H), 1.05 - 1.01 (m, 3H).

[0243]

[0244] Example 14: Preparation of compound (14) (N-(2-(3-(2-aminophenyl)-1-methylureido)ethyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0245] [Reaction Formula 14]

[0246]

[0247] Step (1): Preparation of compound (14-1)

[0248] 5 g of tofacitinib (16.0 mmol, 1 equiv) and 3.24 g of TEA (32.0 mmol, 2 equiv) were dissolved in dichloromethane. 5.84 g of bis(4-nitrophenyl) carbonate (19.2 mmol, 1.2 equiv) was added, and the mixture was stirred at 40°C for 2 hours. 4.54 g of tert-butyl methyl(2-(methylamino)ethyl)carbamate (24.0 mmol, 1.5 eq) was added, and the mixture was stirred at 40°C for 16 hours. After completion of the reaction, the reaction solution was extracted with water and dichloromethane. The organic layer was separated, dried over magnesium sulfate, filtered, and the remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (dichloromethane: methanol = 20:1) to obtain the target compound (14-1). (Yield 39%)

[0249] Step (2): Preparation of compound (14-2)

[0250] 3.3 g of compound (14-1) (7.7 mmol, 1 equiv) was dissolved in 30 mL of ethyl acetate and cooled to 0-5°C. Hydrochloric acid gas was added to the mixture for 2 minutes. After completion of the reaction, the reaction solution was filtered to obtain the target compound (14-2).

[0251] Step (3): Preparation of compound (14-3)

[0252] 900 mg of compound (14-2) (2.1 mmol, 1 equiv) and 320 mg of TEA (3.2 mmol, 1.5 equiv) were dissolved in 20 mL of dichloromethane. 415 mg of 1-isocyanato-2-nitrobenzene (2.5 mmol, 1.2 equiv) was added and stirred at room temperature for 16 hours. After completion of the reaction, the reaction solution was extracted with water and dichloromethane. The organic layer was separated, dried over magnesium sulfate, and filtered. The remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (dichloromethane: methanol = 20:1) to obtain the target compound (14-3). (Yield 36%)

[0253] Step (4): Preparation of compound (14)

[0254] 445 mg of compound (14-3) (0.75 mmol, 1 equiv) was dissolved in 10 mL of methanol. 80 mg of 10% Pd / C was added to the mixture at room temperature, and the mixture was stirred for 2 hours under hydrogen gas at atmospheric pressure. After completion of the reaction, the mixture was purified through a Celite filter and washed with 20 mL of methanol. The filtrate was concentrated under reduced pressure, and the concentrate was purified by silica gel column chromatography (dichloromethane: methanol = 20:1) to obtain the target compound (14). (Yield: 36%)

[0255] 1H-NMR (400 MHz, DMSO-d6) δ 8.25 - 8.18 (m, 1H), 7.73 - 7.50 (m, 1H), 7.40 - 7.20 (m, 1H), 7.06 - 6.82 (m, 2H), 6.80 - 6.65 (m, 2H), 6.51 (d, 1H, J = 7.2 Hz), 4.95 - 4.65 (m, 3H), 4.20 - 4.00 (m, 2H), 3.99 - 3.35 (m, 8H), 3.27 (s, 3H), 3.16 - 2.98 (m, 3H), 2.96 - 2.62 (m, 3H), 2.46 - 2.31 (m, 1H), 1.89 - 1.68 (m, 1H), 1.63 - 1.52 (m, 1H), 1.01 (d, 3H, J = 7.2 Hz).

[0256]

[0257] Example 15: Preparation of compound (15) (N-(2-(3-(2-amino-4-methylphenyl)-1-methylureido)ethyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0258] [Reaction Formula 15]

[0259]

[0260] Step (1): Preparation of compound (15-1)

[0261] 250 mg of compound (14-1) (0.47 mmol, 1 equiv) was dissolved in 20 mL of dichloromethane. 5 mL of TFA (20 V) was added, and the mixture was stirred at room temperature for 2 hours. After completion of the reaction, the mixture was concentrated under reduced pressure to obtain compound (15-1). (Yield: 99%)

[0262] Step (2): Preparation of compound (15-2)

[0263] 202 mg of compound (15-1) (0.47 mmol, 1 equiv) and 84 mg of compound (12-1) were dissolved in 40 mL of anhydrous THF. 47 mg of TEA (0.47, 1 equiv) was added and stirred at room temperature for 16 hours. After completion of the reaction, the concentrate was concentrated under reduced pressure and purified by silica gel column chromatography (dichloromethane: methanol = 20:1) to obtain the target compound, compound (15-2). (Yield: 42%)

[0264] Step (3): Preparation of compound (15)

[0265] 120 mg of compound (15-2) (0.20 mmol, 1 equiv) was dissolved in 15 mL of methanol. 100 mg of 10% Pd / C was added to the mixture at room temperature and stirred for 1 hour under hydrogen gas at atmospheric pressure. After completion of the reaction, the mixture was filtered through a Celite filter, and the filtrate was concentrated under reduced pressure. The concentrate was purified by reverse-phase prep-HPLC to obtain the target compound (15). (Yield: 40%)

[0266] 1 H-NMR (400 MHz, DMSO-d6) δ 8.22 - 8.20 (m, 1H), 7.56 - 7.53 (m, 1H), 7.30 (s, 1H), 6.86 - 6.76 (m, 2H), 6.52 (s, 1H), 6.34 - 6.62 (d, 1H, J = 7.2 Hz), 4.85 (s, 1H), 4.65 (s, 2H), 4.17 - 3.91 (m, 3H), 3.78-3.63 (m, 4h), 3.46 - 3.37 (m, 3H), 3.27 (s, 4H), 3.11 - 3.06 (m, 1H), 2.88 (s, 1H), 2.67 - 2.63 (m, 1H), 2.39 - 2.36 (m, 1H), 2.15 (s, 3H), 1-84 - 1.68 (m, 1H), 1.62 - 1.57 (m, 1H), 1.02 - 1.00 (d, 3H, J = 7.2 Hz).

[0267]

[0268] Example 16: Preparation of compound (16) (N-(2-(3-(2-amino-5-methylphenyl)-1-methylureido)ethyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0269] [Reaction Formula 16]

[0270]

[0271] Step (1): Preparation of compound (16-1)

[0272] 152 mg of 5-methyl-2-nitroaniline (1.0 mmol, 1 equiv) was dissolved in 10 mL of 1, 2-dichloroethane, and the mixture was cooled to 0-5°C. 148 mg of triphosgene (0.5 mmol, 0.5 equiv) was dissolved in 20 mL of 1, 2-dichloroethane, and the mixture was heated to room temperature. The mixture was stirred under reflux for 16 hours. After the reaction was completed, the mixture was concentrated under reduced pressure to obtain compound (16-1). (Yield: 96%)

[0273] Step (2): Preparation of compound (16-2)

[0274] In step (2) of the above Example 15, compound (16-1) was used instead of compound (12-1) and the same procedure was repeated to obtain the target compound, compound (16-2). (Yield: 58%)

[0275] Step (3): Preparation of compound (16)

[0276] In step (3) of the above Example 15, compound (16-2) was used instead of compound (15-2) and the same procedure was repeated to obtain the target compound, compound (16). (Yield: 58%)

[0277] 1 H-NMR (400 MHz, DMSO-d6) δ 8.22 - 8.21 (m, 1H), 7.60 (s, 1H), 7.34 - 7.29 (m, 1H), 6.82 - 6.76 (m, 2H), 6.71 - 6.69 (d, 1H, J = 8 Hz), 6.63 - 6.61 (d, 1H, J = 8 Hz), 4.85 (s, 1H), 4.53 (s, 2H), 4.16 - 3.91 (m, 3H), 3.86 - 3.63 (m, 4H), 3.55 - 3.40 (m, 3H), 3.27 (s, 4H), 3.22 - 2.97 (m, 3H), 2.89 - 2.67 (m, 1H), 2.63 - 2.50 (m, 1H), 2.39 - 2.36 (m, 1H), 2.13 (s, 3H), 1.86 - 1.71 (m, 1H), 1.61 - 1.57 (m, 1H), 1.02 - 1.00 (d, 3H, J = 7.2 Hz).

[0278]

[0279] Example 17: Preparation of compound (17) (N-(2-(3-(2-amino-4-methoxyphenyl)-1-methylureido)ethyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0280] [Reaction Formula 17]

[0281]

[0282] Step (1): Preparation of compound (17-1)

[0283] 1.2 g of compound (14-2) (2.67 mmol, 1 equiv) and 540 mg of TEA (5.34 mmol, 2 equiv) were dissolved in 10 mL of anhydrous THF, and the mixture was cooled to 0-5°C. Compound (13-1) (2.67 mmol, 1 equiv) was added, and the mixture was stirred at room temperature for 16 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the concentrate was purified by silica gel column chromatography (dichloromethane: methanol = 25:1) to obtain the target compound, compound (17-1). (Yield 24%)

[0284] Step (2): Preparation of compound (17)

[0285] 400 mg of compound (17-1) (0.64 mmol, 1 equiv) was dissolved in 20 mL of methanol. 80 mg of 10% Pd / C was added to the mixture at room temperature, and the mixture was stirred for 2 hours under hydrogen gas at atmospheric pressure. After completion of the reaction, the mixture was filtered through a Celite filter, and the filtrate was concentrated under reduced pressure. The concentrate was purified by reverse-phase prep-HPLC to obtain the target compound (17). (Yield: 29%)

[0286] 1 H-NMR (400 MHz, DMSO-d6) δ 8.21 (d, 1H, J = 5.6 Hz), 7.51 (s, 1H), 7.30 (s, 1H), 6.90 - 6.66 (m, 2H), 6.29 (d, 1H, J = 2.7 Hz), 6.10 (dd, 1H, J = 8.4, 2.1 Hz), 4.94 - 4.66 (m, 3H), 4.20 - 3.53 (m, 10H), 3.48 - 3.35 (m, 3H), 3.27 (s, 3H), 3.17 - 2.98 (m, 3H), 2.88 (s, 2H), 2.64 (s, 1H), 2.45 - 2.31 (m, 1H), 1.88 - 1.66 (m, 1H), 1.64 - 1.51 (m, 1H), 1.02 (d, 3H, J = 7.2 Hz).

[0287]

[0288] Example 18: Preparation of compound (18) (N-(2-(3-(2-amino-5-methoxyphenyl)-1-methylureido)ethyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0289] [Reaction Formula 18]

[0290]

[0291] Step (1): Preparation of compound (18-1)

[0292] 500 mg of 5-methoxyl-2-nitroaniline (2.97 mmol, 1 equiv) was dissolved in 10 mL of ethyl acetate, and the mixture was cooled to 0–5°C. 360 mg of TEA (3.57 mmol, 10.2 eq) and 291 mg of triphosgene (0.98 mmol, 0.33 equiv) were added, and the mixture was stirred at room temperature for 1 hour. After completion of the reaction, the mixture was used in the next reaction without purification.

