Compound derivative for inhibiting use1 activity, and pharmaceutical composition for preventing or treating cancer, comprising same

Abstract

The present invention relates to a compound for inhibiting the activity of UBA6-specific E2 conjugating enzyme 1 (USE1), and a pharmaceutical composition for preventing or treating cancer, comprising same. The compound according to the present invention, which has an aryl substituted basepair and oxycarbonylsulfamate as a basic structure, effectively inhibits the activity of USE1 so as to inhibit tumor growth, and thus can be used in the prevention and / or treatment of various cancers.

Description

Compound derivatives that inhibit USE1 activity, and pharmaceutical compositions containing the same for preventing or treating cancer

[0001] The present invention relates to a compound that inhibits USE1 (UBA6-specific E2 conjugating enzyme 1) activity, and a pharmaceutical composition comprising the same for preventing or treating cancer. The compound according to the present invention inhibits the activity or expression of USE1, thereby having a tumor growth inhibitory effect, and thus can be used for preventing and / or treating various cancers.

[0002] Cancer is one of the leading diseases, with 10 million new cases diagnosed worldwide each year. In Korea, cancer is the leading cause of death, accounting for approximately 25.5% of all deaths. This trend is expected to continue. In particular, lung cancer is one of the most common malignant tumors and plays a significant role in cancer mortality worldwide.

[0003] Cancer treatment began with surgery in the 1960s, progressed to radiation therapy in the 1970s, and chemotherapy has been a key treatment modality since the 1980s. However, the current survival rates and quality of life of patients with lung cancer remain inadequate. Therefore, identifying novel therapeutic targets is crucial for the treatment of these tumors.

[0004] USE1 (UBA6-specific E2 conjugating enzyme 1), also known as ubiquitin-conjugating enzyme UBE2Z, is a member of the E2 enzyme family. USE1 contains an E2 catalytic core domain containing an active-site cysteine ​​residue required for the formation of a thioester bond with ubiquitin. Interestingly, for ubiquitin conjugation, USE1 specifically reacts with the UBA6 ubiquitin-activating enzyme, rather than the conventional UBE1 E1 enzyme. A recent study analyzing the protein levels of lung cancer patients revealed that USE1 and UBA6 (ubiquitin-like modifier activating enzyme 6) were elevated in lung cancer tissues compared to surrounding normal tissues.

[0005] That is, overexpression of USE1 significantly increased cancer cell proliferation, migration, and invasion in both cells and animals, and reduction of USE1 levels in lung cancer cells reversed these effects, suggesting that USE1 is a novel target for cancer therapy.

[0006] However, the therapeutic efficacy of USE1 as a lung cancer-specific compound has not yet been confirmed.

[0007] [Prior Art Literature]

[0008] [Patent Document]

[0009] Republic of Korea Patent No. 10-1744397

[0010] The purpose of the present invention is to provide a compound that can effectively inhibit the activity or expression of USE1 (UBA6-specific E2 conjugating enzyme 1).

[0011] In addition, the present invention provides a compound that can effectively inhibit the activity of UBA6, an E1 activating enzyme.

[0012] In addition, the present invention provides a pharmaceutical composition that has an excellent tumor suppression effect by effectively inhibiting the activity of USE1 and / or UBA6, and can be used for the prevention or treatment of various cancers, including lung cancer.

[0013] To achieve the above purpose, the present invention provides a compound represented by the following chemical formula 1 or a pharmaceutically acceptable salt thereof:

[0014] [Chemical Formula 1]

[0015]

[0016] In the above chemical formula 1,

[0017] Y is C or O,

[0018] Ar is selected from the group consisting of the following compounds,

[0019]

[0020] R1 to R4 are each independently selected from the group consisting of hydrogen, hydroxyl group, halogen and C1-C6 alkyl,

[0021] R5 is -O-SO2NR or -NR-SO2NR,

[0022] R6 is hydrogen, COOR” or CN,

[0023] R7 to R9, R 11 are each independently hydrogen or C1-C6 alkyl,

[0024] R 10 is selected from the group consisting of hydrogen, -C≡CR, COR and S-CR',

[0025] R is hydrogen or C1-C6 alkyl,

[0026] R' is halogen,

[0027] R” is hydrogen, C-CR' and C1-C6 alkyl.

[0028] In addition, another embodiment of the present invention provides a pharmaceutical composition for preventing or treating cancer, comprising the above-described compound as an active ingredient.

[0029] The above cancer may be one selected from the group consisting of liver cancer, colon cancer, cervical cancer, kidney cancer, stomach cancer, prostate cancer, breast cancer, brain tumor, lung cancer, uterine cancer, colon cancer, bladder cancer, blood cancer, and pancreatic cancer.

[0030] The cancer may be cervical cancer or lung cancer.

[0031] In addition, another embodiment of the present invention provides a health food composition for preventing or improving cancer, comprising the above-described compound as an active ingredient.

[0032] Additionally, another embodiment of the present invention provides a method of reducing E1 enzyme activity in a sample, comprising contacting the sample with the compound described above.

[0033] The above E1 enzyme may be UBA6 (Ubiquitin-like modifier activating enzyme 6).

[0034] The above sample may contain cancer cells.

[0035] In addition, another embodiment of the present invention provides a method for inhibiting tumor cells, comprising the step of treating cells with the compound described above.

[0036] In addition, another embodiment of the present invention provides a method of treating cancer in a patient in need of cancer treatment, comprising administering to the patient the composition for preventing or treating cancer described above.

[0037] Specific details of other embodiments are included in the detailed description and drawings.

[0038] The compound according to the present invention has an aryl substituted basepair and oxycarbonylsulfamate as its basic structure, and thus has the effect of effectively inhibiting the activity of USE1.

[0039] In addition, the compound according to the present invention has the effect of effectively inhibiting the activity of UBA6, an E1 activating enzyme, based on the UBA6-USE1 enzyme cascade.

[0040] The compound according to the present invention can effectively inhibit the activity of USE1 and / or UBA6 and has an excellent tumor suppression effect, so it can be used as a pharmaceutical composition for preventing or treating various cancers, including lung cancer.

[0041] Figure 1 shows the results of analyzing the cell proliferation ability of KA-01 according to one embodiment of the present invention.

[0042] Figure 2 shows the results of analyzing the cell colony formation ability of KA-01 according to one embodiment of the present invention.

[0043] Figure 3 shows the results of analyzing the wound healing ability of KA-01 according to one embodiment of the present invention.

[0044] Figure 4 shows the results of volume change in cancer tissue treated with KA-01 according to one embodiment of the present invention.

[0045] Figure 5 shows the results of changes in the size of cancer tissue treated with KA-01 according to one embodiment of the present invention.

[0046] Figure 6 shows the results of a Western blot (Charging assay) analysis of compound KA-01 according to the present invention.

[0047] Figure 7 shows the results of Western blot analysis of compound KA-02 according to the present invention.

[0048] Figure 8 shows the results of Western blot analysis of compound KA-03 according to the present invention.

