Heterocyclic compounds as RIPK1 inhibitors

Abstract

The present invention relates to compounds of formula (I), pharmaceutical compositions containing them, processes for their preparation, and their use as receptor interacting protein kinase 1 (RIPK1) inhibitors. The invention also relates to application of the compound or the composition containing the compound to treatment or prevention of related diseases or symptoms mediated by RIPK1. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to heterocyclic compounds, pharmaceutical compositions containing the same, and their use as inhibitors of receptor-interacting protein kinase 1 (RIPK1). More specifically, the present invention provides novel heterocyclic compounds as RIPK1 inhibitors, pharmaceutical compositions containing such compounds, and methods for treating or preventing RIPK1-mediated related diseases or disorders using the compounds. The present invention also relates to methods for preparing the compounds. Background Art

[0002] Receptor-interacting protein kinase 1 (RIPK1) is a serine / threonine protein kinase involved in innate immune signal transduction. RIPK1 is a 76 kDa protein with an N-terminal kinase domain, a C-terminal death domain, and a middle domain with a RHIM (receptor-interacting protein homotypic interaction motif). The C-terminal death domain mediates homodimerization and heterodimerization with other death domain-containing proteins, and the N-terminal kinase domain mediates trans-autophosphorylation to promote self-activation.

[0003] RIPK1 has dual immunomodulatory effects. On the one hand, it can act as a scaffold to promote the activation of the MAPK and NF-κB signaling pathways, thereby promoting inflammation, cell survival, and inhibiting apoptosis. On the other hand, abnormally regulated RIPK1 activity will cause cell necrosis. RIPK1 is a major regulator of the cell determinants of NF-κB signal transduction and death responses, and NF-κB signaling responds to a wide range of inflammatory and pro-death stimuli in human diseases (Degterev, A., et.al. Proc. Natl. Acad. Sci. USA, 2019, 116(20), 9714-9722).

[0004] RIPK1 is widely expressed in various cell types, most abundantly in fat, endothelial, and perivascular cell clusters, and also in immune cell clusters (dendritic cells, macrophages, and T cells). It has been found that the activation of RIPK1 kinase exists in pathological samples of autoimmune diseases and neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and Alzheimer's disease (AD). Anti-tumor necrosis factor-α (TNF-α) drugs have achieved significant clinical success in the treatment of human peripheral inflammatory diseases such as rheumatoid arthritis, colitis, and psoriasis. However, since tumor necrosis factor receptor 2 (TNFR2) mediates nerve regeneration, there are safety concerns in the treatment of central nervous system diseases. RIPK1 inhibitors can safely improve the harmful TNF-α response in the central nervous system without affecting TNFR2. Therefore, RIPK1 inhibitors have the potential to become drugs to replace TNF-α antibodies to make up for the deficiencies of TNF-α antibodies.

[0005] Studies have shown that the RIPK1 small molecule inhibitor Necrostatin-1 (Nec-1), which is known in the art, can effectively block programmed necrosis of cells (Degterev et al. Nat. Chem. Biol. 2005; 1: 112-119.), and has shown effective therapeutic effects in a variety of inflammatory diseases. The brain-penetrating RIPK1 inhibitor DNL-788 is used for amyotrophic lateral sclerosis and so on. These trials have laid the foundation for promoting the clinical application of RIPK1 inhibitors.

[0006] Therefore, the development of novel RIPKl inhibitors with high activity is of great significance in this field, especially highly selective RIPKl inhibitors that can penetrate the blood-brain barrier, providing potential therapeutic drugs for targeting neuroinflammation and cell death leading to various neurological diseases, including Alzheimer's disease, amyotrophic lateral sclerosis (ALS), and multiple sclerosis, as well as acute neurological diseases such as stroke and traumatic brain injury. Summary of the Invention

[0007] The present invention relates to compounds of formula (I), their isomers, prodrugs, solvates, stable isotope derivatives, or pharmaceutically acceptable salts,

[0008]

[0009] Wherein:

[0010] L is a single bond or -(CH2) n -;

[0011] A is a 6-8 membered saturated heterocycle containing 1-3 heteroatoms selected from N, O, S;

[0012] R1 Selected from cyano, -C(O)NH2, halo C1-C8 alkyl, C6-C10 aryl, 5-10 membered heteroaryl, C3-C8 cycloalkyl, 4-8 membered heterocyclic group, wherein the C6-C10 aryl, 5-10 membered heteroaryl, C3-C8 cycloalkyl, 4-8 membered heterocyclic group are unsubstituted or substituted by one or more R x wherein the R x each independently selected from halogen, cyano, C1-C6 alkyl, C3-C6 cycloalkyl, 4-8 membered saturated heterocyclic group, wherein the C1-C6 alkyl, C3-C6 cycloalkyl are unsubstituted or substituted by one or more halogen or cyano;

[0013] R 2 、R 3 、R 4 each independently selected from hydrogen, C1-C8 alkyl, C1-C8 alkoxy, wherein the C1-C8 alkyl and C1-C8 alkoxy are unsubstituted or substituted by one or more halogen or cyano;

[0014] n is an integer selected from 1-5.

[0015] More preferably, the present invention relates to a compound of formula (I) as described above, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, which have the formula (II):

[0016]

[0017] wherein:

[0018] L is a single bond or -(CH2) n -;

[0019] R 1 Selected from cyano, -C(O)NH2, halo C1-C8 alkyl, C6-C10 aryl, 5-10 membered heteroaryl, C3-C8 cycloalkyl, 4-8 membered heterocyclic group, wherein the C6-C10 aryl, 5-10 membered heteroaryl, C3-C8 cycloalkyl, 4-8 membered heterocyclic group are unsubstituted or substituted by one or more R x wherein the R x each independently selected from halogen, cyano, C1-C6 alkyl, C3-C6 cycloalkyl, 4-8 membered saturated heterocyclic group, wherein the C1-C6 alkyl, C3-C6 cycloalkyl are unsubstituted or substituted by one or more halogen or cyano;

[0020] R 2 、R 3 、R 4Each independently selected from hydrogen, C1-C8 alkyl, C1-C8 alkoxy, wherein the C1-C8 alkyl and C1-C8 alkoxy are unsubstituted or substituted by one or more halogens or cyano groups;

[0021] n is an integer selected from 1-3.

[0022] More preferably, the present invention relates to a compound of formula (II) as described above, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, wherein:

[0023] L is a single bond or -CH2-;

[0024] R 1 Selected from cyano, -C(O)NH2, halo C1-C6 alkyl, C6-C10 aryl, 5-8-membered heteroaryl containing 1-3 heteroatoms selected from N, O and S, C3-C8 cycloalkyl, 4-8-membered saturated heterocyclic group containing 1-2 heteroatoms selected from N and O, wherein the C6-C10 aryl, 5-8-membered heteroaryl, C3-C8 cycloalkyl, 4-8-membered saturated heterocyclic group are unsubstituted or substituted by 1-3 R x wherein the R x Each independently selected from fluorine, chlorine, bromine, cyano, C1-C4 alkyl, C3-C6 cycloalkyl, 4-6-membered saturated heterocyclic group; wherein the C1-C4 alkyl and C3-C6 cycloalkyl are unsubstituted or substituted by 1-3 substituents selected from fluorine, chlorine, bromine, cyano;

[0025] R 2 、R 3 、R 4 Each independently selected from hydrogen, C1-C8 alkyl, C1-C8 alkoxy, wherein the C1-C8 alkyl and C1-C8 alkoxy are unsubstituted or substituted by 1-3 halogens or cyano groups.

[0026] Even more preferably, the present invention relates to a compound of formula (II) as described above, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, wherein:

[0027] L is a single bond or -CH2-;

[0028] R 1 Selected from cyano, -C(O)NH2, halo C1-C4 alkyl, phenyl, 5-6-membered heteroaryl containing 1-3 heteroatoms selected from N, O and S, C3-C6 cycloalkyl, 4-6-membered saturated heterocyclic group containing 1 heteroatom selected from N and O, wherein the phenyl, 5-6-membered heteroaryl, C3-C6 cycloalkyl, 4-6-membered saturated heterocyclic group are unsubstituted or substituted by 1-3 R x wherein the Rx Each independently selected from fluorine, chlorine, cyano, C1-C4 alkyl, C3-C6 cycloalkyl, a 4-6 membered saturated heterocyclic group containing 1 heteroatom selected from N and O; wherein the C1-C4 alkyl and C3-C6 cycloalkyl are unsubstituted or substituted with 1-3 substituents selected from fluorine and cyano;

[0029] R 2 、R 3 、R 4 Each independently selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, wherein the C1-C6 alkyl and C1-C6 alkoxy are unsubstituted or substituted with 1-3 fluorine, chlorine, bromine or cyano.

[0030] More preferably, the present invention relates to a compound of formula (II) as described above, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, wherein:

[0031] L is a single bond or -CH2-;

[0032] R 1 Selected from cyano, -C(O)NH2, C1-C4 alkyl substituted with 1-3 fluorine, 5-membered heteroaryl containing 1-3 heteroatoms selected from N, O and S, 4-6 membered saturated heterocyclic group containing 1 O heteroatom, wherein the 5-membered heteroaryl and 4-6 membered saturated heterocyclic group are unsubstituted or substituted with 1-3 R x wherein the R x Each independently selected from fluorine, cyano, methyl, ethyl, C3-C6 cycloalkyl, 4-6 membered saturated heterocyclic group containing 1 O heteroatom; wherein the methyl, ethyl and C3-C6 cycloalkyl are unsubstituted or substituted with 1-3 substituents selected from fluorine and cyano;

[0033] R 2 、R 3 、R 4 Each independently selected from C1-C4 alkyl and C1-C4 alkoxy, wherein the C1-C4 alkyl is unsubstituted or substituted with 1-3 substituents selected from fluorine and cyano.

[0034] Still more preferably, the present invention relates to a compound of formula (II) as described above, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, wherein:

[0035] L is a single bond or -CH2-;

[0036] R 1 Selected from cyano, -C(O)NH2,

[0037]

[0038] R 2 、 R 3 、 R 4 Each independently is selected from methyl, methoxy, monofluoromethyl, difluoromethyl, and cyanomethyl.

[0039] Most preferably, the present invention relates to a compound of formula (II) as described above, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, which are selected from:

[0040]

[0041]

[0042]

[0043] The present invention further relates to a pharmaceutical composition comprising the compound, isomer, prodrug, solvate, stable isotope derivative or pharmaceutically acceptable salt described in any one embodiment of the present invention, optionally one or more other RIPK1 inhibitors, and one or more pharmaceutically acceptable carriers.

[0044] The present invention also relates to the use of the compound, isomer, prodrug, solvate, stable isotope derivative or pharmaceutically acceptable salt described in any one embodiment of the present invention or the pharmaceutical composition according to the present invention in the preparation of a drug for use as an RIPK1 inhibitor; a drug for the treatment or prevention of RIPK1-mediated diseases or disorders or diseases or disorders caused by necroptosis.

[0045] The present invention also relates to the use of the compound, isomer, prodrug, solvate, stable isotope derivative or pharmaceutically acceptable salt described in any one embodiment of the present invention in the preparation of a drug for the treatment or prevention of RIPK1-mediated related diseases or disorders, such as: Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis, acute neurological diseases, inflammatory bowel disease, Crohn's disease, ulcerative colitis, psoriasis, atopic dermatitis, rheumatoid arthritis, spondyloarthritis, gout, systemic lupus erythematosus, non-alcoholic steatohepatitis, alcoholic steatohepatitis, autoimmune hepatitis, autoimmune hepatobiliary diseases, systemic inflammatory response syndrome, cerebrovascular accident, Huntington's disease, cancer (such as pancreatic cancer), bacterial infection, hematological malignancies, solid organ malignancies, etc.

[0046] The present invention also relates to the use of the pharmaceutical composition according to the present invention in the preparation of a medicament, wherein the medicament is used for treating or preventing RIPK1-mediated related diseases or disorders, such as Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis, acute neurological diseases, inflammatory bowel disease, Crohn's disease, ulcerative colitis, psoriasis, atopic dermatitis, rheumatoid arthritis, spondyloarthritis, gout, systemic lupus erythematosus, non-alcoholic steatohepatitis, alcoholic steatohepatitis, autoimmune hepatitis, autoimmune hepatobiliary diseases, systemic inflammatory response syndrome, cerebrovascular accident, Huntington's disease, cancer (such as pancreatic cancer), bacterial infection, hematological malignancies, solid organ malignancies, etc.

[0047] The present invention also relates to a method for treating or preventing RIPK1-mediated related diseases or disorders, which comprises administering to a patient in need thereof a therapeutically effective amount of the compound, isomer, prodrug, solvate, stable isotope derivative or pharmaceutically acceptable salt according to any one embodiment of the present invention, or the pharmaceutical composition according to the present invention, such as Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis, acute neurological diseases, inflammatory bowel disease, Crohn's disease, ulcerative colitis, psoriasis, atopic dermatitis, rheumatoid arthritis, spondyloarthritis, gout, systemic lupus erythematosus, non-alcoholic steatohepatitis, alcoholic steatohepatitis, autoimmune hepatitis, autoimmune hepatobiliary diseases, systemic inflammatory response syndrome, cerebrovascular accident, Huntington's disease, cancer (such as pancreatic cancer), bacterial infection, hematological malignancies, solid organ malignancies, etc.

[0048] Another aspect of the present invention relates to the compound according to any one embodiment of the present invention, or its isomer, prodrug, solvate, stable isotope derivative or pharmaceutically acceptable salt, which is used for treating or preventing RIPK1-mediated related diseases or disorders, such as Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis, acute neurological diseases, inflammatory bowel disease, Crohn's disease, ulcerative colitis, psoriasis, atopic dermatitis, rheumatoid arthritis, spondyloarthritis, gout, systemic lupus erythematosus, non-alcoholic steatohepatitis, alcoholic steatohepatitis, autoimmune hepatitis, autoimmune hepatobiliary diseases, systemic inflammatory response syndrome, cerebrovascular accident, Huntington's disease, cancer (such as pancreatic cancer), bacterial infection, hematological malignancies, solid organ malignancies, etc.

[0049] Another aspect of the present invention relates to a pharmaceutical composition, which comprises the compound, isomer, prodrug, solvate, stable isotope derivative or pharmaceutically acceptable salt described in any embodiment of the present invention, optionally one or more other RIPK1 inhibitors, and one or more pharmaceutically acceptable carriers, diluents and excipients, and is used for treating or preventing related diseases or disorders mediated by RIPK1, such as Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis, acute neurological diseases, inflammatory bowel disease, Crohn's disease, ulcerative colitis, psoriasis, atopic dermatitis, rheumatoid arthritis, spondyloarthritis, gout, systemic lupus erythematosus, non-alcoholic steatohepatitis, alcoholic steatohepatitis, autoimmune hepatitis, autoimmune hepatobiliary diseases, systemic inflammatory response syndrome, cerebrovascular accident, Huntington's disease, cancer (such as pancreatic cancer), bacterial infection, hematological malignancies, solid organ malignancies, etc.