[0293] Step (2): Preparation of compound (18-2)

[0294] In step (1) of the above Example 17, compound (18-1) was used instead of compound (13-1) and the same procedure was repeated to obtain the target compound, compound (18-2). (Yield: 14%)

[0295] Step (3): Preparation of compound (18)

[0296] In step (2) of the above Example 17, compound (18-2) was used instead of compound (17-1) and the same procedure was repeated to obtain the target compound, compound (18). (Yield: 49%)

[0297] 1H-NMR (400 MHz, DMSO-d6) δ 8.26 - 8.16 (m, 1H), 7.69 (s, 1H), 7.45 - 7.19 (m, 1H), 6.85 - 6.60 (m, 3H), 6.56 - 6.48 (m, 1H), 4.96 - 4.76 (m, 1H), 4.57 (s, 2H), 4.20 - 3.50 (m, 10H), 3.50 - 3.35 (m, 3H), 3.30 - 3.20 (m, 3H), 3.18 - 3.00 (m, 3H), 2.96 - 2.78 (m, 2H), 2.74 - 2.56 (m, 1H), 2.44 - 2.31 (m, 1H), 1.90 - 1.65 (m, 1H), 1.64 - 1.50 (m, 1H), 1.01 (d, 3H, J = 8 Hz).

[0298]

[0299] Example 19: Preparation of compound (19) (N-(2-(3-(2-amino-4-hydroxyphenyl)-1-methylureido)ethyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0300] [Reaction Formula 19]

[0301]

[0302] Step (1): Preparation of compound (19-1)

[0303] 1 g of 4-amino-3-nitrophenol (6.49 mmol, 1 equiv) was dissolved in 10 mL of DMF, and the mixture was cooled to 0-5°C. 801 mg of t-BuOK (7.14 mmol, 1.1 equiv) dissolved in 5 mL of THF was added, and the mixture was stirred at 0°C for 20 minutes. Then, 1.2 g of benzyl bromide (7.14 mmol, 1.1 equiv) dissolved in 5 mL of DMF was added dropwise, and the mixture was stirred at 0°C for 20 minutes. After completion of the reaction, 20 mL of NH4Cl aqueous solution was added dropwise, and ethyl acetate was added to perform extraction. The organic layer was separated, dried over magnesium sulfate, and filtered, and the remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (dichloromethane: methanol = 20:1) to obtain the target compound (19-1). (Yield 76%)

[0304] Step (2): Preparation of compound (19-2)

[0305] In step (1) of the above Example 18, compound (19-1) was used instead of 5-methoxyl-2-nitroaniline and the same procedure was repeated to obtain the target compound (19-2).

[0306] Step (3): Preparation of compound (19-3)

[0307] In step (1) of the above Example 17, compound (19-2) was used instead of compound (13-1) and the same procedure was repeated to obtain the target compound, compound (19-3). (Yield: 11%)

[0308] Step (4): Preparation of compound (19)

[0309] In step (2) of the above Example 17, compound (19-3) was used instead of compound (17-1) and the same procedure was repeated to obtain the target compound, compound (19). (Yield: 57%)

[0310] 1H-NMR (400 MHz, DMSO-d6) δ 8.82 - 8.75 (m, 1H), 8.25 - 8.16 (m, 1H), 7.49 - 7.21 (m, 2H), 6.85 - 6.50 (m, 2H), 6.17 - 6.09 (m, 1H), 5.98 - 5.88 (m, 1H), 4.96 - 4.76 (m, 1H), 4.63 (s, 2H), 4.20 - 3.50 (m, 7H), 3.49 - 3.34 (m, 3H), 3.30 - 3.18 (m, 3H), 3.16 - 2.95 (m, 3H), 2.94 - 2.77 (m, 2H), 2.71 - 2.54 (m, 1H), 2.45 - 2.31 (m, 1H), 1.91 - 1.65 (m, 1H), 1.64 - 1.50 (m, 1H), 1.02 (d, 3H, J = 8.0 Hz).

[0311]

[0312] Example 20: Preparation of compound (20) (N-(2-(3-(2-amino-5-hydroxyphenyl)-1-methylureido)ethyl)-4-(((1S,2R)-5-(2-cyanoacetyl)-2-methylcyclohexyl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0313] [Reaction Formula 20]

[0314]

[0315] Step (1): Preparation of compound (20-1)

[0316] 1.5 g of 5-fluoro-2-nitroaniline (9.6 mmol, 1 equiv) and 2.6 g of K2CO3 were dissolved in 40 mL of n-butanol, and the mixture was refluxed at 100°C for 16 hours. After completion of the reaction, the mixture was cooled to room temperature, filtered, and concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (PE: ethyl acetate = 3:1) to obtain the target compound (20-1). (Yield 23%)

[0317] Step (2): Preparation of compound (20-2)

[0318] In step (1) of the above Example 18, compound (20-1) was used instead of 5-methoxyl-2-nitroaniline and the same procedure was repeated to obtain the target compound, compound (19-2).

[0319] Step (3): Preparation of compound (20-3)

[0320] In step (1) of the above Example 17, compound (20-2) was used instead of compound (13-1) and the same procedure was repeated to obtain the target compound, compound (20-3). (Yield: 13%)

[0321] Step (4): Preparation of compound (20)

[0322] In step (2) of the above Example 17, compound (20-3) was used instead of compound (17-1) and the same procedure was repeated to obtain the target compound, compound (20). (Yield: 45%)

[0323] 1H-NMR (400 MHz, DMSO-d6) δ 8.48 (s, 1H), 8.21 (d, 1H, J = 5.6 Hz), 7.61 (s, 1H), 7.42 - 7.22 (m, 1H), 6.81 - 6.52 (m, 3H), 6.33 (dd, 1H, J = 8.4, 2.4 Hz), 4.84 (s, 1H), 4.56 (s, 2H), 4.20 - 3.51 (m, 7H), 3.40 (s, 3H), 3.27 (s, 3H), 3.09 (d, 3H, J = 21.2 Hz), 2.88 (s, 2H), 2.73 - 2.56 (m, 1H), 2.37 (s, 1H), 1.90 - 1.66 (m, 1H), 1.57 (s, 1H), 1.01 (d, 3H, J = 7.2 Hz).

[0324]

[0325] Example 21: Compound (21) Preparation of (N-(2-(3-(2-amino-5-(dimethylamino)phenyl)-1-methylureido)ethyl)-4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide)

[0326] [Reaction Formula 21]

[0327]

[0328] Step (1): Preparation of compound (21-1)

[0329] 181 mg of N1,N1-dimethyl-4-nitrobenzene-1,3-diamine (1.0 mmol, 1 equiv) was dissolved in 10 mL of 1,2-dichloroethane, and the mixture was cooled to 0-5°C. 148 mg of triphosgene (0.5 mmol, 0.5 equiv) was dissolved in 20 mL of 1,2-dichloroethane, and the mixture was heated to room temperature. The mixture was stirred under reflux for 16 hours. After completion of the reaction, the mixture was concentrated under reduced pressure to obtain compound (21-1). (Yield: 97%)

[0330] Step (2): Preparation of compound (21-2)

[0331] In step (2) of the above Example 15, compound (21-1) was used instead of compound (12-1) and the same procedure was repeated to obtain the target compound, compound (21-2) (yield 76%).

[0332] Step (3): Preparation of compound (21)

[0333] 110 mg of compound (21-2) (0.17 mmol, 1 equiv) was dissolved in 10 mL of ethanol and 3 mL of water. 95 mg of Fe (1.7 mmol, 10 equiv) and 91 mg of NH4Cl (1.7 mmol, 10 equiv) were added to the mixture at room temperature, and the mixture was stirred under reflux for 1 hour. After completion of the reaction, the mixture was filtered through a Celite filter, and the filtrate was concentrated under reduced pressure. The concentrate was purified by reverse-phase prep-HPLC to obtain the target compound (21). (Yield: 34%)

[0334] 1H-NMR (400 MHz, DMSO-d6) δ 8.22 - 8.19 (m, 1H), 7.67 (s, 1H), 7.39 - 7.28 (m, 1H), 6.74 - 6.74 (m, 1H), 6.65 - 6.63 (m, 2H), 6.45 - 6.42 (dd, 1H, J = 8.8 Hz, 2.8 Hz), 5.59 - 5.21 (m, 2H), 4.85 (s, 1H), 4.17 - 3.91 (m, 3H), 3.86 - 3.63 (m, 4H), 3.54 - 3.31 (m, 3H), 3.26 (s, 3H), 3.12 - 3.07 (m, 3H), 2.87 - 2.75 (m, 2H), 2.71 (s, 6H), 2.67 - 2.63 (m, 1H), 2.49 - 2.33 (m, 1H), 1.86 - 1.57 (m, 2H), 1.02 - 1.00 (d, 3H, J = 6.8 Hz).

[0335]

[0336] Example 22: Compound (22) (N-(2-(3-(6-amino-1,3-dimethyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-5-yl)-1-methylureido)ethyl)-4-(((3R,4R) -1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide) manufacturing

[0337] [Reaction Formula 22]

[0338]

[0339] Step (1): Preparation of compound (22-1)

[0340] In step (2) of the above Example 15, 5-isocyanato-1,3-dimethyl-6-nitro-1H-benzo[d]imidazol-2(3H)-one was used instead of compound (12-1) and the same procedure was repeated to obtain the target compound, compound (22-1). (Yield: 70%)

[0341] Step (2): Preparation of compound (22)

[0342] In step (3) of the above Example 21, compound (22-1) was used instead of compound (21-2) and the same procedure was repeated to obtain the target compound, compound (22). (Yield: 13%)

[0343] 1 H-NMR (400 MHz, DMSO-d6) δ 8.21 - 8.21 (m, 1H), 7.66 (s, 1H), 7.38 - 7.28 (m, 1H), 6.77 (s, 2H), 6.51 (s, 1H), 4.88 - 4.84 (m, 1H), 4.55 - 4.50 (m, 2H), 4. 18 - 3. 62 (m, 7H), 3.45 - 3.40 (m, 3H), 3.26 - 3.05 (m, 12H), 2. 89 (s, 2H), 2.66 - 2.61 (m, 1H), 2.41 - 2.35 (m, 1H), 1.85 - 1.54 (m, 2H), 1.02 - 1.00 (d, 3H, J = 7.2 Hz).

