Nitrophenyl-acrylamide and its uses

Abstract

Provided herein are nitrophenyl-acrylamide compounds, their preparation and their use.The nitrophenyl-acrylamide compounds described herein have been found to be useful and effective therapeutic agents, including for the treatment of brain tumors, such as glioblastoma.This is surprising, since acrylamide is toxic according to the World Health Organization.Glioblastoma is the most common primary brain tumor and one of the most deadly cancers due to the presence of tumor-proliferating glioma stem cells, and the present invention provides an additional treatment option to current therapy.

Description

[Technical field]

[0001] Related Applications This application claims priority to U.S. Provisional Patent Application No. 63 / 175,476, filed April 15, 2021, the entire contents of which are incorporated herein by reference. [Background technology]

[0002] background Glioblastoma is the most common primary brain tumor and one of the most deadly cancers due to the presence of tumor-proliferating glioma stem cells.Current treatment options include surgical resection, chemotherapy, and radiation.Although these treatments may be temporarily effective, they generally only delay the onset of further symptoms or recurrence, and contain undesirable side effects.Moreover, despite the usefulness of current therapies, recurrence after therapeutic intervention using these current options is unfortunately inevitable.Therefore, additional treatment options are needed. Accordingly, provided herein are compounds useful for treating diseases such as brain tumors, including glioblastoma. [Prior art documents] [Non-patent literature]

[0003] [Non-Patent Document 1] Russian Journal of Physical Chemistry B, 2019, 13, 49-61 Summary of the Invention [Means for solving the problem]

[0004] Abstract In some embodiments, the nitrophenyl-acrylamide compound has the formula: [ka] or a pharma- ceutically acceptable salt thereof is provided herein.

[0005] In some embodiments, the nitrophenyl-acrylamide compound has the formula: [ka] or a pharma- ceutically acceptable salt thereof is provided herein.

[0006] In some embodiments, a method is provided herein. [Brief description of the drawings]

[0007] [Figure 1] Figure 1 shows the results of a qPCR array in two patient-derived glioma stem cells (GSCs) treated with entacapone for 48 hours, showing significant inhibition of various cancer stem cell transcripts.

[0008] [Diagram 2] Figure 2 shows the normalized GSC invasion area of ​​live imaging over 66 h after entacapone treatment (top line is DMSO, bottom line is 40 μM entacapone).

[0009] [Diagram 3] FIG. 3 shows the inhibition of Notch1 protein expression induced by entacapone as described by Example 7.

[0010] [Figure 4] FIG. 4 shows the synthetic scheme for the common core intermediate (CORE) described in Example 8.

[0011] [Diagram 5] FIG. 5 shows a synthetic scheme for compound 1 described in Example 9.

[0012] [Figure 6] FIG. 6 shows a synthetic scheme for compound 20 described in Example 10.

[0013] [Figure 7] FIG. 7 shows a synthetic scheme for compound 29 described in Example 11.

[0014] [Figure 8] FIG. 8 shows a synthetic scheme for compound 31 described in Example 12.

[0015] [Figure 9] FIG. 9 shows a synthetic scheme for compound 33 described in Example 13.

[0016] [Figure 10] FIG. 10 shows a synthetic scheme for compound 35 described in Example 14.

[0017] [Figure 11] FIG. 11 shows the inhibition of glioma stem cell invasion by compound 31 (squares) compared to the control DMSO (circles).

[0018] [Figure 12] FIG. 12 shows the inhibition of glioma stem cell invasion by compound 31 (right side) compared to the control DMSO (left side).

[0019] [Figure 13] FIG. 13 shows the inhibition of the self-renewal capacity of diffuse endogenous pontine glioma cells by compound 31.

[0020] [Figure 14] Figure 14 shows the inhibition of diffuse endogenous pontine glioma cell invasion by compound 31 compared to the control DMSO. Horizontal solid black bars represent 800 μm.

[0021] [Figure 15] FIG. 15 shows quantified inhibition of diffuse endogenous pontine glioma cell invasion by compound 31 compared to the control DMSO. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0022] Detailed Description The nitrophenyl-acrylamide compounds described herein have been found to be useful and effective therapeutic agents, including for the treatment of brain tumors, such as glioblastoma. This is surprising, since acrylamide is toxic, according to the World Health Organization. Moreover, acrylamide is not suitable as a cancer treatment, but rather is a potential carcinogenic chemical. In addition, a derivative of acrylamide, N-(4-nitrophenyl)acrylamide, has an IC of 1 mM in HeLa cells. 50 and have been found to have low in vitro toxicity towards cancer cells (Russian Journal of Physical Chemistry B, 2019, 13, 49-61). Thus, acrylamide, and by extension nitrophenyl-acrylamides, represent a group of compounds that are commonly understood as potentially carcinogenic and ineffective cancer inhibitors.

[0023] The compounds provided herein have also been found to be fat mass and obesity-associated protein (FTO) inhibitors. This is useful because overexpression of METTL3 or inhibition of RNA demethylase FTO suppresses glioma stem cell (GSC) proliferation and self-renewal. Furthermore, inhibition of FTO with the ethyl ester of meclofenamic acid has been shown to suppress tumor progression and substantially extend the life span of GSC-implanted mice. Thus, the compounds described herein are useful as FTO inhibitors or in the treatment of FTO-related diseases.

[0024] It has also been found that the compounds provided herein are Notch1 inhibitors.This is useful because Notch1, a member of the Notch family, is involved in many types of cancer, including breast cancer (in some embodiments, triple-negative breast cancer), leukemia, brain tumors, lung cancer (in some embodiments, non-small cell lung cancer), and many other cancers.Therefore, the compounds described herein are useful as Notch1 inhibitors or in treating Notch1-related diseases. compound / composition

[0025] Thus, provided herein is a nitrophenyl-acrylamide and uses thereof. In some embodiments, the compound has the formula: [ka] or a pharma- ceutically acceptable salt thereof is provided herein. [In the formula, [ka] is a single or double bond; R 1 is C 1~4 is alkyl; R 2 is C 1~4 is alkylene; R 3 is N, O, S, NH, CH, or N-(C 1~4 alkyl); R 4 , H, C 1~4 Alkyl, or (C 1~4 alkyl)-OH; or R 3 and R 4 are combined to form C 2~6 forming a heterocycloalkyl; R 5 is C(O)N(H)(C 1~4 alkyl), C(O)N(H)(C 2~6 Heterocycloalkyl), Heteroaryl, Heteroaryl-(C 1~4 alkyl), C(O)H, CN, pyrrolidinonyl, C 1~4 Alkyl, C 1~4 Haloalkyl, C(O)O(C 1~4 alkyl), C(O)NH2, C(O)N(H)C(O)H, (C 1~4 Alkyl)-OH, C 2~6 Heterocycloalkyl-C(O)H, or O-(C 1~4 alkyl); or R 4 and R 5are combined to form CO, C 3~7 Cycloalkyl, C 2~6 Heterocycloalkyl, pyrrolidinonyl, pyrrolidinonyl-(C 1~4 alkyl), or imidazolidinonyl-OH; R 6 are H, CN, C(O)H, and C 1~4 Alkyl, heteroaryl, O-(C 1~4 alkyl), O-(C 1~4 alkyl)-OH, N(H)-(C 1~4 alkyl), or N(H)-(C 1~4 alkyl)-OH.

[0026] In some embodiments of the formulas provided herein, R 4 , H, C 1~4 Alkyl, or (C 1~4 alkyl)-OH; or R 3 and R 4 are combined to form C 2~6 forming a heterocycloalkyl; R 5 is C(O)N(H)(C 1~4 Alkyl), Heteroaryl, Heteroaryl-(C 1~4 alkyl), C(O)H, CN, pyrrolidinonyl, C 1~4 Alkyl, C 1~4 Haloalkyl, C(O)O(C 1~4 alkyl), C(O)NH2, C(O)N(H)C(O)H, (C 1~4 alkyl)-OH, or O-(C 1~4 alkyl); or R 4 and R 5 are combined to form CO, C 3~7 Cycloalkyl, pyrrolidinonyl, pyrrolidinonyl-(C 1~4 alkyl), or imidazolidinonyl-OH.

[0027] In some embodiments, the compound has the formula: [ka] or a pharma- ceutically acceptable salt thereof is provided herein. [In the formula, R 1 is C 1~4 is alkyl; R 2 is C 1~4 is alkylene; R 7 teeth, [ka] [ka] [is].

[0028] In some embodiments of the formulae provided herein, R 1 is methyl or ethyl.

[0029] In some embodiments of the formulae provided herein, R 2 is methylene.

[0030] In some embodiments of the formulas provided herein, R 3 is N, O, S, NH, CH, or N-(C 1~4 alkyl); R 4 , H, C 1~4 Alkyl, or (C 1~4 alkyl)-OH; R 5 is C(O)N(H)(C 1~4 Alkyl), Heteroaryl, Heteroaryl-(C 1~4 alkyl), C(O)H, CN, pyrrolidinonyl, C 1~4 Alkyl, C 1~4 Haloalkyl, C(O)O(C 1~4 alkyl), C(O)NH2, C(O)N(H)C(O)H, (C 1~4 alkyl)-OH, or O-(C 1~4alkyl).