[0050] According to the present invention, the drug can be in any pharmaceutical dosage form, including but not limited to tablets, capsules, solutions, lyophilized preparations, injections.

[0051] The pharmaceutical preparation of the present invention can be administered in the form of dosage units each containing a predetermined amount of the active ingredient. Such units can contain, for example, from 0.5 mg to 1 g, preferably from 1 mg to 700 mg, particularly preferably from 5 mg to 300 mg of the compound of the present invention, depending on the disease to be treated, the method of administration and the age, weight and condition of the patient, or the pharmaceutical preparation can be administered in the form of dosage units each containing a predetermined amount of the active ingredient. Preferred dosage unit preparations are those containing the active ingredient in the daily dose or divided dose or corresponding fractions as indicated above. In addition, this type of pharmaceutical preparation can be prepared using methods well known in the pharmaceutical field.

[0052] The pharmaceutical preparation of the present invention is suitable for administration by any desired and suitable method, such as by oral (including buccal or sublingual), rectal, nasal, topical (including oral, sublingual or transdermal), vaginal or parenteral (including subcutaneous, intramuscular, intravenous or intradermal) methods. All methods known in the pharmaceutical field can be used to prepare such preparations, for example, by combining the active ingredient with one or more excipients or one or more adjuvants.

[0053] Preparation method

[0054] The present invention also provides a method for preparing the compound described in the present invention.

[0055] Method 1

[0056]

[0057] L, R1 and R 2 、 R 3 and R 4 are defined as described above, and X is chlorine, bromine or iodine;

[0058] Step 1:

[0059] Dissolve compounds (I) and (II) in a solvent (such as N,N - diisopropylethylamine), under the protection of an inert gas (such as nitrogen or argon), add a catalyst (such as copper(I) iodide and bis(triphenylphosphine)palladium(II) dichloride), evacuate the system, displace with an inert gas (such as nitrogen or argon) three times, and react at an oil bath temperature of 60 - 130 °C for 2 - 20 hours to obtain compound (III);

[0060] Step 2:

[0061] Dissolve compound (III) in a solvent (such as dichloromethane or dioxane), add an acid (such as hydrochloric acid or trifluoroacetic acid, etc.), and stir at room temperature for 1 - 5 hours to obtain compound (IV);

[0062] Step 3:

[0063] Dissolve compound (IV), compound (V), a condensing agent (such as 2-(7 - azabenzotriazol - 1 - yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate), and a base (such as triethylamine) in a solvent (such as N,N - dimethylformamide), and stir at room temperature for 2 - 16 hours to obtain compound (VI);

[0064] Step 4:

[0065] Dissolve compound (VI) in a solvent (such as methanol), add a base (such as potassium carbonate), and stir at room temperature for 0.5 - 6 hours to obtain compound (VII);

[0066] Step 5:

[0067] Dissolve compounds (VII) and (VIII) in tetrahydrofuran, under the protection of an inert gas (such as nitrogen or argon), add a base (such as triethylamine), a ligand (such as 2 - dicyclohexylphosphino - 2',4',6'-triisopropylbiphenyl), and a catalyst (such as copper(I) iodide and bis(triphenylphosphine)palladium(II) dichloride or copper(I) iodide and tetrakis(triphenylphosphine)palladium(0)), evacuate the system, and displace with an inert gas (such as nitrogen or argon) three times, and react at an oil bath temperature of 60 - 90 °C for 2 - 20 hours to obtain compound (IX);

[0068] Method 2

[0069]

[0070] L, R1 , R 2 、 R 3 and R 4 are defined as described above;

[0071] Step 1:

[0072] Dissolve compound (I) and (X) in a solvent (such as N,N-dimethylacetamide), under the protection of an inert gas (such as nitrogen or argon), add a base (such as N,N-diisopropylethylamine), a catalyst (such as copper iodide and bis(triphenylphosphine)palladium dichloride), evacuate the system and replace it with an inert gas (such as nitrogen or argon) three times, and react at an oil bath temperature of 60-130 °C for 2-20 hours to obtain compound (XI);

[0073] Step 2:

[0074] Dissolve compound (XI) in a solvent (such as dichloromethane or dioxane), add an acid (such as hydrochloric acid or trifluoroacetic acid, etc.), and stir at room temperature for 1-5 hours to obtain compound (XII);

[0075] Step 3:

[0076] Dissolve compound (XII), compound (V), a condensing agent (such as 2-(7-azabenzotriazol)-N,N,N',N'-tetramethyluronium hexafluorophosphate), and a base (such as triethylamine) in a solvent (such as N,N-dimethylformamide), and stir at room temperature for 2-16 hours to obtain compound (IX); Specific Embodiments

[0077] Definitions

[0078] Unless otherwise stated, the following terms used in the specification and claims have the following meanings. Groups not specifically defined in the present invention have the meanings commonly represented in the art known to those skilled in the art.

[0079] The expression "Cx-Cy" used in the present invention represents the range of the number of carbon atoms, where both x and y are integers. For example, C3-C8 cycloalkyl represents a cycloalkyl having 3-8 carbon atoms.

[0080] In the present invention, the term "alkyl" refers to a saturated aliphatic hydrocarbon group, including straight-chain and branched-chain groups having 1 to 20 carbon atoms, for example, straight-chain and branched-chain groups having 1 to 18 carbon atoms, 1 to 12 carbon atoms, 1 to 8 carbon atoms, 1 to 6 carbon atoms, or 1 to 4 carbon atoms. Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, and various branched isomers thereof. The alkyl group can be optionally substituted or unsubstituted.

[0081] In the present invention, the term "alkoxy" refers to an alkyl-O-group, where the alkyl group has the meaning defined above.

[0082] In the present invention, the term "cycloalkyl" refers to a saturated monocyclic or polycyclic hydrocarbon group, which includes 3 to 12 ring atoms, for example, it can be 3 to 12, 3 to 10, 3 to 8, or 3 to 6 ring atoms, or it can be a 3-, 4-, 5-, or 6-membered ring. Non-limiting examples of the monocyclic group include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, etc. The cycloalkyl group can be optionally substituted or unsubstituted.

[0083] In the present invention, the term "heterocyclic group" refers to a saturated or partially unsaturated monocyclic or polycyclic hydrocarbon group, which includes 3 to 20 ring atoms, for example, it can be 3 to 16, 3 to 12, 3 to 10, 3 to 8, or 4 to 6 ring atoms, where one or more ring atoms are heteroatoms selected from nitrogen, oxygen, or S(O)m (where m is an integer from 0 to 2), but does not include ring moieties of -O-O-, -O-S-, or -S-S-, and the remaining ring atoms are carbon. Preferably, it includes 3 to 10 ring atoms, where 1 to 4 are heteroatoms, more preferably the heterocyclic group ring contains 3 to 8 ring atoms, more preferably includes 4 to 6 ring atoms, and most preferably a 4-membered, 5-membered, or 6-membered ring; where 1 to 4 are heteroatoms, more preferably 1 to 3 are heteroatoms, and most preferably 1 to 2 are heteroatoms. Non-limiting examples of the heterocyclic group include oxetanyl, oxanyl, azetidinyl, morpholinyl, 2-morpholinyl, dihydropyrazolyl, etc. The heterocyclic group can be optionally substituted or unsubstituted.

[0084] In the present invention, the term "aryl" refers to a 6- to 14-membered fully carbon monocyclic or fused polycyclic (i.e., rings sharing adjacent pairs of carbon atoms) group, a polycyclic group having a conjugated π-electron system (i.e., rings thereof having adjacent pairs of carbon atoms), preferably 6- to 10-membered, such as phenyl and naphthyl, and most preferably phenyl. The aryl may be substituted or unsubstituted.

[0085] In the present invention, the term "heteroaryl" refers to a heteroaromatic system containing 1 to 4 heteroatoms and 5 to 14 ring atoms, wherein the heteroatoms include oxygen, sulfur, and nitrogen. Preferably, the heteroaryl contains 1 to 3 heteroatoms, and the heteroatoms contained therein include at least one nitrogen atom. Preferably, the heteroaryl is 5- to 10-membered. More preferably, the heteroaryl is 5- or 6-membered. Preferred heteroaryls are, for example, pyrazolyl, imidazolyl, triazolyl (including 1,2,3-triazolyl, 1,2,4-triazolyl, etc.), thiazolyl, pyrazinyl, oxazolyl, isoxazolyl, pyridyl, etc. The heteroaryl may be optionally substituted or unsubstituted.

[0086] In the present invention, the term "halogen" refers to fluorine, chlorine, bromine, or iodine.

[0087] In the present invention, the term "cyano" refers to -CN.

[0088] In the present invention, "optionally" or "optionally" means that the subsequently described event or circumstance may but need not occur, and this description includes the instances where the event or circumstance occurs or does not occur. For example, "a heterocyclic group optionally substituted by an alkyl group" means that the alkyl group may but need not be present, and this description includes the case where the heterocyclic group is substituted by an alkyl group and the case where the heterocyclic group is not substituted by an alkyl group.

[0089] The substituents include, but are not limited to, the various groups described above.

[0090] The compounds claimed in the present invention include not only the compounds themselves but also the optical isomers or pharmaceutically acceptable salts thereof.

[0091] The "pharmaceutical composition" as described in the present invention refers to a mixture containing one or more compounds described in the present invention or their optical isomers or pharmaceutically acceptable salts and other chemical components. Other components are, for example, pharmaceutically acceptable carriers, diluents, and excipients. The purpose of the pharmaceutical composition is to facilitate the administration to an organism, facilitate the absorption of the active ingredient, and thus exert biological activity.

[0092] When used in the specification, the term "comprising" includes "consisting of".

[0093] The "room temperature" as described in the present invention refers to 15 - 30 °C.

[0094] The "pharmaceutically acceptable salts" of the present invention are discussed in Berge, et al., "Pharmaceutically acceptable salts", J. Pharm. Sci., 66, 1-19 (1977) and are obvious to pharmaceutical chemists. The salts are substantially non-toxic and can provide the desired pharmacokinetic properties, palatability, absorption, distribution, metabolism or excretion, etc.

[0095] The pharmaceutically acceptable salts of the present invention can be synthesized by general chemical methods.

[0096] Generally, the preparation of salts can be achieved by reacting a free base or acid with an equi-chemical equivalent or an excess of an acid (inorganic or organic acid) or base in a suitable solvent or solvent composition.

[0097] The "prodrug" of the present invention refers to a compound that is converted into the original active compound after metabolism in the body. Typically, a prodrug is an inactive substance or less active than the active parent compound, but can provide convenient handling, administration or improved metabolic characteristics.

[0098] The "isomers" of the present invention refer to tautomers, mesomers, racemates, enantiomers, diastereomers, mixtures thereof, etc. of the compounds of formula (I) of the present invention. All these isomers, including stereoisomers such as optical isomers and geometric isomers, are included in the present invention. The geometric isomers include cis-trans isomers.

[0099] The "solvate" of the present invention refers to the association of one or more solvent molecules with the compounds or salts of the present invention. Examples of solvents that form pharmaceutically acceptable solvates include, but are not limited to, water, isopropanol, ethanol, methanol, ethyl acetate, acetic acid, etc.

[0100] The present invention includes any polymorphs of the compounds or salts thereof, as well as any hydrates or other solvates.

[0101] In the present invention, the term "patient" generally refers to mammals, especially humans.

[0102] In the present invention, the term "therapeutically effective amount" refers to the amount of the compounds of the present invention that can effectively treat or prevent related diseases mediated by RIPK1.

[0103] Examples

[0104] The present invention will be further illustrated by the following examples, but the present invention is not limited to the scope of the examples described. The experimental methods without specific conditions in the following examples are carried out according to conventional methods and conditions, or are selected according to the product specifications.

[0105] The structures of all the compounds of the present invention can be identified by nuclear magnetic resonance ( 1 1H NMR) and / or mass spectrometry (MS).

[0106] 1 The 1H NMR chemical shifts (δ) are recorded in PPM (parts per million). NMR is performed on a Bruker AVANCE III - 400 MHz spectrometer. Suitable solvents are selected from chloroform-d (CDCl3), methanol-d (CD3OD), dimethyl sulfoxide-d 6 ) etc., and tetramethylsilane (TMS) is used as the internal standard.

[0107] Low-resolution mass spectrometry (MS) is determined by an Agilent 1260 HPLC / 6120 mass spectrometer, using an Agilent ZORBAX XDB-C18, 4.6×50 mm, 3.5 μm.

[0108] Gradient elution condition 1: 0 min: 95% solvent A1 and 5% solvent B1, 1 - 2 min: 5% solvent A1 and 95% solvent B1; 2.01 - 2.50 min: 95% solvent A1 and 5% solvent B1. The percentages are the volume percentages of a certain solvent in the total solvent volume. Solvent A1: 0.01% formic acid aqueous solution; Solvent B1: acetonitrile solution of 0.01% formic acid; the percentages are the volume percentages of the solute in the solution.

[0109] The thin-layer silica gel plate is Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plate. Column chromatography generally uses Yantai Huanghai 100 - 200 or 200 - 300 mesh silica gel as the carrier.

[0110] Preparative liquid chromatography (prep-HPLC) uses a Waters SQD2 mass spectrometry-guided high-pressure liquid chromatography separation instrument, XBridge-C18; 30X150 mm preparative column, 5 μm;

[0111] Method 1: acetonitrile - water (0.2% formic acid), flow rate 25 mL / min; Method 2: acetonitrile - water (0.8% ammonium bicarbonate), flow rate 25 mL / min;

[0112] The known starting materials of the present invention can be used or synthesized according to methods known in the art, or can be purchased from companies such as Acros Organics, Aldrich Chemical Company, Accela ChemBio Inc, Shanghai Bide Pharmatech, Shanghai Aladdin Chemistry, Shanghai Merck Chemical, J&K Scientific, AnyChem etc.

[0113] Unless otherwise specified in the examples, the solvents used in the reactions are all anhydrous solvents. Among them, anhydrous tetrahydrofuran is commercially available tetrahydrofuran, with sodium chunks as the water remover and benzophenone as the indicator. It is refluxed under argon protection until the solution turns blue-violet, then distilled and collected, and stored at room temperature under argon protection. Other anhydrous solvents are purchased from Energy Chemical and J&K Chemical. Unless otherwise specified, the transfer and use of all anhydrous solvents need to be carried out under nitrogen protection.

[0114] Unless otherwise specified in the examples, the reactions are carried out under an argon or nitrogen atmosphere.

[0115] An argon or nitrogen atmosphere means that the reaction flask is connected to an argon or nitrogen balloon with a volume of about 1 L.

[0116] A hydrogen atmosphere means that the reaction flask is connected to a hydrogen balloon with a volume of about 1 L.

[0117] For the hydrogenation reaction, it is usually evacuated, filled with hydrogen, and this operation is repeated 3 times.