[0344]

[0345] Example 23: Compound (23) ((4-((((2-(3-(2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3- d]pyrimidine-7-carboxamido)ethyl)-3-methylureido)-4-(dimethylamino)phenyl)carbamoyl)oxy)methyl)phenyl)boronic manufacturing of acid)

[0346] [Reaction Formula 23]

[0347]

[0348] Step (1): Preparation of compound (23-1)

[0349] 603 mg of compound (21) (1 mmol, 1 equiv) and 296 mg of 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl carbonochloridate (1 mmol, 1 equiv) were dissolved in 40 mL of dichloromethane. 303 mg of TEA (3 mmol, 3 equiv) was added and stirred at room temperature for 16 hours. After completion of the reaction, water and ethyl acetate were added for extraction. The organic layer was separated, dried over magnesium sulfate, and filtered, and the remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (dichloromethane: methanol = 20:1) to obtain the target compound (23-1). (Yield 46%)

[0350] Step (2): Preparation of compound (23)

[0351] 400 mg of compound (23-1) (0.46 mmol, 1 equiv) was dissolved in 20 mL of MeCN. 10 mg of NH4HCO3 (0.79 mmol, 1.7 equiv) was dissolved in 10 mL of water, added to the mixture, and stirred for 16 hours. After the reaction was completed, water and ethyl acetate were added and extracted. The organic layer was separated, dried over magnesium sulfate, and filtered. The remaining filtrate was concentrated under reduced pressure. The concentrate was purified by reverse-phase prep-HPLC to obtain the target compound (23). (Yield: 30%)

[0352] 1H-NMR (400 MHz, DMSO-d6) δ 8.72 - 8.72 (m, 1H), 8.20 - 8.19 (m, 1H), 8.06 (s, 2H), 7.89 - 7.74 (m, 3H), 7.32 - 7.13 (m, 4H), 6.84 - 6.84 (m, 1H), 6.70 - 6.69 (m, 1H), 6.50 - 6.48 (d, 1H, J = 9.2 Hz), 5.11 5.02 (m, 2H), 4.86 - 4.81 (m, 1H), 4.17 - 3.86 (m, 3H), 3.76 - 3.55 (m, 4H), 3.42 - 3.39 (m, 2H), 3.31 - 2.87 (m, 7H), 2.84 (s, 8H), 2.80 - 2.59 (m, 1H), 2.39 - 2.32 (m, 1H), 1.83 - 1.55 (m, 2H), 1.02 - 1.00 (d, 3H, J = 7.2 Hz).

[0353]

[0354] 실시예 24: 화합물 (24) ((4-((((6-(3-(2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido)ethyl)-3-methylureido)-1,3-dimethyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-5-yl)carbamoyl)oxy)methyl)phenyl)boronic acid)의 제조

[0355] [반응식 24]

[0356]

[0357] 단계 (1): 화합물 (24-1)의 제조

[0358] In step (1) of the above Example 23, compound (22) was used instead of compound (21) and the same procedure was repeated to obtain the target compound, compound (24-1). (Yield 29%)

[0359] Step (2): Preparation of compound (24)

[0360] In step (2) of the above Example 23, compound (24-1) was used instead of compound (23-1) and the same procedure was repeated to obtain the target compound, compound (24). (Yield: 30%)

[0361] 1 H-NMR (400 MHz, DMSO-d6) δ 8.92 - 8.92 (m, 1H), 8.20 -8.19 (m, 1H), 8.07 (s, 2H), 8.00 - 7.63 (m, 3H), 7.38 - 7.22 (m, 4H), 7.10 (s, 1H), 5.16 - 5.05 (m, 2H), 4.87 - 4.76 (m, 1H), 4.17 - 3.90 (m, 3H), 3.76 - 3.57 (m, 4H), 3.44 - 3.38 (m, 2H), 3.29 - 3.20 (m, 10H), 3.09 - 2.99 (m, 3H), 2.86 -2.82 (m, 2H), 2.67 - 2.61 (m, 1H), 2.39 - 2.33 (m, 3H), 1.85 - 1.51 (m, 2H), 0.99 - 0.98 (m, 3H).

[0362]

[0363] Example 25: Compound (25) (4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl Preparation of (2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido)phenyl)carbamate)

[0364] [Reaction Formula 25]

[0365]

[0366] Step (1): Preparation of compound (25-2)

[0367] 4.6 g of Na2CO3 (43 mmol, 10 eq.) was dissolved in 5 mL of toluene, and the mixture was cooled to 0°C. 2.5 g of triphosgene (8.4 mmol, 1.95 eq.) was added, and the mixture was stirred for 30 minutes. 5 mL of toluene containing 1 g of compound (25-1) (4.3 mmol, 1 eq.) was added dropwise to the reaction mixture, and the mixture was warmed to room temperature and stirred for 16 hours. After completion of the reaction, the reaction mixture was filtered, and the filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (ethyl acetate: petroleum ether = 1:20) to obtain the target compound (25-2). (Yield 28%)

[0368] Step (2): Preparation of compound (25)

[0369] Compound (25-2) (160 mg, 0.54 mmol, 1.2 eq) was added to a THF (5 mL) solution containing 200 mg of compound (1) (0.45 mmol, 1 equiv). The mixture was stirred at room temperature for 2 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography to obtain the target compound, compound (25). (Yield: 22%)

[0370] 1H-NMR (400 MHz, CDCl3) δ 11.94 - 11.75 (m, 1H), 8.24 - 8.23 ​​(m, 1H), 7.71 - 7.67 (m, 5H), 7.30 - 7.19 (m, 3H), 7.17 - 7.15 (m, 2H), 6.58 - 6.55 (m, 1H), 5.14 (s, 2H), 5.07 (br, 1H), 4.03 - 3.72 (m, 2H), 3.56 - 3.42 (m, 4H), 3.33 - 3.31 (m, 3H), 2.48 - 2.36 (m, 1H), 1.94 - 1.52 (m, 2H), 1.27 (s, 12H), 1.01 (d, 3H, J = 6.8 Hz).

[0371]

[0372] Example 26: Compound (26) (4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl Preparation of (2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido)-4-methylphenyl)carbamate)

[0373] [Reaction Formula 26]

[0374]

[0375] Step (1): Preparation of compound (26)

[0376] In step (2) of the above Example 25, compound (2) was used instead of compound (1) and the same procedure was repeated to obtain the target compound, compound (26). (Yield 18%)

[0377] 1H-NMR (400 MHz, CDCl3) δ 11.94 - 11.81 (m, 1H), 8.30 - 8.29 (m, 1H), 7.7 - 7.36 (m, 7H), 7.03 - 7.02 (m, 1H), 6.63 - 6.59 (m, 1H), 5.19 (s, 2H), 5.16 - 5.14 (m, 1H), 4.09 - 3.78 (m, 2H), 3.62 - 3.46 (m, 4H), 3.38 - 3.37 (m, 3H), 2.53 - 2.51 (m, 1H), 2.37 - 2.36 (m, 3H), 2.00 - 1.79 (m, 2H), 1.33 (s, 12H), 1.08 - 1.05 (m, 3H).

[0378]

[0379] Example 27: Compound (27) (4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl Preparation of (2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido)-4-methoxyphenyl)carbamate)

[0380] [Reaction Formula 27]

[0381]

[0382] Step (1): Preparation of compound (27)

[0383] In step (2) of the above Example 25, compound (7) was used instead of compound (1) and the same procedure was repeated to obtain the target compound, compound (27). (Yield: 16%)

[0384] 1H-NMR (400 MHz, CDCl3) δ 12.30 - 11.96 (m, 1H), 8.16 (s, 1H), 7.73 - 7.57 (m, 6H), 7.19 - 7.05 (m, 1H), 6.82 - 6.52 (m, 2H), 5.13 - 5.03 (m, 3H), 4.01 - 3.97 (m, 2H), 3.76 (s, 3H), 3.53 - 3.42 (m, 4H), 2.29 (s, 3H), 2.43 - 2.34 (m, 1H), 1.94 - 1.57 (m, 2H), 1.26 (s, 12H), 1.01 - 0.99 (m, 3H).

[0385]

[0386] Example 28: Compound (28) (4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl Preparation of (6-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido)-1,3-dimethyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-5-yl)carbamate)

[0387] [Reaction Formula 28]

[0388]

[0389] Step (1): Preparation of compound (28)

[0390] 133 mg of compound (10) (0.25 mmol, 1 equiv) and 149 mg of 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl carbonochloridate (0.5 mmol, 2 equiv) were dissolved in 4 mL of THF. 85 μL of DIPEA (0.5 mmol, 2 equiv) was added and stirred at 40°C for 16 h. After completion of the reaction, the mixture was cooled to room temperature and concentrated under reduced pressure. The concentrate was recrystallized using dichloromethane and ether. The solid obtained by filtering was purified by silica gel column chromatography (dichloromethane: methanol = 20:1) to obtain the target compound (28). (Yield 54%)

[0391] 1 H-NMR (300 MHz, DMSO-d6) δ 12.02 - 12.02 (m, 1H), 9.37 - 9.36 (m, 1H), 8.23 ​​(s, 1H), 7.88 - 7.38 (m, 5H), 7.16 (s, 1H), 6.91 - 6.84 (m, 1H), 5.13 (s, 1H), 4.91 - 4.78 (m, 2H), 4.55 - 4.54 (m, 1H), 4.20 - 3.72 (m, 5H), 2.73 - 2.69 (m, 2H), 2.20 - 2.09 (m, 3H), 1.97 - 1.85 (m, 3H), 1.74 - 1.47 (m, 3H), 1.29 - 1.23 (m, 10H), 1.16 - 1.14 (m, 1H), 1.07 - 1.01 (m, 3H), 0.85 - 0.81 (t, 1H, J = 6 Hz).

[0392]

[0393] Example 29: Preparation of compound (29) (N-(2-aminophenyl)-8-((3R,4S)-4-ethyl-1-((2,2,2-trifluoroethyl)carbamoyl)pyrrolidin-3-yl)-3H-imidazo[1,2-a]pyrrolo[2,3-e]pyrazine-3-carboxamide)

[0394] [Reaction Formula 29]

[0395]

[0396] Step (1): Preparation of compound (29-1)

[0397] 200 mg of upadacitinib (0.53 mmol, 1 equiv), 216 mg of bis(4-nitrophenyl) carbonate (0.71 mmol, 1.35 equiv), and DMAP (0.74 mmol, 1.4 equiv) were added to 5 mL of dichloromethane and stirred at room temperature for 1 day. 131 mg of tert-butyl (2-aminophenyl)carbamate (0.63 mmol, 1.2 equiv) dissolved in 3 mL of dichloromethane was added dropwise to the mixture, and the mixture was stirred at room temperature for 16 hours. The concentrate was purified by silica gel column chromatography (dichloromethane: methanol = 10:1) to obtain the target compound (29-1). (Yield 63%)

[0398] Step (2): Preparation of compound (29)

[0399] 203 mg of compound (29-1) (0.33 mmol, 1 equiv) was dissolved in 16 mL of dichloromethane, and 4.1 mL of trifluoroacetic acid (20 V) was added to the mixture, followed by stirring at room temperature for 16 hours. After completion of the reaction, the reaction solution was cooled to 0-5°C and neutralized by dropwise addition of NaHCO3 aqueous solution. Dichloromethane was added, and the organic layer was separated, dried over magnesium sulfate, and filtered. The remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (dichloromethane: methanol = 10:1) to obtain the target compound (29). (Yield 81%)

[0400] 1 H-NMR (300 MHz, DMSO-d6) δ 10.76 (s, 1H), 8.88 (s, 1H), 8.09 -8.08 (d, 1H, J = 3 Hz), 7.67 (s, 1H), 7.46 - 7.40 (m, 2H), 7.02 - 6.98 (m, 2H), 6.85 - 6.83 (dd, 1H, J = 6 Hz, 0.9 Hz), 6.67 - 6.64 (td, 1H, J = 6 Hz, 0.9 Hz), 5.14 (s, 2H), 4.42 - 4.41 (m, 1H), 3.89 - 3.80 (m, 4H), 3.72 - 3.69 (m, 1H), 3.27 - 3.25 (m, 1H), 2.61 - 2.59 (m, 1H), 1.09 - 1.05 (m, 1H), 0.82 - 0.77 (m, 1H), 0.65 - 0.63 (m, 3H).