[0049] Figure 9 shows the results of Western blot analysis of compound KA-04 according to the present invention.

[0050] Figure 10 shows the results of Western blot analysis of compound KA-05 according to the present invention.

[0051] Figure 11 shows the results of Western blot analysis of compound KA-06 according to the present invention.

[0052] Figure 12 shows the results of Western blot analysis of compound KA-07 according to the present invention.

[0053] Figure 13 shows the results of Western blot analysis of compound KA-08 according to the present invention.

[0054] Figure 14 shows the results of Western blot analysis of compound KA-09 according to the present invention.

[0055] Figure 15 shows the results of Western blot analysis of compounds KA-10 and KA-11 according to the present invention.

[0056] Figure 16 shows the results of Western blot analysis of compound KA-12 according to the present invention.

[0057] Figure 17 shows the results of Western blot analysis of compound KA-13 according to the present invention.

[0058] Figure 18 shows the results of Western blot analysis of compound KA-14 according to the present invention.

[0059] Figure 19 shows the results of Western blot analysis of compound KA-15 according to the present invention.

[0060] Figure 20 shows the results of Western blot analysis of compounds KA-16 and KA-17 according to the present invention.

[0061] Figure 21 shows the results of Western blot analysis of compound KA-18 according to the present invention.

[0062] Figure 22 shows the results of Western blot analysis of compound KA-19 according to the present invention.

[0063] Figure 23 shows the results of Western blot analysis of compound KA-20 according to the present invention.

[0064] The present invention is susceptible to various modifications and embodiments, and specific embodiments are illustrated and described in detail in the drawings. However, this is not intended to limit the present invention to specific embodiments, but rather to encompass all modifications, equivalents, and alternatives falling within the spirit and technical scope of the present invention. In describing the present invention, detailed descriptions of related known technologies will be omitted if they are deemed to obscure the gist of the present invention.

[0065] The terminology used in this application is only used to describe specific embodiments and is not intended to limit the present invention. The singular expression includes the plural expression unless the context clearly indicates otherwise. In this application, it should be understood that the terms "comprise" or "have" indicate the presence of a feature, number, step, operation, component, part, or combination thereof described in the specification, but do not exclude in advance the possibility of the presence or addition of one or more other features, numbers, steps, operations, components, parts, or combinations thereof.

[0066] Terms such as first, second, etc. may be used to describe various components, but these components should not be limited by these terms. These terms are used solely to distinguish one component from another.

[0067] The present inventors, while studying anticancer agents targeting USE1 based on the fact that overexpression of USE1 significantly increases cell proliferation, migration, and invasion, and that inhibition of USE1 expression in lung cancer cells affects the reduction of lung cancer cells, prepared compounds that inhibit USE1 expression, and found that these compounds can effectively inhibit the activity of USE1 by having an aryl substituted basepair and oxycarbonylsulfamate as their basic structure, thereby completing the present invention.

[0068] In addition, the compound according to the present invention can effectively inhibit the activity of UBA6, an E1 activating enzyme, based on the UBA6-USE1 enzyme cascade, and is therefore more effective in tumor suppression.

[0069] Therefore, the present invention may be a compound capable of effectively inhibiting E1-activating enzymes, particularly UBA6. The compound of the present invention is useful for inhibiting E1 activity in vitro and in vivo, and is useful for treating cell proliferation disorders, particularly cancer, and other disorders associated with E1 activity.

[0070] The compound according to the present invention is capable of inhibiting or reducing the activity level of USE1 and / or UBA6.

[0071] The compound according to the present invention is a compound represented by the following chemical formula (I) or a pharmaceutically acceptable salt thereof:

[0072] [Chemical Formula 1]

[0073]

[0074] In the above chemical formula 1,

[0075] Y is C or O,

[0076] Ar is selected from the group consisting of the following compounds,

[0077]

[0078] R1 to R4 are each independently selected from the group consisting of hydrogen, hydroxyl group, halogen and C1-C6 alkyl,

[0079] R5 is -O-SO2NR or -NR-SO2NR,

[0080] R6 is hydrogen, COOR” or CN,

[0081] R7 to R9, R 11 are each independently hydrogen or C1-C6 alkyl,

[0082] R 10 is selected from the group consisting of hydrogen, -C≡CR, COR and S-CR',

[0083] R is hydrogen or C1-C6 alkyl,

[0084] R' is halogen,

[0085] R” is hydrogen, C-CR' and C1-C6 alkyl.

[0086] And, the compound may be a compound represented by the following chemical formula 2:

[0087] [Chemical Formula 2]

[0088]

[0089] In the above chemical formula 2, R1 to R 11 and Ar are as defined above.

[0090] Additionally, the compound may be represented by the following chemical formula 3.

[0091] [Chemical Formula 3]

[0092]

[0093] In the above chemical formula 3, R1 to R6 and Ar are the same as defined above.

[0094] Additionally, the compound may be represented by the following chemical formula 4.

[0095] [Chemical Formula 4]

[0096]

[0097] In the above chemical formula 4, R1 to R6 and Ar are the same as defined above.

[0098] Additionally, the compound may be represented by the following chemical formula 5.

[0099] [Chemical Formula 5]

[0100]

[0101] In the above chemical formula 5, R1 to R6 and Ar are the same as defined above.

[0102] Additionally, the compound may be selected from the following compounds:

[0103]

[0104]

[0105] In addition, the compound of the above formula I may include, by way of example, the following compounds.

[0106]

[0107]

[0108]

[0109] As described in the experimental examples described below, the inventors of the present invention have confirmed that various derivatives having an aryl substituted basepair and oxycarbonylsulfamate as a basic structure and in which a portion of the amine group of the adenosyl is substituted inhibit cancer cell proliferation (see Fig. 1), inhibit cancer cell cluster formation (see Fig. 2), and have an excellent wound healing inhibition effect (see Fig. 3), and can reduce the volume (see Fig. 4) and size (see Fig. 5) of cancer tissue in xenograft experiments targeting mice.

[0110] In addition, the inventors of the present invention performed Western blot analysis using a charging assay method using various compounds according to the present invention, and as a result, it was confirmed that the compounds according to the present invention can effectively inhibit the production of UBA6-Ub (thioester bond formation through binding of UBA6 and ubiquitin) and USE1-Ub (ubiquitin transferred to UBA6 is transferred to USE1 to form a thioester bond).

[0111] The compounds of the present invention target the E1 activating enzyme to maintain the integrity of cell division and cell signaling, providing a unique opportunity to modulate a critical and diverse biochemical pathway. The E1 activating enzyme functions at the first step of the UBL conjugation pathway, and thus, inhibition of the E1 activating enzyme can specifically modulate the downstream UBL cascade.

[0112] Thus, inhibition of these activating enzymes and subsequent downstream UBL conjugation modulates numerous cellular physiological functions crucial for cell division, cell signaling, and disease mechanisms. Therefore, E1 enzymes, such as UBA6, as regulators of diverse cellular functions, are important therapeutic targets for treating various diseases and disorders.