[0118] Unless otherwise specified in the examples, the reaction temperature is room temperature, and the temperature range is 15°C - 30°C.

[0119] The progress of the reaction in the examples is monitored by thin-layer chromatography (TLC). The developing agent systems used in the reaction are as follows: A: dichloromethane and methanol system; B: petroleum ether and ethyl acetate system. The volume ratio of the solvents is adjusted according to the polarity of the compound.

[0120] The eluent system for column chromatography used to purify the compound and the developing agent system for thin-layer chromatography include: A: dichloromethane and methanol system; B: petroleum ether and ethyl acetate system. The volume ratio of the solvents is adjusted according to the polarity of the compound, and a small amount of triethylamine and acidic or basic reagents can also be added for adjustment.

[0121] The reagents used in the biological experiments of the present invention are as follows: Acetonitrile (chromatographic grade): purchased from Fisher-chemical, product number: A998-4L; Formic acid (chromatographic grade): purchased from Fisher Scientific, product number: A117-50ML; Dimethylacetamide DMA: purchased from Vetec, product number: V900211-500ML; DMSO (chromatographic grade): purchased from Sigma-Aldrich, product number: D5879-1L; Solutol: purchased from Beijing Coupling Technology Co., Ltd., product number: Solutol HS15; Methyl cellulose MC: purchased from Sigma, product number: M6385-100G; Terfenadine: purchased from Sigma, product number: MKBX6318V; Saline: purchased from Shandong Hualu Pharmaceutical Co., Ltd., product number: H37022749; Water: purchased from Watson, product number: GB19298; Warfarin: purchased from Dr. Ehrenstorfer GmbH, product number: 30719; Potassium dihydrogen phosphate: purchased from American Vetec Company, product number: WXBC3341V; Disodium hydrogen phosphate: purchased from American Vetec Company, product number: WXBC3348V.

[0122] The instruments used in the biological experiments of the present invention are as follows: AB SCIEX Triple Quad 5500 and AB SCIEX Triple Quad 4500 liquid chromatography-mass spectrometry (products of AB SCIEX Company, USA), including Triple Quad 5500 and Triple Quad 4500 triple quadrupole tandem mass spectrometers, equipped with ESI source and Analyst 1.6.3 data processing system; The liquid phase part is AB SCIEX liquid phase, equipped with high-pressure infusion pump, auto sampler, and column oven; Electronic balance: Sartorius Company, model: BT25S; Centrifuge: Beckman Company, model: Allegra X-12R; Refrigerator: China Haier Group, model: HYC-198, BCD-248TMPM; Homogenizer: Dinghaoyuan Company, model: TL2020; Vortex mixer: American Labnet Company, model: VX-200; Pipette multichannel (120 μL, 300 μL), purchased from Eppendorf; Single-channel pipette (10 μL, 100 μL, 200 μL, 1000 μL), purchased from Eppendorf;

[0123] Intermediate 1

[0124] 1-(9-Ethynyl-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one

[0125]

[0126] The first step

[0127] 9 - ((Trimethylsilyl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-tert-butyl formate 1a

[0128] Dissolve 9-bromo-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-tert-butyl formate (4.14 g, 12.60 mmol, synthesized with reference to the literature: WO2016075239) in N,N-diisopropylethylamine (50 mL). At room temperature, successively add copper(I) iodide (0.24 g, 1.26 mmol), bis(triphenylphosphine)palladium(II) dichloride (0.88 g, 1.26 mmol) and trimethylsilylacetylene (6.18 g, 63.00 mmol). React at 80 °C for 16 hours under nitrogen protection. Cool the reaction solution to room temperature, filter through diatomaceous earth. Add water (30 mL) to the filtrate, extract with ethyl acetate (20 mL × 3). Wash the combined organic phases with saturated brine (30 mL × 2), dry over anhydrous sodium sulfate, filter to remove the desiccant, and evaporate the solvent under reduced pressure to obtain the crude product. The obtained crude product is purified by silica gel column chromatography (petroleum ether:ethyl acetate = 20:1) to obtain the target product 9 - ((trimethylsilyl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-tert-butyl formate 1a (3.49 g), yield: 80%.

[0129] MS m / z (ESI): 347 [M + 1];

[0130] 1 H NMR (400 MHz, CDCl3) δ 8.50 (s, 1H), 8.33 - 8.26 (m, 1H), 4.58 - 4.46 (m, 2H), 4.30 (t, J = 4.0 Hz, 2H), 3.86 (t, J = 4.0 Hz, 2H), 1.41 (s, 9H), 0.27 (s, 9H).

[0131] The second step

[0132] 9 - ((Trimethylsilyl)ethynyl)-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazine 1b

[0133] 9 - ((Trimethylsilyl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazine tert-Butyl -4(5H)-formate 1a (0.10 g, 0.29 mmol) was dissolved in dichloromethane (10 mL) and trifluoroacetic acid (2 mL), and stirred at room temperature for 30 minutes. Most of the solvent was removed by rotary evaporation under reduced pressure. Saturated aqueous sodium bicarbonate (20 mL) was added for neutralization, and the mixture was extracted with dichloromethane (20 mL x 3). The combined organic phases were dried over anhydrous sodium sulfate, the desiccant was filtered off, and the filtrate was concentrated under reduced pressure to obtain the target product 9-((trimethylsilyl)ethynyl)-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazine 1b (80 mg, crude). MS m / z (ESI): 247 [M+1];

[0134] The third step

[0135] 3,3-Difluoro-2,2-dimethyl-1-(9-((trimethylsilyl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-yl)propan-1-one 1c

[0136] 9-((Trimethylsilyl)ethynyl)-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazine 1b (15 mg, 0.06 mmol) and 3,3-difluoro-2,2-dimethylpropanoic acid (25 mg, 0.18 mmol, synthesis reference: WO2017096301) were dissolved in N,N-dimethylformamide (1.5 mL). 2-(7-Azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (47 mg, 0.12 mmol) and triethylamine (18 mg, 0.18 mmol) were added, and the mixture was stirred at room temperature for 1 hour. After dilution with ethyl acetate (5 mL), it was washed with saturated brine (10 mL x 3). The separated organic phase was dried over anhydrous sodium sulfate, the desiccant was filtered off, and the filtrate was concentrated under reduced pressure to obtain 3,3-difluoro-2,2-dimethyl-1-(9-((trimethylsilyl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-yl)propan-1-one (21 mg, crude), with a yield of 95%.

[0137] MS m / z (ESI): 367 [M+1];

[0138] The fourth step

[0139] 1-(9-Ethynyl-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one (Intermediate 1)

[0140] 3,3-Difluoro-2,2-dimethyl-1-(9-((trimethylsilyl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazin- -4(5H)-yl)propan-1-one 1c (21 mg, 0.06 mmol) was dissolved in methanol (3.0 mL), potassium carbonate (25 mg, 0.18 mmol) was added, and the mixture was stirred at room temperature for 30 minutes. The mixture was filtered through diatomaceous earth. The organic phase was concentrated under reduced pressure to obtain a crude product, which was purified by reverse-phase chromatography column (0 - 20% acetonitrile / (0.1% formic acid) water) to obtain the target product 1-(9-ethynyl-2,3-dihydropyrido[3,4-f][1,4]oxazin- -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one (Intermediate 1) (14.0 mg), with a yield of 80%.

[0141] MS m / z (ESI): 295 [M + 1];

[0142] 1 H NMR (400 MHz, CDCl3) δ 8.52 (s, 1H), 8.38 (s, 1H), 6.10 (t, J = 56.4 Hz, 1H), 4.74 (s, 2H), 4.47 (t, J = 4.4 Hz, 2H), 4.06 (t, J = 4.4 Hz, 2H), 3.37 (s, 1H), 1.36 (s, 6H).

[0143] The synthetic steps of Intermediates 2 - 4 refer to Intermediate 1, where in the third step, different acids were used instead of 3,3-difluoro-2,2-dimethylpropanoic acid.

[0144] Intermediate 2

[0145] 1-(9-Ethynyl-2,3-dihydropyrido[3,4-f][1,4]oxazin- -4(5H)-yl)-3-fluoro-2,2-dimethylpropan-1-one

[0146]

[0147] MS m / z (ESI): 277 [M + 1];

[0148] 1 H NMR (400 MHz, CDCl3) δ 8.54 (s, 1H), 8.40 (s, 1H), 4.73 (s, 2H), 4.47 - 4.43 (m, 2H), 4.42 (d, J = 44.8 Hz, 2H), 4.09 - 4.06 (m, 2H), 3.36 (s, 1H), 1.33 (s, 6H).

[0149] Intermediate 3

[0150] 1-(9-Ethynyl-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-yl)-2,2-dimethylpropan-1-one

[0151]

[0152] MS m / z(ESI): 259 [M+1];

[0153] 1 H NMR(400 MHz, CDCl3) δ 8.52(s, 1H), 8.41(s, 1H), 4.72(s, 2H), 4.51 - 4.39(m, 2H), 4.11 - 4.04(m, 2H), 3.36(s, 1H), 1.28(s, 9H).

[0154] Intermediate 4

[0155] 1-(3,3-Difluorocyclobutyl)-4-iodopyrazole

[0156]

[0157] The first step

[0158] 3-(4-Iodopyrazol-1-yl)cyclobutan-1-one 4a

[0159] Dissolve 4-iodopyrazole (0.50 g, 2.50 mmol) and 3-bromocyclobutan-1-one (0.38 g, 2.50 mmol) in N,N-dimethylformamide (7.0 mL), add cesium carbonate (0.84 g, 2.50 mmol), and react at -10 °C for 1 hour. Filter through diatomaceous earth and concentrate under reduced pressure to obtain the crude product. Purify the crude product by silica gel column chromatography (petroleum ether:ethyl acetate = 3:1) to obtain 3-(4-iodopyrazol-1-yl)cyclobutan-1-one 4a (0.42 g) with a yield of 84%.

[0160] MS m / z(ESI): 263 [M+1];

[0161] 1 H NMR(400 MHz, CDCl3) δ 7.61(s, 1H), 7.59(s, 1H), 5.10 - 5.00(m, 1H), 3.85 - 3.84(m, 2H), 3.65 - 3.53(m, 2H).

[0162] The second step

[0163] 1-(3,3-Difluorocyclobutyl)-4-iodopyrazole (Intermediate 4)

[0164] Dissolve 3-(4-iodopyrazol-1-yl)cyclobutan-1-one 4a (50 mg, 0.02 mmol) in dichloromethane (2.0 mL). Under the condition of an ice-water bath, drip diethylaminosulfur trifluoride (61 mg, 0.04 mmol), and gradually raise the temperature to room temperature and react for 2 hours. Filter with diatomaceous earth and rotary evaporate under reduced pressure to obtain 1-(3,3-difluorocyclobutyl)-4-iodopyrazole (Intermediate 4) (10 mg, crude product).

[0165] MS m / z (ESI): 285 [M+1];

[0166] 1 1H NMR (400 MHz, CDCl3) δ 7.50 (s, 1H), 7.43 (s, 1H), 4.67 - 4.58 (m, 1H), 3.16 - 3.01 (m, 4H).

[0167] Example 1

[0168] 1-(9-((1-(Difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one

[0169]

[0170] Dissolve 1-(9-ethynyl-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one (Intermediate 1) (14 mg, 0.05 mmol) and 1-(difluoromethyl)-4-iodo-1H-pyrazole (25 mg, 0.1 mmol) in tetrahydrofuran (1 mL) and N,N-diisopropylethylamine (1 mL). Add copper(I) iodide (1 mg, 0.005 mmol), 2-bisphosphine-2',4',6'-triisopropylbiphenyl (7 mg, 0.015 mmol) and tetrakis(triphenylphosphine)palladium (6 mg, 0.005 mmol), and stir at 60 °C overnight under nitrogen protection. Cool the reaction to room temperature and concentrate under reduced pressure. The obtained crude product is purified by a reverse-phase chromatography column (37% acetonitrile / (0.1% trifluoroacetic acid) water) to obtain the target compound 1-(9-((1-(difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one (10 mg), with a yield of 63%.

[0171] MS m / z (ESI): 411 [M+1];

[0172] 1 1H NMR (400 MHz, CDCl3) δ 8.57 (s, 1H), 8.41 (s, 1H), 8.04 (s, 1H), 7.81 (s, 1H), 7.87 (t, J = 56.4 Hz, 1H), 6.12 (t, J = 56.4 Hz, 1H), 4.76 (s, 2H), 4.52 - 4.48 (m, 2H), 4.09 - 4.05 (m, 2H), 1.37 (s, 6H).

[0173] The synthesis procedures of Examples 2 to 13 refer to the process of Example 1.

[0174] Example 2

[0175] 1-(9-((2-(Difluoromethyl)thiazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazin- -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one

[0176]

[0177] MS m / z (ESI): 428 [M+1];

[0178] 1 1H NMR (400 MHz, CDCl3) δ 8.60 (s, 1H), 8.41 (s, 1H), 7.75 (s, 1H), 6.86 (t, J = 56.4 Hz, 1H), 6.12 (t, J = 56.4 Hz, 1H), 4.76 (s, 2H), 4.52 (t, J = 4.8 Hz, 2H), 4.07 (t, J = 4.8 Hz, 2H), 1.37 (s, 6H).

[0179] Example 3

[0180] 1-(9-((1-(Difluoromethyl)-1H-imidazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazin- -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one

[0181]

[0182] MS m / z (ESI): 411 [M+1];

[0183] 11H NMR (400 MHz, CDCl3) δ 8.70 - 8.45 (broad, 2H), 7.81 (singlet, 1H), 7.41 - 7.37 (multiplet, 1H), 7.02 (triplet, J = 60.8 Hz, 1H), 6.04 (triplet, J = 56.4 Hz, 1H), 4.69 (singlet, 2H), 4.45 - 4.35 (multiplet, 2H), 4.01 - 3.96 (multiplet, 2H), 1.29 (singlet, 6H).

[0184] Example 4

[0185] 1-(9-((1-(Difluoromethyl)-1H-imidazol-5-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one

[0186]

[0187] MS m / z (ESI): 411 [M + 1];

[0188] 1 1H NMR (400 MHz, CDCl3) δ 8.55 (singlet, 1H), 8.42 (singlet, 1H), 7.95 (singlet, 1H), 7.42 (singlet, 1H), 7.25 (triplet, J = 60.4 Hz, 1H), 6.11 (triplet, J = 56.4 Hz, 1H), 4.76 (singlet, 2H), 4.51 - 4.48 (multiplet, 2H), 4.10 - 4.07 (multiplet, 2H), 1.37 (singlet, 6H).

[0189] Example 5

[0190] 3,3-Difluoro-1-(9-((1-(fluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-2,2-dimethylpropan-1-one

[0191]

[0192] MS m / z (ESI): 393 [M + 1];

[0193] 11H NMR (400 MHz, CDCl3) δ 8.60 (s, 1H), 8.38 (s, 1H), 7.87 (s, 1H), 7.80 (s, 1H), 6.11 (t, J = 56.8 Hz, 1H), 5.96 (d, J = 52.4 Hz, 2H), 4.75 (s, 2H), 4.49 (t, J = 4.8 Hz, 2H), 4.07 (t, J = 4.8 Hz, 2H), 1.37 (s, 6H).