[0401]

[0402] Example 30: Preparation of compound (30) (N-(2-amino-4-(dimethylamino)phenyl)-8-((3R,4S)-4-ethyl-1-((2,2,2-trifluoroethyl)carbamoyl)pyrrolidin-3-yl)-3H-imidazo[1,2-a]pyrrolo[2,3-e]pyrazine-3-carboxamide)

[0403] [Reaction Formula 30]

[0404]

[0405] Step (1): Preparation of compound (30-1)

[0406] In step (1) of the above Example 29, tert-butyl (2-amino-5-(dimethylamino)phenyl)carbamate was used instead of tert-butyl (2-aminophenyl)carbamate, and the same procedure was repeated to obtain the target compound (30-1). (Yield 52%)

[0407] Step (2): Preparation of compound (30)

[0408] In step (2) of the above Example 29, compound (30-1) was used instead of compound (29-1) and the same procedure was repeated to obtain the target compound, compound (30). (Yield: 91%)

[0409] 1 H-NMR (300 MHz, DMSO-d6) δ 10.46 (s, 1H), 8.87 (s, 1H), 8.06 - 8.06 (d, 1H, J = 1.8 Hz), 7.66 (s, 1H), 7.38 - 7.37 (d, 1H, J = 2.1 Hz), 7.11 - 7.10 (d, 1H, J = 4.2 Hz), 7.00 - 6.98 (t, 1H, J = 3 Hz), 6.19 - 6.19 (d, 1H, J = 1.2 Hz), 6.08 - 6.07 (dd, 1H, J = 3 Hz, 1.5 Hz), 4.96 (s, 2H), 4.42 - 4.41 (m, 1H), 3.91 - 3.79 (m, 4H), 3.72 - 3.69 (m, 1H), 3.27 - 3.24 (m, 1H), 2.85 (s, 6H). 2.62 - 2.61 (m, 1H), 1.09 - 1.03 (m, 1H), 0.82 - 0.77 (m, 1H), 0.65 - 0.63 (t, 3H, J = 6 Hz).

[0410]

[0411] Example 31: Compound (31) (N-(6-amino-1,3-dimethyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-5-yl)-8-((3R,4S)-4-ethyl-1-((2,2,2 -trifluoroethyl)carbamoyl)pyrrolidin-3-yl)-3H-imidazo[1,2-a]pyrrolo[2,3-e]pyrazine-3-carboxamide) manufacturing

[0412] [Reaction Formula 31]

[0413]

[0414] Step (1): Preparation of compound (31-1)

[0415] 1.5 g of 5-amino-1,3-dimethyl-6-nitro-1H-benzo[d]imidazol-2(3H)-one (6.75 mmol, 1 equiv) was dissolved in a methanol (150 mL) solution, and 750 mg of 10% Pd / C (10 wt.) was added. The mixture was stirred for 18 hours under hydrogen gas at atmospheric pressure. The mixture was warmed to room temperature and stirred for 18 hours. After completion of the reaction, the reaction solution was filtered through Celite and washed with 50 mL of methanol. The filtrate was concentrated under reduced pressure to obtain the target compound (31-1).

[0416] Step (2): Preparation of compound (31-2)

[0417] 1.3 g of compound (31-1) (6.75 mmol, 1 eq.) and (Boc)2 (1.77 g, 8.1 mmol, 1.2 eq.) were dissolved in 100 mL of dichloromethane, and the mixture was cooled to 0°C. 120 mg of NBS (0.675 mmol, 0.1 eq.) was added, and the mixture was warmed to room temperature and stirred for 18 hours. After completion of the reaction, the reaction solution was filtered, and the filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (100% ethyl acetate) to obtain the target compound (31-2). (2 steps, yield 25%)

[0418] Step (3): Preparation of compound (31-3)

[0419] To a solution of 570 mg of Upadacitinib (1.5 mmol, 1 eq.) and NPC (500 mg, 1.65 mmol, 1.1 eq.) in dichloromethane (40 mL) was added 0.52 mL of triethylamine (3.75 mmol, 2.5 eq.), and the mixture was stirred under reflux for 18 hours. After completion of the reaction, the reaction solution was cooled to room temperature, and the target compound (31-3) was obtained without a purification process.

[0420] Step (4): Preparation of compound (31-4)

[0421] Compound (31-2) (450 mg, 1.5 mmol, 1 eq.) was added to the reaction solution of the above step (3), and the mixture was stirred for 18 hours. After the reaction was completed, the reaction solution was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography to obtain the target compound, compound (31-4). (Yield 37%)

[0422] Step (5): Preparation of compound (31)

[0423] 735 mg of compound (31-4) (1.05 mmol, 1 equiv) was dissolved in dichloromethane (30 mL), and 6 mL of TFA was added. The mixture was stirred at room temperature for 2 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and then Na2CO3 aqueous solution was added to make it alkaline. Dichloromethane was added and extracted. The organic layer was separated, dried over magnesium sulfate, and filtered. The remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography to obtain the target compound (31). (Yield: 85%)

[0424] 1 H-NMR (400 MHz, DMSO-d6) δ 10.75 (s, 1H), 8.89 (s, 1H), 8.09 (d, 1H, J=4.0 Hz), 7.68 (s, 1H), 7.41 (d, 1H, J=4.0 Hz), 7.24 (s, 1H), 7.01 (t, 1H, J=6.0 Hz), 4.93 (s, 2H), 4.44 - 4.40 (m, 1H), 3.90 - 3.79 (m, 4H), 3.73 - 3.69 (m, 1H), 3.29 - 3.25 (m, 7H), 2.60 - 2.57 (m, 1H), 1.11 - 1.05 (m, 1H), 0.85 - 0.79 (m, 1H), 0.64 (t, 3H, J=7.2 Hz).

[0425]

[0426] Example 32: Compound (32) ((4-((((6-(8-((3R,4S)-4-ethyl-1-((2,2,2-trifluoroethyl)carbamoyl)pyrrolidin-3-yl)-3H-imidazo[1,2-a]pyrrolo[2,3-e]p yrazine-3-carboxamido)-1,3-dimethyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-5-yl)carbamoyl)oxy)methyl)phenyl)boronic manufacturing of acid)

[0427] [Reaction Formula 32]

[0428]

[0429] Step (1): Preparation of compound (32)

[0430] In step (2) of the above Example 25, compound (31) was used instead of compound (1) and the same procedure was repeated to obtain the target compound, compound (32). (Yield 28%)

[0431] 1 H-NMR (400 MHz, DMSO-d6) δ 11.57 (s, 1H), 9.44 (s, 1H), 8.59 - 8.57 (m, 1H), 8.10 - 8.00 (m, 2H), 7.90 - 7.69 (m, 3H), 7.51 - 6.84 (m, 6H), 5.45 (s, 1H), 4.41 - 4.34 (m, 1H), 3.88 - 3.66 (m, 5H), 3.36 - 3.25 (m, 9H), 2.60 - 2.54 (m, 1H), 1.13 - 1.05 (m, 1H), 0.84 - 0.77 (m, 1H), 0.64 (t, 1H, J=7.2 Hz).

[0432]

[0433] Example 33: Preparation of compound (33) (2-(9-fluoro-11,17-dihydroxy-10,13,16-trimethyl-3-oxo-6,7,8,9,10,11,12,13,14,15,16,17-dodecahydro-3H-cyclopenta[a]phenanthren-17-yl)-2-oxoethyl (2-aminophenyl)carbamate)

[0434] [Reaction Formula 33]

[0435]

[0436] Step (1): Preparation of compound (33-1)

[0437] 5.0 g of dexamethasone (12.76 mmol, 1 eq.) and pyridine (3.09 mL, 38.27 mmol, 3 eq.) were dissolved in 40 mL of dichloromethane, and the mixture was cooled to 0°C. A solution of 3.33 g of 4-nitrophenyl carbonochloridate (16.58 mmol, 1.3 eq.) dissolved in dichloromethane (10 mL) was added dropwise, and the mixture was warmed to room temperature and stirred for 1 hour. The mixture was warmed to room temperature and stirred for 18 hours. After the reaction was completed, water was added to the reaction solution, stirred, and the organic layer was separated. The separated organic layer was washed successively with 80 mL of 0.1 M HCl and saturated aqueous NaHCO3 solution, dried over magnesium sulfate, filtered, and the remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (ethyl acetate: petroleum ether = 1:1) to obtain the target compound (33-1). (Yield 87%)

[0438] Step (2): Preparation of compound (33)

[0439] 500 mg (0.9 mmol, 1 Eq) of compound (33-1), 0.59 mL (5.39 mmol, 6 Eq) of NMM, and 291 mg (2.69 mmol, 3 Eq) of o-Phenylenediamine were added to 8 mL of DMF, and the mixture was stirred at room temperature for 3 days. After the reaction was completed, water and ethyl acetate were added to the reaction solution to extract it. The organic layer was separated, dried over magnesium sulfate, and filtered, and the remaining filtrate was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography (ethyl acetate: petroleum ether = 3:1) to obtain the target compound (33). (Yield 54%)

[0440] 1H-NMR (400 MHz, DMSO-d6) δ 8.83 (s, 1H), 7.29 (d, 1H, J=10.4 Hz), 7.16 (d, 1H, J=6.0 Hz), 6.88 (t, 1H, J=7.2 Hz), 6.70 (dd, 1H, J=7.6, 1.2 Hz), 6.53 (t, 1H, J=7.2 Hz), 6.23 (d, 1H, J=8.0 Hz), 6.01 (s, 1H), 5.42 (d, 1H, J=4.0 Hz), 5.14 (s, 1H), 5.04 - 4.79 (m, 4H), 4.16 (br, 1H), 2.91 (br, 1H), 2.65 - 2.63 (m, 1H), 2.41 - 2.31 (m, 2H), 2.20 - 2.15 (m, 2H), 1.91 - 1.77 (m, 1H), 1.69 - 1.58 (m, 2H), 1.49 (s, 3H), 1.41 - 1.36 (m, 1H), 1.11 - 1.02 (m, 1H), 0.91 (s, 3H), 0.80 (d, 3H, J=7.2 Hz).