[0113] As used herein, the term "El," "El enzyme," or "El activating enzyme" refers to any one of a family of ATP-dependent activating enzymes involved in activating or promoting ubiquitin or ubiquitin-like ("UBL") conjugation to a target molecule. The El activating enzyme acts through the formation of an adenylation / thioester intermediate to transfer the appropriate UBL to the respective E2 conjugating enzyme via a transthiolation reaction. The activated UBL-E2 then promotes the ultimate conjugation of the UBL to the target protein. A variety of cellular proteins that play a role in cell signaling, the cell cycle, and protein turnover are substrates for UBL conjugation regulated through E1 activating enzymes (e.g., UBLs). Unless otherwise specified, the term "E1 enzyme" is intended to mean any E1 activating enzyme protein, examples of which include, but are not limited to, nedd8 activating enzyme (NAE). (APPBP1 / Uba3)), ubiquitin-activating enzyme (UAE (Uba1)), sumo-activating enzyme (SAE (Aos1 / Uba2)), or ISG15-activating enzyme (Ube1L), Ub-activating enzyme (UBA6 (Ubiquitin-like modifier activating enzyme 6)), and also human UBA6.

[0114] The term "E1 enzyme inhibitor" or "inhibitor of an E1 enzyme" is used to mean a compound having a structure as defined herein that can interact with the E1 enzyme and inhibit its enzymatic activity. Inhibiting E1 enzyme activity means reducing the ability of the E1 enzyme to activate ubiquitin analogue (UBL) conjugation (e.g., ubiquitination) to a substrate peptide or protein. In various embodiments, the reduction in E1 enzyme activity is at least about 50%, at least about 75%, at least about 90%, at least about 95%, or at least about 99%.

[0115] In various embodiments, the concentration of E1 enzyme inhibitor required to reduce E1 enzyme activity is less than about 1 μM, less than about 500 nM, less than about 100 nM, less than about 50 nM, or less than about 10 nM.

[0116] In some embodiments, the inhibition is selective, i.e., the E1 enzyme inhibitor reduces the ability of one or more E1 enzymes (e.g., UBA6) to catalyze UBL conjugation to a substrate peptide or protein at a concentration of the inhibitor lower than that required to produce another, unrelated biological effect. In some such embodiments, the E1 enzyme inhibitor reduces the activity of one E1 enzyme at a concentration lower than that required to reduce the enzymatic activity of a different E1 enzyme. In other embodiments, the E1 enzyme inhibitor also reduces the enzymatic activity of another E1 enzyme, preferably an enzyme associated with the regulation of a signaling pathway involved in cancer (e.g., UBA6).

[0117] Another embodiment of the present invention is a pharmaceutical composition for preventing or treating cancer, comprising the above-described compound as an active ingredient.

[0118] The 'cancer' encompassed in the present invention includes one or more diseases selected from the group consisting of gastric cancer, colon cancer, breast cancer, lung cancer, non-small cell lung cancer, bone cancer, pancreatic cancer, skin cancer, head or cervical cancer, cutaneous or intraocular melanoma, uterine cancer, ovarian cancer, colon cancer, rectal cancer, anal cancer, fallopian tube carcinoma, endometrial carcinoma, cervical carcinoma, vaginal carcinoma, vulvar carcinoma, Hodgkin's disease, esophageal cancer, small intestine cancer, endocrine cancer, thyroid cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, chronic or acute leukemia, lymphocytic lymphoma, bladder cancer, kidney or ureter cancer, renal cell carcinoma, renal pelvic carcinoma, central nervous system tumor, primary CNS lymphoma, spinal cord tumor, brainstem glioma, and pituitary adenoma, and is not limited thereto, but may be more preferably cervical cancer or lung cancer.

[0119] The pharmaceutical composition according to the present invention may further comprise an appropriate carrier, excipient or diluent commonly used in the manufacture of pharmaceutical compositions.

[0120] Examples of carriers, excipients or diluents usable in the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate or mineral oil.

[0121] The pharmaceutical composition according to the present invention can be formulated and used in the form of oral formulations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, external preparations, suppositories, and sterile injection solutions, respectively, according to conventional methods.

[0122] When formulating, it is usually prepared using diluents or excipients such as fillers, bulking agents, binders, wetting agents, disintegrants, and surfactants. Solid preparations for oral administration include tablets, pills, powders, granules, and capsules, and these solid preparations are prepared by mixing the compound with at least one excipient, such as starch, calcium carbonate, sucrose or lactose, or gelatin.

[0123] In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid preparations for oral administration include suspensions, solutions, emulsions, and syrups. In addition to the commonly used simple diluents, such as water and liquid paraffin, various excipients may be included, such as wetting agents, sweeteners, flavoring agents, and preservatives.

[0124] The dosage of the pharmaceutical composition according to the present invention may vary depending on the patient's age, sex, and weight, but may be administered once or several times a day at a dosage of 0.01 ng / kg to 10 g / kg.

[0125] Additionally, the dosage may be increased or decreased depending on the route of administration, severity of the disease, gender, body weight, age, etc. Therefore, the dosage does not limit the scope of the present invention in any way.

[0126] The composition of the present invention can be administered orally or parenterally (e.g., intravenously, subcutaneously, or locally in the abdominal cavity) depending on the intended method, and the dosage range varies depending on the patient's weight, age, sex, health condition, diet, administration time, administration method, excretion rate, and severity of the disease.

[0127] In addition, another embodiment of the present invention provides a health food composition for preventing or improving cancer containing the above-described compound as an active ingredient.

[0128] The health food composition of the present invention may be provided in the form of a powder, granule, tablet, capsule, syrup, or beverage. The health food composition may be used in combination with other foods or food additives in addition to the active ingredient, and may be appropriately used according to conventional methods. The amount of the active ingredient mixed may be appropriately determined depending on the intended use, such as prevention, health, or therapeutic treatment.

[0129] The effective dosage of the active ingredient contained in the above health food composition may be used in accordance with the effective dosage of the above pharmaceutical composition, but in the case of long-term intake for the purpose of health and hygiene or health control, it may be below the above range. It is certain that the active ingredient may be used in an amount exceeding the above range because there is no problem in terms of safety.

[0130] There are no special restrictions on the types of the above health foods, and examples include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages, and vitamin complexes.

[0131] The present invention may be better understood by the following examples, which are provided for illustrative purposes only and are not intended to limit the scope of protection defined by the appended claims.

[0132] [Example]

[0133] [Manufacturing Example 1]

[0134] Synthesis of KA-01, KA-02, and KA-03

[0135] Synthesis of KA-01

[0136]

[0137] KA-01:((2R,3S,4R,5R)-5-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methyl (tert-butoxycarbonyl)sulfamate

[0138] Step 1: The starting material (2R,3R,4S,5R)-2-(6-chloro-9H-purin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (0.4 g, 1.4 mmol) was dissolved in n-BuOH (1 ml), and 3-ethynylaniline (0.16 ml, 1.5 mmol) and DIPEA (0.48 ml, 2.8 mmol) were added at room temperature. The reaction mixture was reacted at 105 °C for 48 h. Water was added to the reaction mixture to stop the reaction, and the mixture was extracted with ethyl acetate. Anhydrous Na2SO4 was added to the organic layer, filtered, and the solvent was removed under reduced pressure. Recrystallization under CH2Cl2 / n-hex conditions gave 173 mg (68% yield) of the target compound.

