Polysubstituted benzamide compound, and preparation method, application and pharmaceutical composition thereof

Abstract

The invention belongs to the technical field of pharmacy, and relates to a polysubstituted benzamide compound as well as a preparation method, application and a pharmaceutical composition thereof. The polysubstituted benzamide compound is shown as a formula (I), and is a PKMYT1 inhibitor. # imgabs0 #

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical technology and relates to a multi-substituted benzamide compound, a preparation method, uses thereof, and a pharmaceutical composition. Background Art

[0002] Membrane-associated tyrosine and threonine kinase (Myt1 kinase, also known as PKMYT1) is encoded by the PKMYT1 gene and belongs to the Wee kinase family. It was initially discovered in Xenopus laevis and was able to phosphorylate Thr14 and Tyr15 on Cdc2. The inhibitory phosphorylation of Cdc2 is very important for controlling cell mitosis. The entry of cells from the G2 phase into the M phase is initiated by the M-phase promoting factor (MPF), which consists of the cdc2 protein kinase and cyclin B. Cells ensure that mitosis occurs only after the early stages of the cell cycle are completed by appropriately regulating MPF. During the interphase of mitosis (G1 phase, S phase, and G2 phase), the phosphorylation of Cdc2 at Tyr15 and Thr14 inhibits its activity. When transitioning from the G2 phase to the M phase, Cdc2 is dephosphorylated at Tyr15 and Thr14, enabling MPF to phosphorylate its mitotic substrates.

[0003] Studies have shown that the premature activation of Cdc2 leads to mitotic catastrophe and cell death. The inhibition of PKMYT1 leads to the premature activation of Cdc2, which may kill rapidly proliferating tumor cells. In addition, studies have shown that tumor cells are more dependent on the G2-M checkpoint for DNA damage repair and maintaining genomic integrity, thereby maintaining the survival of tumor cells. Inhibiting PKMYT1 will force cells to enter mitosis prematurely, resulting in the death of tumor cells, thereby reducing the drug resistance of tumor cells to traditional DNA-damaging chemotherapy drugs.

[0004] Therefore, the research and development of PKMYT1 inhibitors have important significance in the targeted therapy of tumors. Summary of the Invention

[0005] The object of the present invention is to provide a new PKMYT1 inhibitor, which has strong inhibitory activity against the PKMYT1 kinase, and thus has a preventive and / or therapeutic effect on related diseases mediated by PKMYT1, especially tumors.

[0006] To solve the technical problems of the present invention, the present invention provides the following technical solutions:

[0007] The first aspect of the technical solution of the present invention is to provide a compound of formula (I) or a pharmaceutically acceptable salt thereof:

[0008]

[0009] Wherein: Ring A is selected from 6-membered aryl or heteroaryl;

[0010] Each R1 is independently selected from cyano, halogen, aldehyde group, acetyl group, C1-C3 alkyl, halo-C1-C3 alkyl, C1-C3 alkoxy;

[0011] n is an integer of 0, 1, 2, 3, 4 or 5;

[0012] R2 is selected from hydrogen, halogen, C1-C3 alkyl, halo-C1-C3 alkyl;

[0013] R3 is selected from hydrogen, halogen, C1-C3 alkyl, halo-C1-C3 alkyl;

[0014] R4 is selected from hydrogen, halogen, C1-C3 alkyl, halo-C1-C3 alkyl;

[0015] R5 is selected from hydrogen, halogen, C1-C3 alkyl, halo-C1-C3 alkyl.

[0016] Preferably, the present invention provides a compound represented by formula (I) or a pharmaceutically acceptable salt thereof: Wherein: Ring A is selected from phenyl, pyridyl, pyrimidinyl, pyridone group;

[0017] Each R1 is independently selected from fluorine, chlorine, methyl, trifluoromethyl, acetyl group;

[0018] n is an integer of 0, 1, 2, 3, 4 or 5;

[0019] R2 is selected from hydrogen, methyl;

[0020] R3 is selected from hydrogen, methyl;

[0021] R4 is selected from hydrogen;

[0022] R5 is selected from hydrogen.

[0023] Specifically, the preferred compounds according to the present invention are as follows:

[0024]

[0025]

[0026]

[0027] The second aspect of the technical solution of the present invention is to provide a preparation method of the compound, which includes the following steps:

[0028]

[0029] (1) Using compound A as a starting material, reacting with 4-pyridineboronic acid through Suzuki reaction to prepare compound B;

[0030] (2) Compound B is hydrolyzed with concentrated sulfuric acid to prepare Compound C;

[0031] (3) Compound C is prepared by bromination reaction to obtain Compound D;

[0032] (4) Compound D is prepared by Suzuki reaction to obtain Compound E (R6 is methoxy) or the compound shown in formula (I) (R6 is hydroxyl) or a pharmaceutically acceptable salt thereof;

[0033] (5) Compound E reacts with aluminum trichloride to remove the methyl group to prepare the compound shown in formula (I) or a pharmaceutically acceptable salt thereof;

[0034]

[0035] (6) Using Compound F as the starting material, it is hydrolyzed with concentrated sulfuric acid to prepare Compound G;

[0036] (7) Compound G and bis(pinacolato)diboron are subjected to Miyaura borylation reaction to prepare Compound H;

[0037] (8) Compound H and 3-bromo-2,4-dimethylphenol are subjected to Suzuki reaction to prepare Compound I;

[0038] (9) Compound I is prepared by bromination reaction to obtain Compound J;

[0039] (10) Compound J and a substituted A-ring boronic acid pinacol ester or boric acid are subjected to Suzuki reaction to prepare the compound shown in formula (I) or a pharmaceutically acceptable salt thereof.

[0040] The third aspect of the technical solution of the present invention is to provide a pharmaceutical composition, which comprises the compound or a pharmaceutically acceptable salt thereof, and optionally a pharmaceutically acceptable carrier and / or excipient; preferably, the pharmaceutical composition further comprises one or more drug active ingredients for preventing and / or treating tumors other than the compound or a pharmaceutically acceptable salt thereof.

[0041] In another aspect, the present invention also provides a pharmaceutical preparation, which comprises at least one of the compounds or a pharmaceutically acceptable salt thereof, and optionally a pharmaceutically acceptable carrier, diluent or excipient; preferably, the pharmaceutical preparation is selected from the following pharmaceutical dosage forms: parenteral administration preparations, such as injection solutions or suspensions; enteral administration preparations, such as oral preparations, such as tablets or capsules; topical administration preparations, such as lotions, gels, ointments, emulsions, nasal administration preparations, suppositories, transdermal administration preparations or ophthalmic preparations.