[0441]

[0442] Example 34: Preparation of compound (34) (2-(9-fluoro-11,17-dihydroxy-10,13,16-trimethyl-3-oxo-6,7,8,9,10,11,12,13,14,15,16,17-dodecahydro-3H-cyclopenta[a]phenanthren-17-yl)-2-oxoethyl (2-amino-4-methylphenyl)carbamate)

[0443] [Reaction Formula 34]

[0444]

[0445] Step (1): Preparation of compound (34-2)

[0446] In step (2) of the above Example 33, compound (34-1) was used instead of o-Phenylenediamine and the same procedure was repeated to obtain the target compound, compound (34-2). (Yield 30%)

[0447] Step (2): Preparation of compound (34)

[0448] In step (2) of the above Example 1, compound (34-2) was used instead of compound (1-1) and the same procedure was repeated to obtain the target compound, compound (34). (Yield: 14%)

[0449] 1 H-NMR (400 MHz, DMSO-d6) δ 8.75 (s, 1H), 7.29 (d, 1H, J=10.0 Hz), 7.02 (s, 1H,), 6.51 (s, 1H), 6.35 (d, 1H, J=7.6 Hz), 6.23 (d, 1H, J=10.0 Hz), 6.01 (s, 1H), 5.42 (d, 1H, J=4.4 Hz), 5.15 (s, 1H), 5.03 - 4.78 (m, 4H), 4.16 (br, 1H), 2.91 (br, 1H), 2.67 - 2.58 (m, 1H), 2.34 - 2.31 (m, 2H), 2.19 - 2.12 (m, 5H), 1.91 - 1.79 (m, 1H), 1.68 - 1.58 (m, 2H), 1.49 (s, 3H), 1.37 - 1.34 (m, 1H), 1.10 - 1.07 (m, 1H), 0.90 (s, 3H), 0.81 - 0.79 (m, 3H).

[0450]

[0451] Example 35: Preparation of compound (35) ((1R,2R,E)-1-((2S,5S,11S,14S,17S,20S,23R,26S,29S,32S)-5-ethyl-11,17,26,29-tetraisobutyl-14,32-diisopropyl-1,7,10,16,20,23,25,28,31-nonamethyl-3,6,9,12,15,18,21,24,27,30,33-undecaoxo-1,4,7,10,13,16,19,22,25,28,31-undecaazacyclotritriacontan-2-yl)-2-methylhex-4-en-1-yl (2-aminophenyl)carbamate)

[0452] [Reaction Formula 35]

[0453]

[0454] Step (1): Preparation of compound (35-1)

[0455] To a solution of 120 mg of cyclosporine (0.1 mmol, 1 eq.) dissolved in toluene (15 mL), triphosgene (2.97 g, 10 mmol, 100 eq.) and triethylamine (10 mg, 0.1 mmol, 1 eq.) were added, and the mixture was stirred at 40°C for 16 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain the target compound (35-1).

[0456] Step (2): Preparation of compound (35)

[0457] Compound 35-1 (120 mg, 0.1 mmol, 1 eq.), o-Phenylenediamine (108 mg, 1 mmol, 10 eq.), and triethylamine (100 mg, 1.0 mmol, 10 eq.) were added to 5 mL of DMF, and the mixture was stirred at 20°C for 16 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography to obtain the target compound (35). (Yield 10%) Figure 3 shows NMR data of compound (35) prepared in Example 35.

[0458] MS (m / z): 1337.7 (M+H).

[0459]

[0460] Comparative example (negative control): Preparation of compound (36)

[0461] [Reaction Formula 36]

[0462]

[0463] Step (1): Preparation of compound 36

[0464] In step (1) of the above Example 1, o-toluidine was used instead of tert-butyl (2-aminophenyl)carbamate and the same procedure was repeated to obtain the target compound, 4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-(o-tolyl)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamide. (Yield 46%)

[0465] 1H-NMR (300 MHz, DMSO-d6) δ 11.85 - 11.83 (m, 1H), 8.43 - 8.41 (d, 1H, J = 6 Hz), 8.14 - 8.05 (m, 2H), 7.78 - 7.77 (m, 1H), 7.33 - 7.25 (m, 2H), 7.13 - 7.08 (t, 1H, J = 6Hz), 6.96 - 6.92 (m, 1H), 4.92 (s, 1H), 4.20 - 3.67 (m, 5H), 3.57 - 3.37 (m, 2H), 2.44 - 2.39 (m, 4H), 1.91 - 1.58 (m, 3H), 1.04 - 1.01 (d, 3H, J = 9 Hz).

[0466]

[0467] Experimental Example 1: Confirmation of the nitric oxide scavenging ability of a nitric oxide-sensitive pharmaceutical composition (DAF-2 Assay)

[0468] The NO scavenging ability of the nitric oxide-sensitive pharmaceutical compositions comprising the compounds according to Examples 1 to 22, 29 to 31, and 33 to 35 was evaluated through the DAF-2 Assay. Using the NO donor SNAP (S-Nitroso-N-Acetyl-D,L-Penicillamine), which can release NO under hydrophilic conditions, the amount of NO was confirmed by reacting the compositions with DAF-2, which exhibits fluorescence when bound to NO.

[0469] Specifically, to prepare 5 mM of DAF-2, 100 mM of SNAP, and 5 mM of the compound synthesized in the above example, a reagent stock solution was prepared using DMSO (Dimethyl sulfoxide), and the final concentrations for the reaction were 10 μM of DAF-2, 100 μM of SNAP, and the concentrations in Table 1 were added to PBS (Phosphate buffered saline) solution, and the reaction was performed in an incubator at 37°C for 3 hours. In order to relatively compare the amount of remaining NO, the NO removal ability was calculated using the following Equation 1 and is shown in Table 10 (N / T: Not Tested).

[0470] [Formula 1]

[0471] NO removal ability (%) = 100 - [(fluorescence value with added example compound - background fluorescence value) / (fluorescence value without added compound - background fluorescence value)] × 100

[0472]

[0473] Example Concentration (μM) NO Removal Ability (%) (Mean ± SD) Example Concentration (μM) NO Removal Ability (%) (Mean ± SD) Example Concentration (μM) NO Removal Ability (%) (Mean ± SD) 1140.50 ± 2.57 11187.24 ± 0.22 2112.57 ± 1.66 569.02 ± 0.12 586.37 ± 0.05 538.13 ± 1.01 1080.8 ± 5.69 1088.57 ± 0.97 1066.27 ± 0.14 2187.53 ± 0.12 12183.14 ± 2.29 22114.83 ± 6.5 158 7.76±0.06586.44±1.93536.86±2.091089.24±0.041088.19±1.751048.96±3.123171.48±0.9913179.02±1.762913.68±7.63581.67±0 .40583.57±3.2451.71±4.161084.28±0.251087.80±1.251025.18±3.1441N / T141-3.19±0.9130183.14±2.29585.52±0.965-3.82±0.3 5586.44±1.931097.86±0.2102.82±1.551088.19±1.7551N / T151N / T31179.61±3.60546.06±0.7451.48±1.89590.34±0.981097.53±0 .14104.47±0.491092.21±0.9861N / T1615.29±0.373312.22±4.76547.9±0.595N / T51.72±4.281049.15±1.491014.47±1.01101.52±10 .2271N / T1711.04±0.60341-0.67±6.17587.87±0.3750.28±0.845-0.15±16.191090.49±0.54102.73±2.11100.76±5.8181N / T18132.91±0.89351-4.87±8.61576.82±0.31562.36±0.415-18.88±14.931087.82±0.271071.19±0.0610-32.35±12.0291N / T1919.38±1.99Vitamin B12 (positive control) 13.18 ± 1.03 567.01 ± 0.16 529.83 ± 1.29 58.98 ± 2.46 108 1.32 ± 2.94 105 1.5 ± 0.52 1012.37 ± 3.73 101 N / T 20 15 1.77 ± 2.9610041.09±1.57596.19±0.07596.99±0.401,00099.48±0.951097.58±0.021098.66±0.06.

[0474]

[0475] As a positive control, vitamin B12 (Cyanocobalamin), known to have NO scavenging ability, was used.

[0476] Referring to Table 10 above, it was confirmed that the compounds synthesized in the examples had a higher nitric oxide removal ability than the positive control, vitamin B12.

[0477]

[0478] Experimental Example 2: Confirmation of the nitric oxide scavenging ability of a nitric oxide-sensitive pharmaceutical composition capable of scavenging reactive oxygen species (DAF-2 Assay)

[0479] The nitric oxide-sensitive pharmaceutical composition comprising the compound according to Examples 23 to 28 and 32 may have the ability to scavenge reactive oxygen species (ROS). The NO scavenging ability of the pharmaceutical composition was evaluated through the DAF-2 Assay. Using the NO donor SNAP capable of releasing NO under hydrophilic conditions and H2O2 capable of releasing ROS, the amount of NO was confirmed by reacting with DAF-2, which exhibits fluorescence when bound to NO after treatment of the composition.

[0480] Specifically, the compound synthesized in the above example was added to a 100 μM SNAP solution at the concentration shown in Table 2 below and reacted, then 10 μM DAF-2 was added and reacted in an incubator at 37°C for 3 hours. In order to relatively compare the amount of remaining NO, the NO removal ability was calculated using Equation 1 above and shown in Table 11 below.

[0481] Additionally, the H2O2 (10 M) stock solution was added to the reaction solution to obtain a final concentration of 100 μM SNAP and 1 mM H2O2, and the compound synthesized in the above example was added at the concentration shown in Table 2 below and reacted. The reaction was performed in the same manner, and the NO removal ability was calculated using the above formula 1 and shown in Table 11 below (N / T: Not Tested).

[0482] Example Concentration (μM) NO Removal Ability (%) (Mean ± SD) Example Concentration (μM) NO Removal Ability (%) (Mean ± SD) Without H2O2 With H2O2 Without H2O2 With H2O2 23 121.08 ± 0.77 25.76 ± 6.11 27 1N / TN / T 5 39.33 ± 8.29 64.84 ± 1.94 5N / TN / T 10 52.96 ± 6.2 383.96 ± 0.29 1059.31 ± 0.55 97.09 ± 0.01 24 116.43 ± 4.86 34.66 ± 3.80 28 13.73 ± 0.57 76.23 ± 0.94 5 27.84 ± 7.68 43.58 ± 6.00 5 13.25 ± 0.69 98.0 1±0.041040.54±3.9653.52±3.091014.18±0.6298.34±0.05251N / TN / T3216.42±10.864.05±3.575N / TN / T56.84± 5.2129.92±7.091042.36±1.9189.96±0.19109.26±5.5132.06±3.99261N / TN / T5N / TN / T1042.98±2.5192.96±0.04

[0483]

[0484] Referring to Table 11, the compounds of the above examples are compounds containing an active oxygen scavenger, and it was confirmed that the nitrogen monoxide removal ability was improved when H2O2 was present rather than when it was not present.

[0485]

[0486] Experimental Example 3: Nitric oxide sensitivity analysis of a nitric oxide-sensitive pharmaceutical composition (HPLC)

[0487] It was confirmed whether a nitric oxide-sensitive pharmaceutical composition containing a compound according to the above example could release a low-molecular-weight compound having pharmaceutical activity by separating a nitric oxide scavenger in an environment where nitric oxide is present.