[0031] In some embodiments of the formulae provided herein, R 3 is N, NH, or O. In some embodiments, R 3 is O or S.

[0032] In some embodiments, [ka] is a single bond, and R 3 is N, O, or S.

[0033] In some embodiments, [ka] is a double bond, and R 3 is NH or CH.

[0034] In some embodiments, R 4 is H or C 1~4 In some embodiments, R 4 is H, methyl, or ethyl.

[0035] In some embodiments, R 3 and R 4 are combined to form C 2~6 In some embodiments, R 3 and R 4 are combined to form a C having 1, 2 or 3 heteroatoms independently selected from N, O, or S. 2~6 In some embodiments, R 3 and R 4 are combined to form a C having 1 or 2 heteroatoms independently selected from N or O 2~5 It forms a heterocycloalkyl.

[0036] In some embodiments, R 4 and R5 are combined to form CO, C 3~7 Cycloalkyl, pyrrolidinonyl, pyrrolidinonyl-(C 1~4 alkyl), or imidazolidinonyl-OH. In some embodiments, R 4 and R 5 are combined to form CO, C 3~4 Cycloalkyl, pyrrolidinonyl, pyrrolidinonyl-(C 1~2 alkyl), or imidazolidinonyl-OH.

[0037] In some embodiments, R 7 teeth, [ka] It is.

[0038] In some embodiments, R 7 teeth, [ka] It is.

[0039] In some embodiments, R 7 teeth, [ka] It is.

[0040] In some embodiments, R 7 teeth, [ka] It is.

[0041] In some embodiments, R 7 teeth, [ka] [ka] It is.

[0042] In some embodiments, R 7 teeth, [ka] It is.

[0043] In some embodiments, R 1 is ethyl, and R 2 is methylene.

[0044] In some embodiments of the formulas provided herein, C 1~4 Alkyl refers to methyl, ethyl, propyl, isopropyl, cyclopropyl, butyl, isobutyl, or cyclobutyl.

[0045] In some embodiments, the compound provided herein is a compound selected from Table 1, or a pharma- ceutically acceptable salt thereof. [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5] [Table 1-6]

[0046] In some embodiments, C 1~4 Alkylene refers to methylene, ethylene, propylene, isopropylene, butylene, or isobutylene.

[0047] In some embodiments, halo refers to one or more of fluorine, chlorine, bromine, or iodine. 1~4 Haloalkyl is C 1~4 In some embodiments, C refers to mono-, di-, or tri-haloalkyl. 1~4 Haloalkyl is C 1~4 Fluoroalkyl or C 1~4 In some embodiments, C 1~4 Fluoroalkyl is C 1~4 Monofluoroalkyl, C 1~4 Difluoroalkyl, or C 1~4 Refers to trifluoroalkyl.

[0048] In some embodiments, heteroaryl is a C aryl group having 1, 2, 3, 4, 5, or 6 heteroatoms independently selected from N, O, or S. 3~15 Heteroaryl (i.e., C 3~10 or C 3~9 ) groups. In some embodiments, heteroaryl is monocyclic. In some embodiments, heteroaryl is bicyclic or tricyclic, and the cyclic rings are fused or linked by bonds. In some embodiments, heteroaryl refers to furanyl, pyridinyl, pyrimidinyl, pyrazinyl, triazinyl, pyrrolyl, pyrazolyl, or imidazolyl.

[0049] In some embodiments, the compounds provided herein are selected from the compounds shown in Table 1, or a pharma- ceutically acceptable salt thereof.

[0050] In some embodiments, the present application provides a composition comprising one or more of the compounds provided herein.In some embodiments, the composition is a pharmaceutical composition that further comprises a pharma- ceutically acceptable carrier.

[0051] In some embodiments, the compounds or compositions provided herein can be contained in a container, and optionally the container reduces or blocks the transmission of visible or ultraviolet light through the container. Compounds or compositions contained in such containers can decompose at a slower rate than if they were contained in a container that is transparent to visible or ultraviolet light.

[0052] The compounds described herein also include isotopically labeled compounds in which one or more atoms are replaced by an atom having the same atomic number but an atomic mass or mass number different from the atomic mass or mass number predominantly found in nature. Examples of isotopes suitable for inclusion in the compounds described herein include: 2 H, 3 H, 11 C. 13 C. 14 C. 36 Cl, 18 F, 123 I, 125 I, 13 N, 15 N, 15 O. 17 O. 18 O. 32 P, and 35 In some embodiments, isotope-labeled compounds are useful in drug or substrate tissue distribution studies. In another embodiment, substitution with heavy isotopes such as deuterium may result in greater metabolic stability (e.g., increased in vivo half-life or reduced dosage requirements). In yet another embodiment, 11 C. 18 F, 15 O and 13 Substitution with positron emitting isotopes, such as N, is useful in Positron Emission Topography (PET) studies for examining substrate receptor occupancy. Isotopically labeled compounds are prepared by any suitable method or process that employs an appropriately isotopically labeled reagent in place of a non-labeled reagent otherwise used.

[0053] In some embodiments, the compounds described herein are labeled by other means, including but not limited to, the use of chromophores or fluorescent moieties, bioluminescent labels, or chemiluminescent labels.

[0054] The compounds described herein, and other related compounds having different substituents, can be prepared using the techniques and materials described herein and as described in, for example, Fieser and Fieser's Reagents for Organic Synthesis, Volumes 1-17 (John Wiley and Sons, 1991); Rodd's Chemistry of Carbon Compounds, Volumes 1-5 and Supplementals (Elsevier Science Publishers, 1989); Organic Reactions, Volumes 1-40 (John Wiley and Sons, 1991), Larock's Comprehensive Organic Transformations (VCH Publishers Inc., 1989), March, Advanced Organic Chemistry 4 th Ed., (Wiley 1992); Carey and Sundberg, Advanced Organic Chemistry 4 th Ed., Vols. A and B (Plenum 2000, 2001), and Green and Wuts, Protective Groups in Organic Synthesis 3 rd Ed., (Wiley 1999), all of which are incorporated by reference for such disclosures. The general methods for preparing the compounds as described herein are modified by the use of appropriate reagents and conditions to introduce the various moieties found in the formulas as provided herein.

[0055] The compounds described herein are synthesized using any suitable procedure, starting from compounds available from commercial sources or prepared using the procedures described herein.

[0056] In some embodiments, the compounds described herein can be prepared by synthetic methods, including any of the synthetic schemes depicted in the Examples or FIG.

[0057] In some embodiments, reactive functional groups such as hydroxyl, amino, imino, thio, or carboxy groups are protected to avoid their undesired participation in reactions. Protecting groups are used to block some or all of the reactive moieties to prevent such groups from participating in chemical reactions until the protecting group is removed. In another embodiment, each protecting group is separable by different means. Protecting groups that are cleaved under completely disparate reaction conditions meet the requirement of differential separation.

[0058] In some embodiments, the protecting groups are removed by acid, base, reducing conditions (e.g., by hydrogenolysis), or oxidative conditions. Groups such as trityl, dimethoxytrityl, acetal, and t-butyldimethylsilyl are acid labile and are used in the presence of amino groups protected with Cbz groups, which are removable by hydrogenolysis, and Fmoc groups, which are base labile, to protect carboxy and hydroxy reactive moieties. Carboxylic acid and hydroxy reactive moieties are blocked with base labile groups, such as, but not limited to, methyl, ethyl, and acetyl, in the presence of amines blocked with acid labile groups, such as t-butyl carbamate, or carbamates, which are stable to both acid and base, but removable by hydrolysis. method

[0059] In some embodiments, provided herein is a method of inhibiting fat mass obesity related protein (FTO) activity in a cell, comprising contacting the cell with an effective amount of entacapone or a pharma- ceutical acceptable salt thereof.

[0060] In some embodiments, provided herein is a method of inhibiting FTO activity in a cell, comprising contacting the cell with an effective amount of a compound provided herein.

[0061] In some embodiments, provided herein is a method of inhibiting FTO activity in a subject in need thereof, comprising administering to the subject an effective amount of entacapone or a pharma- ceutically acceptable salt thereof.

[0062] In some embodiments, provided herein is a method of inhibiting FTO activity in a subject in need thereof, comprising administering to the subject an effective amount of a compound provided herein.

[0063] In some embodiments, provided herein is a method of inhibiting Notch1 activity in a cell, comprising contacting said cell with an effective amount of entacapone or a pharma- ceutically acceptable salt thereof.

[0064] In some embodiments, provided herein is a method of inhibiting Notch1 activity in a cell, comprising contacting the cell with an effective amount of a compound provided herein.

[0065] In some embodiments, provided herein is a method of inhibiting Notch1 activity in a subject in need thereof, comprising administering to the subject an effective amount of entacapone or a pharma- ceutically acceptable salt thereof.

[0066] In some embodiments, provided herein is a method for inhibiting Notch1 activity in a subject in need thereof, comprising administering to the subject an effective amount of a compound provided herein.