[0042] On the other hand, the present invention also provides the use of the compound or its pharmaceutically acceptable salt, or the pharmaceutical composition in the preparation of a drug for preventing and / or treating tumors. In other words, the present invention provides a method for preventing and / or treating tumors, which method comprises administering to a subject in need thereof a prophylactically and / or therapeutically effective amount of the compound or its pharmaceutically acceptable salt, or the pharmaceutical composition.

[0043] Some of the terms used in the present invention are defined as follows, and other undefined terms have the meanings well known to those skilled in the art.

[0044] "Halogen" refers to fluorine, chlorine, bromine or iodine.

[0045] "Halogenated" means that any hydrogen in the substituent can be replaced by one or more identical or different halogens. "Halogen" is defined as above.

[0046] When any variable (e.g., R a , R b ) appears more than once in the composition or structure of a compound, its definition in each case is independent. For example, if a group is substituted by two R b , then each R b has an independent option.

[0047] "C m -C n " means having an integer number of carbon atoms in the range of m - n. For example, "C1 - C3" means that the group can have 1 carbon atom, 2 carbon atoms, or 3 carbon atoms.

[0048] "Alkyl" refers to a hydrocarbon group of the general formula C n H 2n+1 . The alkyl can be straight-chain or branched-chain. For example, the term "C1 - C3 alkyl" should be understood to mean a straight-chain or branched-chain saturated monovalent hydrocarbon group having 1, 2, or 3 carbon atoms. Examples of such groups include, but are not limited to: methyl, ethyl, propyl, isopropyl.

[0049] "C 1-3 alkyl" means an alkyl containing 1 to 3 carbon atoms. Examples of such groups include, but are not limited to: methyl, ethyl, propyl, isopropyl.

[0050] "C 1-3 alkoxy" means -O-alkyl, wherein the alkyl contains 1 to 3 carbon atoms and is straight-chain, branched-chain or cyclic. Examples of such groups include, but are not limited to: methoxy, ethoxy, n-propoxy, isopropoxy or cyclopropoxy.

[0051] Aryl refers to a monocyclic or bicyclic aromatic carbocyclic group, which usually has 6 - 10 carbon atoms; for example, phenyl or naphthyl.

[0052] Heteroaryl refers to a 5- to 10-membered aromatic heterocyclic group, including but not limited to: 5-membered heteroaryl groups: furyl, thienyl, pyrrolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, imidazolyl, pyrazolyl, triazolyl (1,2,4-triazolyl, 1,3,4-triazolyl or 1,2,3-triazolyl), thiadiazolyl (1,3,4-thiadiazolyl, 1,2,5-thiadiazolyl, 1,2,3-thiadiazolyl or 1,2,4-thiadiazolyl) and oxadiazolyl (1,3,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,2,3-oxadiazolyl or 1,2,4-oxadiazolyl); and 6-membered heteroaryl groups: pyridyl, pyrimidinyl, pyrazinyl and pyridazinyl; and bicyclic groups such as benzofuryl, benzothienyl, indazolyl, purinyl, quinolinyl, isoquinolinyl, phthalazinyl, naphthyridinyl, quinoxalinyl (chinocalinyl), quinazolinyl, cinnolinyl, pteridinyl, indolizinyl, indolyl, isoindolyl.

[0053] The term "inhibitor" refers to a compound or agent that can inhibit the biological function of a target protein or polypeptide, such as inhibiting the activity or expression of a target protein or polypeptide.

[0054] The term "tumor" includes but is not limited to solid tumors and hematological tumors, such as glioma, thyroid cancer, breast cancer, small cell lung cancer, non-small cell carcinoma, gastric cancer, colon cancer, gastrointestinal stromal tumor, pancreatic cancer, cholangiocarcinoma, ovarian cancer, endometrial cancer, prostate cancer, renal cancer, anaplastic large cell lymphoma, leukemia, multiple myeloma, mesothelioma and melanoma and combinations thereof, etc.

[0055] The term "effective amount" refers to the amount of the compounds or pharmaceutical compositions described herein that is sufficient to achieve the intended applications described below, and the intended applications include but are not limited to the treatment of diseases. The effective amount may vary according to the following: the intended application (in vivo or in vitro); or the individual and disease conditions being treated, such as the weight and age of the individual, the severity of the disease; the mode of administration, etc. The effective amount can be readily determined by a person of ordinary skill in the art.

[0056] "Optionally" means that the subsequent described event or circumstance may but does not have to occur, and this description includes the cases where the event or circumstance occurs or does not occur. For example, "alkyl optionally substituted by halogen" means that halogen may but does not have to be present, and this description includes the case where the alkyl is substituted by halogen and the case where the alkyl is not substituted by halogen.

[0057] The compounds of the present invention also include their isotopically labeled compounds. The term "isotopically labeled compound" refers to a compound in which one or more atoms are replaced by atoms having the same atomic number but an atomic mass number different from the atomic mass number typically found in nature. Examples of isotopes suitable for the present invention include, but are not limited to: isotopes of hydrogen 2 H and 3 H; isotopes of carbon 11 C, 13 C and 14 C; isotopes of chlorine 36 Cl; isotopes of fluorine 18 F; isotopes of iodine 123 I and 125 I; isotopes of nitrogen 13 N and 15 N; isotopes of oxygen 15 O, 17 O and 18 O; isotopes of phosphorus 32 P and isotopes of sulfur 35 S.

[0058] Also included within the scope of the present invention are various solvates and hydrates of the compounds or salts thereof of the present invention, as well as their polymorphs.

[0059] The term "solvate" refers to a compound that further includes a solvent, either stoichiometric or non-stoichiometric, bound by non-covalent intermolecular forces. The solvate may be the disclosed compound or a pharmaceutically acceptable salt thereof. When the solvent is water, the solvate is a "hydrate". Pharmaceutically acceptable solvates and hydrates are complexes that may include, for example, from 1 to about 100, or from 1 to about 10, or from 1 to about 2, about 3 or about 4 solvent or water molecules. The term "polymorph" refers to a compound that exists in two or more different crystalline forms.

[0060] Prodrugs of the compounds described in the present invention are also included within the scope of the present invention. Certain derivatives of the compounds described in the present invention have weak or no pharmacological activity by themselves, but when these derivatives are administered in vivo or to the body, they can be converted into the compounds described in the present invention with pharmacological activity, for example, by means of hydrolysis cleavage, etc. These derivatives are called "prodrugs". Further information on the use of prodrugs can be found in Pro-drugs as Novel Delivery Systems, Vol. 14, ACS Symposium Series (T. Higuchi and W. Stella) and Bioreversible Carriers in Drug Design, Pergamon Press, 1987 (ed. E. B. Roche, American Pharmaceutical Association).