[0139] Step 2: The starting material (2R,3R,4S,5R)-2-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (50 mg, 0.136 mmol) was dissolved in CH3CN (1.5 ml), and burgess-type reagent (Org. Process Res. Dev.2015,19, 1299-1307) (44 mg, 0.15 mmol) was added, and the mixture was reacted at 50 °C for 20 h. Water was added to the reaction mixture to stop the reaction, and the mixture was extracted with ethyl acetate. Anhydrous Na2SO4 was added to the organic layer, filtered, and the solvent was removed under reduced pressure. Recrystallization under CH2Cl2 / n-hex conditions gave 23 mg (31% yield) of the target compound.

[0140] 1H NMR (300 MHz, MeOD) δ8.46 (s, 1H), 8.40 (s, 1H), 8.07 (t, J = 1.9 Hz, 1H), 7.83 - 7.77 (m, 1H), 7.34 (t, J = 7.9 Hz, 1H), 7.20 (dt, J = 7.7, 1.3 Hz, 1H), 6.12 (d, J = 5.0 Hz, 1H), 4.71 (t, J = 5.1 Hz, 1H), 4.54 (dd, J = 6.2, 3.7 Hz, 2H), 4.43 (t, J = 4.8 Hz, 1H), 4.34 (q, J = 3.9 Hz, 1H), 3.50 (s, 1H), 1.42 (s, 9H)

[0141] KA-02:((2R,3S,4R,5R)-5-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methyl ((2,2,2-trifluoroethoxy)carbonyl)sulfamate

[0142]

[0143] Starting from (2R,3R,4S,5R)-2-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol, the target compound was obtained in the same manner as KA-01 using another burgess-type reagent (Adv. Synth. Catal.2013,355, 3363-3368) in step 2 of KA-01 synthesis.

[0144] 1H NMR (300 MHz, MeOD) δ8.60 (s, 1H), 8.45 (s, 1H), 8.08-8.07 (m, 1H), 7.83-7.79 (m, 1H), 6.16 (d, J = 6.0 Hz, 1H), 4.73 - 4.67 (m, 1H), 4.53 - 4.42 (m, 2H), 4.41-4.38(m, 1H), 4.33-4.30 (m, 3H), 3.50 (s, 1H)

[0145] KA-03:((2R,3S,4R,5R)-5-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methyl cyanosulfamate

[0146]

[0147] Starting from (2R,3R,4S,5R)-2-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol, the target compound was obtained in the same manner as KA-01 using another burgess-type reagent (Adv. Synth. Catal.2013,355, 3363-3368) in step 2 of KA-01 synthesis.

[0148] 1H NMR (300 MHz, MeOD) δ 8.49 (s, 1H), 8.46 (s, 1H), 8.07 (s, 1H), 7.82 (d, J = 8.4 Hz, 1H), 7.34 (t, J = 8.0 Hz, 1H), 7.19 (d, J = 7.6 Hz, 1H), 6.16 (d, J = 5.5 Hz, 1H), 4.72 - 4.64 (m, 1H), 4.47 - 4.37 (m, 2H), 4.36-4.33 (m, 1H), 3.69 (s, 1H), 3.49 (s, 1H)

[0149] Synthesis of KA-04

[0150] KA-04:((2R,3S,4R,5R)-5-(2-chloro-6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methyl (tert-butoxycarbonyl)sulfamate

[0151]

[0152] The target compound was obtained using (2R,3R,4S,5R)-2-(2-chloro-6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol as a starting material in the same manner as step 2 of KA-01.

[0153] 1H NMR (300 MHz, MeOD) δ8.46 (s, 1H), 8.01 (s, 1H), 7.86 (d, J = 8.4 Hz, 1H), 7.38 (t, J = 8.0 Hz, 1H), 7.24 (d, J = 7.8 Hz, 1H), 6.09 (d, J) = 5.1 Hz, 1H), 4.67 (t, J = 5.1 Hz, 1H), 4.50 - 4.31 (m, 4H), 3.53 (s, 1H)

[0154] Synthesis of KA-05

[0155] KA-05: ((2R,3S,4R,5R)-3,4-dihydroxy-5-(6-((3-methoxyphenyl)amino)-9H-purin-9-yl)tetrahydrofuran-2-yl)methyl (tert-butoxycarbonyl)sulfamate

[0156]

[0157] The target compound was obtained using (2R,3S,4R,5R)-2-(hydroxymethyl)-5-(6-((3-methoxyphenyl)amino)-9H-purin-9-yl)tetrahydrofuran-3,4-diol as a starting material in the same manner as step 2 of KA-01.

[0158] LC / MS [M+H] + = 553

[0159] Synthesis of KA-06

[0160] KA-06:((2R,3S,4R,5R)-5-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methyl (methoxycarbonyl)sulfamate

[0161]

[0162] Using (2R,3R,4S,5R)-2-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol as a starting material, the target compound was obtained in the same manner as KA-01 using another burgess-type reagent (CAS 29684-56-8) in step 2 of KA-01 synthesis.

[0163] 1H NMR (300 MHz, MeOD) δ8.60 (s, 1H), 8.45 (s, 1H), 8.08 (t, J = 1.6 Hz, 1H), 7.81 (d, J = 8.3 Hz, 1H), 7.34 (t, J = 7.9 Hz, 1H), 7.19 (d, J) = 7.6 Hz, 1H), 6.16 (d, J = 5.9 Hz, 1H), 4.74 - 4.69 (m, 1H), 4.41 (d, J = 4.9 Hz, 1H), 4.31 (s, 2H), 3.70 (s, 1H), 3.60 (s, 3H), 3.49 (s, 1H)

[0164] Synthesis of KA-07

[0165] KA-07:((2R,3S,4R,5R)-3,4-dihydroxy-5-(6-((3-((trifluoromethyl)thio)phenyl)amino)-9H-purin-9-yl)tetrahydrofuran-2-yl)methyl (tert-butoxycarbonyl)sulfamate

[0166]

[0167] The target compound was obtained using 2R,3S,4R,5R)-2-(hydroxymethyl)-5-(6-((3-((trifluoromethyl)thio)phenyl)amino)-9H-purin-9-yl)tetrahydrofuran-3,4-diol as a starting material in the same manner as step 2 of KA-01.