[0067] In some embodiments, provided herein is a method of treating cancer in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of entacapone or a pharma- ceutically acceptable salt thereof.

[0068] In some embodiments, provided herein is a method of treating cancer in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound provided herein.

[0069] In some embodiments of the methods described herein, the cancer is glioblastoma. In some embodiments, the cancer is diffuse intrinsic pontine glioma.

[0070] In some embodiments of the methods described herein, the cancer comprises brain cancer or tumor, leukemia, breast cancer, lung cancer, colon cancer, pancreatic cancer, ovarian cancer, prostate cancer, or kidney cancer.

[0071] Provided herein are compounds useful in treating the diseases associated with the RNA demethylase activity of FTO.Thus, in some embodiments, provided herein are methods for treating systemic solid tumors, metabolic diseases, obesity, diabetes, or neurodegenerative disorders.

[0072] In some embodiments, provided herein is a method of treating an FTO-related disease in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of entacapone or a pharma- ceutically acceptable salt thereof.

[0073] In some embodiments, provided herein is a method of treating an FTO-related disease in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound provided herein.

[0074] In some embodiments, provided herein is a method of treating a Notch1-associated disease in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of entacapone or a pharma- ceutically acceptable salt thereof.

[0075] In some embodiments, provided herein is a method for treating a Notch1-associated disease in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound provided herein.

[0076] In some embodiments of these methods, the compound administered is a compound of Table 1 or a pharma- ceutically acceptable salt thereof. In some embodiments of these methods, the method includes administration or use of a compound selected from compound 31 or a pharma- ceutically acceptable salt thereof.

[0077] In some embodiments of these methods, the compound is administered or provided as a composition, hi some embodiments, the compound is administered or delivered as a pharmaceutical composition that further comprises a pharma- ceutically acceptable carrier.

[0078] In some embodiments of these methods, entacapone or a pharma- ceutically acceptable salt thereof is administered or provided as a composition, hi some embodiments, entacapone or a pharma- ceutically acceptable salt thereof is administered or provided as a pharmaceutical composition further comprising a pharma- ceutically acceptable carrier.

[0079] In some embodiments of these methods, the FTO-related disease is obesity, diabetes, Alzheimer's disease, or cancer.

[0080] In some embodiments of the methods provided herein, the cancer is brain cancer. In some embodiments, the cancer is glioblastoma. In some embodiments, the cancer is breast cancer, leukemia, brain cancer, or lung cancer. In some embodiments, the breast cancer is triple-negative breast cancer. In some embodiments, the lung cancer is non-small cell lung cancer.

[0081] In some embodiments, the cell is a brain cell. In some embodiments, the cell is a glioma stem cell. In some embodiments, the cell contacting is in a subject. In some embodiments, the cell contacting is in vitro. In some embodiments, the cell is a cell that began as a hematopoietic stem cell (e.g., a blood cell), a breast cell, a lung cell, a colon cell, a pancreatic cell, an ovarian cell, a prostate cell, or a kidney cell. In some embodiments, the blood cell is a pluripotent stem cell. In some embodiments, the pluripotent stem cell is a lymphoid progenitor cell or a myeloid progenitor cell. In some embodiments, the lymphoid progenitor cell is a natural killer cell, a T lymphocyte, or a B lymphocyte. In some embodiments, the myeloid progenitor cell is a neutrophil, a basophil, an eosinophil, a monocyte, a platelet, or a red blood cell. In some embodiments, the monocyte is a macrophage.

[0082] The subjects considered herein are typically humans. However, the subject may be any mammal for which treatment is desired. Thus, the methods described herein can be applied to both human and veterinary applications.

[0083] In some embodiments of these methods, the administered active agent may be administered in combination with a second active agent, which may be any compound described herein, or the second active agent may be selected from any agent suitable for any of the uses described herein.

[0084] In some embodiments of these methods, the active agent is provided in a form suitable for the desired route of administration, which may vary according to the needs of the situation.For example, in some embodiments, oral administration can be utilized, and the agent can be formulated as a solid dosage form, gel, liquid, paste, spray, or another suitable form for oral administration.In some embodiments, administration is oral, pulmonary, or intravenous.In some embodiments, administration is systemic, and the compound crosses the blood-brain barrier to exert its activity.In some embodiments, administration is local injection.In some embodiments, administration is intracerebral administration.In some embodiments, administration is intracranial administration.

[0085] In some embodiments, the subject has a refractory disease, hi some embodiments, the refractory disease comprises a refractory cancer. Administration

[0086] The formulation of therapeutic compositions and their subsequent administration (dosing) are within the skill of those skilled in the art. Dosing depends on the severity and responsiveness of the condition being treated, and the course of treatment may last from several days to several months, or until sufficient reduction of the condition is achieved. From the measurement of drug accumulation in the patient's body, the optimal dosing schedule can be calculated.

[0087] Those skilled in the art can readily determine optimal dosages, dosing methods and repetition rates. Optimal dosages may vary depending on the relative potency of the individual compounds, and generally will be determined based on the EC50 concentration found to be effective in in vitro and in vivo animal models. 50The repetition rate of dosing can be estimated based on the value. In general, the dosage can range from 0.01 μg to 1 g / kg of body weight and can be administered once or multiple times a day, week, month, or year, or even once every 2 to 20 years. Those skilled in the art can easily estimate the repetition rate of dosing based on the residence time and concentration of the drug measured in bodily fluids or tissues. After successful treatment, it may be desirable to subject the patient to maintenance therapy to prevent recurrence of the condition, in which the compound is administered at a maintenance dose ranging from 0.01 μg to 1 g / kg of body weight once or multiple times a day, up to once every 20 years.

[0088] In some embodiments, the compounds described herein are administered alone, i.e., without another, distinct active agent. In some embodiments, the compounds described herein are administered in combination with another (i.e., one or more) distinct active agents.

[0089] In some embodiments of these methods, the active agent is provided in a form suitable for the desired route of administration, which may vary according to the needs of the situation.For example, in some embodiments, oral administration may be utilized, and the agent may be formulated as a solid dosage form, gel, liquid, paste, spray, or another suitable form for oral administration.In some embodiments, the administration is oral, pulmonary, or intravenous.In some embodiments, the administration is comprehensive, and the compound crosses the blood-brain barrier to exert its activity.In some embodiments, the administration is a local injection.In some embodiments, the administration is intracerebral administration.In some embodiments, the administration is intracranial administration.

[0090] In some embodiments, the compound is administered in a therapeutically effective amount or dosage. The amount of compound that corresponds to a therapeutically effective amount will vary depending on the type of disease, the stage of the disease, the age of the patient being treated, and other factors.

[0091] The amount of the compound described herein should provide effective treatment of the specific disease described herein, but the amount administered is preferably not excessively toxic to the patient (i.e., the amount is preferably within the toxicity limit as established by clinical practice guidelines).In some embodiments, the total dosage administered is limited to prevent excessive toxicity or to obtain more effective treatment of the disease described herein, or both.Typically, the amount considered herein is per day.However, half-day and 2-day or 3-day cycles are also contemplated.

[0092] Various dosing regimens can be used for treatment. In some embodiments, a daily dosage such as any of the above exemplary dosages is administered once, twice, three times, or four times a day, for 3, 4, 5, 6, 7, 8, 9, or 10 days. Depending on the stage and severity of the disease to be treated, shorter treatment periods (e.g., up to 5 days) can be used with higher dosages, or longer treatment periods (e.g., 10 days or more, or several weeks, or a month, or longer) can be used with lower dosages. In some embodiments, a daily dosage is administered once or twice a day, every other day.

[0093] The compounds described herein, or their pharmaceutically acceptable salts, can be administered in pure form or in suitable pharmaceutical compositions, via any of the accepted modes of administration or agents known in the art.The compounds can be administered, for example, orally, nasally, parenterally (intravenously, intramuscularly, or subcutaneously), topically, transdermally, intravaginally, intravesically, intracisternally, or rectally.Dosage forms can be, for example, solid, semi-solid, lyophilized powder, or liquid dosage forms, such as, for example, tablets, pills, soft elastic or hard gelatin capsules, powders, solutions, suspensions, suppositories, aerosols, or the like, in unit dosage forms suitable for easy administration of precise dosages.A particular route of administration is oral, particularly one that allows for convenient daily dosage regimens to be adjusted according to the severity of the disease to be treated.

[0094] The auxiliary and adjuvant agents may include, for example, preservatives, wetting agents, suspending agents, sweeteners, flavoring agents, perfumes, emulsifiers, and dispensing agents. Prevention of microbial action is generally achieved by various antibacterial and antifungal agents. Isotonicity agents may also be included. Prolonged absorption of injectable pharmaceutical forms can be achieved by using agents that delay absorption. The auxiliary agents may also include wetting agents, emulsifying agents, pH buffering agents, and antioxidants.

[0095] Solid dosage forms can be prepared with coatings and shells, such as enteric coatings and others known in the art. They can contain pacifying agents and can be of a composition that releases the active agent (i.e., the compounds described herein) in a delayed manner in a certain part of the intestinal tract. The compounds described herein can also be in microencapsulated form, if appropriate, with one or more of the above-mentioned carriers.