[0194] Example 6

[0195] 3,3 - Difluoro - 2,2 - dimethyl - 1-(9 - ((1-(2,2,2 - trifluoroethyl)-1H - pyrazol - 4 - yl)ethynyl)-2,3 - dihydropyrido[3,4 - f][1,4]oxazine -4(5H)-yl)propan - 1 - one

[0196]

[0197] MS m / z (ESI): 443 [M + 1];

[0198] 1 1H NMR (400 MHz, CDCl3) δ 8.58 (s, 1H), 8.41 (s, 1H), 7.75 (s, 1H), 7.74 (s, 1H), 6.12 (t, J = 56.4 Hz, 1H), 4.75 (s, 4H), 4.48 (t, J = 4.8 Hz, 2H), 4.06 (t, J = 4.8 Hz, 2H), 1.37 (s, 6H).

[0199] Example 7

[0200] 2-(4 - ((4-(3,3 - Difluoro - 2,2 - dimethylpropanoyl)-2,3,4,5 - tetrahydropyrido[3,4 - f][1,4]oxazine -9 - yl)ethynyl)-1H - pyrazol - 1 - yl)acetonitrile

[0201]

[0202] MS m / z (ESI): 400 [M + 1];

[0203] 11H NMR (400 MHz, CDCl3) δ 8.48 (s, 1H), 8.31 (s, 1H), 7.72 (s, 1H), 7.68 (s, 1H), 6.04 (t, J = 56.4 Hz, 1H), 5.02 (s, 2H), 4.68 (s, 2H), 4.48 - 4.36 (m, 2H), 4.08 - 3.93 (m, 2H), 1.30 (s, 6H).

[0204] Example 8

[0205] 1-(9-(1-(3,3-Difluorocyclobutyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-yl)-3,3-difluoro-2,2-dimethyl-1-propanone

[0206]

[0207] MS m / z (ESI): 451 [M + 1];

[0208] 1 1H NMR (400 MHz, CDCl3) δ 8.70 - 8.20 (br, 2H), 7.66 (s, 1H), 7.61 (s, 1H), 6.04 (t, J = 56.4 Hz, 1H), 5.29 - 5.27 (m, 1H), 4.78 - 4.69 (m, 2H), 4.43 - 4.34 (m, 2H), 4.02 - 3.98 (m, 2H), 3.24 - 3.01 (m, 4H), 1.29 (s, 6H).

[0209] Example 9

[0210] 3,3-Difluoro-2,2-dimethyl-1-(9-((1-(trifluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-yl)propan-1-one

[0211]

[0212] MS m / z (ESI): 429 [M + 1];

[0213] 11H NMR (400 MHz, CDCl3) δ 8.60 - 8.24 (br, 2H), 7.97 (s, 1H), 7.83 (s, 1H), 6.04 (t, J = 56.4 Hz, 1H), 4.76 - 4.63 (m, 2H), 4.48 - 4.34 (m, 2H), 4.07 - 3.95 (m, 2H), 1.30 (s, 6H).

[0214] Example 10

[0215] 3,3 - Difluoro - 2,2 - dimethyl - 1-(9 - ((1 - (oxiran - 3 - yl)-1H - pyrazol - 4 - yl)ethynyl)-2,3 - dihydropyrido[3,4 - f][1,4]oxazin -4(5H)-yl)propan - 1 - one

[0216]

[0217] MS m / z (ESI): 417 [M + 1];

[0218] 1 1H NMR (400 MHz, CDCl3) δ 8.57 - 8.41 (br, 2H), 7.75 (s, 1H), 7.70 (s, 1H), 6.05 (t, J = 56.4 Hz, 1H), 5.46 - 5.31 (m, 1H), 5.01 - 4.99 (m, 4H), 4.79 - 4.63 (m, 2H), 4.42 - 4.39 (m, 2H), 4.02 - 3.98 (m, 2H), 1.30 (s, 6H).

[0219] Example 11

[0220] 4 - ((4 - (3,3 - Difluoro - 2,2 - dimethylpropionyl)-2,3,4,5 - tetrahydropyrido[3,4 - f][1,4]oxazin -9 - yl)ethynyl)-1 - methyl - 1H - pyrazole - 3 - carbonitrile

[0221]

[0222] MS m / z (ESI): 400 [M + 1];

[0223] 1 1H NMR (400 MHz, CDCl3) δ 8.57 - 8.41 (br, 2H), 7.62 (s, 1H), 6.01 (t, J = 56.4 Hz, 1H), 4.76 (s, 2H), 4.58 - 4.51 (m, 2H), 4.06 - 4.00 (m, 2H), 3.94 (s, 3H), 1.30 (s, 6H).

[0224] Example 12

[0225] 2-(4-((4-Pivaloyl)-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazine -9-yl)ethynyl)-1H-pyrazol-1-yl)acetonitrile

[0226]

[0227] MS m / z(ESI):364[M+1];

[0228] 1 H NMR(400MHz,CDCl3)δ8.47(s,1H),8.34(s,1H),7.72(s,1H),7.68(s,1H),5.02(s,2H),4.67(s,2H),4.45 - 4.30(m,2H),4.08 - 3.83(m,2H),1.22(s,9H).

[0229] Example 13

[0230] 1-(9-((1-(Difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-yl)-3-fluoro-2,2-dimethylpropan-1-one

[0231]

[0232] MS m / z(ESI):393[M+1];

[0233] 1 H NMR(400MHz,CDCl3)δ8.48(s,1H),8.33(s,1H),7.97(s,1H),7.74(s,1H),7.11(t,J = 60.0Hz,1H),4.70 - 4.68(m,2H),4.42 - 4.41(m,2H),4.31 - 4.29(m,2H),4.04 - 4.01(m,2H),1.27(s,3H),1.26(s,3H).

[0234] Example 14

[0235] 1-(9-(3-(1H-Imidazol-1-yl)prop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-yl)-2,2-dimethylpropan-1-one

[0236]

[0237] The first step

[0238] 9-Bromo-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazine Hydrochloride 14-1

[0239] Dissolve compound 9-bromo-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-tert-butyl formate (synthesis reference: WO2018213632) (0.35 g, 1.28 mmol) in methanol (4 mL), add dioxane hydrochloride (4 M, 4 mL), and stir at room temperature for 1 hour. Concentrate under reduced pressure to obtain the target compound 9-bromo-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazine Hydrochloride 14-1 (0.31 g, crude product), and use it directly in the next step without purification. MS m / z (ESI): 229&231 [M+1].

[0240] The second step

[0241] 1-(9-Bromo-2,3-dihydropyrido[3,4-f][1,4]oxazin -4(5H)-yl)-2,2-dimethylpropan-1-one 14-2

[0242] Dissolve 9-bromo-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazine Hydrochloride 14-1 (0.50 g, 1.89 mmol) in dichloromethane (10 mL), add triethylamine (0.95 g, 9.43 mmol) and pivaloyl chloride (0.28 g, 2.26 mmol) to the reaction solution, and stir at room temperature for 2 hours. Add water (20 mL) to the reaction solution, extract with dichloromethane (30 mL×3), wash the combined organic phases with saturated brine (35 mL×2), dry over anhydrous sodium sulfate, filter to remove the desiccant, and concentrate under reduced pressure to obtain the crude product. Silica gel column chromatography (30% ethyl acetate dissolved in petroleum ether) gives the target product 1-(9-bromo-2,3-dihydropyrido[3,4-f][1,4]oxazin -4(5H)-yl)-2,2-dimethylpropan-1-one 14-2 (0.47 g), with a yield of 80%.

[0243] MS m / z (ESI): 313&315 [M+1];

[0244] 1H NMR (400MHz, CDCl3) δ 8.49 (s, 1H), 8.31 (s, 1H), 4.64 (s, 2H), 4.47 (t, J = 4.0Hz, 2H), 4.03 (t, J = 4.0Hz, 2H), 1.21 (s, 9H).

[0245] Step 3

[0246] 1-(9-(3-(1H-imidazol-1-yl)prop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine -4(5H)-yl)-2,2-dimethylpropan-1-one 14

[0247] Compound 1-(9-bromo-2,3-dihydropyrido[3,4-f][1,4]oxazepine -4(5H)-yl)-2,2-dimethylpropane-1-one 14-2 (30 mg, 0.10 mmol) was dissolved in N,N-diisopropylethylamine (3 mL) and N,N-dimethylacetamide (3 mL), and cuprous iodide (4 mg, 0.02 mmol), tetrakis(triphenylphosphine)palladium (23 mg, 0.02 mmol) and 1-(prop-2-yn-1-yl)-1H-imidazole (synthesis reference: Chem. Commun., (2012), 48(5), 675-677) (53 mg, 0.5 mmol) were added in sequence, and the mixture was reacted at 80°C for 8 hours under nitrogen protection. The reaction solution was cooled to room temperature, filtered through diatomaceous earth, diluted with water (10 mL), and extracted with ethyl acetate (10 mL×3). The organic phases were combined and washed with saturated brine (15 mL×2), dried over anhydrous sodium sulfate, filtered to remove the desiccant, and desolventized under reduced pressure to obtain a crude product. The target compound 1-(9-(3-(1H-imidazol-1-yl)prop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine was purified by TLC (dichloromethane:methanol=20:1). -4(5H)-yl)-2,2-dimethylpropan-1-one (18 mg), yield 48%.

[0248] MS m / z(ESI):339[M+1];

[0249] 11H NMR (400 MHz, CDCl3) δ 8.40 (s, 1H), 8.34 (s, 1H), 7.66 (s, 1H), 7.09 - 7.06 (m, 1H), 7.04 - 7.00 (m, 1H), 4.94 (s, 2H), 4.65 (s, 2H), 4.47 (t, J = 4.4 Hz, 2H), 4.03 (t, J = 4.4 Hz, 2H), 1.21 (s, 9H).

[0250] The synthesis procedures of Examples 15 to 19 refer to Example 14.

[0251] Example 15

[0252] 1-(9-(3-(1H-Imidazol-1-yl)prop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one

[0253]

[0254] MS m / z (ESI): 375 [M+1];

[0255] 1 1H NMR (400 MHz, CD3OD) δ 9.11 (s, 1H), 8.59 (s, 1H), 8.52 (s, 1H), 7.80 (s, 1H), 7.64 (s, 1H), 6.10 (t, J = 56.4 Hz, 1H), 4.88 (s, 2H), 4.47 (t, J = 4.8 Hz, 2H), 4.03 (t, J = 4.8 Hz, 2H), 3.30 (s, 2H), 1.33 (s, 6H).

[0256] Example 16

[0257] 1-(9-(3-(1H-1,2,4-Triazol-1-yl)prop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one

[0258]

[0259] MS m / z (ESI): 376 [M+1];

[0260] 11H NMR (400 MHz, CDCl3) δ 8.51 (s, 1H), 8.42 - 8.39 (m, 2H), 8.00 (s, 1H), 6.10 (t, J = 56.4 Hz, 1H), 5.25 (s, 2H), 4.74 (s, 2H), 4.51 - 4.39 (m, 2H), 4.11 - 4.01 (m, 2H), 1.36 (s, 6H).

[0261] Example 17

[0262] 1-(9-(3-(1H-Pyrazol-1-yl)prop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one

[0263]

[0264] MS m / z (ESI): 375 [M + 1];

[0265] 1 1H NMR (400 MHz, CDCl3) δ 8.52 (s, 1H), 8.39 (s, 1H), 7.70 - 7.64 (m, 1H), 7.61 - 7.52 (m, 1H), 6.39 - 6.28 (m, 1H), 6.10 (t, J = 56.4 Hz, 1H), 5.23 (s, 2H), 4.73 (s, 2H), 4.53 - 4.32 (m, 2H), 4.16 - 3.91 (m, 2H), 1.36 (s, 6H).

[0266] Example 18

[0267] 3,3-Difluoro-1-(9-(3-(4-fluoro-1H-imidazol-1-yl)prop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazin -4(5H)-yl)-2,2-dimethylpropan-1-one

[0268]

[0269] MS m / z (ESI): 393 [M + 1];

[0270] 11H NMR (400 MHz, CDCl3) δ 8.71 - 8.55 (broad, 2H), 7.22 (singlet, 1H), 6.56 (doublet, J = 8.0 Hz, 1H), 6.03 (triplet, J = 56.4 Hz, 1H), 4.92 - 4.86 (multiplet, 2H), 4.71 - 4.63 (multiplet, 2H), 4.39 (singlet, 2H), 4.02 - 3.98 (multiplet, 2H), 1.29 (singlet, 6H).

[0271] Example 19

[0272] 3-(4-Pivaloyl-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepin -9-yl)propiolamide

[0273]

[0274] MS m / z (ESI): 302 [M + 1];

[0275] 1 1H NMR (400 MHz, DMSO-d6) δ 8.45 (singlet, 2H), 8.16 (singlet, 1H), 7.71 (singlet, 1H), 4.47 (singlet, 2H), 4.03 (triplet, J = 4.4 Hz, 2H), 3.99 (triplet, J = 4.4 Hz, 2H), 1.17 (singlet, 9H).

[0276] Example 20

[0277] 1-(9-((1-(Difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-2-methoxy-2-methylpropan-1-one

[0278]

[0279] The first step

[0280] tert-Butyl 9-ethynyl-2,3-dihydropyrido[3,4-f][1,4]oxazepine-4(5H)-carboxylate 20 - 1

[0281] Compound 9-((Trimethylsilyl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine tert-Butyl -4(5H)-nicotinate 1a (1.20 g, 3.47 mmol) and potassium carbonate (1.92 g, 13.87 mmol) were added to methanol (20 mL), and the mixture was stirred at room temperature for 1 hour. The reaction mixture was diluted with 25 mL of water and extracted with ethyl acetate (25 mL × 3). The combined organic phases were washed with saturated brine (50 mL × 2), dried over anhydrous sodium sulfate. The desiccant was removed by filtration, and the solvent was evaporated under reduced pressure to obtain the crude product. The target product 9-ethynyl-2,3-dihydropyrido[3,4-f][1,4]oxazepine was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 4:1). tert-Butyl -4(5H)-nicotinate 20-1 (0.73 g), yield 76%.

[0282] MS m / z (ESI): 275 [M+1];

[0283] 1 1H NMR (400 MHz, CDCl3) δ 8.65 (s, 1H), 8.45 (s, 1H), 4.73 - 4.60 (m, 2H), 4.42 (s, 2H), 4.11 - 3.90 (m, 2H), 3.48 (s, 1H), 1.56 (s, 9H).