[0061] The compounds described in the present invention include pharmaceutically acceptable salts thereof. The term "pharmaceutically acceptable salt" refers to a salt that is pharmaceutically acceptable and has the pharmacological activity required of the parent compound. Pharmaceutically acceptable salts are described in detail by Berge et al. in J. Pharma. Sci., 1977, 66, 1-19, which is incorporated herein by reference. The compounds described in the present invention may contain sufficient acidic groups, sufficient basic groups, or both types of functional groups, and accordingly react with some inorganic or organic bases, or inorganic and organic acids to form pharmaceutically acceptable salts. Examples of pharmaceutically acceptable salts include sulfates, pyrosulfates, bisulfates, sulfites, bisulfites, phosphates, monohydrogen phosphates, dihydrogen phosphates, metaphosphates, pyrophosphates, hydrochlorides, hydrobromides, hydroiodides, acetates, propionates, caprates, octanoates, acrylates, formates, isobutyrates, hexanoates, heptanoates, propiolates, oxalates, malonates, succinates, suberates, sebacates, fumarates, maleates, butyne-1,4-dioates, hexyne-1,6-dioates, benzoates, chlorobenzoates, methylbenzoates, dinitrobenzoates, hydroxybenzoates, methoxybenzoates, phthalates, sulfonates, xylenesulfonates, phenylacetates, phenylpropionates, phenylbutyrates, citrates, lactates, γ-hydroxybutyrates, glycolates, tartrates, methanesulfonates, propanesulfonates, naphthalene-1-sulfonates, naphthalene-2-sulfonates, and mandelates.

[0062] When the compounds of the present invention are used as drugs, they are usually administered in the form of pharmaceutical compositions. Therefore, pharmaceutical compositions comprising the compounds of the present invention and pharmaceutically acceptable carriers, diluents or excipients are also included in the scope of the present invention. The carriers, adjuvants, and excipients used herein include any and all solvents, diluents or other liquid excipients, dispersants or suspending agents, surfactants, isotonic agents, thickeners or emulsifiers, preservatives, solid adhesives, lubricants, etc. suitable for the desired specific dosage form. In Remington: The Science and Practice of Pharmacy, 21st edition, 2005, ed. DB Troy, Lippincott Williams & Wilkins, Philadelphia, and Encyclopedia of Pharmaceutical Technology, eds. J. Swarbrick and JC Boylan, 1988-1999, Marcel Dekker, New York, various carriers for preparing pharmaceutically acceptable compositions and known techniques for their preparation are disclosed, and their contents are all incorporated herein by reference.

[0063] The composition of the invention can be administered by any route suitable for the condition to be treated. In particular, it can be administered parenterally, for example in the form of an injectable solution or suspension; enterally, for example orally, for example in the form of tablets or capsules; topically, for example in the form of a lotion, gel, ointment or cream or in the form of a nasal or suppository. Topical application is, for example, application to the skin. Another form of topical administration is administration to the eye.

[0064] The pharmaceutical composition can be administered in solid, semisolid, liquid or gaseous form, or can be in a dried powder, such as a lyophilized form. The pharmaceutical composition can be packaged in a form that is convenient for delivery, including, for example, solid dosage forms such as capsules, sachets, cachets, gelatin, paper, tablets, suppositories, pellets, pills, lozenges and pastilles. The type of packaging will generally depend on the route of administration. Implantable sustained-release formulations are also contemplated, as well as transdermal formulations.

[0065] Some examples of materials that can serve as pharmaceutically acceptable carriers include, but are not limited to: ion exchangers, alumina, aluminum stearate, lecithin, serum proteins (such as human serum albumin), buffering substances (such as phosphates), glycine, sorbic acid or potassium sorbate, mixtures of partial glycerides of saturated vegetable fatty acids, water, salts or electrolytes (such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts), colloidal silica, magnesium trisilicate, polyvinylpyrrolidone, polyacrylates, waxes, polyethylene-polypropylene block copolymers, lanolin, sugars (such as lactose, glucose and sucrose), starches (such as corn starch and potato starch), cellulose and its derivatives, such as sodium carboxymethylcellulose, ethyl cellulose and cellulose acetate; tragacanth powder; malt; gelatin; talc; excipients, such as cocoa butter and suppository waxes; oils, such as peanut oil, cottonseed oil; safflower oil; sesame oil; olive oil; corn oil and soybean oil; diols, such as propylene glycol or polyethylene glycol; esters, such as ethyl oleate and ethyl laurate; agar; buffering agents, such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethanol; and phosphate buffer solutions, as well as other non-toxic compatible lubricants, such as sodium lauryl sulfate and magnesium stearate. Colorants, release agents, coating agents, sweetening agents, flavoring agents and fragrances, preservatives and antioxidants may also be present in the composition, according to the judgment of the formulator.

[0066] The compounds of the present invention can be used alone or in combination with other therapeutic agents for treating the diseases or disorders (such as cancer) described in the present invention. In certain embodiments, the compounds of the present invention are combined with a second compound having anti-high-proliferation properties or used for treating highly proliferative diseases (such as cancer) in a pharmaceutical combination formulation, or in an administration regimen as combination therapy. The second compound in the pharmaceutical combination formulation or the dosing regimen preferably has an activity complementary to that of the compounds of the present invention so that they do not have an adverse effect on each other. Such compounds are appropriately present in the combination in an amount effective for the intended purpose. In one embodiment, the compounds of the present invention are combined with other anti-tumor drugs.The anti-tumor drugs include: alkylating agents, including but not limited to cyclophosphamide, nitrogen mustard, melphalan, chlorambucil, carmustine; metal platinum compounds, including but not limited to carboplatin, cisplatin, oxaliplatin; topoisomerase inhibitors, including but not limited to topotecan, camptothecin, topotecan, irinotecan; antibiotics, including but not limited to annamycin, actinomycin D, daunorubicin, doxorubicin, mitoxantrone, bleomycin, plicamycin; anti-microtubule or anti-mitotic agents, including but not limited to paclitaxel, vinorelbine, docetaxel, doxorubicin; anti-metabolites, including but not limited to fluorouracil, methotrexate, cytarabine, mercaptopurine, thioguanine, and gemcitabine; antibodies, including but not limited to Herceptin, bevacizumab; hormones, including but not limited to letrozole, vorazole, tamoxifen, toremifene, fulvestrant, flutamide, nilutamide, triptorelin; kinase inhibitors, EGFR kinase inhibitors, including but not limited to gefitinib, erlotinib, lapatinib, afatinib; VEGFR inhibitors, including but not limited to sorafenib, regorafenib, sunitinib, cabozantinib, pazopanib, vandetanib, axitinib; ALK inhibitors, including but not limited to crizotinib, ceritinib, alectinib; Bcr-Abl inhibitors, including but not limited to imatinib, ponatinib, nilotinib, dasatinib; BTK inhibitors, including but not limited to ibrutinib; B-RAF inhibitors, including but not limited to vemurafenib; cyclin-dependent kinase CDK4 / 6 inhibitors, palbociclib; mTOR inhibitors, including but not limited to rapamycin, everolimus; deacetylase inhibitors, including but not limited to vorinostat; PD1 / PDL1 antibodies, Keytruda (pembrolizumab), Opdivo (nivolumab). Detailed implementation manners

[0067] The following are specific embodiments of the present invention, which further describe the technical solutions of the present invention. However, the protection scope of the present invention is not limited to these embodiments. Any changes or equivalent substitutions that do not deviate from the concept of the present invention are included in the protection scope of the present invention.