[0488] The HPLC analysis sample was prepared as follows. 28 mg of NaNO2 (0.41 mmol, 16.4 equiv) was dissolved in 0.3 mL of water, and the mixture was cooled to 0 to 5°C. 0.2 mL of sulfuric acid (0.1 mmol, 4 equiv) was added to the mixture, and 12.2 mg (0.025 mmol, 1 equiv) of the compound (4) prepared in Example 4, dissolved in 0.2 mL of acetone, was added. The mixture was stirred at room temperature for 1 hour. After completion of the reaction, water and dichloromethane were added to the reaction solution and extracted. The organic layer was separated, dried over magnesium sulfate, filtered, and the remaining filtrate was concentrated under reduced pressure.

[0489] The above analysis sample was subjected to HPLC (Agilent 1200, USA) under the conditions shown in Table 12 below, and the results are shown in Figure 2.

[0490] DetectorExternal spectrophotometerDetection wavelengthUV 280 nmColumnYMC ODS-A (250 × 4.6 mm, S-5㎛, 12nm)Column temperature35℃Injection amount10 μLFlow rate1 mL / minMobile phase time(min)WaterAcetonitrile08020200100300100318020508020

[0491]

[0492] HPLC test results confirmed that the nitric oxide-sensitive pharmaceutical composition can separate the nitric oxide scavenger and release a low molecular weight compound having pharmaceutical activity in an environment where an excess of nitric oxide is present.

[0493] Specifically, referring to Fig. 2a, the peak of compound (4) of Example 4, which is a nitric oxide-sensitive pharmaceutical composition, was detected at 14.2 minutes. On the other hand, referring to Fig. 2b, in an environment where an excess of nitric oxide was present, the peak of compound (4) disappeared and a peak detected at 7.7 minutes was generated. Fig. 2c shows the results of an HPLC test conducted on tofacitinib, a small-molecular compound having pharmaceutical activity, in which a peak was detected at 7.7 minutes. Therefore, Figs. 2a to 2c are results showing that compound (4) of Example 4 reacted with an excess of nitric oxide to release tofacitinib.

[0494]

[0495] Experimental Example 4: Confirmation of the nitric oxide scavenging ability of a nitric oxide-sensitive pharmaceutical composition in immune cells.

[0496] Raw 264.7 cell line was cultured at 8 × 10 4 Cells were seeded in a 48-well plate at a density of 10 cells / well and cultured in DMEM medium containing 10% FBS, 100 U / mL penicillin, and 100 μg / mL streptomycin at 37°C.

[0497] After overnight incubation of the above medium, the medium was replaced with fresh medium containing 10 μM of a nitric oxide-sensitive pharmaceutical composition comprising the compounds of Examples 1 to 30 and 500 ng / mL of LPS, and incubated again for 24 hours.

[0498] Finally, to measure the amount of NO produced, the NO2 present in the cell culture medium was measured using Griess reagent (Sulfanilamide, N-1-naphthylethylenediamine Dihydrochloride). -The amount was measured. Specifically, 100 μL of Griess reagent was mixed with 50 μL of cell culture medium and 50 μL of PBS, reacted in a well plate for 10 minutes, and then the absorbance at 540 nm was measured. The NO scavenging ability was calculated according to Equation 2 below, and the results are shown in Table 13 below (Examples 12 and 13 are SD values ​​derived from more than three independent experiments, and the remaining examples are representative values ​​from more than three independent experiments).

[0499] [Formula 2]

[0500] NO removal ability (%) = 100 - [(NO value of the group to which the example compound was added - NO value of the group not treated with anything) / (NO value of the group stimulated with LPS - NO value of the group not treated with anything)] × 100

[0501] Example (10 μM) NO removal ability (%) (Mean ± SD) Example (10 μM) NO removal ability (%) (Mean ± SD) 177.4 ± 0.5 2 16 4 9.3 ± 0.23 27 8.1 ± 0.21 17 29.6 ± 2.95 39 5.4 ± 0.11 18 55.8 ± 1.17 4 6 8.7 ± 0.74 19 54.1 ± 2.5 15 8 4.1 ± 0.01 20 59.5 ± 0.11 6 8.5 ± 0.11 21 6 1.2 ± 0.15 7 8 8.2 ± 0.06 22 3 1.3 ± 0.88 8 7 2.6 ±0.312316.8±0.70978.9±0.21249.8±0.641093.7±0.112563.2±0.561148.7±0.012673.7±0.331231.7±0.152759.3±0.061334.7±0.282849.8±0.431446.0±2.522990.1±0.741554.0±0.593091.6±0.68

[0502]

[0503] Referring to Table 13 above, it was confirmed that when the compound according to the above example was treated, the concentration of nitric oxide was also reduced within immune cells.

[0504]

[0505] Experimental Example 5: Confirmation of the inhibitory effect of a nitric oxide-sensitive pharmaceutical composition on the secretion of inflammatory cytokines in immune cells.

[0506] RAW 264.7 cell line was cultured at 8 × 10 4 Cells were seeded in 48-well plates at a density of 10 cells / well and cultured in DMEM medium containing 10% FBS, 100 U / mL penicillin, and 100 μg / mL streptomycin at 37°C.

[0507] After overnight incubation, the medium was replaced with fresh medium containing 10 μM of a nitric oxide-sensitive pharmaceutical composition comprising compounds of Examples 1 and 11 to 13 and 500 ng / mL of LPS, and incubated for another 24 hours.

[0508] Finally, after obtaining the supernatant, the amount of IL-6 (Interlukin-6) expression was analyzed using a sandwich enzyme-linked immunosorbent assay (ELISA). Specifically, 50 μL of the standard solution and the diluted supernatant were added to each well of a well plate coated with an antibody specific for IL-6 to bind IL-6 and the immobilized antibody, and then washed with a washing buffer, reacted with HRP as a secondary antibody at room temperature for 30 minutes, washed three times with the washing buffer, and developed using TMB (3,3', 5,5'-TetraMethyl Benzidine) as a substrate, and the reaction was stopped using 1 M sulfuric acid. The degree of color development was measured at 450 nm absorbance. Based on the degree of color development, the degree of decrease in IL-6 expression was calculated using the following Equation 3 and is shown in Table 14 below.

[0509] [Formula 3]

[0510] IL-6 reduction ability (%) = 100 - [(IL-6 quantitative value of the group to which the example compound was added - IL-6 quantitative value of the group not treated with anything) / (IL-6 quantitative value of the LPS-stimulated group - IL-6 quantitative value of the group not treated with anything)] × 100

[0511]

[0512] Example (10 μM) IL-6 reduction ability (%) (Mean ± SD) Example (10 μM) IL-6 reduction ability (%) (Mean ± SD) 193.6 ± 0.60 126 1.68 ± 1.85 114 3.6 ± 1.87 136 3.10 ± 1.23

[0513]

[0514] Referring to Table 14 above, it was confirmed that when the compound according to the above example was treated, the secretion of inflammatory cytokines in immune cells was suppressed.

[0515]

[0516] Experimental Example 6: Confirmation of the nitric oxide scavenging ability, immune cell proliferation inhibition effect, and inflammatory cytokine secretion inhibition effect of a nitric oxide-sensitive pharmaceutical composition.

[0517] Experimental Example 6-1: Confirmation of nitric oxide removal ability in spleen cells

[0518] Spleens were isolated from 8-week-old C57BL / 6 mice, and splenocytes were isolated by physical methods.

[0519] Isolated splenocytes were cultured at 5 × 10 in RPMI 1640 medium containing 10% FBS, 100 U / mL penicillin, and 100 μg / mL streptomycin. 6After diluting to a concentration of cells / mL, 500 μL / well was dispensed into 48-well plates and 100 μL / well into 96-well plates, and cultured in an incubator for 72 hours. When dispensing splenocytes, an immune response was induced by treating with Concanavalin A (5 μg / mL), and at this time, 10 μM of a pharmaceutical composition including a compound according to the example was added together. After 72 hours, the supernatant was obtained and the remaining NO level was confirmed through grease analysis, which was calculated using Equation 4 below, and the results are shown in Table 15 below.

[0520] [Formula 4]

[0521] NO removal ability (%) = 100 - [(NO value of the group to which the example compound was added - NO value of the group not treated with anything) / (NO value of the group stimulated with Concanavalin A - NO value of the group not treated with anything)] × 100

[0522]

[0523] Experimental Example 6-2: Confirmation of IFN-γ reduction in spleen cells

[0524] NO is an important inflammatory mediator and can increase levels of inflammatory cytokines. Concanavalin A (5 μg / mL), which induces an immune response in splenocytes, and the compounds of the examples were treated in a concentration-dependent manner to determine whether levels of IFN-γ, a representative inflammatory cytokine, were reduced. Supernatants were collected 72 hours after the treatment and analyzed using a quantitative enzyme-linked immunosorbent assay (QELISA), as shown in Table 15.

[0525] Experimental Example 6-3: Confirmation of the ability to inhibit cell proliferation of spleen cells

[0526] In the above experimental example 6-1, the cell proliferation ability of spleen cells was confirmed through CCK-8 analysis instead of Greek analysis, and is shown in Table 15 below.

[0527] Example (10 μM) NO removal ability (%) (Mean ± SD) IFN-γ reduction ability (%) (Mean ± SD) Cell proliferation inhibition ability (%) (Mean ± SD) 198.1 ± 0.09 91.5 ± 1.37 55.01 ± 8.48 294.3 ± 0.27 95.2 ± 0.39 90.43 ± 4.69 398.1 ± 0.34 99.1 ± 0.6 176.17 ± 2.45 491.0 ± 0.01 98.9 ± 0.03 79.51 ± 6.67 594.1 ± 0.5 296.1 ± 0.45 60.47 ± 5.48 1 077.0±0.8793.0±2.9976.35±2.451167.7±0.8457.6±15.5349.71±3.781264.6±3.6597.0±0.0564.00±6.171366.9±5.0689.7±0.5716.15±2.701531.6±0.1686.5±0.2922.62±6.361627.1±1.20 81.4±0.5024.96±5.672152.4±0.6798.1±0.0348.34±3.722245.9±0.5141.8±3.1419.40±0.212319.5±0.6568.0±0.4511.80±2.082478.3±0.5095.5±0.1567.17±4.6725100.0±0.0098.7±0.048 1.36±7.9426100.0±0.0097.4±0.1075.58±0.962777.6±1.2474.6±2.4181.14±9.032892.0±0.56100.0±0.0080.69±7.122988.5±0.47100.0±0.0078.03±6.903098.1±0.0991.5±1.3755.01±8.48

[0528]

[0529] Referring to Table 15 above, it was confirmed that when the compound according to the above example was treated, nitric oxide removal, inflammatory cytokine secretion, and cell proliferation were inhibited in spleen cells.

[0530]

[0531] Experimental Example 7: JAK 1, 2, 3 Activity Assay of Nitric Oxide-Sensitive Pharmaceutical Composition

[0532] The JAK activity of the compound according to the example was confirmed using the JAK Human TK Kinase Enzymatic Radiometric Kinase Profiler Assay service of Eurofins.

[0533] After reacting with the JAK 1, 2, and 3 active sites using radiation-coupled ATP, the activity was confirmed. The JAK 1, 2, and 3 activities according to each concentration are shown in Tables 16 and 17 below. The closer the JAK activity is to 100, the lower the JAK activity of the compound according to the example.