[0284] The second step

[0285] 9-((1-(Difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine tert-Butyl -4(5H)-nicotinate 20-2

[0286] Compound 9-ethynyl-2,3-dihydropyrido[3,4-f][1,4]oxazepine tert-Butyl -4(5H)-carboxylate 20-1 (0.73 g, 2.36 mmol) and 1-(difluoromethyl)-4-iodo-1H-pyrazole (0.65 g, 2.36 mmol) were dissolved in N,N-diisopropylethylamine (5 mL) and tetrahydrofuran (5 mL). Copper(I) iodide (59 mg, 0.30 mmol), tetrakis(triphenylphosphine)palladium (0.35 g, 0.30 mmol), and 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl (0.43 g, 0.9 mmol) were added successively. The reaction was carried out at 60 °C for 15 h under nitrogen protection. After the reaction was cooled to room temperature, it was quenched with 25 mL of water, extracted with ethyl acetate (25 mL × 3), the combined organic phases were washed with saturated brine (25 mL × 2), dried over anhydrous sodium sulfate, the desiccant was filtered off, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 4:1) to obtain the target product 9-((1-(difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine tert-Butyl -4(5H)-carboxylate 20-2 (0.60 g), yield 57%.

[0287] MS m / z (ESI): 391 [M + 1];

[0288] 1 1H NMR (400 MHz, CDCl3) δ 8.78 - 8.11 (br, 2H), 7.96 (s, 1H), 7.73 (s, 1H), 7.12 (t, J = 53.8 Hz, 1H), 4.55 - 4.48 (m, 2H), 4.36 - 4.17 (m, 2H), 3.93 - 3.75 (m, 2H), 1.35 (s, 9H);

[0289] The third step

[0290] 9-((1-(difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepine Trifluoroacetate 20-3

[0291] Compound 9-((1-(difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine tert-Butyl -4(5H)-carboxylate 20-2 (0.6 g, 1.54 mmol) was dissolved in trifluoroacetic acid (2 mL) and dichloromethane (2 mL), and stirred at room temperature for 1 h. The solvent was concentrated under reduced pressure to obtain the target compound 9-((1-(difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepine Trifluoroacetate 20-3 (440 mg, crude), used directly in the next step without purification. MS m / z (ESI): 291 [M+1];

[0292] The fourth step

[0293] 1-(9-((1-(Difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-2-methoxy-2-methylpropan-1-one 20

[0294] Compound 9-((1-(Difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepine Trifluoroacetate 20-3 (40 mg, 0.14 mmol) was dissolved in N,N-dimethylformamide (2 mL), and N,N-diisopropylethylamine (35 mg, 0.27 mmol), 2-methoxy-2-methylpropanoic acid (33 mg, 0.27 mmol) and 2-(7-azabenzotriazol)-N,N,N',N'-tetramethyluronium hexafluorophosphate (78 mg, 0.21 mmol) were added. The mixture was stirred at room temperature for 1 hour. 10 mL of water was added to quench the reaction mixture, and the mixture was extracted with ethyl acetate (10 mL × 3). The combined organic phases were washed with saturated brine (20 mL × 2), dried over anhydrous sodium sulfate, the desiccant was filtered off, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was purified by preparative liquid chromatography (acetonitrile: water (containing 0.1% formic acid) = 3:1) to obtain the target product 1-(9-((1-(difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-2-methoxy-2-methylpropan-1-one 20 (13 mg), yield 24%.

[0295] MS m / z (ESI): 391 [M+1];

[0296] 1 H NMR (400 MHz, CDCl3) δ 8.62 - 8.19 (br, 2H), 7.97 (s, 1H), 7.73 (s, 1H), 7.12 (t, J = 56.4 Hz, 1H), 5.11 (s, 1H), 4.65 (s, 1H), 4.58 - 4.23 (m, 3H), 3.98 (s, 1H), 3.12 - 2.93 (m, 3H), 1.39 (s, 3H), 1.35 (s, 3H).

[0297] The synthetic procedures of Examples 21 to 23 refer to the process of Example 20.

[0298] Example 21

[0299] 1-(9-((1-(Difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazin -4(5H)-yl)-2,2-dimethylpropan-1-one

[0300]

[0301] MS m / z(ESI): 375 [M+1];

[0302] 1 H NMR(400 MHz, CDCl3) δ 8.84 (s, 1H), 8.40 (s, 1H), 7.97 (s, 1H), 7.73 (s, 1H), 7.12 (t, J = 60.0 Hz, 1H), 4.67 (s, 2H), 4.42 - 4.30 (m, 2H), 4.05 - 4.00 (m, 2H), 1.20 (s, 9H).

[0303] Example 22

[0304] 1-(9-((1-(Difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazin -4(5H)-yl)-3-fluoro-2-(fluoromethyl)-2-methylpropan-1-one

[0305]

[0306] MS m / z(ESI): 411 [M+1];

[0307] 1 H NMR(400 MHz, CDCl3) δ 8.50 (m, 2H), 8.06 (s, 1H), 7.83 (s, 1H), 7.21 (t, J = 64 Hz, 1H), 4.79 (s, 2H), 4.78 - 4.56 (m, 3H), 4.56 - 4.47 (m, 3H), 4.16 - 4.09 (m, 2H), 1.39 (s, 3H).

[0308] Example 23

[0309] 4-(9-((1-(Difluoromethyl)-1H-pyrazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazin -4(5H)-yl)-3,3-dimethyl-4-oxobutanenitrile

[0310]

[0311] MS m / z (ESI): 400 [M+1];

[0312] 1 H NMR (400 MHz, CDCl3) δ 8.48 (s, 1H), 8.31 (s, 1H), 7.97 (s, 1H), 7.73 (s, 1H), 7.12 (t, J = 60.4 Hz, 1H), 4.69 (s, 2H), 4.48 - 4.34 (m, 2H), 4.11 - 3.99 (m, 2H), 2.58 (s, 2H), 1.41 (s, 6H).

[0313] Example 24

[0314] 1-(9-(3,3-Difluoroprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-2,2-dimethylpropan-1-one

[0315]

[0316] The first step

[0317] 9-(3-((tert-Butyldimethylsilyl)oxy)prop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine tert-Butyl 9-(3-((tert-butyldimethylsilyl)oxy)prop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine-4(5H)-carboxylate 24-1

[0318] Dissolve tert-butyl 9-bromo-2,3-dihydropyrido[3,4-f][1,4]oxazepine-4(5H)-carboxylate 1a (0.90 g, 2.74 mmol) and dimethyl tert-butyl(2-propynyloxy)silane (1.86 g, 10.91 mmol) in N,N-dimethylacetamide (6 mL) and N,N-diisopropylethylamine (6 mL). Add copper(I) iodide (0.14 g, 0.73 mmol) and tetrakis(triphenylphosphine)palladium (0.63 g, 0.55 mmol). The reaction mixture is purged with nitrogen twice and stirred at 80 °C for 8 hours. Cool the reaction mixture to room temperature and concentrate it under reduced pressure. Purify the crude product by silica gel column chromatography (petroleum ether:ethyl acetate = 8:1) to obtain tert-butyl 9-(3-((tert-butyldimethylsilyl)oxy)prop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine -4(5H)-carboxylate 24-1 (0.90 g), with a yield of 76%.

[0319] MS m / z (ESI): 419 [M+1];

[0320] The second step

[0321] 9-(3-Hydroxyprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-tert-Butyl carboxylate 24-2

[0322] Dissolve 9-(3-((tert-Butyldimethylsilyl)oxy)prop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-tert-Butyl carboxylate 24-1 (120 mg, 0.29 mmol) in tetrahydrofuran (4 mL), and dropwise add tetrabutylammonium fluoride (0.15 mL, 0.15 mmol, 1 M solution in tetrahydrofuran) under an ice-water bath, and react for 20 minutes under an ice-water bath. Add water (15 mL) to quench the reaction, extract with ethyl acetate (10 mL×3), combine the organic phases and wash with saturated brine (15 mL×2), dry over anhydrous sodium sulfate, filter to remove the desiccant, and concentrate under reduced pressure to obtain the crude product. Purify by preparative thin-layer chromatography (petroleum ether:ethyl acetate = 20:1) to obtain the target compound 9-(3-Hydroxyprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-tert-Butyl carboxylate 24-2 (70 mg), with a yield of 36%.

[0323] MS m / z (ESI): 305 [M+1];

[0324] 1 H NMR (400 MHz, CDCl3) δ 8.53 (s, 1H), 8.35 - 8.29 (m, 1H), 4.67 - 4.40 (m, 4H), 4.31 (t, J = 4.8 Hz, 2H), 3.86 (t, J = 4.8 Hz, 2H), 1.41 (s, 9H);

[0325] The third step

[0326] 9-(3-Oxoprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-tert-Butyl carboxylate 24-3

[0327] 9-(3-Hydroxyprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine tert-Butyl 9-(3-oxoprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine-4(5H)-carboxylate (0.28 g, 0.86 mmol) was dissolved in dry dichloromethane (10 mL). The reaction mixture was cooled to 0 °C in an ice-water bath, and Dess-Martin periodinane (0.73 g, 1.72 mmol) was added. The reaction was carried out at room temperature for 30 minutes. The reaction was quenched by adding saturated aqueous sodium bicarbonate solution (15 mL), and the mixture was extracted with dichloromethane (10 mL × 3). The combined organic phases were washed with saturated brine (15 mL × 2), dried over anhydrous sodium sulfate, filtered to remove the desiccant, and concentrated under reduced pressure to obtain the crude product. The crude product was purified by preparative thin-layer chromatography (petroleum ether:ethyl acetate = 20:1) to give the target compound 9-(3-oxoprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine tert-Butyl 9-(3-oxoprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine-4(5H)-carboxylate (120 mg), yield 46%.

[0328] MS m / z (ESI): 303 [M+1];

[0329] 1 H NMR (400 MHz, CDCl3) δ 9.46 (s, 1H), 8.61 (s, 1H), 8.35 - 8.29 (m, 1H), 4.56 - 4.43 (m, 2H), 4.28 (t, J = 4.8 Hz, 2H), 3.87 (t, J = 4.8 Hz, 2H), 1.42 (s, 9H);

[0330] The fourth step

[0331] 9-(3,3-Difluoroprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine tert-Butyl 9-(3,3-difluoroprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine-4(5H)-carboxylate

[0332] Under nitrogen protection, 9-(3-oxoprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine tert-Butyl 9-(3,3-difluoroprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine-4(5H)-carboxylate 24-3 (0.12 g, 0.40 mmol) was dissolved in dry dichloromethane (5 mL). The mixture was cooled to 0 °C, and diethylaminosulfur trifluoride (0.19 g, 1.20 mmol) diluted with dichloromethane (2 mL) was slowly added dropwise to the reaction solution. The reaction was carried out at 0 °C for 1 hour. The reaction was quenched with saturated aqueous sodium bicarbonate solution (10 mL), and extracted with dichloromethane (10 mL × 3). The combined organic phases were washed with saturated brine (15 mL × 2), dried over anhydrous sodium sulfate, filtered to remove the desiccant, and concentrated under reduced pressure to obtain the crude product. The crude product was purified by preparative thin-layer chromatography (petroleum ether:ethyl acetate = 20:1) to obtain the target compound 9-(3,3-difluoroprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine tert-Butyl 9-(3,3-difluoroprop-1-yn-1-yl)-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepine-4(5H)-carboxylate 24-4 (66 mg), yield 51%.

[0333] MS m / z (ESI): 325 [M + 1];

[0334] 1 1H NMR (400 MHz, CDCl3) δ 8.55 (s, 1H), 8.35 - 8.29 (m, 1H), 6.45 (t, J = 56.4 Hz, 1H), 4.56 - 4.43 (m, 2H), 4.28 (t, J = 4.8 Hz, 2H), 3.87 (t, J = 4.8 Hz, 2H), 1.42 (s, 9H);

[0335] Step 5

[0336] 9-(3,3-Difluoroprop-1-yn-1-yl)-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepine trifluoroacetate 24-5

[0337] 9-(3,3-Difluoroprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine tert-Butyl 9-(3,3-difluoroprop-1-yn-1-yl)-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepine-4(5H)-carboxylate 24-4 (0.13 g, 0.41 mmol) was dissolved in dichloromethane (5 mL), and trifluoroacetic acid (1.0 mL) was added at room temperature. The mixture was stirred at room temperature for 1 hour and concentrated under reduced pressure to dryness to obtain 9-(3,3-difluoroprop-1-yn-1-yl)-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepine trifluoroacetate 24-5 (0.10 g, crude product). MS m / z (ESI): 225 [M + 1];

[0338] Step 6

[0339] 1-(9-(3,3-difluoroprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-2,2-dimethylpropan-1-one 24

[0340] Dissolve 9-(3,3-difluoroprop-1-yn-1-yl)-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepine 24-5(6 mg, 0.03 mmol) in dichloromethane (2 mL). Add triethylamine (0.010 g, 0.08 mmol) and pivaloyl chloride (5 mg, 0.04 mmol) to the reaction solution, and stir at room temperature for 2 hours. Add water (10 mL) to the reaction solution, extract with dichloromethane (10 mL×3), wash the combined organic phases with saturated brine (20 mL×2), dry over anhydrous sodium sulfate, filter off the desiccant, and evaporate the solvent under reduced pressure to obtain the crude product. The target product 1-(9-(3,3-difluoroprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-2,2-dimethylpropan-1-one 24 (3 mg) was obtained by high performance liquid chromatography, with a yield of 31%.

[0341] MS m / z (ESI): 309 [M+1];

[0342] 1 1H NMR (400 MHz, CDCl3) δ 8.49 (s, 1H), 8.41 (s, 1H), 6.37 (t, J = 56.4 Hz, 1H), 4.67 (s, 2H), 4.38 (t, J = 4.8 Hz, 2H), 4.02 (t, J = 4.8 Hz, 2H), 1.21 (s, 9H).

[0343] The synthesis procedure of Example 25 refers to the process of Example 24.

[0344] Example 25

[0345] 1-(9-(3,3-difluoroprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one

[0346]

[0347] MS m / z (ESI): 345 [M+1];

[0348] 11H NMR (400 MHz, CDCl3) δ 8.54 (s, 1H), 8.44 (s, 1H), 6.44 (t, J = 56.4 Hz, 1H), 6.10 (t, J = 56.4 Hz, 1H), 4.74 (s, 2H), 4.56 (t, J = 4.8 Hz, 2H), 4.03 (t, J = 4.8 Hz, 2H), 1.36 (s, 6H).