[0068] Preparation method

[0069] The compounds described in the present invention can be synthesized according to the synthetic schemes herein and / or techniques well known in the art. For example, the compounds provided by the present invention can be prepared according to the following general synthetic methods.

[0070] In a general synthetic method, the compound shown in formula (I) is prepared according to Method-1.

[0071] Method-1

[0072]

[0073] Specifically, in Method-1, the polysubstituted benzamide compounds described in the present invention can be prepared through 4 or 5 steps of reactions. Starting from compound A, reacting with 4-pyridineboronic acid through Suzuki reaction to prepare compound B; hydrolyzing compound B with concentrated sulfuric acid to prepare compound C; subjecting compound C to bromination reaction to prepare compound D; preparing compound E (R6 is methoxy) or the compound shown in formula (I) (R6 is hydroxyl) through Suzuki reaction of compound D; reacting compound E with aluminum trichloride to remove the methyl group to prepare the compound shown in formula (I).

[0074] In another general synthetic method, the compound shown in formula (I) is prepared according to Method-2.

[0075] Method-2

[0076]

[0077] Specifically, in Method-2, the polysubstituted benzamide compounds described in the present invention can be prepared through 5 steps of reactions. Starting from compound F, hydrolyzing with concentrated sulfuric acid to prepare compound G; reacting compound G with bis(pinacolato)diboron through Miyaura borylation reaction to prepare compound H; reacting compound H with 3-bromo-2,4-dimethylphenol through Suzuki reaction to prepare compound I; subjecting compound I to bromination reaction to prepare compound J; reacting compound J with substituted A-ring boronic acid pinacol ester or boric acid through Suzuki reaction to prepare the compound shown in formula (I).

[0078] The compounds described in the present invention can be synthesized according to one or more of the synthetic schemes herein and / or techniques well known in the art. Those skilled in the art will recognize that the synthetic methods of certain embodiments described in detail in the present invention can be readily adapted to synthesize other embodiments. In some embodiments, the compounds described herein can be prepared by appropriate combinations of synthetic methods well known in the art. Many starting materials and other reagents are commercially available from suppliers such as Alfa Aesar (China) Chemical Co., Ltd., or can be readily prepared using synthetic methods commonly used in the art.

[0079] 1 1H NMR spectra were recorded on an instrument operating at 400 MHz or 500 MHz. 1 1H NMR spectra were obtained in solution (reported in ppm), using CDCl3 (7.26 ppm) or DMSO-d6 (2.50 ppm) or the internal standard tetramethylsilane (0.00 ppm) as a reference standard. When reporting peak multiplicities, the following abbreviations were used: s (singlet), d (doublet), t (triplet), q (quartet), m (multiplet), br (broad), dd (doublet of doublets), dt (doublet of triplets). The coupling constants given are in Hertz (Hz).

[0080] In the following preparation methods and examples, "PE" refers to petroleum ether, "EA" refers to ethyl acetate, "MeOH" refers to methanol, "DMSO-d6" refers to deuterated dimethyl sulfoxide, "DCM" refers to dichloromethane, "DCE" refers to 1,2-dichloroethane, "NBS" refers to N-bromosuccinimide, "Pd(dppf)Cl2" refers to dichlorobis(1,1'-bis(diphenylphosphino)ferrocene)palladium(II), "Pd(dtbpf)Cl2" refers to dichlorobis(1,1'-bis(di-tert-butylphosphino)ferrocene)palladium(II), "M" refers to molarity, "rt" refers to room temperature, "h" refers to hour, "mL" refers to milliliter, "μL" refers to microliter, "mmol" refers to millimole, "μM" refers to micromole, "nM" refers to nanomole, and "°C" refers to degrees Celsius.

[0081] Preparation of Intermediate 2-Amino-3-bromo-5-(pyridin-4-yl)benzamide

[0082] Step 1: Synthesis of 2-Amino-5-(pyridin-4-yl)benzonitrile

[0083]

[0084] 2-Amino-5-bromobenzonitrile (9.85 g, 50 mmol), 4-pyridineboronic acid (8.60 g, 70 mmol), 200 mL of dioxane, 75 mL of 2 M aqueous potassium carbonate, and Pd(dppf)Cl2 (1.83 g, 2.5 mmol) were successively added to a reaction flask. The flask was purged with argon and reacted at 100 °C for 10 h. The reaction mixture was filtered through diatomaceous earth and extracted with EA (300 mL × 3). The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, concentrated, and purified by silica gel column chromatography (DCM / MeOH = 25:1, v / v) to obtain 8.08 g of a brown solid with a yield of 82.9%.

[0085] 1 H NMR (400 MHz, DMSO-d6) δ 8.57–8.51 (m, 2H), 7.95 (d, J = 2.3 Hz, 1H), 7.82 (dd, J = 8.9, 2.3 Hz, 1H), 7.68–7.62 (m, 2H), 6.90 (d, J = 8.8 Hz, 1H), 6.45 (s, 2H).

[0086] MS (ESI+) m / z: 195.98 [M+H] + .

[0087] Step 2: Synthesis of 2-Amino-5-(pyridin-4-yl)benzamide

[0088]

[0089] 2-Amino-5-(pyridin-4-yl)benzonitrile (2.02 g, 10.4 mmol) was added to 25 mL of concentrated sulfuric acid at 0 °C and reacted overnight at rt. The reaction mixture was poured into ice water, and the pH was adjusted to 7 with saturated sodium bicarbonate solution. The mixture was extracted with EA (150 mL × 3), and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to obtain 1.79 g of a yellow solid with a yield of 81.3%.

[0090] 1 H NMR (500 MHz, DMSO-d6) δ 8.55–8.50 (m, 2H), 8.05 (d, J = 2.2 Hz, 2H), 7.72–7.64 (m, 3H), 7.23 (s, 1H), 6.98 (s, 2H), 6.82 (d, J = 8.6 Hz, 1H). MS (ESI+) m / z: 213.96 [M+H] + .