[0168] 1H NMR (300 MHz, MeOD) δ8.49 (s, 1H), 8.48 (s, 1H), 8.38 - 8.37 (m, 1H), 8.06 - 8.00 (m, 1H), 7.49 (t, J = 7.9 Hz, 1H), 7.40 (d, J = 7.7 Hz, 1H), 6.14 (d, J = 5.3 Hz, 1H), 4.73 (t, J = 5.2 Hz, 1H), 4.50 - 4.44 (m, 2H), 4.43 (d, J = 4.6 Hz, 1H), 4.34 (t, J = 3.7 Hz, 1H), 1.42 (s, 9H)

[0169] Synthesis of KA-08

[0170] KA-08:((2R,3R,4S,5R)-5-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-4-fluoro-3-hydroxytetrahydrofuran-2-yl)methyl (tert-butoxycarbonyl)sulfamate

[0171]

[0172] The target compound was obtained using (2R,3R,4S,5R)-5-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-4-fluoro-2-(hydroxymethyl)tetrahydrofuran-3-ol as a starting material in the same manner as step 2 of KA-01.

[0173] 1H NMR (300 MHz, MeOD) δ8.47 (s, 1H), 8.34 (d, J = 2.4 Hz, 1H), 8.06 (d, J = 1.9 Hz, 1H), 7.80 (dt, J = 8.3, 1.7 Hz, 1H), 7.35 (t, J = 7.9) Hz, 1H), 7.20 (dt, J = 7.6, 1.3 Hz, 1H), 6.59 (dd, J = 17.3, 3.9 Hz, 1H), 5.18 (dt, J = 51.9, 3.5 Hz, 1H), 4.64 (dd, J = 4.3, 2.8 Hz, 1H), 4.56 (d, J = 5.0 Hz, 2H), 4.27 (q, J = 4.8 Hz, 1H), 3.50 (s, 1H), 1.43 (s, 9H)

[0174] Synthesis of KA-09

[0175] KA-09: ((2R,3R,4S,5R)-4-fluoro-3-hydroxy-5-(6-((3-((trifluoromethyl)thio)phenyl)amino)-9H-purin-9-yl)tetrahydrofuran-2-yl)methyl (tert-butoxycarbonyl)sulfamate

[0176]

[0177] The target compound was obtained using (2R,3R,4S,5R)-4-fluoro-2-(hydroxymethyl)-5-(6-((3-((trifluoromethyl)thio)phenyl)amino)-9H-purin-9-yl)tetrahydrofuran-3-ol as a starting material in the same manner as step 2 of KA-01.

[0178] 1H NMR (300 MHz, MeOD) δ8.52 (s, 1H), 8.43 (d, J = 2.3 Hz, 1H), 8.40 (t, J = 1.7 Hz, 1H), 8.08 - 8.03 (m, 1H), 7.51 (t, J = 7.9 Hz, 1H), 7.42 (d, J = 7.7 Hz, 1H), 6.61 (dd, J = 17.1, 3.9 Hz, 1H), 5.29 - 5.08 (m, 1H), 4.69 - 4.60 (m, 1H), 4.43 (d, J = 5.2 Hz, 2H), 4.28 (t, J = 4.7 Hz, 1H), 1.45 (s, 9H)

[0179] Synthesis of KA-10, KA-11, and KA-12

[0180] KA-10:tert-butyl (N-(((2R,3S,4R,5R)-5-(4-((3-ethynylphenyl)amino)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methyl)sulfamoyl)carbamate

[0181]

[0182] The target compound was obtained using (2R,3S,4R,5R)-2-(aminomethyl)-5-(4-((3-ethynylphenyl)amino)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)tetrahydrofuran-3,4-diol as a starting material in the same manner as step 2 of KA-01.

[0183] 1H NMR (300 MHz, DMSO) δ10.92 (s, 1H), 10.92 (s, 1H), 8.55 (s, 1H), 8.46 (s, 1H), 8.17 (s, 1H), 7.98 (d, J = 8.5 Hz, 1H), 7.35 (t, J = 8.0 Hz, 1H), 7.15 (d, J = 7.6 Hz, 1H), 5.92 (d, J = 6.6 Hz, 1H), 5.54 (d, J = 6.2 Hz, 1H), 5.34 (d, J = 4.4 Hz, 1H), 4.74 (q, J = 6.0 Hz, 1H), 4.18 (s, 1H), 4.17 - 4.07 (m, 2H), 3.17 (d, J = 5.2 Hz, 1H), 1.75 (s, 1H), 1.37 (s, 9H)

[0184] KA-11: methyl (N-(((2R,3S,4R,5R)-5-(4-((3-ethynylphenyl)amino)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methyl)sulfamoyl)carbamate

[0185]

[0186] Using (2R,3S,4R,5R)-2-(aminomethyl)-5-(4-((3-ethynylphenyl)amino)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)tetrahydrofuran-3,4-diol as a starting material, the target compound was obtained in the same manner as KA-01 using another burgess-type reagent (CAS 29684-56-8) in step 2 of KA-01 synthesis.

[0187] 1H NMR (300 MHz, DMSO) δ10.14 (s, 1H), 8.55 (s, 1H), 8.48 (s, 1H), 8.18 (t, J = 1.9 Hz, 1H), 7.98 (dd, J = 8.4, 2.2 Hz, 1H), 7.35 (t, J = 8.0 Hz, 1H), 7.15 (d, J = 7.6 Hz, 1H), 4.74 (q, J = 6.1 Hz, 1H), 4.18 (s, 1H), 4.14 - 4.07 (m, 2H), 3.56 (s, 3H), 3.05 (q, J = 7.3 Hz, 2H)

[0188] KA-12:tert-butyl (N-(((2R,3S,4R,5R)-5-(4-((3-ethynylphenyl)amino)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-3,4-dihydroxy-3-methyltetrahydrofuran-2-yl)methyl)sulfamoyl)carbamate

[0189]

[0190] The target compound was obtained using (2R,3S,4R,5R)-2-(aminomethyl)-5-(4-((3-ethynylphenyl)amino)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-3-methyltetrahydrofuran-3,4-diol as a starting material in the same manner as step 2 of KA-01.

[0191] 1H NMR (300 MHz, MeOD) δ8.31 (s, 1H), 7.96 (s, 1H), 7.82 - 7.75 (m, 1H), 7.67 (d, J = 3.7 Hz, 1H), 7.34 (t, J = 7.9 Hz, 1H), 7.20 (d, J = 7.3 Hz, 1H), 6.82 (d, J = 3.8 Hz, 1H), 6.36 (d, J = 7.9 Hz, 1H), 4.40 (d, J = 8.1 Hz, 1H), 4.37 - 4.35 (m, 2H), 4.22 (t, J = 3 Hz, 1H), 3.50 (s, 1H), 1.52 (s, 3H), 1.47 (s, 9H)

[0192] Synthesis of KA-13 and KA-14

[0193] KA-13:((2R,3S,4R,5R)-5-(4-((3-ethynylphenyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methyl (tert-butoxycarbonyl)sulfamate

[0194]

[0195] The starting material (2R,3R,4S,5R)-2-(4-((3-ethynylphenyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol was synthesized by referring to reference (J. Med. Chem. 1981, 24, 1165-1172), and the target compound was obtained by the same method as step 2 of KA-01.