[0534] Example Concentration (nM) 0.1110100 Activity of JAK 1 1101102457111081031051011210911010381131101131068915102909966161011049859219710510285229197847723949010083 Positive Control (Tofacitinib) 10769173

[0535]

[0536] Example Concentration (nM) 0.1 1 1 0 1 0 Activity of JAK 2 1 0 0 9 2 6 9 1 6 1 1 9 1 9 9 8 9 9 1 2 1 0 8 1 1 1 1 0 2 9 2 1 3 1 2 0 1 0 7 1 0 2 1 0 4 Positive Control (Tofacitinib) 1 0 9 8 7 3 5 4 Activity of JAK 3 1 9 4 9 5 5 1 6 1 1 1 0 3 1 0 3 9 8 9 4 1 2 1 1 4 1 0 7 1 1 1 8 1 3 1 1 3 1 0 9 1 1 2 9 2 Positive Control (Tofacitinib) 9 6 8 0 2 7 5

[0537]

[0538] Referring to Table 16 above, the activity of the nitric oxide-sensitive pharmaceutical composition against JAK 1 was found to be very low compared to the positive control, tofacitinib. In particular, as shown in Table 17, the positive control, tofacitinib, was found to have high activity at 100 nM not only against JAK 1 but also against JAK 2 and 3, whereas compounds (11) to (13) were found to have very low activity.

[0539]

[0540] Experimental Example 8: Dose escalation experiment of a nitric oxide-sensitive pharmaceutical composition

[0541] Compound (1) and the positive control, tofacitinib, were orally administered once a day to experimental animals in increasing doses, and the maximum tolerated dose was compared in mice.

[0542] Specifically, the experimental animals were 12 6-week-old C57BL / 6 male mice. The experimental animals were housed in polycarbonate cages in groups of 4 each, and were allowed free access to food and water. The animal breeding room was maintained at a temperature of approximately 20±1℃ and a humidity of approximately 50±5%, and the lighting cycle was set to a constant 12-hour cycle. The experimental animals were allowed to acclimate to the animal room environment for 1 week and then randomly divided into 3 groups of 4 each based on body weight. The experimental groups and the administered compounds for each experimental group were set as shown in Table 18 below.

[0543] Experimental group Compound dosage (mg / kg)Normal Example Compound (1)2004008001,600 Positive control group Tofacitinib2004008001,600

[0544]

[0545] The compound of Table 18 above was administered orally once a day as a single dose, starting with a low dose of 200 mg / kg on the first day and increasing the dose daily by 2 times, to 400 mg / kg on the second day, 800 mg / kg on the third day, and 1,600 mg / kg on the fourth day, and the presence or absence of death was observed.

[0546] In the group administered with increasing doses of compound (1), no deaths occurred for 24 hours after the dose was increased to 1,600 mg / kg. In contrast, in the positive control group administered with tofacitinib, two deaths occurred before the fourth day of administration after the third day of administration with 800 mg / kg. Therefore, the maximum tolerated dose of tofacitinib, the positive control, was confirmed to be 400 mg / kg, and the maximum tolerated dose of compound (1) of the example was confirmed to be 1,600 mg / kg.

[0547]

[0548] Experimental Example 9: Evaluation of hematological toxicity after single oral administration of a nitric oxide-sensitive pharmaceutical composition.

[0549] Compounds (1) and (4) were administered orally as a single dose to experimental animals and compared with tofacitinib in terms of hematological toxicity responses.

[0550] The preparation of experimental animals was the same as in the previous experimental example, and the experimental groups and administered compounds for each experimental group were set as shown in Table 19 below.

[0551] Experimental group Compound dose (mg / kg)Normal Example Compound (1) 200800 Compound (4) 200800 Positive control group Tofacitinib 200800

[0552]

[0553] The compounds in Table 19 were administered orally once a day as a single dose. Two days after administration, whole blood was collected by cardiac puncture during autopsy. At this time, blood was placed in EDTA (ethylene diamine tetra acetate) tubes to prevent blood clotting, and a general blood test (XN-1000, SYSMEX, Japan) was performed to evaluate hematological toxicity, which is shown in Table 20 below.

[0554] Experimental group dose (mg / kg) WBC (% of normal) WBC Differential Counting (NEU) (% of normal) WBC Differential Counting (LYM) (% of normal) Normal 100±5.6 100±16.4 100±7.0 Example (compound (1)) 2009 4.6±7.7 105.9±18.1 91.9±5.2 8008 9.6±16.6 64.4±4.5 92.6±19.7 Example (compound (4))200104.0±2.6108.9±22.7103.5±4.480094.8±18.765.3±18.699.0±18.4Positive control group (Tofacitinib)20080.7±28.668.3±15.781.1±31.380062.1±4.7*56.4±7.9*61.2±4.6*

[0555] * p<0.05, ** p<0.01, *** p<0.001 compared to the Normal group, measured by student t-test.

[0556]

[0557] Toxicological comparison of the test substances was performed by evaluating hematological toxicity after single administration of a low concentration of 200 mg / kg and a high concentration of 800 mg / kg. In all groups administered with compound (1) and compound (4), there was no statistically significant difference from the normal control group, and no hematological toxicity reaction was observed.

[0558] Only in the group administered with the high dose of 800 mg / kg tofacitinib, a positive control group, hematological toxicity was observed, with statistically significant decreases in total white blood cell (WBC) and neutrophil (NEU) and lymphocyte (LYM) counts (p<0.05).

[0559] Therefore, when the same high concentration of 800 mg / kg compound was administered orally to mice as a single dose, the hematological toxicity results were compared, and it was confirmed that the hematological toxicity of the example compound was lower than that of tofacitinib.

[0560]

[0561] Experimental Example 10: Evaluation of the efficacy of a nitric oxide-sensitive pharmaceutical composition for inflammatory bowel disease (DSS-induced ulcerative colitis)

[0562] Experimental Example 10-1: Evaluation of the efficacy of compound (1)

[0563] Compound (1) of the example was orally administered once a day to experimental animals with inflammatory bowel disease, and the disease alleviation efficacy was compared with that of tofacitinib as a positive control group.

[0564] The experimental animals were prepared in the same manner as in the previous experiment, except that they were randomly divided into four groups of five animals each.

[0565] To establish an inflammatory bowel disease model, 2.5% DSS powder was mixed with drinking water and administered ad libitum. The mice were fed for 5–6 days during the disease induction period, after which normal drinking water was provided. The experimental groups and the administered compounds for each experimental group were set as shown in Table 21 below.

[0566] Experimental group Compound dose (mg / kg) Drug administration period (treatment period) Normal--From 3 days after the start of disease induction until the end of the experiment Example Compound (1) 25 Positive control group Tofacitinib 25

[0567]

[0568] The compounds administered in Table 21 above were administered orally once daily. Bloody stool, diarrhea, and weight loss were quantified according to the criteria in Table 22 below, and the total was calculated as a disease activity index (DAI), which is shown in Figure 4.

[0569] Score Bloody stools Degree of diarrhea Rate of loss compared to initial body weight 0 None None Less than 1% 1 Small bloody stools and dry anus Loose stools not stained by the anus 1% or more but less than 5% 2 Large bloody stools and blood observed around the anus Very loose stools stained by the anus 5% or more but less than 10% 3 Reddish-brown bloody stools and blood stains around the anus Wet anus and long, liquid stools 10% or more but less than 15% 4 Blood or blood clots Diarrhea 15% or more

[0570]

[0571] Referring to Figure 4, statistically significant efficacy was observed for tofacitinib (p<0.01) and compound (1) (p<0.01) on day 7 of the test. The efficacy of compound (1) in alleviating inflammatory bowel disease was confirmed to be on average equivalent to that of tofacitinib.

[0572]

[0573] Experimental Example 10-2: Efficacy Evaluation of Compound (4)

[0574] In the above Experimental Example 10-1, the efficacy of compound (4) on inflammatory bowel disease was confirmed in the same manner as in Experimental Example 10-1 except that compound (4) was used instead of compound (1), DSS was administered for up to 5 days, and the drug was administered from the 6th day, and the results are shown in Fig. 5.

[0575] Referring to Figure 5, on the 9th day of the test, the inflammatory bowel disease alleviation effect of tofacitinib was not observed, and statistically significant efficacy was observed only in the compound (4) administration group (p<0.05).

[0576] As seen in the results of Figures 4 and 5, the inflammatory bowel disease alleviation efficacy of compounds (1) and (4) was confirmed to be equivalent to or superior to tofacitinib.

[0577]

[0578] Experimental Example 11: Evaluation of the Efficacy of a Nitric Oxide-Sensitive Pharmaceutical Composition for Rheumatoid Arthritis

[0579] Experimental Example 11-1: Evaluation of the efficacy of compound (1)

[0580] Compound (1) of the example and tofacitinib were orally administered once daily to experimental animals with rheumatoid arthritis disease, and their disease alleviation efficacy was compared.

[0581] The experimental animals were 30 DBA / 1 male mice, 8 weeks old. The experimental animals were housed in polycarbonate cages, 6 animals each, and were allowed free access to food and water. The animal breeding room was maintained at a temperature of approximately 20±1℃ and a humidity of approximately 50±5%, and the lighting cycle was set to a constant 12-hour cycle. The experimental animals were allowed to acclimate to the animal room environment for 1 week and then randomly divided into 5 groups. Normal mice were kept at 5 animals to minimize the number of experimental animals. The compound (1) administration group consisted of 5 to 6 animals, the minimum number for calculating statistics due to the limited drug dose.

[0582] To establish a rheumatoid arthritis model, 100 μL of a 1:1 mixture of 2 mg / mL chick type II collagen and 1 mg / mL complete Freund's adjuvant (CFA) was administered intradermally to the tails of mice. After 19 days, 100 μL of an emulsion of 1:1 mixture of 2 mg / mL chick type II collagen and incomplete Freund's adjuvant (IFA) was administered intradermally to the tails of mice. The experimental groups and the administered compounds for each experimental group were set as shown in Table 23 below.

[0583] Experimental group Compound dosage (mg / kg) Drug administration period Normal--From 19 days after the start of disease induction to the end of the experiment Disease induction group (RA)--Example Compound (1) 30 Example Compound (1) 60 Positive control group Tofacitinib 40

[0584]

[0585] The test substance administration in Table 23 above was administered orally daily according to the drug administration period and administration frequency, and the rheumatism score was determined by digitizing each of the four scores according to the criteria in Table 24 below and adding them all together to determine the score, and the results are shown in Figure 6.

[0586] Rheumatoid Score Symptoms 0 Normal, no difference from normal animals, no symptoms. 1 Inflammation of 1-2 toes, no swelling of the instep and ankle. 2 Inflammation of 3 or more toes, but no swelling or mild overall swelling of the feet. 3 Swelling of the entire foot. 4 Severe swelling of the entire foot and instep, or swelling of the ankles and toes to the point where foot function is impaired.