[0091] Step 3: Synthesis of 2-Amino-3-bromo-5-(pyridin-4-yl)benzamide

[0092]

[0093] 2-Amino-5-(pyridin-4-yl)benzamide (535 mg, 2.5 mmol) was dissolved in 10 mL of glacial acetic acid, and NBS (463 mg, 2.6 mmol) was added portionwise. The reaction was carried out overnight at rt. The reaction solution was diluted with methanol, concentrated, and purified by silica gel column chromatography (DCM / MeOH = 10:1, v / v) to give 700 mg of a pale yellow solid in a yield of 95.6%.

[0094] 1 H NMR (400 MHz, DMSO-d6) δ 8.59–8.53 (m, 2H), 8.20 (s, 1H), 8.11–8.04 (m, 2H), 7.78–7.72 (m, 2H), 7.48 (s, 1H), 7.03 (s, 2H). MS(ESI+) m / z: 291.89, 293.86 [M+H] + .

[0095] Example 1: Synthesis of 2-Amino-3-(3-hydroxyphenyl)-5-(pyridin-4-yl)benzamide

[0096]

[0097] 2-Amino-3-bromo-5-(pyridin-4-yl)benzamide (50 mg, 0.17 mmol), 3-hydroxyphenylboronic acid (34 mg, 0.24 mmol), 2 mL of dioxane, 250 μL of 2 M aqueous potassium carbonate, and Pd(dppf)Cl2 (14 mg, 0.02 mmol) were successively added to a reaction flask. The atmosphere was replaced with argon, and the reaction was carried out at 100 °C for 10 h. The reaction solution was concentrated, and the product was purified by preparative thin-layer chromatography (DCM / MeOH = 10:1, v / v) to give 34 mg of a yellow solid in a yield of 65.6%.

[0098] 1 H NMR (400 MHz, DMSO-d6) δ 9.56 (s, 1H), 8.56–8.50 (m, 2H), 8.13 (s, 1H), 8.05 (d, J = 2.2 Hz, 1H), 7.77–7.71 (m, 2H), 7.53 (d, J = 2.2 Hz, 1H), 7.35 (s, 1H), 7.30 (t, J = 7.8 Hz, 1H), 6.89–6.77 (m, 3H), 6.53 (s, 2H).

[0099] MS(ESI+) m / z: 305.99 [M+H] + .

[0100] Example 2: Synthesis of 2-Amino-3-(3-hydroxy-2-methylphenyl)-5-(pyridin-4-yl)benzamide

[0101]

[0102] Step 1: Synthesis of 2-Amino-3-(3-methoxy-2-methylphenyl)-5-(pyridin-4-yl)benzamide

[0103]

[0104] 2-Amino-3-bromo-5-(pyridin-4-yl)benzamide (293 mg, 1 mmol), 3-methoxy-2-methylphenylboronic acid (232 mg, 1.4 mmol), 20 mL of dioxane, 1.5 mL of 2 M aqueous potassium carbonate solution, and Pd(dppf)Cl2 (73 mg, 0.1 mmol) were successively added to a reaction flask. After purging with argon, the reaction was carried out at 100 °C for 10 h. The reaction mixture was concentrated, and preparative thin-layer chromatography (DCM / MeOH = 10:1, v / v) was used to obtain 120 mg of a pale yellow solid with a yield of 36.0%.

[0105] 1 H NMR (400 MHz, DMSO-d6) δ 8.51 (d, J = 5.3 Hz, 2H), 8.14 (s, 1H), 8.09 (d, J = 2.2 Hz, 1H), 7.76–7.70 (m, 2H), 7.46 (d, J = 2.2 Hz, 1H), 7.34 (s, 1H), 7.29 (t, J = 7.9 Hz, 1H), 7.03 (dd, J = 8.4, 1.1 Hz, 1H), 6.80 (dd, J = 7.7, 1.1 Hz, 1H), 6.27 (s, 2H), 1.94 (s, 3H).

[0106] MS (ESI+) m / z: 334.08 [M+H] + .

[0107] Step 2: Synthesis of 2-Amino-3-(3-hydroxy-2-methylphenyl)-5-(pyridin-4-yl)benzamide

[0108]

[0109] 2-Amino-3-(3-methoxy-2-methylphenyl)-5-(pyridin-4-yl)benzamide (53 mg, 0.16 mmol) was added to 10 mL of DCE at 0 °C, and the mixture was heated under reflux for 4 h. The reaction was quenched with 5 mL of water and 10 mL of saturated sodium bicarbonate solution, and the mixture was extracted with EA (20 mL × 3). The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. Preparative thin-layer chromatography (DCM / MeOH = 9:1, v / v) was used to obtain 16 mg of a pale yellow solid with a yield of 31.3%.

[0110] 1 1H NMR (400 MHz, DMSO-d6) δ 9.41 (s, 1H), 8.48 (d, J = 5.6 Hz, 2H), 8.09 (s, 1H), 8.05 (d, J = 2.3 Hz, 1H), 7.75–7.69 (m, 2H), 7.43 (d, J = 2.2 Hz, 1H), 7.30 (s, 1H), 7.06 (t, J = 7.8 Hz, 1H), 6.83 (d, J = 8.0 Hz, 1H), 6.59 (dd, J = 7.6, 1.3 Hz, 1H), 6.23 (s, 2H), 1.87 (s, 3H).

[0111] MS (ESI+) m / z: 320.02 [M+H] + .

[0112] Example 3: Synthesis of 2-Amino-3-(5-hydroxy-2-methylphenyl)-5-(pyridin-4-yl)benzamide

[0113]

[0114] Step 1: Synthesis of 2-Amino-3-(5-methoxy-2-methylphenyl)-5-(pyridin-4-yl)benzamide

[0115]

[0116] 2-Amino-3-bromo-5-(pyridin-4-yl)benzamide (117 mg, 0.4 mmol), 3-methoxy-2-methylphenylboronic acid (232 mg, 0.56 mmol), 10 mL of dioxane, 600 μL of 2 M aqueous potassium carbonate solution, and Pd(dppf)Cl2 (30 mg, 0.04 mmol) were successively added to a reaction flask. After purging with argon, the reaction was carried out at 100 °C for 10 h. The reaction mixture was concentrated, and preparative thin-layer chromatography (DCM / MeOH = 9:1, v / v) was used to obtain 75 mg of a pale yellow solid with a yield of 56.2%.

[0117] 1 1H NMR (400 MHz, DMSO-d6) δ 8.55–8.49 (m, 2H), 8.14 (s, 1H), 8.09 (d, J = 2.2 Hz, 1H), 7.77–7.71 (m, 2H), 7.49 (d, J = 2.2 Hz, 1H), 7.35 (s, 1H), 7.26 (d, J = 8.5 Hz, 1H), 6.91 (dd, J = 8.4, 2.8 Hz, 1H), 6.75 (d, J = 2.8 Hz, 1H), 6.29 (s, 2H), 2.03 (s, 3H).