[0196] 1H NMR (300 MHz, MeOD) δ8.46 (s, 1H), 8.25 (s, 1H), 8.01 (s, 1H), 7.81 (d, J = 8.1 Hz, 1H), 7.39 (t, J = 7.9 Hz, 1H), 7.28 (d, J = 7.6 Hz, 1H), 6.40 (d, J = 2.8 Hz, 1H), 4.76 - 4.70 (m, 1H), 4.66 (t, J = 5.3 Hz, 1H), 4.39 - 4.21 (m, 2H), 4.24 - 4.15 (m, 1H), 3.55 (s, 1H), 1.40 (s, 9H)

[0197] KA-14:((2R,3S,4R,5R)-5-(4-((3-ethynylphenyl)amino)-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methyl (tert-butoxycarbonyl)sulfamate

[0198]

[0199] The starting material (2R,3R,4S,5R)-2-(4-((3-ethynylphenyl)amino)-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol was synthesized by referring to reference (J. Med. Chem. 1981, 24, 1165-1172), and the target compound was obtained by the same method as step 2 of KA-01.

[0200] 1H NMR (300 MHz, MeOD) δ8.48 (d, J = 3.1 Hz, 1H), 8.13 (s, 1H), 8.01 (s, 1H), 7.78 (d, J = 8.3 Hz, 1H), 7.41 (t, J = 7.9 Hz, 1H), 7.41 (t, J = 7.9 Hz, 1H), 6.34 (d, J = 3.1 Hz, 1H), 4.73 - 4.68 (m, 1H), 4.60 (t, J = 5.2 Hz, 1H), 4.43 (t, J = 6.9 Hz, 1H), 4.30 (t, J = 5.4 Hz, 2H), 3.58 (s, 1H), 1.41 (s, 9H)

[0201] Synthesis of KA-16 and KA-17

[0202] KA-16:((2R,3R,4S,5R)-5-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-4-fluoro-3-hydroxytetrahydrofuran-2-yl)methyl ((2,2,2-trifluoroethoxy)carbonyl)sulfamate

[0203]

[0204] Starting from (2R,3R,4S,5R)-5-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-4-fluoro-2-(hydroxymethyl)tetrahydrofuran-3-ol, the target compound was obtained in the same manner as in KA-01 using another burgess-type reagent (Adv. Synth. Catal.2013,355, 3363-3368) in step 2 of KA-01.

[0205] LC / MS [M+H] + = 575

[0206] KA-17:((2R,3R,4S,5R)-4-fluoro-3-hydroxy-5-(6-((3-((trifluoromethyl)thio)phenyl)amino)-9H-purin-9-yl)tetrahydrofuran-2-yl)methyl ((2,2,2-trifluoroethoxy)carbonyl)sulfamate

[0207]

[0208] Starting from (2R,3R,4S,5R)-4-fluoro-2-(hydroxymethyl)-5-(6-((3-((trifluoromethyl)thio)phenyl)amino)-9H-purin-9-yl)tetrahydrofuran-3-ol, the target compound was obtained in the same manner as in KA-01 using another burgess-type reagent (Adv. Synth. Catal.2013,355, 3363-3368) in step 2 of KA-01.

[0209] 1H NMR (300 MHz, MeOD) δ8.51 (s, 1H), 8.43 (d, J = 2.2 Hz, 1H), 8.39 (s, 1H), 8.08 - 8.02 (m, 1H), 7.50 (t, J = 7.9 Hz, 1H), 7.42 (d, J = 7.8 Hz, 1H), 6.60 (dd, J = 17.2, 3.9 Hz, 1H), 5.18 (dt, J = 51.9, 3.4 Hz, 2H), 4.68 - 4.58 (m, 1H), 4.52 (q, J = 8.9 Hz, 2H), 4.39 (d, J = 5.2 Hz, 2H), 4.26 (q, J = 4.8 Hz, 1H)

[0210] Synthesis of KA-18 and KA-19

[0211] KA-18: ((2R,3R,4S,5R)-5-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-4-fluoro-3-hydroxytetrahydrofuran-2-yl)methyl (isopropoxycarbonyl)sulfamate

[0212]

[0213] Starting from (2R,3R,4S,5R)-5-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-4-fluoro-2-(hydroxymethyl)tetrahydrofuran-3-ol, the target compound was obtained in the same manner as KA-01 using another burgess-type reagent (Org. Process Res. Dev.2015,19, 1299-1307) in step 2 of KA-01.

[0214] 1H NMR (300 MHz, MeOD) δ8.46 (d, J = 0.9 Hz, 1H), 8.41 - 8.37 (m, 1H), 8.07 (d, J = 2.0 Hz, 1H), 7.81 (d, J = 8.3 Hz, 1H), 7.35 (t, J = 7.9) Hz, 1H), 7.20 (d, J = 7.7 Hz, 1H), 6.58 (dd, J = 17.2, 3.9 Hz, 1H), 5.12 (dd, J = 51.8, 3.1 Hz, 1H), 4.62 (d, J = 14.6 Hz, 1H), 4.65 - 4.57 (m, 1H), 4.45 - 4.38 (m, 2H), 4.27 - 4.23 (m, J = 4.9 Hz, 1H), 3.64 (d, J = 2.1 Hz, 1H), 3.50 (s, 1H), 1.21 (d, J = 6.2 Hz, 6H)

[0215] KA-19: ((2R,3R,4S,5R)-4-fluoro-3-hydroxy-5-(6-((3-((trifluoromethyl)thio)phenyl)amino)-9H-purin-9-yl)tetrahydrofuran-2-yl)methyl (isopropoxycarbonyl)sulfamate

[0216]

[0217] Starting from (2R,3R,4S,5R)-4-fluoro-2-(hydroxymethyl)-5-(6-((3-((trifluoromethyl)thio)phenyl)amino)-9H-purin-9-yl)tetrahydrofuran-3-ol, the target compound was obtained in the same manner as KA-01 using another burgess-type reagent (Org. Process Res. Dev.2015,19, 1299-1307) in step 2 of KA-01.

[0218] LC / MS [M+H] + = 611

[0219] Synthesis of KA-20

[0220] KA-20:((2R,3R,5R)-5-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-4,4-difluoro-3-hydroxytetrahydrofuran-2-yl)methyl (tert-butoxycarbonyl)sulfamate

[0221]

[0222] The target compound was obtained using (2R,3R,5R)-5-(6-((3-ethynylphenyl)amino)-9H-purin-9-yl)-4,4-difluoro-2-(hydroxymethyl)tetrahydrofuran-3-ol as a starting material in the same manner as step 2 of KA-01.

[0223] LC / MS [M+H] + = 567

[0224] [Comparative Example]: Preparation of a previously known compound

[0225] As a comparative example, a previously known compound was prepared using the same or similar method as the above-described example.

[0226] UBA6i is a compound (I-01) known from patent WO2014 / 022744 A1, and its structure is as follows.