[0096] Liquid dosage forms for oral administration include pharma- ceutically acceptable emulsions, solutions, suspensions, syrups, and elixirs.Such dosage forms are prepared, for example, by dissolving, dispersing, etc., a compound described herein and an optional pharmaceutical carrier, thereby forming a solution or suspension.

[0097] In general, depending on the intended mode of administration, a pharma- ceutically acceptable composition will contain from about 1% to about 99% by weight of a compound described herein and 99% to 1% by weight of a pharma- ceutically acceptable carrier. In one example, the composition may be from about 5% to about 75% by weight of a compound described herein, with the remainder being a suitable pharmaceutical carrier.

[0098] Actual methods for preparing such dosage forms are known, or will be apparent, to those skilled in the art. See, e.g., Remington's Pharmaceutical Sciences, 18th Ed. (Mack Publishing Company, Easton, Pa., 1990). kit

[0099] In other embodiments, a kit is provided. The kit comprises a package containing the compound described herein, or a combination thereof, or a composition. In some embodiments, the package can be a box or wrapping. The packaging materials used in packaging pharmaceutical products are well known to those skilled in the art. Examples of pharmaceutical packaging materials include, but are not limited to, bottles, tubes, inhalers, pumps, bags, vials, containers, syringes, and any packaging materials suitable for the selected formulation and intended mode of administration and treatment.

[0100] The kit may include additional items in the package, such as a pipette, either attached to the outside of the package or located within the package.

[0101] The kit may further include instructions for the approved use and for administering the compounds described herein to patients. The kit may include labeling or product inserts for the compounds. The kit may include active agents in solid or liquid phase in a package. The kit may include buffers for preparing solutions for carrying out the methods described herein, and pipettes for transferring liquids from one container to another. definition

[0102] Certain terms, whether used alone or as part of a phrase or another term, are defined below.

[0103] The articles "a" and "an" refer to one or to more than one of the grammatical object of the article.

[0104] Numerical values ​​on a scale are subject to measurement errors that limit their accuracy. For this reason, all numerical values ​​provided herein should be understood to be modified by the term "about" unless otherwise indicated. Thus, the last decimal place of a numerical value provided herein indicates its precision. If no other tolerance is indicated, the maximum tolerance is ascertained by applying rounding rules to the last decimal place, or to the last significant digit if no decimals are present in the given numerical value.

[0105] The term "alkyl" refers to a branched or straight-chain saturated hydrocarbon.

[0106] As used herein, the term "remission" refers to a decrease in the severity of at least one indicator of a pathology or disease.In certain embodiments, remission includes a delay or slowing down of the progression of one or more indicators of a pathology or disease.The severity of indicators can be determined by subjective or objective measures known to those skilled in the art.

[0107] The term "aryl" refers to a carbocyclic aromatic system containing one, two, three or more rings.

[0108] "C n~m The term "alkyl" refers to a moiety containing n to m carbon atoms, where n and m are integers.

[0109] The term "cycloalkyl" refers to an alkyl moiety that includes cyclic alkyl moieties that contain one, two, three or more rings.

[0110] The terms "composition" and "pharmaceutical composition" refer to a mixture of at least one compound described herein and a pharma- ceutical acceptable carrier. Pharmaceutical compositions facilitate the administration of a compound to a patient or subject. There are multiple techniques for administering a compound, including but not limited to intravenous, oral, aerosol, parenteral, ocular, pulmonary, and topical administration.

[0111] As used herein, "effective amount" or "therapeutically effective amount" refers to the amount of a therapeutic compound, such as a compound described herein, administered to a subject as a single dose or as part of a series of doses that is effective to produce a desired therapeutic effect.In general, a therapeutically effective amount can be initially estimated in cell culture assays or in mammalian animal models, such as non-human primates, mice, rabbits, dogs, or pigs.Animal models can also be used to determine appropriate concentration ranges and routes of administration.Such information can then be used to determine the dosage and route useful for administration in non-human and human subjects.

[0112] The term "haloalkyl" refers to an alkyl moiety substituted with one or more halogens.

[0113] The terms "halogen" and "halo" refer to one or more atoms independently selected from F, Br, Cl, or I.

[0114] The term "heteroaryl" refers to an aryl moiety containing at least one ring heteroatom selected from O, S, or N, and each ring may independently contain 1, 2, 3, or 4 ring heteroatoms independently selected from O, S, or N.

[0115] The term "heterocycloalkyl" refers to an alkyl moiety that contains one, two, three, or more rings and at least one heteroatom selected from O, S, or N, and each ring may contain 1, 2, 3, or 4 ring heteroatoms independently selected from O, S, or N.

[0116] As used herein, the term "package" means any container suitable for containing a compound or composition described herein.

[0117] The term "pharmaceutical acceptable carrier" refers to a pharmaceutically acceptable substance, composition or carrier, such as a liquid filler, solid filler, stabilizer, dispersant, suspending agent, diluent, excipient, thickener, solvent, or encapsulating material, involved in carrying or transporting at least one compound described herein in or to a patient so that the compound can perform its intended function. A given carrier must be "acceptable" in the sense of being compatible with the other ingredients of a particular formulation containing the compound described herein, and not harmful to the patient. Other ingredients that may be included in the pharmaceutical compositions described herein are known in the art and are described, for example, in "Remington's Pharmaceutical Sciences" (Genaro (Ed.), Mack Publishing Co., 1985), the entire contents of which are incorporated herein by reference.

[0118] As used herein, the term "pharmaceutical acceptable salt" refers to the derivative of the compounds described herein, in which the parent compound is modified by converting an existing acid or base moiety into its salt form. Lists of suitable salts can be found in Remington's Pharmaceutical Sciences, 17th ed., Mack Publishing Company, Easton, Pa., 1985, p. 1418 and Journal of Pharmaceutical Science, 66, 2 (1977), each of which is incorporated herein by reference in its entirety.

[0119] The term "refractory disease" refers to a disease that continues to progress during treatment with a pharmaceutical ingredient other than the compound provided herein, that responds partially to other treatments, or that responds only temporarily to other treatments. This term can apply to each of the diseases mentioned herein.

[0120] As used herein, the term "treatment" refers to the application of one or more specific procedures used for the amelioration of disease. In certain embodiments, the specific procedure is the administration of one or more pharmaceutical agents. "Treatment" of an individual (e.g., a mammal, such as a human, a domestic pet, or a livestock animal) or cell is any kind of intervention used in an attempt to change the natural course of the individual or cell. Treatment includes, but is not limited to, the administration of a pharmaceutical composition, and may be performed prophylactically or after the onset of a pathological event or contact with a pathogenic agent. Treatment includes any desired effect on the symptoms or pathology of a disease or condition, and may include, for example, a minor change or improvement in one or more measurable markers of the disease or condition being treated. Also included are "prophylactic" treatments, which may be directed to slowing down the rate of progression of the disease or condition being treated, delaying the onset of the disease or condition, or reducing the severity of its onset. EXAMPLES

[0121] The following examples further illustrate the aspects of the compounds and methods provided herein.However, these examples are in no way intended to be limiting of the teachings or disclosures described herein.These examples are provided for illustrative purposes.Unless otherwise specified, all starting materials for the synthesis preparation procedures provided herein are obtained from commercial suppliers and used without purification. Example 1 High FTO expression correlates with an aggressive GSC phenotype.

[0122] To investigate whether fat mass and obesity associated protein (FTO) expression correlates with glioma stem cell (GSC) phenotype, raw transcriptome data from 44 GSCs were clustered with high- and low-expression of FTO using a median expression cutoff. Differential expression analysis between high and low FTO groups was performed using DEseq with an FC of 1.5 and p<0.05. Differential analysis was performed on the top and bottom 25% FTO expression samples. Eleven high-expressing FTO samples were compared with eleven low-expressing samples. KEGG pathway enrichment was performed on upregulated genes in the high FTO group, and significant functional categories found were: FoxO signaling; stem cell pluripotency regulation; sphingolipid metabolism; ether lipid metabolism; and phospholipase D signaling metabolism.

[0123] Next, gene set enrichment analysis was performed on 22 high and 22 low FTO expressing GSCs. Hallmark gene set signatures were used to evaluate differences in high vs low FTO GSC groups. The top 10 upregulated gene sets are listed in Table 2. [Table 2]

[0124] The epithelial-mesenchymal transition signature set was highly enriched with a nominal p-value of <0.01. These data indicated that high FTO expression correlated with an aggressive GSC phenotype. Example 2 The FTO inhibitor entacapone inhibits GSC self-renewal.

[0125] To examine whether treatment with entacapone affects the ability of GSCs to self-renew, serial dilutions of GSCs from two patients with glioblastoma were cultured for 14 days in the presence of 40 μM entacapone or DMSO (control). A 40 μM dose of entacapone corresponds to 120 mg of dry drug, which is considerably lower than the daily dose currently approved for treating Parkinson's disease (800-1600 mg). This showed that entacapone significantly reduced the tumorsphere formation frequency of GSCs from two patients with glioblastoma, as investigated by in vitro limiting dilution assay (ELDA), a widely used method to determine the self-renewal capacity of stem cells. Example 3 Virtual screening of compound activity.