[0349] Example 26

[0350] 3-(4-Pivaloyl-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazine -9-yl) propionitrile

[0351]

[0352] The first step

[0353] 9-(Cyanoethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-tert-butyl formate 26-1

[0354] Dissolve 9-(3-hydroxyprop-1-yn-1-yl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-tert-butyl formate 24-2 (40 mg, 0.13 mmol) in a mixed solution of acetonitrile and water (volume ratio 9:1, 3 mL). Sequentially add 2,2,6,6-tetramethylpiperidine oxide (4 mg, 0.03 mmol), ammonium acetate (41 mg, 0.53 mmol) and iodobenzene diacetate (92 mg, 0.29 mmol). React at room temperature for 0.5 hour, extract with ethyl acetate (10 mL × 3), combine the organic phases and wash with saturated brine (10 mL × 2), dry over anhydrous sodium sulfate, filter to remove the desiccant, and evaporate the solvent under reduced pressure to obtain the crude product. The crude product is purified by preparative thin-layer chromatography (petroleum ether:ethyl acetate = 20:1) to obtain the target product 9-(cyanoethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazine -4(5H)-tert-butyl formate 26-1 (26 mg), yield 67%.

[0355] MS m / z (ESI): 300 [M+1];

[0356] 11H NMR (400 MHz, CDCl3) δ 8.53 (s, 1H), 8.35 - 8.29 (m, 1H), 4.73 - 4.61 (m, 2H), 4.50 (t, J = 4.4 Hz, 2H), 4.12 (t, J = 4.4 Hz, 2H), 1.34 (s, 9H);

[0357] The second step

[0358] 3-(2,3,4,5-Tetrahydropyrido[3,4-f][1,4]oxazepin -9-yl)propiolonitrile 26-2

[0359] Dissolve 9-(cyanoethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine -4(5H)-carboxylic acid tert-butyl ester 26-1 (26 mg, 0.01 mmol) in dichloromethane (3 mL), and add trifluoroacetic acid (1 mL) under an ice bath. Stir at room temperature for 0.5 hour. After the reaction is completed, directly concentrate under reduced pressure. The obtained crude product is dissolved in dichloromethane and washed with saturated aqueous sodium bicarbonate solution (3 × 5 mL). The organic phase is dried over anhydrous sodium sulfate, filtered, and concentrated by rotary evaporation to obtain 3-(2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepin -9-yl)propiolonitrile 26-2 (30 mg, crude product). MS m / z (ESI): 200 [M+1];

[0360] The third step

[0361] 3-(4-Pivaloyl-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepin -9-yl)propiolonitrile 26

[0362] Dissolve 3-(2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepin -9-yl)propiolonitrile 26-2 (30 mg, 0.13 mmol) in dichloromethane (5 mL). Add triethylamine (0.07 g, 0.65 mmol) and pivaloyl chloride (0.02 g, 0.17 mmol) to the reaction solution and stir at room temperature for 2 hours. Add water (15 mL) to the reaction solution, extract with dichloromethane (20 mL × 3). The combined organic phases are washed with saturated brine (25 mL × 2), dried over anhydrous sodium sulfate, filtered to remove the desiccant, and concentrated under reduced pressure to obtain the crude product. The target product 3-(4-pivaloyl-2,3,4,5-tetrahydropyrido[3,4-f][1,4]oxazepin -9-yl)propiolonitrile 26 (30 mg) is obtained by high performance liquid chromatography, with a yield of 63%.

[0363] MS m / z (ESI): 284 [M+1];

[0364] 1 H NMR (400 MHz, CDCl3) δ 8.59 (s, 1H), 8.52 (s, 1H), 4.74 (s, 2H), 4.53 (t, J = 4.4 Hz, 2H), 4.16 (t, J = 4.4 Hz, 2H), 1.28 (s, 9H).

[0365] Example 27a, 27b and 27c

[0366] 1-(9-(((1-(Difluoromethyl)-1H-1,2,3-triazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 27a

[0367] 1-(9-((1-(Difluoromethyl)-1H-1,2,3-triazol-5-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 27b

[0368] 1-(9-((2-(Difluoromethyl)-2H-1,2,3-triazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 27c

[0369]

[0370] The first step

[0371] 3,3-Difluoro-2,2-dimethyl-1-(9-((1-((2-(Trimethylsilyl)ethoxy)methyl)-1H-1,2,3-triazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazin -4(5H)-yl)propan-1-one 27-1

[0372] 1-(9-Ethynyl-2,3-dihydropyrido[3,4-f][1,4]oxazin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one intermediate 1 (100 mg, 0.39 mmol) and 4-bromo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-1,2,3-triazole (272 mg, 0.98 mmol, synthesis reference: WO2021087127) were dissolved in tetrahydrofuran (2 mL) and N,N-diisopropylethylamine (2 mL). Copper(I) iodide (11.4 mg, 0.06 mmol), tetrakis(triphenylphosphine)palladium(0) (69.3 mg, 0.06 mmol) and 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl (57.2 mg, 0.12 mmol) were added. Under nitrogen protection, the reaction was carried out at 60 °C for 5 hours. After cooling to room temperature, most of the solvent was removed by concentration under reduced pressure. Ethyl acetate (10 mL) was added to dissolve the residue, and the organic phase was washed with saturated brine (5 mL × 3), dried over anhydrous sodium sulfate, filtered to remove the desiccant, and concentrated under reduced pressure to obtain the crude product. The crude product was purified by preparative thin-layer chromatography (petroleum ether / ethyl acetate = 1:1) to obtain the target product 3,3-difluoro-2,2-dimethyl-1-(9-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-1,2,3-triazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine -4(5H)-yl)propan-1-one 27-1 (100 mg), yield: 52%; MS m / z (ESI): 492 [M + 1];

[0373] The second step

[0374] 1-(9-((1H-1,2,3-triazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 27-2

[0375] 3,3-Difluoro-2,2-dimethyl-1-(9-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-1,2,3-triazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine -4(5H)-yl)propan-1-one 27-1 (100 mg, 0.20 mmol) was added to trifluoroacetic acid (1 mL) and dichloromethane (2 mL), and the mixture was stirred at room temperature for 1 hour. The solvent was removed by evaporation under reduced pressure to obtain the crude product. The residue was added to an aqueous sodium bicarbonate solution (5 mL), and the mixture was extracted with dichloromethane (5 mL × 3). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the desiccant, and the solvent was removed by evaporation under reduced pressure to obtain the target product 1-(9-((1H-1,2,3-triazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepine -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 27-2 (58.0 mg, crude), yield: 79%, the product was used directly in the next step without purification. MS m / z (ESI): 362 [M+1];

[0376] The third step

[0377] 1-(9-(((1-(difluoromethyl)-1H-1,2,3-triazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 27a

[0378] 1-(9-((1-(difluoromethyl)-1H-1,2,3-triazol-5-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 27b

[0379] 1-(9-((2-(difluoromethyl)-2H-1,2,3-triazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 27c

[0380] 1-(9-((1H-1,2,3-triazol-4-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 27-2 (58 mg, 0.16 mmol), cesium carbonate (104.3 mg, 0.32 mmol) and N,N-dimethylformamide (2 mL) were mixed, and diethyl bromofluoromethylphosphonate (64.1 mg, 0.24 mmol) was added dropwise. The mixture was stirred at room temperature for 1 hour. Cesium carbonate was removed by filtration, and the filtrate was purified by preparative liquid chromatography (acetonitrile / H2O = 20%-40%) to obtain three compounds. The first peak obtained (1.0 mg), yield: 2%.

[0381] MS m / z (ESI): 412 [M+1];

[0382] 11H NMR (400 MHz, CD3OD) δ 8.80 (s, 1H), 8.58 (s, 1H), 8.45 (s, 1H), 8.00 (t, J = 58.4 Hz, 1H), 6.13 (t, J = 56.4 Hz, 1H), 4.88 - 4.79 (m, 2H), 4.77 - 4.63 (m, 2H), 4.24 - 4.05 (m, 2H), 1.36 (s, 6H);

[0383] The second peak obtained (1.1 mg), yield: 2%.

[0384] MS m / z (ESI): 412 [M+1];

[0385] 1 1H NMR (400 MHz, CD3OD) δ 8.80 (s, 1H), 8.58 (s, 1H), 8.45 (s, 1H), 8.00 (t, J = 58.4 Hz, 1H), 6.13 (t, J = 56.4 Hz, 1H), 4.88–4.79 (m, 2H), 4.77–4.63 (m, 2H), 4.24–4.05 (m, 2H), 1.36 (s, 6H);

[0386] The third peak obtained (1.9 mg), yield: 3%.

[0387] MS m / z (ESI): 412 [M+1];

[0388] 1 1H NMR (400 MHz, CD3OD) δ 8.62 (s, 1H), 8.50 (s, 1H), 8.17 (s, 1H), 7.78 (t, J = 58.4 Hz, 1H), 6.13 (t, J = 56.4 Hz, 1H), 4.79 - 4.66 (m, 4H), 4.26 - 4.08 (m, 2H), 1.36 (s, 6H).

[0389] Example 28

[0390] 1-(9-((3-(Difluoromethyl)-1H-pyrazol-5-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin-4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one

[0391]

[0392] The first step

[0393] Methyl 3-bromo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole-5-carboxylate 28-1

[0394] Methyl 3-bromo-1H-pyrazole-5-carboxylate (4.00 g, 19.60 mmol) was dissolved in N,N-dimethylformamide (50.0 mL). At 0 °C, cesium carbonate (12.80 g, 39.30 mmol) and 2-(trimethylsilyl)ethoxymethyl chloride (6.50 g, 39.20 mmol) were added successively. After the addition, the reaction was carried out at room temperature for 3 hours. The mixture was poured into water (100 mL), and the aqueous phase was extracted with ethyl acetate (30 mL × 3). The combined organic phases were washed with saturated brine (50 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was filtered off, and the solvent was removed under reduced pressure to obtain the crude product. Purification by silica gel column chromatography (petroleum ether / ethyl acetate = 100:1 - 100:5) gave the target product methyl 3-bromo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole-5-carboxylate 28-1 (4.00 g). Yield: 61%.

[0395] MS m / z (ESI): 335&337 [M+1];

[0396] 1 H NMR (400 MHz, CDCl3) δ 6.92 (s, 1H), 5.83 (s, 2H), 3.93 (s, 3H), 3.69 - 3.61 (m, 2H), 0.95 - 0.87 (m, 2H), 0.00 (s, 9H);

[0397] The second step

[0398] (3-Bromo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-5-yl)methanol 28-2

[0399] Methyl 3-bromo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole-5-carboxylate 28-1 (0.60 g, 1.80 mmol) was dissolved in ethanol (10.0 mL). At 0 °C, anhydrous calcium chloride (0.40 g, 3.60 mmol) and sodium borohydride (0.14 g, 3.70 mmol) were added successively. After the addition, the reaction was carried out at room temperature for 12 hours. The mixture was poured into water (50 mL), and the aqueous phase was extracted with ethyl acetate (30 mL × 3). The combined organic phases were washed with saturated brine (30 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was filtered off, and the solvent was removed under reduced pressure to obtain the crude product (3-bromo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-5-yl)methanol 28-2 (0.59 g).

[0400] MS m / z (ESI): 307&309 [M+1];

[0401] 1 1H NMR (400 MHz, CDCl3) δ 6.33 (s, 1H), 5.48 (s, 2H), 4.69 (s, 2H), 3.64 - 3.53 (m, 2H), 0.95 - 0.83 (m, 2H), 0.00 (s, 9H);

[0402] The third step

[0403] 3 - Bromo - 1 - ((2 - (trimethylsilyl)ethoxy)methyl)-1H - pyrazole - 5 - carbaldehyde 28 - 3

[0404] Dissolve (3 - bromo - 1 - ((2 - (trimethylsilyl)ethoxy)methyl)-1H - pyrazol - 5 - yl)methanol 28 - 2 (0.60 g, 1.96 mmol) in dichloromethane (5.0 mL), add manganese dioxide (0.85 g, 9.77 mmol), and react at room temperature for 12 hours after addition. Filter the mixture to remove manganese dioxide, and remove the solvent under reduced pressure to obtain the crude product 3 - bromo - 1 - ((2 - (trimethylsilyl)ethoxy)methyl)-1H - pyrazole - 5 - carbaldehyde 28 - 3 (0.50 g).

[0405] MS m / z (ESI): 305&307 [M + 1];

[0406] 1 1H NMR (400 MHz, CDCl3) δ 9.92 (s, 1H), 6.99 (s, 1H), 5.78 (s, 2H), 3.69 - 3.58 (m, 2H), 0.96 - 0.86 (m, 2H), 0.00 (s, 9H);

[0407] The fourth step

[0408] 3 - Bromo - 5 - (difluoromethyl)-1 - ((2 - (trimethylsilyl)ethoxy)methyl)-1H - pyrazole 28 - 4

[0409] Dissolve 3 - bromo - 1 - ((2 - (trimethylsilyl)ethoxy)methyl)-1H - pyrazole - 5 - carbaldehyde 28 - 3 (0.50 g, 1.64 mmol) in dichloromethane (5.0 mL), add diethylaminosulfur trifluoride (0.53 g, 3.29 mmol) at - 78 °C under nitrogen protection, and react at room temperature for 12 hours after addition. Pour the mixture into saturated sodium bicarbonate aqueous solution (30 mL), extract the aqueous phase with ethyl acetate (10 mL × 3), combine the organic phases and wash with saturated brine (10 mL × 2). Dry the organic phase with anhydrous sodium sulfate, filter to remove the desiccant, and remove the solvent under reduced pressure to obtain the crude product, and obtain the target product 3 - bromo - 5 - (difluoromethyl)-1 - ((2 - (trimethylsilyl)ethoxy)methyl)-1H - pyrazole 28 - 4 (0.30 g) by preparative liquid phase.

[0410] MS m / z (ESI): 327 & 329 [M+1];

[0411] 1 H NMR (400 MHz, CDCl3) δ 6.86 (t, J = 54.4 Hz, 1H), 6.61 (s, 1H), 5.50 (s, 2H), 3.65 - 3.51 (m, 2H), 0.96 - 0.86 (m, 2H), 0.00 (s, 9H);

[0412] The fifth step

[0413] 3 - Bromo - 5 - (difluoromethyl) - 1H - pyrazole 28 - 5

[0414] Dissolve 3 - bromo - 5 - (difluoromethyl) - 1 - ((2 - (trimethylsilyl)ethoxy)methyl) - 1H - pyrazole 28 - 4 (0.10 g, 0.31 mmol) in dichloromethane (2.0 mL), add trifluoroacetic acid (2.0 mL), and react for 1 hour at room temperature after addition. Concentrate under reduced pressure to obtain the crude product 3 - bromo - 5 - (difluoromethyl) - 1H - pyrazole 28 - 5 (0.10 g, crude product).