[0227]

[0228] [Experimental Example 1]: Cell Proliferation Analysis

[0229] The degree of cell proliferation was analyzed using the CellTiter-Glo Luminescent Cell Viability Assay Kit (G9241, Promega). 2 x 10 4 A549 cells were cultured in 96-well cell culture dishes, and analyses were conducted from the first day to the third day. Before analysis, the CellTiter-Glo reagent and the cell culture dish to be used for the analysis were exposed to room temperature for 30 minutes to adjust the temperature to room temperature.

[0230] 100 μl of the reagent diluted 1 / 10 in PBS and KA-01 according to the present invention were treated at 1 μM, 3 μM, and 10 μM, respectively, and reacted at room temperature for 5 minutes in an orbital shaker. After reacting for an additional 10 minutes at room temperature, the luminescence intensity was measured using a GloMax®96 Microplate Luminometer (Promega).

[0231] The results are as shown in Fig. 1. Fig. 1 shows the results of analyzing the cell proliferation ability of KA-01 according to one embodiment of the present invention. As shown therein, it was confirmed that treatment with KA-01 according to the present invention could inhibit cell proliferation in a concentration-dependent manner.

[0232] [Experimental Example 2]: Cell Colony Formation Analysis

[0233] A549 cells were washed once with PBS solution and trypsinized for 1 minute at 37°C in a CO2 incubator. The detached A549 cells were centrifuged at 1,200 rpm for 5 minutes. After mixing the cells with the culture solution, 1,000 cells per ml were prepared using a hemacytometer. 1 ml of the prepared A549 cells was added to a 6-well plate to which 2 ml of culture had been added in advance, and KA-01 according to the present invention was treated at 1 μM, 3 μM, and 10 μM, respectively, and mixed well. After culturing the cells for 10 to 20 days in a 37°C CO2 incubator, the culture solution was removed, washed once with PBS, and fixed with a 3.7% PFA (EMS, 15710-S) solution for 5 minutes. After staining with a 0.05% crystal violet (Sigma-Aldrich, C3886) solution for 30 minutes, the dye on the plate was removed using tap water, and the plate was scanned or photographed after waiting for it to dry.

[0234] The results are as shown in Fig. 2. Fig. 2 shows the results of analyzing the cell colony formation ability of KA-01 according to one embodiment of the present invention. As shown therein, it was confirmed that treatment with KA-01 according to the present invention could inhibit cell colony formation in a concentration-dependent manner.

[0235] [Experimental Example 3]: Analysis of wound healing ability

[0236] To investigate the migration ability of A549 cells, a wound healing assay was performed. A549 cells were washed once with phosphate-buffered saline (PBS) solution and trypsinized for 1 min in a CO2 incubator at 37°C. The detached A549 cells were centrifuged at 1,200 rpm for 5 min, remixed with culture medium, and counted at 500,000 cells per ml using a hemacytometer.

[0237] After that, one or two lines were drawn on the bottom of a 6-well plate, and 1 ml of culture solution was first added to each well. 1 ml of the prepared A549 cells were added, and KA-01 according to the present invention was treated at 1 μM, 3 μM, and 10 μM, respectively, and the plate was shaken evenly. After that, the cells were cultured in a CO2 incubator at 37°C. When the cell density was 80 to 90%, a line was carefully drawn using a 200P tip, and the cells were carefully washed twice using the culture solution. After that, a 100x magnification 0-hour photograph was obtained using a microscope (NiKon eclipse Ti-U). When obtaining the next photograph, the cells were washed once with the warmed culture solution, and then the photograph was obtained, and the photograph was analyzed using the Image J program.

[0238] The results are as shown in Fig. 3. Fig. 3 shows the results of analyzing the wound healing ability of KA-01 according to one embodiment of the present invention. As shown therein, it was found that treatment with KA-01 according to the present invention resulted in excellent wound healing ability in a concentration-dependent manner.

[0239] [Experimental Example 4]: Cancer Cell Xenograft Analysis

[0240] Male BALB / c nude mice, 4 to 6 weeks old, were randomly assigned to experimental groups. 1 x 10 per mouse was injected subcutaneously. 7 A549 cells were injected in a volume of 100 μl. The volume of the grown cancer tissue was monitored after 7 days (long axis x short axis x shortened / 2), and about 50 mm 3 When the condition reached a certain level, treatment trials began.

[0241] Vehicle was used as a negative control group, MLN4924 at a dose of 60 mg / kg was used as a positive control group, and KA-01 according to the present invention at doses of 12.5 mg / kg, 25 mg / kg, and 50 mg / kg was administered intraperitoneally at intervals of 2 to 3 days. The volume and size of the tumor tissue and the body weight of the mice were measured and monitored simultaneously with the drug injection.

[0242] The results are as shown in Figs. 4 and 5. Fig. 4 shows the results of changes in the volume of cancer tissue treated with KA-01 according to one embodiment of the present invention. Fig. 5 shows the results of changes in the size of cancer tissue treated with KA-01 according to one embodiment of the present invention. As shown here, it was confirmed that when KA-01 according to the present invention was treated, the volume and size of cancer tissue decreased in a concentration-dependent manner.

[0243] [Experimental Example 5]: Western Blot (Charging Assay) Analysis

[0244] 5 x 10 A549 cells 5 After being grown in a 6-well cell culture dish, each of the compounds according to the present invention, including KA-01, was treated at various concentrations.

[0245] Afterwards, the cells were lysed using MES lysis buffer (50 mM MES pH 4.5, 150 mM NaCl, 0.5 mM, 0.5 mM β-mercaptoethanol, 0.2% NP-40, protease inhibitors) and proteins were extracted by centrifugation. Then, the extracted proteins were mixed with NuPAGE™LDS Sample Buffer to prepare samples for analysis. Then, 20 μg protein samples were subjected to electrophoresis through non-reducing Bolt™ Bis-Tris Plus Mini Protein Gels (4-12%). Some samples were used in the experiment in a reduced state by treating them with 0.5 mM β-mercaptoethanol before electrophoresis. The separated proteins were reacted with primary antibodies and secondary antibodies conjugated with HRP (horseradish peroxidase), and chemiluminescence was confirmed using ECL solution (Clarity Western ECL Substrate).

[0246] The results are as shown in Figs. 6 to 23. Figure 6 shows the results of a Western blot (Charging assay) analysis of compound KA-01 according to the present invention, Figure 7 shows the results of a Western blot analysis of compound KA-02 according to the present invention, Figure 8 shows the results of a Western blot analysis of compound KA-03 according to the present invention, Figure 9 shows the results of a Western blot analysis of compound KA-04 according to the present invention, Figure 10 shows the results of a Western blot analysis of compound KA-05 according to the present invention, Figure 11 shows the results of a Western blot analysis of compound KA-06 according to the present invention, Figure 12 shows the results of a Western blot analysis of compound KA-07 according to the present invention, Figure 13 shows the results of a Western blot analysis of compound KA-08 according to the present invention, Figure 14 shows the results of a Western blot analysis of compound KA-09 according to the present invention, Figure 15 shows the results of a Western blot analysis of compounds KA-10 and KA-11 according to the present invention, and Figure 16 shows the results of a Western blot analysis of compound KA-12 according to the present invention. The results of the blot analysis are as follows: FIG. 17 is a result of the Western blot analysis of compound KA-13 according to the present invention; FIG. 18 is a result of the Western blot analysis of compound KA-14 according to the present invention; FIG. 19 is a result of the Western blot analysis of compound KA-15 according to the present invention; FIG. 20 is a result of the Western blot analysis of compounds KA-16 and KA-17 according to the present invention; FIG. 21 is a result of the Western blot analysis of compound KA-18 according to the present invention; FIG. 22 is a result of the Western blot analysis of compound KA-19 according to the present invention; and FIG. 23 is a result of the Western blot analysis of compound KA-20 according to the present invention.