[0126] Molecular dynamics and binding free energy calculations were used to evaluate the propensity of selected compounds provided herein to inhibit FTO. The GBVI / WSA dG of selected compounds provided herein are listed in Table 3.

[0127] The rsynth value was also determined for selected compounds provided herein and is listed in Table 3. The rsynth value is a calculated synthetic feasibility score. The synthetic feasibility score is the fraction of atoms of the new structure that ultimately appear in the retrosynthetic fragments found in the starting material database. A value of 1 indicates that the molecule is highly likely to be synthetically feasible. As the rsynth value approaches 0, the likelihood of synthetic feasibility or ease of synthesis decreases. That is, as the rsynth value approaches 0, the difficulty of synthesizing the compound is predicted to increase.

[0128] MOE Dock (Molecular Operating Environment (MOE), 2018.01; Chemical Computing Group Inc., 1010 Sherbooke St. West, Suite #910, Montreal, QC, Canada, H3A 2R7. 2018.) was used for molecular docking of the protein with entacapone. The crystal structure of FTO was downloaded from RSDB (PDB CODE: 3LFM), and the 2D structure of the molecule was obtained from ChemBioDraw and converted to 3D in MOE by energy minimization. The protonation state and hydrogen coordination of the target were then optimized by QuickPrep module at a pH of 7.0 and a temperature of 300 K. Prior to docking, the implicit model of the force field and reaction field (R-field) of AMBER10:EHT was selected. The binding site of FTO was identified by reference (Han, Z.; Niu, T.; Chang, J.; Lei, X.; Zhao, M.; Wang, Q.; Cheng, W.; Wang, J.; Feng, Y.; Chai, J., Crystal structure of the FTO protein reveals basis for its substrate specificity. Nature 2010, 464 (7292), 1205-9), and the natural ligand 3DT was defined as the binding site. The docking workflow followed the "induced fit" protocol, where the side chains of the receptor pocket were allowed to move with the ligand conformation, with constraints on their positions. The weight used to anchor the side chain atoms to their native positions was 10. All docked poses of the molecule were first ranked by London dG scoring, and then force field refinement was performed on the top 30 poses, followed by GBVI / WSA dG rescoring. The best ranked pose was selected as the final binding mode.

[0129] The binding models of the compounds with the natural ligands were analyzed using the structure-based drug design module in MOE. The compounds were minimized and aligned by GBVI / WSA dG. [Table 3] Example 4 Entacapone inhibits the expression of cancer stem cell genes.

[0130] GSCs were treated with 40 μM entacapone for 48 h to assess whether entacapone affects the expression of stemness-related transcripts. A qPCR array with 84 validated cancer stem cell markers was performed (Qiagen-RT-Profiler PCR Array). It was found that treatment with entacapone resulted in significant inhibition of several cancer stem cell transcripts (Figure 1). [ka] Example 5 Live imaging of GSC invasion.

[0131] GSCs in Matrigel 3D matrix were treated with 40 μM entacapone or DMSO (control) and live imaging was performed at regular time intervals using an Incucyte live cell imaging system. The normalized invasion area was quantified after normalizing the invasion area with the area occupied by the GSC sphere. Figure 2 shows the normalized GSC invasion over 66 hours of live imaging after entacapone treatment. These data indicate that entacapone inhibits the invasion of GSCs. Example 6 Assessment of gene expression correlated with FTO expression.

[0132] A set of stem-related genes was collected from the cancer stemness panel. These genes were evaluated for correlation to FTO expression using the TCGA HG-UG133A Affymetrix platform. A significant correlation between FTO and Notch1 expression in GSCs was observed (R=0.53, p-val=1.13e-39). Example 7 Notch1 inhibition assay.

[0133] To determine whether inhibition of FTO with entacapone leads to inhibition of Notch1 protein expression in GSCs, GSCs from two patients with glioblastoma (two different GSC lines) were treated with 40 μM entacapone for 48 hours and Western blots for Notch1 were performed (Figure 3). These data indicate that entacapone induces inhibition of Notch1 protein expression. Example 8 Synthetic preparation of common core intermediates.

[0134] The compounds provided herein comprise a common core moiety. [ka] Includes.

[0135] The synthetic preparation of the compounds provided herein involves the use of a common core intermediate, (E)-2-cyano-3-(3,4-dihydroxy-5-nitrophenyl)acrylic acid (CORE). The synthetic scheme for CORE is shown in Figure 4 and described herein.

[0136] A solution of 3,4-dihydroxy-5-nitrobenzaldehyde (10.0 g, 54.6 mmol), 2-cyanoacetic acid (9.4 g, 109.2 mmol) and piperidine (3 mL) in pyridine (50 mL) was heated at 90° C. overnight. The mixture was concentrated to dryness and the residue was washed with acetone (50 mL) and EtOAc (30 mL×2) to give (E)-2-cyano-3-(3,4-dihydroxy-5-nitrophenyl)acrylic acid (5.5 g, 40%) as a yellow solid. The compound was analyzed by LC / MS and NMR (Hz) NMR (Hz) δ 1.0-1.5 Hz, 1.0 Hz, ... 1 It was characterized by 1 H NMR spectroscopy. Example 9 Synthetic preparation of compound 1.

[0137] Compound 1 is prepared according to the synthetic scheme shown in FIG. 5 as follows.

[0138] Synthesis of compound 1-2: To a solution of compound 1-1 (846 mg, 4.52 mmol) in MeOH (20 mL), TEA (457 mg, 4.52 mmol) was added and the mixture was stirred at room temperature for 20 min. Benzyl ethyl (2-oxoethyl) carbamate (1.00 g, 4.52 mmol) was added and stirring was continued for 3 h. Sodium triacetoxyborohydride (3.84 g, 18.2 mmol) was added and stirring was continued at room temperature for 16 h. The mixture was poured into cold water (50 mL), extracted with EtOAc (30 mL×3), and the combined organic phase was washed with water (100 mL), brine (100 mL), dried over Na2SO4, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (DCM / MeOH=50 / 1, v / v) to give compound 1-2 (1.20 g, 52%) as a colorless oil. Compounds were analyzed by LC / MS and 1 It was characterized by 1 H NMR spectroscopy.

[0139] Synthesis of compound 1-3: To a solution of compound 1-2 (500 mg, 1.71 mmol) in ethanol (5 mL), 10% Pd / C (200 mg) was added, and the mixture was heated at 60° C. under H2 atmosphere for 16 h. The mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography (DCM / MeOH=20 / 1, v / v) to give compound 1-3 (200 mg, 74%) as a colorless oil. The compound was analyzed by LC / MS and 1 It was characterized by 1 H NMR spectroscopy.

[0140] Synthesis of compound 1: To a solution of (E)-2-cyano-3-(3,4-dihydroxy-5-nitrophenyl)acrylic acid (316 mg, 1.26 mmol) and DIPEA (489 mg, 3.78 mmol) in anhydrous DMF (5 mL) at 0° C., HOBT (204 mg, 1.51 mmol) and EDCI.HCl (363 mg, 1.89 mmol) were added and the mixture was stirred for 30 min. Then, a solution of compound 1-3 (200 mg, 1.26 mmol) in anhydrous DMF (2 mL) was added and stirring was continued at room temperature for 16 h. The mixture was directly purified by C18, RP column (Biotage, ACN / HO=20%, buffered with 0.1% HCOOH) followed by preparative HPLC (Agilent 10 prep-C18, 10 μm, 250×21.2 mm column, eluted with a gradient of MeCN in water with 0.1% formic acid at a flow rate of 20 mL / min) to give compound 1 (52.0 mg, 11%) as a red solid. The compound was identified by LC / MS and 1 It was characterized by 1 H NMR spectroscopy. Example 10 Synthetic preparation of compound 20.

[0141] Compound 20 is prepared according to the synthetic scheme shown in FIG.

[0142] Synthesis of 20-3: To a suspension of NaH (60% suspension in oil, 438 mg, 18.3 mmol) in THF (15 mL) at 0° C., a solution of compound 20-1 (1.50 g, 12.2 mmol) in THF (2 mL) was added dropwise, and the mixture was heated at 70° C. for 1 h. The mixture was cooled to room temperature, compound 20-2 (2.61 g, 13.4 mmol) was added, and stirring was continued for 16 h at room temperature. The mixture was poured into cold water (50 mL), extracted with EtOAc (30 mL×2), and the combined organic extracts were washed with water (50 mL), brine (100 mL), dried over Na2SO4, and concentrated to dryness. The residue was purified by silica gel chromatography (petroleum ether / EtOAc=30 / 1, v / v) to give compound 20-3 (2.50 g, 87%) as a colorless oil. The compound was characterized by LC / MS and HPLC. 1 It was characterized by 1 H NMR spectroscopy.