[0118] MS(ESI+) m / z: 334.07 [M+H] + .

[0119] Step 2: Synthesis of 2-Amino-3-(5-hydroxy-2-methylphenyl)-5-(pyridin-4-yl)benzamide

[0120]

[0121] 2-Amino-3-(5-methoxy-2-methylphenyl)-5-(pyridin-4-yl)benzamide (75 mg, 0.225 mmol) was added to 10 mL of DCE at 0 °C and refluxed for 4 h. The reaction was quenched with 5 mL of water and 10 mL of saturated sodium bicarbonate solution, extracted with EA (30 mL × 3), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to give 41 mg of a pale yellow solid with a yield of 57.1%.

[0122] 1 H NMR (400 MHz, DMSO-d6) δ 9.30 (s, 1H), 8.55–8.49 (m, 2H), 8.13 (s, 1H), 8.09 (d, J = 2.3 Hz, 1H), 7.78–7.72 (m, 2H), 7.47 (d, J = 2.2 Hz, 1H), 7.34 (s, 1H), 7.13 (d, J = 8.3 Hz, 1H), 6.73 (dd, J = 8.2, 2.6 Hz, 1H), 6.58 (d, J = 2.6 Hz, 1H), 6.26 (s, 2H), 1.98 (s, 3H).

[0123] MS(ESI+) m / z: 320.05 [M+H] + .

[0124] Preparation of Intermediate 2-Amino-5-bromo-3-(3-hydroxy-2,6-dimethylphenyl)benzamide

[0125] Step 1: Synthesis of 2-Amino-3-bromobenzamide

[0126]

[0127] 2-Amino-3-bromobenzonitrile (1.58 g, 8 mmol) was added to 16 mL of concentrated sulfuric acid at 0 °C and reacted overnight at rt. The reaction solution was poured into ice water, and the pH was adjusted to 7 with saturated sodium bicarbonate solution. It was extracted with EA (100 mL × 3), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to give 1.58 g of a pale yellow solid with a yield of 91.9%.

[0128] 11H NMR (500 MHz, DMSO-d6) δ 7.93 (s, 1H), 7.62–7.56 (m, 1H), 7.55–7.50 (m, 1H), 7.35 (s, 1H), 6.67 (s, 2H), 6.50 (t, J = 7.8 Hz, 1H). MS (ESI+) m / z: 214.89, 216.88 [M+H] + .

[0129] Step 2: Synthesis of 2-Amino-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide

[0130]

[0131] 2-Amino-3-bromobenzamide (5.22 g, 24.3 mmol), bis(pinacolato)diboron (7.42 g, 29.2 mmol), potassium acetate (7.15 g, 72.9 mmol), 240 mL of dioxane, and Pd(dppf)Cl2 (1.78 g, 0.243 mmol) were successively added to a reaction flask. After purging with argon, the mixture was reacted at 100 °C for 10 h. The reaction mixture was concentrated, filtered through diatomaceous earth, extracted with EA (200 mL × 3), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The residue was purified by flash column chromatography (PE / EA = 3:2, v / v) to obtain 5.36 g of a white solid. The crude product obtained was used for the next reaction without further purification.

[0132] HRMS (ESI+) m / z: 263.1556 [M+H] + .

[0133] Step 3: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)benzamide

[0134]

[0135] 2-Amino-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide (3.91 g, 14.9 mmol), 3-bromo-2,4-dimethylphenol (2.49 g, 12.4 mmol), 75 mL of dioxane, 2 M aqueous potassium carbonate solution (18.6 mL, 37.2 mmol), and Pd(dtbpf)Cl2 (808 mg, 1.24 mmol) were successively added to a reaction flask. After purging with argon, the mixture was reacted at 100 °C for 24 h. The reaction mixture was concentrated, filtered through diatomaceous earth, extracted with EA (150 mL × 3), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The residue was purified by flash column chromatography (DCM / MeOH = 20:1, v / v) to obtain 1.45 g of a brown solid with a yield of 45.7%.

[0136] 1 1H NMR (400 MHz, DMSO-d6) δ 9.13 (s, 1H), 7.82 (s, 1H), 7.57 (dd, J = 8.0, 1.6 Hz, 1H), 7.16 (s, 1H), 6.94 (dt, J = 8.1, 0.7 Hz, 1H), 6.86 (dd, J = 7.2, 1.5 Hz, 1H), 6.74 (d, J = 8.1 Hz, 1H), 6.62 (dd, J = 7.9, 7.2 Hz, 1H), 5.73 (s, 2H), 1.84 (s, 3H), 1.77 (s, 3H).

[0137] HRMS (ESI+) m / z: 257.1309 [M+H] + .

[0138] Step 4: Synthesis of 2-Amino-5-bromo-3-(3-hydroxy-2,6-dimethylphenyl)benzamide

[0139]

[0140] 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)benzamide (1.35 g, 5.3 mmol) was dissolved in 50 mL of glacial acetic acid, and NBS (943 mg, 5.3 mmol) was added portionwise. The reaction was carried out at rt for 1 h. The reaction solution was diluted with methanol, concentrated, and the residue was purified by flash column chromatography (DCM / MeOH = 40:1, v / v) to give 1.39 mg of a pale yellow solid in a yield of 77.4%.

[0141] 1 1H NMR (400 MHz, DMSO-d6) δ 9.19 (s, 1H), 7.96 (s, 1H), 7.76 (d, J = 2.4 Hz, 1H), 7.30 (s, 1H), 7.01–6.92 (m, 2H), 6.76 (d, J = 8.1 Hz, 1H), 5.86 (s, 2H), 1.85 (s, 3H), 1.78 (s, 3H).

[0142] HRMS (ESI+) m / z: 335.0381, 337.0359 [M+H] + .

[0143] Example 4: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-(pyridin-4-yl)benzamide

[0144]

[0145] 2-Amino-5-bromo-3-(3-hydroxy-2,6-dimethylphenyl)benzamide (100 mg, 0.3 mmol), 4-pyridineboronic acid (52 mg, 0.42 mmol), 5 mL of dioxane, 450 μL of 2 M aqueous potassium carbonate solution, and Pd(dppf)Cl2 (22 mg, 0.1 mmol) were added to a reaction flask. After purging with argon, the mixture was reacted at 100 °C for 10 h. The reaction mixture was concentrated, filtered through diatomaceous earth, extracted with EA (30 mL × 3), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The residue was purified by preparative thin-layer chromatography (DCM / MeOH = 100:7.5, v / v) to give 35 mg of a pale yellow solid with a yield of 35.0%.