[0415] MS m / z (ESI): 197 & 199 [M+1];

[0416] 1 H NMR (400 MHz, CDCl3) δ 6.71 (t, J = 54.8 Hz, 1H), 6.52 (s, 1H);

[0417] The sixth step

[0418] 1 - (9 - ((5 - (difluoromethyl) - 1H - pyrazol - 3 - yl)ethynyl) - 2,3 - dihydropyrido[3,4 - f][1,4]oxazine -4(5H) - yl) - 3,3 - difluoro - 2,2 - dimethylpropan - 1 - one 28

[0419] Dissolve 1 - (9 - ethynyl - 2,3 - dihydropyrido[3,4 - f][1,4]oxazine -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one (Intermediate 1) (50.0 mg, 0.17 mmol), 3-bromo-5-(difluoromethyl)-1H-pyrazole (100.0 mg, 0.307 mmol) were dissolved in N,N-dimethylformamide (2 mL) and N,N-diisopropylethylamine (2 mL). Copper(I) iodide (3.0 mg, 0.016 mmol), 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl (7 mg, 0.015 mmol) and tetrakis(triphenylphosphine)palladium(0) (12 mg, 0.01 mmol) were added, and the mixture was stirred at 100 °C overnight under nitrogen protection. The reaction was cooled to room temperature, and the mixture was poured into water (50 mL). The aqueous phase was extracted with ethyl acetate (15 mL × 3), and the combined organic phases were washed with saturated brine (30 mL × 2). The organic phase was dried over anhydrous sodium sulfate and the desiccant was removed by filtration. The obtained crude product was purified by preparative silica gel plate (2:1 = ethyl acetate:petroleum ether) to obtain the target compound 1-(9-((5-(difluoromethyl)-1H-pyrazol-3-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 28 (9.0 mg), yield 13%.

[0420] MS m / z (ESI): 411 [M+1];

[0421] 1 H NMR (400 MHz, CDCl3) δ 8.60 (s, 1H), 8.40 (s, 1H), 6.81 - 6.53 (m, 2H), 6.03 (t, J = 56.4 Hz, 1H), 4.70 (s, 2H), 4.45 (s, 2H), 4.02 (s, 2H), 3.42 (s, 6H).

[0422] Example 29

[0423] 1-(9-((4-(difluoromethyl)-1H-imidazol-2-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one

[0424]

[0425] The first step

[0426] 1-((2-(Trimethylsilyl)ethoxy)methyl)-1H-imidazole-4-carbaldehyde 29-1

[0427] Dissolve 1H-imidazole-4-carbaldehyde (0.96 g, 10.00 mmol) in dry tetrahydrofuran (15 mL). Under a nitrogen atmosphere in an ice bath, add sodium hydride (0.48 g, 11.00 mmol, 60% wt dispersed in mineral oil) portionwise. Stir at 0 °C for 20 minutes, and slowly add 2-(trimethylsilyl)ethoxymethyl chloride (1.80 g, 12.00 mmol) to the reaction solution. React at room temperature for 15 hours, quench the reaction with saturated aqueous ammonium chloride solution, and extract the organic phase with ethyl acetate (3 × 20 mL). Dry the organic phase over anhydrous sodium sulfate, filter, and concentrate by rotary evaporation. Purify by silica gel column chromatography (petroleum ether:ethyl acetate = 2:1) to obtain 1-((2-(trimethylsilyl)ethoxy)methyl)-1H-imidazole-4-carbaldehyde 29-1 (1.30 g), with a yield of 59%. MS m / z (ESI): 227 [M+1];

[0428] The second step

[0429] 2-Bromo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-imidazole-4-carbaldehyde 29-2

[0430] Dissolve 1-((2-(trimethylsilyl)ethoxy)methyl)-1H-imidazole-4-carbaldehyde 29-1 (1.30 g, 5.75 mmol) in carbon tetrachloride (30 mL). Add azobisisobutyronitrile (0.05 g, 0.29 mmol) and N-succinimide (1.13 g, 6.33 mmol) at room temperature, then heat to 85 °C in a nitrogen atmosphere and stir for 16 hours. Cool to room temperature, quench the reaction solution with saturated aqueous sodium sulfite solution and saturated aqueous sodium bicarbonate solution, extract the organic phase with dichloromethane (3 × 20 mL), dry the organic phase over anhydrous sodium sulfate, filter, and concentrate by rotary evaporation. Purify by silica gel column chromatography (petroleum ether:ethyl acetate = 4:1) to obtain 2-bromo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-imidazole-4-carbaldehyde 29-2 (0.41 g), with a yield of 23.3%. MS m / z (ESI): 305&307 [M+1];

[0431] The third step

[0432] 2-Bromo-4-(difluoromethyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-imidazole 29-3

[0433] 2-Bromo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-imidazole-4-carbaldehyde 29-2 (0.15 g, 0.50 mmol) was dissolved in dry dichloromethane (10 mL) under a nitrogen atmosphere. Diethylaminosulfur trifluoride (0.24 g, 1.50 mmol) was slowly added dropwise at -78 °C. Then the reaction mixture was stirred overnight at room temperature, quenched with saturated aqueous sodium bicarbonate, and the organic phase was extracted with dichloromethane and dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated under reduced pressure to obtain the crude product 2-bromo-4-(difluoromethyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-imidazole 29-3 (0.15 g). MS m / z (ESI): 327&329 [M+1];

[0434] The fourth step

[0435] 1-(9-((4-(Difluoromethyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-imidazol-2-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 29-4

[0436] 1-(9-Ethynyl-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one intermediate 1 (60 mg, 0.20 mmol) and 2-bromo-4-(difluoromethyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-imidazole 29-3 (70 mg, 0.20 mmol) were dissolved in dry 1,4-dioxane (2 mL) and N,N-diisopropylethylamine (2 mL). Tetrakis(triphenylphosphine)palladium (10 mg, 0.01 mmol), copper(I) iodide (2 mg, 0.01 mmol) and 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl (5 mg, 0.01 mmol) were added at room temperature. The reaction system was purged with nitrogen three times and then reacted at 100 °C for 15 hours. After cooling to room temperature, the reaction mixture was concentrated under reduced pressure and the residue was purified by silica gel column chromatography (dichloromethane:methanol = 25:1) to obtain 1-(9-((4-(difluoromethyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-imidazol-2-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 29-4 (0.02 g) with a yield of 19%. MS m / z (ESI): 541 [M+1];

[0437] The fifth step

[0438] 1-(9-((4-(Difluoromethyl)-1H-imidazol-2-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 29

[0439] Dissolve 1-(9-((4-(difluoromethyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-imidazol-2-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 29 (20 mg, 0.04 mmol) in dichloromethane (3 mL), and add trifluoroacetic acid (3 mL) under an ice bath. Stir at room temperature for 5 hours. After the reaction is completed, directly concentrate under reduced pressure. The obtained crude product is dissolved in dichloromethane, washed with saturated aqueous sodium bicarbonate solution (3×5 mL), the organic phase is dried over anhydrous sodium sulfate, filtered, and concentrated by rotary evaporation. Purify by silica gel column (dichloromethane:methanol = 20:1) to obtain 1-(9-((4-(difluoromethyl)-1H-imidazol-2-yl)ethynyl)-2,3-dihydropyrido[3,4-f][1,4]oxazepin -4(5H)-yl)-3,3-difluoro-2,2-dimethylpropan-1-one 29 (6 mg), with a yield of 37%.

[0440] MS m / z(ESI): 411 [M+1].

[0441] 1 1H NMR (400 MHz, CDCl3) δ 8.60 - 8.24 (m, 2H), 7.32 - 7.25 (m, 1H), 6.77 - 6.49 (m, 1H), 6.15 - 5.87 (m, 1H), 4.81 - 4.68 (m, 2H), 4.60 - 4.46 (m, 2H), 4.08 - 3.98 (m, 2H), 1.29 (s, 6H).

[0442] Biological experiments

[0443] Example 30: Detection of RIPK1 kinase inhibitory activity

[0444] Experimental method: This experiment uses the homogeneous ADP-Glo kinase activity detection method to test the inhibitory effect of the compound on RIPK1 kinase activity, and obtain the half inhibitory concentration IC of the compound on RIPK1 kinase activity 50。First, dilute the enzyme reaction buffer. Dilute the enzyme reaction buffer stock solution (5x) in the ADP-Glo kinase assay kit purchased from Promega with deionized water, and add DTT at a final concentration of 1 mM and MnCl2 at a final concentration of 5 mM. The compound is serially diluted 4-fold with DMSO and then diluted 40-fold with the enzyme reaction buffer; the recombinant human RIPK1 protein is purchased from SignalChem, and the recombinant monkey RIPK1 protein is purchased from Abcam. Add 4 μL of the compound solution and 2 μL of the RIPK1 kinase solution diluted with the reaction buffer to a 384-well plate. The final concentrations of human and monkey RIPK1 are 1 ng / μL and 4 ng / mL, respectively. After mixing evenly, incubate at room temperature for 30 minutes; then add 2 μL of the ATP solution diluted with the reaction buffer, incubate at room temperature for 2 hours, add 5 μL of ADP-Glo to each well, continue to incubate at room temperature for 40 minutes, and finally add 10 μL of the Kinase detetion detection reagent in the kit to each well, continue to incubate at room temperature for 30 minutes, and detect the chemiluminescence signal of each well with an Envision multimode microplate reader (Perkin Elmer, Waltham, MA).

[0445] Example 31 Experiment on TNF-α-induced p-RIPK1 activation in different species

[0446] Test method: In this experiment, the ultrasensitive electrochemiluminescence method of Meso Scale Discovery (MSD) was used to test the inhibitory effect of the compound on the p-RIPK1 level, and the half-maximal inhibitory concentration IC 50 。

[0447] Peripheral blood mononuclear cells (PBMCs) from healthy humans and cynomolgus monkeys were purchased from Shanghai Aoneng Biotechnology Co., Ltd. Bone marrow cells (BMDMs) from rats and mice were isolated from the femurs and tibias of rats or mice. The compound was serially diluted 5-fold to 8 concentration points with the highest final concentration of 5000 nM, and added to a culture plate containing 0.5 - 1×10 5 cells. The cells were derived from PBMCs or BMDMs, and then TNF-α, Z-VAD-FMK, and SM-164 were added respectively. The cells were co-incubated in an incubator at 37 °C and 5% CO2 for 3 hours, and DMSO treatment was used as a control. Then, the cells were washed with PBS and cell lysate was added, and the cells were lysed on ice for 30 minutes. After that, the cells were transferred to a centrifuge tube and centrifuged at high speed at 12,000 rpm for 10 minutes. After centrifugation, the cell lysate was collected, and protein quantification was performed using the BCA method.

[0448] The level of p-RIPK in the cell lysate was measured using the Meso Scale Discovery (MSD) method. The biotinylated RIPK1 capture antibody was diluted in PBS and added to a 96-well streptavidin plate from MSD, and incubated overnight at 4°C. It was blocked with the blocking solution from MSD for 2 hours, and then incubated with 20 - 40 μL of the lysate at room temperature for 2 hours. The Phospho-RIPK1 antibody (CellSignaling #31122) was diluted in the blocking solution and added as the detection antibody, and incubated at room temperature for 1 hour; the SULFO-labeled goat anti-rabbit antibody (MSD, R32AB-1) was diluted 500 - 1000 times in the blocking solution, and continued to be incubated at room temperature for 45 minutes. Then, 100 μL of 2-fold diluted detection buffer (R92TC-3) was added, and after incubation, the electrochemical signal was read on the MSD Meso Sector imager S600.

[0449] Table 1: Detection results of the inhibitory effect on RIPK1 enzyme

[0450]

[0451] (The reference compound is Example 29 in Patent US11203600B2)

[0452] The structure of the compound in Example 29 of Patent US11203600B2 is as follows:

[0453]

[0454] The data in Table 1 show that Compound 1 and Compound 7 in this application have significantly better inhibitory effects on the RIPK1 enzyme than Example 29 (US11203600B2).

[0455] Example 32: Necroptosis of I2.1 cells

[0456] I2.1 cells are Jurkat cells with FADD gene knockout and are suitable for studying the necroptosis of cells under TNF-α induction conditions.

[0457] Experimental method: I2.1 cells (purchased from ATCC, catalog number CRL-2572; cultured in a cell incubator at 37°C with 5% CO2 in 1640 medium, seeded on a 384-well white plate, the compound was serially diluted in a 4-fold gradient, and the final starting concentration was 500 nM. 40 ng / mL of TNF-α and the test drug were co-incubated with the cells for 24 hours, where Example 29 (US11203600B2) was used as the positive control, and the control wells were treated with DMSO (D5879-500ML, purchased from Sigma).

[0458] The viability of the cells was determined using the Cell-TiterGlo kit based on the change in ATP content. For the determination of cell viability, the value represents the percentage of cell viability in the drug-treated wells compared to the control wells, where a higher value indicates stronger cell viability (drug action viability calculation = drug-treated wells / control wells * 100%).

[0459] Example 3 Programmed necrosis of HT-29 cells

[0460] Experimental method: HT-29 cells (purchased from Nanjing Kebai Biotechnology Co., Ltd., catalog number CBP30001L) were cultured in a cell incubator at 37 °C with 5% CO2 in 1640 medium. The cells were seeded on a 384-well white plate. The compound was serially diluted in a 4-fold gradient, with a final starting concentration of 500 nM. 100 ng / mL TNF-α, Q-VD-Oph (10 μM), SM-164 (1 μM), and the test drug were co-incubated with the cells for 48 hours. Example 29 (US11203600B2) was used as a positive control, and the control wells were treated with DMSO.

[0461] The viability of the cells was determined using the Cell-TiterGlo kit based on the viability assay of ATP. For the determination of cell viability, the value represents the percentage of cell viability in the drug-treated wells compared to the control wells, where a higher value indicates stronger cell viability (drug action viability calculation = drug-treated wells / control wells * 100%).

[0462] Example 34 Programmed necrosis of L-929 cells

[0463] Experimental method: L-929 cells (purchased from Nanjing Kebai Biotechnology Co., Ltd., catalog number CBP60878) were cultured in a cell incubator at 37 °C with 5% CO2 in MEM medium. The cells were seeded on a 384-well white plate. The compound was serially diluted in a 4-fold gradient, with a final starting concentration of 10,000 nM. 40 ng / mL TNF-α, Q-VD-Oph (10 μM), and the test drug were co-incubated with the cells for 24 hours. Example 29 was used as a positive control, and the control wells were treated with DMSO. The viability of the cells was determined using the Cell-TiterGlo kit based on the viability assay of ATP. For the determination of cell viability, the value represents the percentage of cell viability in the drug-treated wells compared to the control wells, where a higher value indicates stronger cell viability (drug action viability calculation = drug-treated wells / control wells * 100%).

[0464] Table 2: Detection results of the cell activity of the compounds of the present invention

[0465]

[0466]

[0467] The data in Table 2 indicate that some compounds of this application still maintain inhibitory activity in L929 cells of rodents, and their activities against HT29 and I2.1 are significantly better than those of Example 29 (US11203600B2).

[0468] Example 35: Pharmacokinetic study in SD rats and determination of blood-brain barrier permeability

[0469] The contents of plasma, brain tissue and CSF samples were measured by LC-MS / MS to obtain the ratio K of CSF concentration to plasma sample concentration p,CSF , the ratio K of CSF concentration to free plasma sample concentration puu,CSF , the ratio K of brain tissue to plasma exposure p,Brain and the ratio K of free brain tissue to free plasma exposure puu,Brain , so as to quickly screen out compounds with blood-brain penetrability. At the same time, Phoenix WinNolin was used to calculate the pharmacokinetic parameters of plasma to screen out compounds with better pharmacokinetic properties.