[0143] Synthesis of 20-4: To a solution of compound 20-3 (2.50 g, 10.5 mmol) in DCM (5 mL), TFA (3.60 g, 31.6 mmol) was added dropwise and the mixture was heated for 3 h at 40° C. The mixture was concentrated to dryness to give compound 20-4 (1.80 mg, 88%) as a yellow oil, which was used as such in the next step.

[0144] Synthesis of 20-5: To a solution of compound 20-4 (1.50 g, 8.28 mmol) in MeOH (5 mL) at 0° C., oxalyl chloride (1.4 mL, 16.6 mmol) was added dropwise and the mixture was stirred for 2 h at room temperature. The mixture was concentrated to dryness to give compound 20-5 (1.30 mg, 80%) as a yellow oil, which was used as such in the next step.

[0145] Synthesis of 20-6: To a solution of compound 20-5 (1.30 g, 6.66 mmol) in THF (2 mL), a 2 M solution of ethylamine in THF (35 mL) was added, and the mixture was heated at 80° C. for 16 h in a sealed tube. The mixture was concentrated to dryness, and the residue was purified by silica gel chromatography (petroleum ether / EtOAc=20 / 1, v / v) to give compound 20-6 (1.10 g, 79%) as a yellow oil. The compound was analyzed by LC / MS and HPLC. 1 It was characterized by 1 H NMR spectroscopy.

[0146] Synthesis of 20-7: To a solution of compound 20-6 (500 mg, 2.40 mmol) in anhydrous THF (5 mL) at -20 °C, borane-tetrahydrofuran complex (12.0 mL, 24.0 mmol) was added slowly via syringe, and the mixture was allowed to warm to room temperature and stirred overnight. The reaction was carefully quenched by the addition of methanol (2 mL) and 1M aqueous HCl (10 mL), and the mixture was stirred at room temperature for 2 h, then concentrated to dryness. The residue was diluted with saturated aqueous Na2CO3 (10 mL), extracted with EtOAc (30 mL x 3), and the combined organic layers were dried over Na2SO4 and concentrated to dryness to give compound 20-7 (348 mg, 75%) as a colorless oil. The compound was analyzed by LC / MS and HPLC. 1It was characterized by 1 H NMR spectroscopy.

[0147] Synthesis of compound 20: To a solution of (E)-2-cyano-3-(3,4-dihydroxy-5-nitrophenyl)acrylic acid (322 mg, 1.29 mmol) and DIPEA (333 mg, 2.57 mmol) in DMF (5 mL), HOBT (209 mg, 1.54 mmol) and EDCI.HCl (370 mg, 1.93 mmol) were added and the mixture was stirred for 30 min. Then, a solution of compound 20-7 (250 mg, 1.29 mmol) in DMF (1 mL) was added and stirring was continued at room temperature overnight. The mixture was diluted with water (50 mL) and extracted with EtOAc (30 mL x 2), and the combined organic layers were washed with brine (50 mL x 2), dried over Na2SO4, and concentrated under reduced pressure. The residue was purified by preparative HPLC (Agilent 10 prep-C18, 10 μm, 250×21.2 mm column, eluted with a gradient of MeOH in water containing 0.1% formic acid at a flow rate of 20 mL / min) to give compound 5 (72.9 mg, 13%) as a red solid. 1 It was characterized by 1 H NMR spectroscopy. Example 11 Synthetic preparation of compound 29.

[0148] Compound 29 is prepared according to the synthetic scheme shown in FIG.

[0149] Synthesis of compound 29-2: To a solution of compound 29-1 (5.0 g, 22.4 mmol) in DCM (15 mL) at 0° C., Dess-Martin reagent (11.4 g, 26.9 mmol) was added, and the mixture was warmed to room temperature and stirred overnight. The mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography (petroleum ether / EtOAc=5 / 1, v / v) to give compound 29-2 (3.9 g, 79%) as a light yellow oil. The compound was analyzed by LC / MS and HPLC. 1 It was characterized by 1 H NMR spectroscopy.

[0150] Synthesis of compound 29-3: To a solution of L-alaninamide hydrochloride (2.2 g, 17.62 mmol) in MeOH (10 mL), TEA (1.8 g, 17.6 mmol) was added and the mixture was stirred at room temperature for 10 min. Compound 29-2 (3.9 g, 17.6 mmol) was added and stirred for 1 h, followed by Na(CN)3BH (4.4 g, 70.5 mmol). The mixture was then stirred overnight. The reaction was quenched by the addition of water (50 mL) and the mixture was extracted with EtOAc (40 mL x 3). The combined organic layers were washed with water (100 mL x 3), brine (50 mL), dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by silica gel chromatography (DCM / MeOH = 20 / 1, v / v) to give compound 29-3 (2.4 g, 46%) as a yellow oil. Compounds were analyzed by LC / MS and 1 It was characterized by 1 H NMR spectroscopy.

[0151] Synthesis of compound 29-4: To a solution of compound 29-3 (2.4 g, 8.18 mmol) and TEA (2.48 g, 24.5 mmol) in DCM (15 mL) at 0 °C, Boc2O (3.85 g, 16.4 mmol) was added slowly, and the mixture was stirred at room temperature overnight. The mixture was diluted with water and extracted with DCM (40 mL x 3), and the combined organic layer was washed with water (50 mL x 3), brine (50 mL), dried over Na2SO4, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (DCM / MeOH = 20 / 1, v / v) to give compound 29-4 (1.9 g, 59%) as a light yellow oil. The compound was characterized by LC / MS and NMR (Hz) NMR (Hz) δ 1.0-1.5 Hz, 1.5 Hz, 1.0 ... 1 It was characterized by 1 H NMR spectroscopy.

[0152] Synthesis of compound 29-5: A mixture of compound 29-4 (1.9 g, 5.59 mmol) and 10% Pd / C (220 mg, 2.07 mmol) in MeOH was stirred at room temperature under H2 atmosphere overnight. The mixture was filtered and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel chromatography (DCM / MeOH=10 / 1, buffered with 0.1% NH3·H2O) to give compound 29-5 (350 mg, 28%) as a light yellow oil. The compound was analyzed by LC / MS and HPLC.1 It was characterized by 1 H NMR spectroscopy.

[0153] Synthesis of compound 29-6: To a solution of (E)-2-cyano-3-(3,4-dihydroxy-5-nitrophenyl)acrylic acid (337.5 mg, 1.35 mmol) and DIPEA (523.6 mg, 4.05 mmol) in DCM (10 mL) at 0 °C, HOBT (218.9 mg, 1.62 mmol) and EDCI.HCl (388.2 mg, 2.03 mmol) were added. The mixture was stirred for 30 min. A solution of compound 29-5 (350 mg, 1.35 mmol) in DCM (3 mL) was added and the mixture was warmed to room temperature and stirred overnight. The mixture was concentrated under reduced pressure, and the residue was purified by silica gel chromatography (DCM / MeOH=10 / 1, v / v) followed by preparative HPLC (Agilent 10 prep-C18, 10 μm, 250×21.2 mm column, eluted with a gradient of MeCN in water containing 0.1% formic acid at a flow rate of 20 mL / min) to give compound 29-6 (40 mg, 6%) as a red oil. The compound was identified by LC / MS and HPLC. 1 It was characterized by 1 H NMR spectroscopy.

[0154] Synthesis of compound 29: A solution of compound 29-6 (40 mg, 0.08 mmol) in HCOOH (2 mL) was stirred at room temperature for 3 h and then concentrated under reduced pressure to give compound 22 (29 mg, 60%) as a red solid. The compound was analyzed by LC / MS and 1 It was characterized by 1 H NMR spectroscopy. Example 12 Synthetic preparation of compound 31.

[0155] Compound 31 is prepared according to the synthetic scheme shown in FIG.

[0156] Synthesis of compound 31-2: To a solution of compound 31-1 (3.54 g, 20.0 mmol) in DMF (50 mL), 1-bromo-3-fluoropropane (2.82 g, 20.0 mmol) and K2CO3 (5.53 g, 40.0 mmol) were added, and the mixture was stirred at room temperature overnight. The mixture was concentrated under reduced pressure, and the residue was purified by silica gel chromatography (Pet. ether / EtOAc=10 / 1) to give compound 31-2 (2.21 g, 47%) as a colorless oil. The compound was analyzed by LC / MS and 1 It was characterized by 1 H NMR spectroscopy.

[0157] Synthesis of compound 31-3: To a solution of compound 31-2 (2.01 g, 8.50 mmol) in anhydrous DMF (15 mL) at 0° C., NaH (60% suspension in oil, 1.02 g, 25.5 mmol) was added and the mixture was stirred at 0° C. for 30 min. Then, iodoethane (2.65 g, 17.0 mmol) was added dropwise and the mixture was warmed to room temperature and stirred for 2 h. The reaction was quenched with water and the mixture was concentrated under reduced pressure. The residue was purified by silica gel chromatography (petroleum ether / EtOAc=20 / 1) to give compound 31-3 (1.28 g, 57%) as a colorless oil. The compound was analyzed by LC / MS and HPLC. 1 It was characterized by 1 H NMR spectroscopy.