[0146] 1 H NMR (400 MHz, DMSO-d6) δ 9.17 (s, 1H), 8.54–8.48 (m, 2H), 8.13 (s, 1H), 8.09 (d, J = 2.2 Hz, 1H), 7.76–7.70 (m, 2H), 7.39 (d, J = 2.2 Hz, 1H), 7.33 (s, 1H), 6.98 (d, J = 8.2 Hz, 1H), 6.78 (d, J = 8.2 Hz, 1H), 6.12 (s, 2H), 1.88 (s, 3H), 1.82 (s, 3H).

[0147] HRMS (ESI+) m / z: 334.1552 [M+H] + .

[0148] Example 5: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-phenylbenzamide

[0149]

[0150] According to the method in Example 4, the title compound was prepared from phenylboronic acid.

[0151] 1 H NMR (400 MHz, DMSO-d6) δ 9.15 (s, 1H), 8.07 (s, 1H), 7.92 (d, J = 2.2 Hz, 1H), 7.71–7.64 (m, 2H), 7.38 (dd, J = 8.4, 7.2 Hz, 2H), 7.29–7.18 (m, 3H), 6.97 (d, J = 8.2 Hz, 1H), 6.76 (d, J = 8.1 Hz, 1H), 5.85 (s, 2H), 1.89 (s, 3H), 1.82 (s, 3H).

[0152] MS (ESI+) m / z: 333.11 [M+H] + .

[0153] Example 6: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-(pyridin-3-yl)benzamide

[0154]

[0155] According to the method in Example 4, the title compound was prepared from 3-pyridineboronic acid.

[0156] 1 H NMR(400MHz,DMSO-d6)δ9.16(s,1H),8.93(dd,J=2.4,0.9Hz,1H),8.44(dd,J=4.7,1.6Hz,1H),8.07(ddd,J=8.1,2.4,1.6Hz,2H),7.99(d,J=2.2Hz,1H),7.39(ddd,J=8.0,4.8,0.9Hz,1H),7.29(d,J=2.2Hz,2H),6.97(d,J=8.2Hz,1H),6.77(d,J=8.1Hz,1H),5.97(s,2H),1.89(s,3H),1.83(s,3H).

[0157] HRMS(ESI+)m / z:334.1540[M+H] + .

[0158] Example 7: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-(pyrimidin-5-yl)benzamide

[0159]

[0160] According to the method in Example 4, the title compound was prepared from 5-pyrimidineboronic acid.

[0161] 1 H NMR(400MHz,DMSO-d6)δ9.16(d,J=5.3Hz,3H),9.04(s,1H),8.07(d,J=2.2Hz,2H),7.41(d,J=2.1Hz,1H),7.34(s,1H),6.97(d,J=8.1Hz,1H),6.77(d,J=8.2Hz,1H),6.09(s,2H),1.89(s,3H),1.82(s,3H).

[0162] HRMS(ESI+)m / z:335.1496[M+H] + .

[0163] Example 8: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-(3-methylpyridin-4-yl)benzamide

[0164]

[0165] According to the method in Example 4, the title compound was prepared from 3-methyl-4-pyridineboronic acid.

[0166] 1 H NMR(400MHz,DMSO-d6)δ9.17(s,1H),8.45–8.37(m,2H),7.96(s,1H),7.68(d,J=2.1Hz,1H),7.34(d,J=5.0Hz,1H),7.25(s,1H),7.00–6.93(m,2H),6.76(d,J=8.2Hz,1H),5.98(s,2H),2.30(s,3H),1.90(s,3H),1.83(s,3H).

[0167] HRMS(ESI+)m / z:348.1717[M+H] + .

[0168] Example 9: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-(2-methylpyridin-4-yl)benzamide

[0169]

[0170] According to the method in Example 4, the title compound was prepared from 2-methyl-4-pyridineboronic acid.

[0171] 1 H NMR(400MHz,DMSO-d6)δ9.17(s,1H),8.38(dd,J=5.5,0.7Hz,1H),8.12(s,1H),8.07(d,J=2.2Hz,1H),7.62(d,J=1.8Hz,1H),7.54(dd,J=5.3,1.9Hz,1H),7.38(d,J=2.2Hz,1H),7.33(s,1H),6.97(d,J=8.2Hz,1H),6.77(d,J=8.2Hz,1H),6.10(s,2H),2.47(s,3H),1.88(s,3H),1.81(s,3H).

[0172] HRMS(ESI+)m / z:348.1696[M+H] + .

[0173] Example 10: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-(2-fluoropyridin-4-yl)benzamide

[0174]

[0175] According to the method in Example 4, the title compound was prepared from 2-fluoro-4-pyridineboronic acid.

[0176] 1 H NMR(400MHz,DMSO-d6)δ9.18(s,1H),8.19–8.13(m,3H),7.73(dt,J=5.5,1.9Hz,1H),7.56(d,J=1.5Hz,1H),7.47(d,J=2.2Hz,1H),7.37(s,1H),6.98(d,J=8.2Hz,1H),6.78(d,J=8.2Hz,1H),6.24(s,2H),1.88(s,3H),1.82(s,3H).

[0177] HRMS(ESI+)m / z:352.1464[M+H] + .

[0178] Example 11: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-(2-chloropyridin-4-yl)benzamide

[0179]

[0180] According to the method in Example 4, the title compound was prepared from 2-chloro-4-pyridineboronic acid.

[0181] 1 H NMR(400MHz,DMSO-d6)δ9.18(s,1H),8.32(d,J=5.4Hz,1H),8.19–8.11(m,2H),7.92(d,J=1.6Hz,1H),7.77(dd,J=5.4,1.7Hz,1H),7.47(d,J=2.2Hz,1H),7.37(s,1H),6.98(d,J=8.1Hz,1H),6.78(d,J=8.1Hz,1H),6.25(s,2H),1.88(s,3H),1.81(s,3H).

[0182] HRMS(ESI+)m / z:368.1150[M+H] + .

[0183] Example 12: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-[2-(trifluoromethyl)pyridin-4-yl]benzamide

[0184]

[0185] According to the method in Example 4, the title compound was prepared from 2-(trifluoromethyl)pyridine-4-boronic acid pinacol ester.

[0186] 1 H NMR(400MHz,DMSO-d6)δ9.18(s,1H),8.66(d,J=5.3Hz,1H),8.27(d,J=1.8Hz,1H),8.20(d,J=2.2Hz,2H),8.05(dd,J=5.3,1.8Hz,1H),7.56(d,J=2.2Hz,1H),7.43–7.38(m,1H),6.98(d,J=8.2Hz,1H),6.79(d,J=8.1Hz,1H),6.28(s,2H),1.89(s,3H),1.82(s,3H).

[0187] MS(ESI+)m / z:402.10[M+H] + .