[0247] As shown in Fig. 6, compound KA-01 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was hardly observed when KA-01 was used at a concentration of 3.0 μM or higher. In Fig. 6, the control is the result using DMSO.

[0248] As shown in Fig. 7, compound KA-02 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was hardly observed when KA-02 was used at a concentration of 10.0 μM or higher.

[0249] As shown in Fig. 8, compound KA-03 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was not significantly observed when KA-03 was used at a concentration of 100 μM or higher.

[0250] As shown in Fig. 9, compound KA-04 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was hardly observed when KA-04 was used at a concentration of 30.0 μM or higher.

[0251] As shown in Fig. 10, compound KA-05 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was hardly observed when KA-05 was used at a concentration of 100.0 μM or higher.

[0252] As shown in Fig. 11, the compound KA-06 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, it was confirmed that the production of USE1-Ub was inhibited when KA-06 was used at a concentration of 50.0 μM or higher.

[0253] As shown in Fig. 12, compound KA-07 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was hardly observed when KA-07 was used at a concentration of 3.0 μM to 10.0 μM or higher.

[0254] As shown in Fig. 13, compound KA-08 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was hardly observed when KA-08 was used at a concentration of 0.5 μM or higher.

[0255] As shown in Fig. 14, compound KA-09 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was significantly reduced when KA-09 was used at a concentration of 30.0 μM or higher.

[0256] As shown in FIG. 15, compound KA-10 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was significantly reduced when KA-10 was used at a concentration of 30.0 μM or higher.

[0257] As shown in FIG. 15, compound KA-11 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was significantly reduced when KA-11 was used at a concentration of 30.0 μM or higher.

[0258] As shown in Fig. 16, compound KA-12 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was significantly reduced when KA-12 was used at a concentration of 30.0 μM or higher.

[0259] As shown in Fig. 17, compound KA-13 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was significantly reduced when KA-13 was used at a concentration of 30.0 μM or higher.

[0260] As shown in Fig. 18, compound KA-14 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was significantly reduced when KA-14 was used at a concentration of 30.0 μM or higher.

[0261] As shown in Fig. 19, compound KA-15 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was hardly observed when KA-15 was used at a concentration of 10.0 μM or higher.

[0262] As shown in Fig. 20, compound KA-16 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was significantly reduced when KA-16 was used at a concentration of 10.0 μM or higher.

[0263] As shown in Fig. 20, compound KA-17 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was significantly reduced when KA-17 was used at a concentration of 10.0 μM or higher.

[0264] As shown in Fig. 21, compound KA-18 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was hardly observed when KA-18 was used at a concentration of 10.0 μM or higher.

[0265] As shown in Fig. 22, compound KA-19 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was hardly observed when KA-19 was used at a concentration of 10.0 μM or higher.

[0266] As shown in Fig. 23, compound KA-20 according to the present invention was able to effectively inhibit the production of UBA6-Ub and USE1-Ub, and among them, USE1-Ub was hardly observed when KA-20 was used at a concentration of 10.0 μM or higher.

[0267] Although the present invention has been illustrated and described above with respect to specific preferred embodiments, it will be apparent to those skilled in the art that the present invention may be variously modified and changed without departing from the technical features or scope of the present invention as defined by the following claims.

Claims

1. A compound represented by the following chemical formula 1 or a pharmaceutically acceptable salt thereof: [Chemical Formula 1] In the above chemical formula 1, Y is C or O, Ar is selected from the group consisting of the following compounds, R 1 Inland R 4 are each independently hydrogen, hydroxyl group, halogen and C 1 -C 6 is selected from the group consisting of alkyl, R 5 is -O-SO 2 NR or -NR-SO 2 NR 2 And, R 6 is hydrogen, COOR” or CN, R 7 Inland R 9 , R 11 are each independently hydrogen or C 1 -C 6 It is alkyl, R 10 is selected from the group consisting of hydrogen, -C≡CR, COR and S-CR', R is hydrogen or C 1 -C 6 It is alkyl, R' is halogen, R” is hydrogen, C-CR' and C 1 -C 6 It's alkyl.

2. In paragraph 1, The compound is a compound represented by the following chemical formula 2: [Chemical formula 2] In the above chemical formula 2, R 1 Inland R 11 and Ar are as defined in the first paragraph above.

3. In paragraph 1, The compound is a compound represented by the following chemical formula 3: [Chemical Formula 3] In the above chemical formula 3, R 1 Inland R 6 and Ar are as defined in the first paragraph above.

4. In paragraph 1, The compound above is a compound represented by the following chemical formula 4: [Chemical Formula 4] In the above chemical formula 4, R 1 Inland R 6 and Ar are as defined in the first paragraph above.

5. In paragraph 1, The compound above is a compound represented by the following chemical formula 5: [Chemical Formula 5] In the above chemical formula 5, R 1 Inland R 6 and Ar are as defined in the first paragraph above.

6. In paragraph 1, The compound is selected from the following compounds:

7. A pharmaceutical composition for preventing or treating cancer, comprising a compound of any one of claims 1 to 6 as an active ingredient.

8. In paragraph 7, A pharmaceutical composition for preventing or treating cancer, wherein the cancer is at least one selected from the group consisting of liver cancer, colon cancer, cervical cancer, kidney cancer, stomach cancer, prostate cancer, breast cancer, brain tumor, lung cancer, uterine cancer, colon cancer, bladder cancer, blood cancer, and pancreatic cancer.

9. In paragraph 7, A pharmaceutical composition for preventing or treating cancer, wherein the cancer is cervical cancer or lung cancer.

10. A health food composition for preventing or improving cancer, comprising a compound of any one of claims 1 to 6 as an effective ingredient.

11. A method for reducing E1 enzyme activity in a sample, comprising contacting the sample with a compound of any one of claims 1 to 6.

12. In paragraph 11, A method for reducing E1 enzyme activity in a sample, wherein the E1 enzyme is UBA6 (Ubiquitin-like modifier activating enzyme 6).

13. In paragraph 11, A method for reducing E1 enzyme activity in a sample, wherein the sample contains cancer cells.

14. A method for inhibiting tumor cells, comprising a step of treating cells with a compound of any one of claims 1 to 6.

15. A method for preventing or treating cancer, comprising administering to a patient a composition for preventing or treating cancer of Article 7.

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