[0158] Synthesis of compound 31-4: To a solution of compound 31-3 (280 mg, 1.05 mmol) in EtOAc (2 mL) was added a 4 M solution of HCl in EtOAc (2 mL) and the mixture was stirred for 1 h. The mixture was concentrated under reduced pressure to give compound 31-4 (165.3 mg, 78%), which was used as it was in the next step.

[0159] Synthesis of compound 31: To a solution of compound 31-4 (165.3 mg, 0.822 mmol) and (E)-2-cyano-3-(3,4-dihydroxy-5-nitrophenyl)acrylic acid (250.2 mg, 1.00 mmol) in DMF (5 mL), HOBT (202.8 mg, 1.50 mmol), EDCI (287.6 mg, 1.5 mmol) and DIPEA (387.6 mg, 3.0 mmol) were added and the mixture was stirred at 30° C. for 16 h. The mixture was diluted with EtOAc, washed with water, brine, dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by preparative HPLC (Agilent 10 prep-C18, 10 μm, 250×21.3 mm column, eluted with a gradient of ACN in water containing 0.1% formic acid at a flow rate of 20 mL / min) to give compound 4 (90 mg, 28%) as a yellow solid. 1 It was characterized by 1 H NMR spectroscopy. (Example 13) Synthetic preparation of compound 33.

[0160] Compound 33 is prepared according to the synthetic scheme shown in FIG.

[0161] Synthesis of 33-1: To a solution of compound 33-1 (10.0 g, 43.0 mmol) in Et2O (200 mL) at 0 °C, MeOH (cat) and LiBH4 (2 M solution in THF, 28.0 mL, 56.0 mmol) were added. The mixture was stirred at 0 °C for 1.5 h, then warmed to room temperature and stirred for an additional 1.5 h. The reaction was quenched with water (150 mL) and the mixture was extracted with EtOAc (100 mL x 3). The combined organic layers were washed with water (150 mL x 3), brine (50 mL), dried over Na2SO4, and concentrated under reduced pressure to give compound 33-2 (7.0 g, 86%) as a colorless oil. The compound was characterized by LC / MS and NMR (Hz) NMR (Hz) δ 1.0, 1.5, ... 1 It was characterized by 1 H NMR spectroscopy.

[0162] Synthesis of 33-3: To a solution of 33-2 (3.5 g, 18.4 mmol) in DCM (30 mL), TEA (5.12 mL, 36.8 mmol) and 4-nitrophenyl carbonochloridate (4.45 g, 22.1 mmol) were added, and the mixture was stirred at room temperature for 2 h. The mixture was diluted with water (50 mL), extracted with DCM (20 mL x 3), and the combined organic layer was washed with water (30 mL x 3), brine (15 mL), dried over Na2SO4, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. ether / EtOAc = 10 / 1) to give compound 33-3 (3.0 g, 46%) as a colorless oil. The compound was characterized by LC / MS and NMR (Hz) NMR (Hz): δ 1.0, 1.5, ... 1 It was characterized by 1 H NMR spectroscopy.

[0163] Synthesis of compound 33-4: To a solution of tert-butyl (2-aminoethyl) (ethyl) carbamate (528 mg, 2.81 mmol) and DIPEA (781 mg, 5.62 mmol) in DCM (10 mL) at 0 °C, 33-3 (1 g, 2.81 mmol) was added, and the mixture was stirred at room temperature for 2 h. The mixture was diluted with water (20 mL), extracted with DCM (10 mL × 3), and the combined extracts were washed with water (20 mL), brine (20 mL), dried over Na2SO4, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. ether / EtOAc = 2 / 1, v / v) to give compound 33-4 (600 mg, 53%) as a colorless oil, which was used as such in the next step.

[0164] Synthesis of compound 33-5: To a solution of compound 33-4 (300 mg, 0.74 mmol) in DCM (5 mL) at 0° C., a 4 M solution of HCl in dioxane (1 mL) was added and the mixture was stirred at room temperature for 20 h. The mixture was concentrated under reduced pressure to give compound 33-5 (150 mg, 89%). The compound was used in the next step without purification.

[0165] Synthesis of compound 33: To a solution of (E)-2-cyano-3-(3,4-dihydroxy-5-nitrophenyl)acrylic acid (240 mg, 0.96 mmol) and DIPEA (435 mg, 3.36 mmol) in DMF (5 mL) at room temperature, HOBT (260 mg, 1.92 mmol) and EDCI.HCl (276 mg, 1.44 mmol) were added and the mixture was stirred for 30 min. Compound 33-5 (200 mg, 0.88 mmol) was added and stirring was continued overnight. The mixture was diluted with water (20 mL) and extracted with DCM (10 mL x 3), and the combined organic layers were washed with water (20 mL), brine (20 mL), dried over Na2SO4, and concentrated under reduced pressure. The residue was purified by preparative HPLC (Agilent 10 prep-C18, 10 μm, 250×21.2 mm column, eluted with a gradient of MeOH in water containing 0.1% formic acid at a flow rate of 20 mL / min) to give compound 13 (75 mg, 20%) as a red oil. 1 It was characterized by 1 H NMR spectroscopy. Example 14 Synthetic preparation of compound 35.

[0166] Compound 35 is prepared according to the synthetic scheme shown in FIG.

[0167] Synthesis of 35-2: To a suspension of NaH (60% suspension in oil, 480 mg, 12.0 mmol) in DMF (20 mL) at 0 °C, a solution of compound 35-1 (1.20 g, 6.0 mmol) in DMF (5 mL) was added and the mixture was stirred for 1 h. Ethyl 2-bromoacetate (1.50 g, 9.0 mmol) was added and the mixture was warmed to room temperature and stirred for 16 h. The mixture was poured into cold water (100 mL), extracted with EtOAc (30 mL x 3), and the combined organic layers were washed with water (100 mL x 3), brine (100 mL), dried over Na2SO4, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (petroleum ether / EtOAc = 30 / 1, v / v) to give compound 35-2 (900 mg, 52%) as a colorless oil. The compound was characterized by LC / MS and HPLC. 1 It was characterized by 1 H NMR spectroscopy.

[0168] Synthesis of 35-3: A mixture of 40% aqueous ethanamine solution (5.00 g) in ethanol (5 mL) and compound 35-2 (900 mg, 3.13 mmol) was heated at 80° C. for 16 h in a 25 mL sealed tube. The mixture was concentrated under reduced pressure, and the residue was purified by silica gel chromatography (EtOAc / petroleum ether=1 / 5, v / v) to give compound 35-3 (700 mg, 78%) as a white solid. The compound was analyzed by LC / MS and NMR (Hz) NMR (Hz) δ 1.00-1.00 (m, Hz) ... 1 It was characterized by 1 H NMR spectroscopy.

[0169] Synthesis of 35-4: To a solution of compound 35-3 (700 mg, 2.45 mmol) in anhydrous THF (10 mL) at -20 °C, borane-methyl sulfide complex (1.0 mL, 10 mmol) was added slowly via syringe, and the mixture was allowed to warm to room temperature and stirred overnight. The reaction was carefully quenched by adding methanol (2 mL) and 1M aqueous HCl (10 mL), and the mixture was stirred at room temperature for 20 h, then concentrated under reduced pressure. The residue was diluted with saturated aqueous Na2CO3 (10 mL), extracted with EtOAc (20 mL x 5), and the combined organic layers were dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by silica gel chromatography (DCM / MeOH = 30 / 1, buffered with 0.5% NH3·H2O) to give compound 35-4 (320 mg, 48%) as a colorless oil. The compound was analyzed by LC / MS and HPLC. 1 It was characterized by 1 H NMR spectroscopy.

[0170] Synthesis of compound 35-5: To a solution of (E)-2-cyano-3-(3,4-dihydroxy-5-nitrophenyl)acrylic acid (240 mg, 0.96 mmol) and DIPEA (435 mg, 3.36 mmol) in DCM (20 mL) at 0 °C, HOBT (260 mg, 1.92 mmol) and EDCI·HCl (276 mg, 1.44 mmol) were added and the mixture was stirred for 30 min. A solution of compound 35-4 (260 mg, 0.95 mmol) in DCM (3 mL) was added and the mixture was warmed to room temperature and stirred for 2 h. The mixture was diluted with water (20 mL), the layers were separated, and the aqueous layer was extracted with DCM (10 mL × 3). The combined organic layers were washed with water (20 mL), brine (20 mL), dried over Na2SO4, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (DCM / MeOH=50 / 1, v / v) followed by preparative TLC (DCM / MeOH=20 / 1, v / v) to give compound 35-5 (150 mg, 31%) as a red solid. 1 It was characterized by 1 H NMR spectroscopy.

[0171] Synthesis of compound 35: To a solution of compound 35-5 (170 mg, 0.34 mmol) in DCM (2 mL) at 0 °C, a 4 M solution of HCl in dioxane (0.2 mL) was added and the mixture was stirred at room temperature for 20 h. The solvent was removed under reduced pressure and the residue was purified by C18, RP column (Biotage, ACN / H2O, 0-15%, buffered with 0.1% HCl) followed by preparative HPLC (Agilent 10 prep-C18, 10 μm, 250 × 21.2 mm column, eluted with a gradient of MeCN in water with 0.1% HCl at a flow rate of 20 mL / min) to give compound 20 (17.6 mg, 12%) as a red solid. The compound was characterized by LC / MS and NMR (Hz) NMR (Hz): δ 1.0, 1.5, ... 1 It was characterized by 1 H NMR spectroscopy. Example 15 Inhibition of the RNA demethylase activity of FTO.