[0188] Example 13: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-(2-acetylpyridin-4-yl)benzamide

[0189]

[0190] According to the method in Example 4, the title compound was prepared from 2-acetylpyridine-4-boronic acid pinacol ester.

[0191] 1 H NMR(400MHz,DMSO-d6)δ9.18(s,1H),8.65(dd,J=5.2,0.7Hz,1H),8.30–8.23(m,2H),8.17(d,J=2.2Hz,1H),7.99(dd,J=5.3,2.0Hz,1H),7.43(d,J=2.2Hz,1H),7.36(s,1H),6.98(d,J=8.2Hz,1H),6.78(d,J=8.2Hz,1H),6.19(s,2H),2.66(s,3H),1.89(s,3H),1.82(s,3H).

[0192] HRMS(ESI+)m / z:376.1651[M+H] + .

[0193] Example 14: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-(4-fluorophenyl)benzamide

[0194]

[0195] According to the method in Example 4, the title compound was prepared from 4-fluorophenylboronic acid.

[0196] 1 H NMR(400MHz,DMSO-d6)δ9.15(s,1H),8.06(s,1H),7.89(d,J=2.2Hz,1H),7.75–7.67(m,2H),7.26(s,1H),7.24–7.16(m,3H),6.96(d,J=8.2Hz,1H),6.76(d,J=8.2Hz,1H),5.85(s,2H),1.89(s,3H),1.82(s,3H).

[0197] MS(ESI+)m / z:351.10[M+H] + .

[0198] Example 15: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-(2-methylpyrimidin-5-yl)benzamide

[0199]

[0200] According to the method in Example 4, the title compound was prepared from (2-methylpyrimidin-5-yl)boronic acid.

[0201] 1 H NMR(500MHz,DMSO-d6)δ9.18(s,1H),9.03(s,2H),8.08(s,1H),8.01(s,1H),7.38–7.34(m,2H),6.97(d,J=8.2Hz,1H),6.77(d,J=8.1Hz,1H),6.04(s,2H),2.62(s,3H),1.89(s,3H),1.82(s,3H).

[0202] MS(ESI+)m / z:349.13[M+H] + .

[0203] Example 16: Synthesis of 2-Amino-3-(3-hydroxy-2,6-dimethylphenyl)-5-(1-methyl-2-oxo-1,2-dihydropyridin-4-yl)benzamide

[0204]

[0205] According to the method in Example 4, the title compound was prepared from (1-methyl-2-oxo-1,2-dihydropyridin-4-yl)boronic acid.

[0206] 1 H NMR (500 MHz, DMSO-d6) δ 9.18 (s, 1H), 8.16 (s, 1H), 8.03–7.98 (m, 1H), 7.66 (d, J = 7.2 Hz, 1H), 7.31 (s, 1H), 7.26 (d, J = 2.2 Hz, 1H), 6.97 (d, J = 8.2 Hz, 1H), 6.79–6.73 (m, 2H), 6.64 (dd, J = 7.2, 2.1 Hz, 1H), 6.15 (s, 2H), 3.41 (s, 3H), 1.87 (s, 3H), 1.80 (s, 3H).

[0207] HRMS (ESI+) m / z: 364.1646 [M+H] + .

[0208] Pharmacological activity evaluation

[0209] Example 17: Kinase activity test of PKMYT1

[0210] The in vitro kinase activity assay was used to evaluate the effect of the compounds of the present invention on PKMYT1. The Lanthascreen Assay was used to detect the inhibitory effect of small molecule inhibitors on the PKMYT1 kinase. Terbium was labeled on the monoclonal antibody of the polypeptide substrate as the "donor" fluorescent molecule, and the emission light of terbium was about 480 nm; while the "acceptor" fluorescent molecule was labeled on the polypeptide substrate, and the emission light of terbium corresponding to the "acceptor" was about 520 nm. When the polypeptide substrate was phosphorylated, it could bind to the antibody, resulting in energy transfer, which could be recognized by a device capable of reading time-resolved fluorescence. The PKMYT1 kinase reaction system was 20 μL, which included 2 nM PKMYT1, small molecule inhibitors with concentration gradients, 10 mM MgCl2, 2 mM DTT, 95 μM ATP (the measured Km value), 0.02 μM Fluorescein-PolyGT, 0.01% Tween-20, 0.01% BSA, and 50 mM HEPES at pH 7.5. The enzyme and the inhibitor were added to a 384-well plate and incubated at room temperature for 10 minutes, then the substrate and ATP were added to start the reaction. After 10 minutes, 20 μL of the corresponding antibody and EDTA mixture was added to terminate the reaction, and it was left at room temperature for 60 minutes. The Envision instrument read the data, and a curve was plotted with the Log concentration of the inhibitor as the X-axis and the inhibition rate as the Y-axis. According to the formula Y = Bottom + (Top - Bottom) / (1 + (IC 50 / X)^HillSlope), the IC 50 value was obtained.

[0211] Table 1: Inhibitory activity against PKMYT1 kinase.

[0212]

[0213]

Claims

1. A compound of formula (I) or a pharmaceutically acceptable salt thereof: Wherein: Ring A is selected from 6-membered aryl or heteroaryl; Each R1 is independently selected from cyano, halogen, aldehyde, acetyl, C1-C3 alkyl, halo-C1-C3 alkyl, C1-C3 alkoxy; n is an integer of 0, 1, 2, 3, 4 or 5; R2 is selected from hydrogen, halogen, C1-C3 alkyl, halo-C1-C3 alkyl; R3 is selected from hydrogen, halogen, C1-C3 alkyl, halo-C1-C3 alkyl; R4 is selected from hydrogen, halogen, C1-C3 alkyl, halo-C1-C3 alkyl; R5 is selected from hydrogen, halogen, C1-C3 alkyl, halo-C1-C3 alkyl.

2. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein: Ring A is selected from phenyl, pyridyl, pyrimidinyl, pyridone; Each R1 is independently selected from fluorine, chlorine, methyl, trifluoromethyl, acetyl; n is an integer of 0, 1, 2, 3, 4 or 5; R2 is selected from hydrogen, methyl; R3 is selected from hydrogen, methyl; R4 is selected from hydrogen; R5 is selected from hydrogen.

3. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 2, wherein the compound is selected from:

4. A pharmaceutical composition comprising at least one compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-3 and optionally a pharmaceutically acceptable carrier and / or excipient.

5. The pharmaceutical composition according to claim 4, which further comprises a pharmaceutically active ingredient other than the compound or a pharmaceutically acceptable salt thereof.

6. Use of the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-3, or the pharmaceutical composition according to any one of claims 4 or 5 in the preparation of a medicament for preventing and / or treating diseases related to PKMYT1.

7. Use of the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-3, or the pharmaceutical composition according to any one of claims 4 or 5 in the preparation of a medicament for preventing and / or treating tumors.

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