[0587]

[0588] Referring to Figure 6, from the 28th day of disease induction, the tofacitinib administration group and the compound (1) administration group were observed to have similar efficacy in alleviating rheumatoid symptoms on average. From the 33rd day of disease induction (p<0.01) to the end of the experiment, a statistically significant rheumatoid arthritis alleviation effect was observed in the tofacitinib 40 mg / kg administration group. In the compound (1) 30 mg / kg administration group, an alleviation effect was observed from the 35th day of disease induction (p<0.05), and from the 42nd to the 47th day of disease induction, a statistically significant alleviation effect was observed again from the 49th day to the end of the experiment (p<0.05). In the compound (1) 30 mg / kg administration group, the average rheumatoid score was observed to be equivalent to that of the tofacitinib 40 mg / kg administration group, and it was confirmed that the part where no statistical alleviation effect was observed from day 42 to day 47 of disease induction was caused by inter-individual differences.

[0589] In the compound (1) 60 mg / kg administration group, an efficacy of alleviating rheumatoid arthritis equivalent to or greater than that of the tofacitinib 40 mg / kg administration group, which is the same level of tofacitinib dose contained in compound (1) 60 mg / kg, was observed from the 33rd day of disease induction (p<0.01) to the end of the experiment.

[0590] Therefore, when compound (1) was orally administered once daily at concentrations of 30 mg / kg and 60 mg / kg during the entire experimental period, it was confirmed to alleviate rheumatoid arthritis to a level equivalent to or greater than that of tofacitinib 40 mg / kg.

[0591]

[0592] Experimental Example 11-2: Evaluation of the efficacy of compound (11)

[0593] Example compound (11) was orally administered once daily to rheumatoid arthritis experimental animals, and the disease alleviation efficacy was compared with that of tofacitinib as a positive control group.

[0594] The experimental animals were 31 DBA / 1 male mice, 8 weeks old. The experimental animals were housed in polycarbonate cages, 5 each, and were allowed free access to food and water. The animal breeding room was maintained at a temperature of approximately 20±1℃ and a humidity of approximately 50±5%, and the lighting cycle was set to a constant 12-hour light / dark cycle. The experimental animals were allowed to acclimate to the animal room environment for 1 week and then randomly divided into 5 groups based on body weight. The normal mice were kept at 5 to minimize the number of experimental animals. The compound (11) administration group consisted of 5 animals, the minimum number for calculating statistics due to the limited drug dose. The remaining groups were composed of 7 animals each, with 2 extra animals added to account for experimental error.

[0595] To establish a rheumatoid arthritis model, 100 μL of a 1:1 mixture of 2 mg / mL chick type II collagen and 1 mg / mL complete Freund's adjuvant (CFA) was intradermally administered to the tails of mice. After 19 days, 100 μL of an emulsion of 1:1 mixture of 2 mg / mL chick type II collagen and incomplete Freund's adjuvant (IFA) was intradermally administered to the tails of mice.

[0596] The experimental groups and the compounds administered to each experimental group were set as shown in Table 25 below.

[0597] Experimental group Compound administered Dose (mg / kg) Dosage frequency Drug administration period (treatment period) Normal---From 19 days after the start of disease induction to the end of the experiment Disease induction group (RA)---Example Compound (11) 20 Once a day (QD) Positive control group Tofacitinib 10 Twice a day (BID) Positive control group Dexamethasone 1 Once a day (QD)

[0598]

[0599] Oral administration was performed daily according to the dosage, frequency, and duration of administration in Table 25 above. The arthritis scores for four feet were quantified and summed according to the criteria in Table 24 above, and the scores are shown in Figure 7.

[0600] Referring to Figure 7, from day 28 of the test, the compound (11) administration group and tofacitinib were observed to have similar efficacy in alleviating rheumatoid symptoms on average. Statistically significant rheumatoid arthritis alleviation efficacy was observed in the tofacitinib administration group on day 39 (p<0.05) and day 40 (p<0.01). In the compound (11) administration group, no statistical difference was confirmed due to intra-group variation, but alleviation efficacy equivalent to tofacitinib was observed for the entire 40 days.

Claims

1. A compound of the following chemical formula 1, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof: [Chemical Formula 1] In the above chemical formula 1, X is a low molecular weight compound containing an amine group or a hydroxyl group in the compound molecular structure and having pharmaceutical activity, Y is hydrogen, C1-C6 alkyl group, C1-C6 alkoxy group, hydroxy group, nitro group, amino group, halogen, thiol group, cyano group and is selected from the group consisting of, Z is , , and is selected from the group consisting of, n and m are each independently integers from 0 to 4, The above R1 and R2 are each independently a C1-C6 alkyl group, a C1-C6 alkoxy group, a hydroxy group, a nitro group, an amino group, a halogen, a thiol group, a cyano group, a C3-C 14 Aryl group, C3-C 14 Heteroaryl group, -NR 11 R 12 , -NR 12 C(=O)R 12 , -C(=O)R 11 and -C(=O)OR 11 is selected from the group consisting of, The above R 11 and the above R 12 are each independently hydrogen, halogen, CF3, C1-C6 alkyl group, C3-C 14 Aryl group and C3-C 14 It is selected from the group consisting of heteroaryl groups, The above aryl group or heteroaryl group is unsubstituted or has at least one substituent independently selected from the group consisting of a halogen, a hydroxyl group, a thiol group, a cyano group, a C1-C6 alkyl group, and a C1-C6 alkoxy group, When the above R1 and R2 are independently two or more, they are the same or different from each other, and two adjacent R1 or R2 may or may not be combined to form a fused ring. The above R3 and R4 are each independently selected from the group consisting of hydrogen, halogen, hydroxyl group, thiol group, cyano group, C1-C6 alkyl group and C1-C6 alkoxy group, or R3 and R4 are combined to form a fused ring.

2. In paragraph 1, The above X is a compound which is a kinase inhibitor, an anticancer agent or an anti-inflammatory agent, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof.

3. In paragraph 2, A compound, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof, wherein the kinase inhibitor is selected from the group consisting of Netarsudil, Fostamatinib, Belumosudil, Tofacitinib, Upadacitinib, Baricitinib, Filgotinib, Abrocitinib, Delgocitinib, Oclacitinib, Peficitinib, and Ruxolitinib.

4. In paragraph 2, The above anticancer drugs are Doxorubicin, Cyclophosphamide, Cisplatin, Oxaliplatin, 5-Fu (5-Fluorouracil), Gemcitabine, Paclitaxel, Docetaxel, Irinotecan, MMAE (Monomethyl auristatin E), Crizotinib, Osimertinib, Sorafenib, Ibrutinib, Ruxolitinib, Vemurafenib, Ceritinib, Alectinib, Brigatinib, Lorlatinib, Capmatinib, Gefitinib Gefitinib, Erlotinib, Lapatinib, Icotinib, Afatinib, Neratinib, Dacomitinib, Almonertinib, Tucatinib, Midostaurin, Gilteritinib, Quizartinib, Pexidartinib, Sunitinib, Pazopanib, Vandetanib, Axitinib, Cabozantinib, Regorafenib, Apatinib, Lenvatinib, Tivozanib, Fruquintinib (Fruquintinib), Nintedanib, Anlotinib, Erdafitinib, Pemigatinib, Avapritinib,Ripretinib, Pralsetinib, Larotrectinib, Entrectinib, Imatinib, Dasatinib, Nilotinib, Bosutinib, Radotinib, Ponatinib, Acalabrutinib, Zanubrutinib, Fedratinib, Dabrafenib, Encorafenib, Trametinib, Cobimetinib, Binimetinib, Selumetinib, Palbociclib, Ribociclib Abemaciclib, Idelalisib, Copanlisib, Duvelisib, Alpelisib, Tazemetostat, Vorinostat, Belinostat, Tucidinostat, Panobinostat, Enasidenib, Ivosidenib, Venetoclax, Vismodegib, Sonidegib, Glasdegib, Bortezomib, Carfilzomib, Ixazomib, Olaparib, Rucaparib, Niraparib Talazoparib, Umbralisib, Trilaciclib, Infigratinib, Mobocertinib, Asciminib,A compound selected from the group consisting of futibatinib, pacritinib and everolimus, or a solvate, hydrate, stereoisomer or pharmaceutically acceptable salt thereof.

5. In paragraph 2, A compound, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof, wherein the anti-inflammatory agent is selected from the group consisting of dexamethasone, methotrexate, cyclosporine, acetaminophen, etodolac, piroxicam, and aceclofenac.

6. In paragraph 1, A compound selected from the group consisting of the following chemical formulas 2 to 6, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof: [Chemical Formula 2] [Chemical Formula 3] [Chemical Formula 4] [Chemical Formula 5] [Chemical Formula 6] In the above chemical formulas 2 to 6, X, R1, R2, R3, R4, R 11 , R 12 , the definitions of n and m are the same as those in the above chemical formula 1.

7. In paragraph 1, A compound selected from the group consisting of compounds (1) to (35), or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof: [Compound (1)] [Compound (2)] [Compound (3)] [Compound (4)] [Compound (5)] [Compound (6)] [Compound (7)] [Compound (8)] [Compound (9)] [Compound (10)] [Compound (11)] [Compound (12)] [Compound (13)] [Compound (14)] [Compound (15)] [Compound (16)] [Compound (17)] [Compound (18)] [Compound (19)] [Compound (20)] [Compound (21)] [Compound (22)] [Compound (23)] [Compound (24)] [Compound (25)] [Compound (26)] [Compound (27)] [Compound (28)] [Compound (29)] [Compound (30)] [Compound (31)] [Compound (32)] [Compound (33)] [Compound (34)] [Compound (35)] .

8. A pharmaceutical composition for the prevention or treatment of a disease involving the generation of nitric oxide, comprising a compound according to paragraph 1, or a solvate, hydrate, stereoisomer, or pharmaceutically acceptable salt thereof.

9. In paragraph 8, The above diseases involving the production of nitric oxide are Inflammatory diseases selected from the group consisting of inflammatory diseases caused by viral infection, inflammatory diseases caused by bacterial infection, sepsis, and degenerative inflammatory diseases; Neurological diseases selected from the group consisting of Alzheimer's disease, amyotrophic sclerosis, Huntington's disease, Parkinson's disease, and stroke; Cardiovascular disease such as hypertension or heart failure; An autoimmune disease selected from the group consisting of rheumatoid arthritis, ulcerative bowel disease, Crohn's disease, systemic lupus erythematosus, psoriasis, multiple sclerosis, and juvenile idiopathic arthritis; Allergic diseases selected from the group consisting of asthma, rhinitis and atopic dermatitis; A cancer selected from the group consisting of lung cancer, breast cancer, colon cancer, stomach cancer, liver cancer, brain cancer, pancreatic cancer, thyroid cancer, skin cancer, bone marrow cancer, lymphoma, uterine cancer, cervical cancer, ovarian cancer, kidney cancer, and melanoma; A pharmaceutical composition for the prevention or treatment of diseases involving nitric oxide production, such as obesity; myelofibrosis; or hepatic encephalopathy.

10. In paragraph 8, A pharmaceutical composition for the prevention or treatment of a disease involving the generation of nitric oxide, wherein the pharmaceutical composition is administered orally, intraarticularly, intravenously, subcutaneously, intramuscularly, intraperitoneally, intranasally, intrapulmonary, or rectally.

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