[0172] To determine the inhibitory activity of compounds provided herein against FTO and compare them with entacapone, FTO activity assay was carried out. This assay measures the ability of FTO to demethylate m6A methylated RNA substrate. The assay is specific to FTO and is not affected by the function of ALKBH5, another RNA demethylase in eukaryotic cells. This experiment showed that compound 1 and compound 31 exhibit significantly higher FTO inhibitory activity compared to entacapone (see Table 4). For example, at 20 μM, entacapone exhibits less than 20% reduction in FTO activity, while compound 1 and compound 31 exhibit about 40% or more inhibition of FTO activity. Even at a double concentration of 40 μM, entacapone exhibits lower inhibition of FTO activity than 20 μM compound 31. [Table 4] Example 16 IC of compound 31 in adult glioblastoma 50 Decision.

[0173] Glioma stem cells were incubated with increasing concentrations of compound 31, and the percentage of viable cells at 48 hours was quantified using Cell Titer Glo (Promega). This experiment demonstrated an IC of 114.5 μM compound 31. 50 and an IC of 70 μM 10 In the functional assays described below, IC 10 was used. (Example 17) Inhibition of glioma stem cell invasion by compound 31.

[0174] Patient-derived glioma stem cells (GSCs) were incubated with 70 μM compound 31 or DMSO (control) for 72 hours, and GSC invasion into the surrounding extracellular matrix was quantified in real time using an Incucyte Cell Imager. As shown in Figures 11 and 12, compound 31 induced significant inhibition of GSC invasion (p<0.05). GSC invasion is one of the hallmarks of glioblastoma. (Example 18) Inhibition of the self-renewal ability of pediatric DIPG cells by compound 31.

[0175] A limiting dilution assay (LDA) was used to determine the effect of FTO inhibition with compound 31 on the self-renewal ability of diffuse intrinsic pontine glioma (DIPG) cells (a type of brain tumor found in the brainstem region known as the pons). In this assay, DIPG cells are seeded in 96-well plates at various numbers (from 2000 cells per well to 1 cell per well) and the ability of the cells to self-renew and form clonal tumorspheres is measured over a two-week period. The results (Figure 13) showed that compound 31 significantly inhibited the self-renewal ability of DIPG cells, a hallmark of this disease. (Example 19) Inhibition of pediatric DIPG cell invasion by compound 31.

[0176] Patient-derived DIPG cells were incubated with 10 μM compound 31 or DMSO (control) for 72 hours, and DIPG invasion into the surrounding extracellular matrix was quantified in real time using an Incucyte Cell Imager. As shown in Figures 14 and 15, compound 31 induces significant inhibition of DIPG invasion (p<0.05).

[0177] It should be understood that whenever values ​​and ranges are provided herein, all values ​​and ranges subsumed within those values ​​and ranges, or combinations of those values ​​and ranges, are intended to be encompassed within the scope of the aspects and embodiments provided herein. Furthermore, all values ​​that fall within those ranges, as well as the upper or lower limits of a range of values, are also contemplated by the application.

[0178] Unless otherwise defined, all technical and scientific terms used herein are given the meaning commonly known to one of ordinary skill in the art.

[0179] Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments provided herein which equivalents are intended to be encompassed by the following claims.

Claims

1. A compound having the formula: 【Chemistry 17】 or a pharma- ceutically acceptable salt thereof [In the formula, 【Chemistry 18】 is a single bond or a double bond; R １ is C １～４ is alkyl; R ２ is C １～４ is alkylene; R ３ is N, O, S, NH, CH, or N-(C １～４ alkyl); R ４ is H, C １～４ Alkyl, or (C １～４ alkyl)-OH; Or R ３ and R ４ are combined to form C ２～６ forming a heterocycloalkyl; R ５ is C(O)N(H)(C １～４ alkyl), C(O)N(H)(C ２～６ Heterocycloalkyl), heteroaryl, heteroaryl-(C １～４ alkyl), C(O)H, CN, pyrrolidinonyl, C １～４ Alkyl, C １～４ Haloalkyl, C(O)O(C １～４ alkyl), C(O)NH ２ , C(O)N(H)C(O)H, (C １～４ alkyl)-OH,C ２～６ Heterocycloalkyl-C(O)H, or O-(C １～４ alkyl); Or R ４ and R ５ are combined to form CO, C ３～７ Cycloalkyl, C ２～６ Heterocycloalkyl, pyrrolidinonyl, pyrrolidinonyl-(C １～４ alkyl), or imidazolidinonyl-OH; R ６ is H, CN, C(O)H, C １～４ Alkyl, heteroaryl, O-(C １～４ alkyl), O-(C １～４ alkyl)-OH, N(H)-(C １～４ alkyl), or N(H)-(C １～４ alkyl)-OH.

2. R １ The compound of claim 1 , wherein is methyl or ethyl.

3. R ２ The compound of claim 1 , wherein is methylene.

4. 【Chemical 19】 is a single bond, R ３ The compound of claim 1 , wherein is N, O, or S. 【Request 5】 【Chemical 20】 is a double bond, R ３ is NH or CH.

6. R ４ is H or C １～４ The compound of claim 1 , wherein the aryl group is an alkyl group.

7. 2. The compound of claim 1, wherein each instance of heteroaryl independently refers to furanyl, pyridinyl, pyrimidinyl, pyrazinyl, triazinyl, pyrrolyl, pyrazolyl, or imidazolyl.

8. R ３ and R ４ Combined, C ２～６ The compound of claim 1 forming a heterocycloalkyl.

9. R ４ and R ５ Combined, CO, C ３～７ Cycloalkyl, pyrrolidinonyl, pyrrolidinonyl-(C １～４ 2. The compound of claim 1, which is formed as follows:

10. R ３ is N, O, S, NH, CH, or N-(C １～４ alkyl); R ４ But, H, C １～４ Alkyl, or (C １～４ alkyl)-OH; R ５ But C(O)N(H)(C １～４ alkyl), heteroaryl, heteroaryl-(C １～４ alkyl), C(O)H, CN, pyrrolidinonyl, C １～４ Alkyl, C １～４ Haloalkyl, C(O)O(C １～４ alkyl), C(O)NH ２ , C(O)N(H)C(O)H, (C １～４ alkyl)-OH, or O-(C １～４ 2. The compound of claim 1, wherein

11. Below formula: 【Chemistry 21】 or a pharma- ceutically acceptable salt thereof. [In the formula, R ７ teeth, 【Chemistry 22-1】 【Chemistry 22-2】 is.] 【Request 12】 【Chemical 23-1】 【Chemistry 23-2】 【Chemistry 23-3】 2. The compound of claim 1, selected from:

13. A composition comprising a compound according to one of claims 1 to 12.

14. The composition of claim 13, which is a pharmaceutical composition further comprising a pharma- ceutically acceptable carrier.

15. A composition comprising a compound according to one of claims 1 to 12, or a pharmaceutical composition comprising a compound according to one of claims 1 to 12 and a pharma- ceutically acceptable carrier, or a composition comprising entacapone or a pharma- ceutically acceptable salt thereof, for treating cancer in a subject in need thereof.

16. The composition of claim 15 , wherein the cancer comprises a brain tumor.

17. 16. The composition of claim 15, wherein the cancer comprises glioblastoma or diffuse intrinsic pontine glioma.

18. 16. The composition of claim 15, wherein the cancer comprises brain cancer or tumor, leukemia, breast cancer, lung cancer, colon cancer, pancreatic cancer, ovarian cancer, prostate cancer, or kidney cancer.

19. A composition comprising a compound according to any one of claims 1 to 12, or a pharmaceutical composition comprising a compound according to any one of claims 1 to 12 and a pharma- ceutically acceptable carrier, or a composition comprising entacapone or a pharma- ceutically acceptable salt thereof, for use in a method for inhibiting fat mass obesity-related protein (FTO) or Notch1 activity in a cell, the method comprising contacting the cell with entacapone, or the pharma- ceutically acceptable salt, the compound, or the composition.

20. the cell is a brain cell (i.e., a glioma stem cell), a blood cell, a breast cell, a lung cell, a colon cell, a pancreatic cell, an ovarian cell, a prostate cell, or a kidney cell, and optionally the contacting is in a subject; or 20. The composition of claim 19, wherein the contacting is in vitro, and optionally the cell is a brain cell (i.e., a glioma stem cell), a blood cell, a breast cell, a lung cell, a colon cell, a pancreatic cell, an ovarian cell, a prostate cell, or a kidney cell.

21. 13. A composition of the compound according to claim 1, optionally contained within a container, the container optionally reducing or blocking the transmission of visible or ultraviolet light through the container, or optionally comprising a pharma- ceutically acceptable carrier.

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