[0470] Experimental conditions and procedures:

[0471] 1. Drug preparation

[0472] Weigh an appropriate amount of the test substance into a weighing bottle, and sequentially add 5% DMA / 10% Solutol / 85% Saline, and stir at room temperature until a homogeneous solution is obtained for gavage administration.

[0473] 2. Drug administration

[0474] Prepare 24 male Sprague-Dawley (SD) rats in each group (N = 3 / time point), weigh them. The administration dose is 5 mg / kg, and the administration volume is 10 mL / kg. The rats are given gavage administration. Start timing after administration, and collect plasma and brain tissue at 0.25 h, 0.5 h, 1 h, 2 h, 3 h, 5 h, 8 h and 24 h, and collect cerebrospinal fluid CSF at 1 h and 5 h.

[0475] 3. LC-MS / MS method

[0476] According to the exact molecular weight of the compound, find the corresponding parent ion and fragment ion, and optimize to find suitable mass spectrometry conditions and liquid phase methods.

[0477] 4. Preparation of standard curve and quality control samples

[0478] Preparation of the working solution: Weigh an appropriate amount of the test sample accurately, dissolve it in an appropriate amount of DMSO, and prepare a standard stock solution with a concentration of 2 mg / mL. Using the standard stock solution, add acetonitrile-water (50:50, v / v) to prepare a series of concentration test sample standard curves and quality control working solutions.

[0479] Plasma standard curve Preparation of plasma samples and quality control plasma samples: Take 20 μL of blank plasma from SD rats, add 2 μL of a series of concentration test sample standard curves and quality control working solutions to obtain standard curve plasma samples and quality control plasma samples.

[0480] Brain tissue homogenate standard curve Preparation of brain tissue samples and quality control brain tissue samples: Take 20 μL of blank brain tissue homogenate from SD rats, add 2 μL of a series of concentration test sample standard curves and quality control working solutions to obtain standard curve brain tissue samples and quality control brain tissue samples.

[0481] Standard curve Preparation of CSF samples and quality control CSF samples: Take 5 μL of blank CSF from SD rats, add 5 μL of test sample standard curves and quality control working solutions to obtain standard curve CSF samples and quality control CSF samples.

[0482] 5. Sample analysis process

[0483] Plasma: Take 20 μL of plasma sample, add 2 μL of acetonitrile-water (50:50, v / v), place it in a 1.1 mL high tube, add 200 μL of internal standard solution (5 ng / mL terfenadine), vortex for 1 minute, centrifuge at 3000 rpm for 10 minutes at 4 °C, take the supernatant, dilute it with water, vortex and mix well, and then inject for analysis. When measuring the sample concentration, process the standard curve samples and quality control samples in the same way.

[0484] Brain tissue samples: Take a part of the tissue sample and place it in a 2 mL centrifuge tube, weigh and record the weight, add pure water at a ratio of 1:3 (g:mL), homogenize at 1800 rpm for 10 minutes. Take 20 μL of the tissue homogenate sample and place it in a 1.1 mL high tube, add 200 μL of internal standard solution (5 ng / mL terfenadine), vortex for 1 minute, centrifuge at 3000 rpm for 20 minutes at 4 °C, take the supernatant, dilute it with water, vortex and mix well, and then inject for analysis. When measuring the sample concentration, process the standard curve samples and quality control samples in the same way as the samples.

[0485] Cerebrospinal fluid CSF: Take 5 μL of CSF sample, add 5 μL of acetonitrile-water (50:50, v / v), place it in a 1.1 mL high tube, add 200 μL of internal standard solution (5 ng / mL terfenadine), vortex for 1 minute, centrifuge at 3000 rpm for 10 minutes at 4 °C, take the supernatant, dilute it with water, vortex and mix well, and then inject for analysis. When measuring the sample concentration, process the standard curve samples and quality control samples in the same way.

[0486] 6. Sample detection

[0487] Detect the sample using a suitable LC-MS / MS method.

[0488] 7. Data processing

[0489] The main computerized systems used in this experiment:

[0490] Microsoft Office Excel: 2010, for entering, calculating, and statistically analyzing data;

[0491] Analysis 1.6.3, for data acquisition and processing;

[0492] Phoenix WinNolin 8.3.1, for calculating pharmacokinetic parameters

[0493] GraphPad Prism 6.0, for drug concentration-time curves

[0494] Table 3: Pharmacokinetic results of the RIPK1 inhibitor of the present invention in SD rats

[0495]

[0496] It can be seen from the pharmacokinetic data in Table 3 that Compound 1 has good pharmacokinetic properties.

[0497] Table 4: Results of brain permeability determination of the RIPK1 inhibitor of the present disclosure in SD rats

[0498]

[0499] It can be seen from the data in Table 4 that Compound 1 of the present application shows good blood-brain barrier penetration and cerebrospinal fluid distribution and can be used to treat central nervous system diseases.

[0500] Pharmacokinetic study of Example 36 in beagle dogs

[0501] Determine the concentration of the compound in beagle dog plasma by LC-MS / MS, and use Phoenix WinNolin to calculate the pharmacokinetic parameters of the plasma to screen out compounds with better pharmacokinetic properties.

[0502] Experimental conditions and procedures:

[0503] 1. Drug preparation

[0504] Weigh an appropriate amount of the test substance into a weighing bottle, and successively add 10% DMA / 10% PEG400 / 80% saline. Stir at room temperature until a homogeneous solution is obtained for intravenous administration. Weigh an appropriate amount of the test substance into a weighing bottle, and successively add PEG-400 / 0.5% MC (v / v, 4 / 6). Stir at room temperature until a homogeneous solution is obtained for gavage administration.

[0505] 2. Administration

[0506] Prepare 3 male beagle dogs in each group and weigh them. The administration dose is 1 mg / kg, and the administration volume is 1 mL / kg. Administer the drug intravenously to the beagle dogs. Start timing after administration. Collect plasma before administration, at 0.083 h, 0.25 h, 1 h, 2 h, 4 h, 8 h, and 24 h.

[0507] Prepare 3 male beagle dogs in each group and weigh them. The administration dose is 3 mg / kg, and the administration volume is 5 mL / kg. Administer the drug by gavage to the beagle dogs. Start timing after administration. Collect plasma before administration, at 0.25 h, 0.5 h, 1 h, 2 h, 4 h, 8 h, and 24 h.

[0508] 3. LC-MS / MS method

[0509] Based on the exact molecular weight of the compound, find the corresponding parent ion and fragment ion, and optimize to find suitable mass spectrometry conditions and liquid phase methods.

[0510] 4. Preparation of standard curve and quality control samples

[0511] Preparation of working solution: Accurately weigh an appropriate amount of the test substance, dissolve it in an appropriate amount of DMSO, and prepare a standard stock solution with a concentration of 2 mg / mL. Use the standard stock solution and add acetonitrile water (50:50, v / v) to prepare a series of concentration test substance standard curves and quality control working solutions.

[0512] Preparation of plasma standard curve plasma samples and quality control plasma samples: Take 20 μL of beagle dog blank plasma, add 2 μL of a series of concentration test substance standard curve and quality control working solutions to obtain standard curve plasma samples and quality control plasma samples.

[0513] 5. Sample analysis process

[0514] Plasma: Take 20 μL of plasma sample, add 2 μL of acetonitrile water (50:50, v / v), place it in a 1.1 mL high tube, add 200 μL of internal standard solution (5 ng / mL terfenadine), vortex for 1 minute, centrifuge at 3000 rpm for 10 minutes at 4°C, take the supernatant, dilute it with water, vortex and mix well, and then inject for analysis. When determining the sample concentration, process the standard curve samples and quality control samples in the same way.

[0515] 6. Sample detection

[0516] Detect the sample using a suitable LC-MS / MS method.

[0517] 7. Data processing

[0518] The main computerized systems used in this experiment:

[0519] Microsoft Office Excel: 2010, for entering, calculating, and statistically analyzing data;

[0520] Analysis 1.6.3, for data acquisition and processing;

[0521] Phoenix WinNolin 8.3.1, for calculating pharmacokinetic parameters

[0522] GraphPad Prism 6.0, for drug concentration-time curves

[0523] Table 5: Pharmacokinetic results of the RIPK1 inhibitor of the present invention in beagle dogs

[0524]

[0525] From the pharmacokinetic data in Table 5, it can be seen that in beagle dogs, all the pharmacokinetic properties of Compound 1 are far superior to those of Example 29 (US11203600B2).

[0526] Determination of plasma protein binding rate in Example 37

[0527] Use a 96-well equilibrium dialysis device (RED Device Inserts) to determine protein binding in plasma. By measuring the degree of binding of the test article to plasma proteins of different species, the binding rates of the test article to plasma proteins of different species at incubation concentrations of 0.100, 0.500, and 2.50 μM after 5 hours of incubation were investigated.

[0528] Experimental conditions and procedures:

[0529] 1. Preparation of reaction solutions for various species

[0530] Accurately weigh a certain amount of the test article and the reference substance (warfarin), and prepare a stock solution with a certain concentration using DMSO. Dilute the stock solutions of the test article and the reference substance with DMSO to obtain working solutions with concentrations of 250 μM, 50 μM, and 10 μM, respectively. Take 5 μL of the test solution and mix it with 495 μL of blank plasma of various species. The concentrations of the test article and the reference substance in the resulting plasma are 2.50 μM, 0.500 μM, and 0.100 μM, respectively.

[0531] 2. Determination of plasma protein binding rate

[0532] Add 200 μL of plasma reaction solutions of various species into the sample chamber (side A) of the equilibrium dialysis tube, and add 350 μL of 100 mM phosphate buffer (pH 7.4) into the dialysis solution chamber (side B) of the equilibrium dialysis tube, in duplicate (n = 2). Then place the base plate into a constant temperature shaking incubator at 37 °C and incubate with horizontal shaking at 140 rpm. After 5 hours, take 20 μL of samples from the sample chamber (side A) and the buffer chamber (side B) respectively.

[0533] Add 20 μL of phosphate buffer to the 20 μL of sample taken from the sample chamber (side A), vortex at 1000 rpm for 1 minute, and then add 400 μL of termination solution (10 ng / mL terfenadine) to the sample on side A.

[0534] Add 20 μL of plasma of the corresponding species to the 20 μL of sample taken from the dialysis solution chamber (side B). Vortex at 1000 rpm for 1 minute, and then add 400 μL of termination solution (10 ng / mL terfenadine) to the samples on both sides of B.

[0535] After refrigerating at 4 °C for 30 minutes, centrifuge at 3000 rpm for 15 minutes at 4 °C. After centrifugation, take the supernatant, dilute it with water, vortex and mix, and then perform LC-MS / MS detection.

[0536] 3. LC-MS / MS Method

[0537] According to the exact molecular weight of the compound, find the corresponding parent ion and fragment ions, and optimize to find suitable mass spectrometry conditions and liquid phase methods.

[0538] 4. Data Analysis

[0539] The plasma protein binding rate is calculated by the following formula:

[0540]

[0541]

[0542] where C D and C R are the peak area ratios of the test substance and the internal standard measured in the supply chamber and the receiving chamber after incubation, respectively, and f b and f u are the protein binding rate and the free percentage, respectively.

[0543] The recovery rate is calculated by the following formula:

[0544]

[0545] where C I is the peak area ratio of the test substance and the internal standard measured in the 0 hr sample.

[0546] Table 6: Results of determination of plasma protein binding rate of the RIPK1 inhibitor of the present invention

[0547]

[0548] As can be seen from Table 6, 12.8% - 20.6% of Compound 1 of the present invention application is in the free state in plasma of various species. Combining the data in Table 3 and Table 4, it can be considered that Compound 1 has good brain permeability and can be used for the treatment of central nervous system diseases.

[0549] Based on the above experiments, the HT-29 cell activity of Compound 1 of the present invention application is 4.9 times that of reference compound Example 29 (US11203600B2), and the I2.1 cell activity is 8.0 times that of Example 29 (US11203600B2). The exposure amount of Compound 1 of the present invention application at the same dose in the PO experiment of SD rats is close to that of reference compound Example 29 (US11203600B2), and the exposure amount at the same dose in the PO experiment of beagle dogs is 15.6 times that of reference compound Example 29 (US11203600B2). It is expected that in the treatment of various diseases mediated by RIPK1, the efficacy of Compound 1 of the present application is better than that of reference compound Example 29 (US11203600B2).

[0550] From the above experimental results, it can be seen that the example compounds of the present invention can effectively inhibit the activity of RIPK1, have small species differences, and have good plasma protein binding rate and brain permeability. They can be used for the treatment or prevention of related diseases or disorders mediated by RIPK1, especially central nervous system diseases, such as rheumatoid arthritis, ulcerative colitis, psoriasis, Alzheimer's disease, multiple sclerosis, amyotrophic lateral sclerosis, etc.

Claims

1. A compound of formula (I), its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, wherein: L is a single bond or -(CH2) n -; A is a 6- to 8-membered saturated heterocycle containing 1 to 3 heteroatoms selected from N, O, and S; R 1 Selected from cyano, -C(O)NH2, halo C1-C8 alkyl, C6-C10 aryl, 5-10 membered heteroaryl, C3-C8 cycloalkyl, 4-8 membered heterocyclic group, wherein the C6-C10 aryl, 5-10 membered heteroaryl, C3-C8 cycloalkyl, 4-8 membered heterocyclic group are unsubstituted or substituted by one or more R x wherein the R x are each independently selected from halogen, cyano, C1-C6 alkyl, C3-C6 cycloalkyl, 4-8 membered saturated heterocyclic group, wherein the C1-C6 alkyl, C3-C6 cycloalkyl are unsubstituted or substituted by one or more halogen or cyano; R 2 、R 3 、R 4 are each independently selected from hydrogen, C1-C8 alkyl, C1-C8 alkoxy, wherein the C1-C8 alkyl and C1-C8 alkoxy are unsubstituted or substituted by one or more halogens or cyano groups; n is an integer selected from 1 to 5.

2. The compound according to claim 1, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, which has the formula (II): wherein: L is a single bond or -(CH2) n -; R 1 Selected from cyano, -C(O)NH2, halogenated C1-C8 alkyl, C6-C10 aryl, 5-10 membered heteroaryl, C3-C8 cycloalkyl, 4-8 membered heterocyclic group, wherein the C6-C10 aryl, 5-10 membered heteroaryl, C3-C8 cycloalkyl, 4-8 membered heterocyclic group are unsubstituted or substituted by one or more R x wherein the R x each independently selected from halogen, cyano, C1-C6 alkyl, C3-C6 cycloalkyl, 4-8 membered saturated heterocyclic group, wherein the C1-C6 alkyl, C3-C6 cycloalkyl are unsubstituted or substituted by one or more halogen or cyano; R 2 、R 3 、R 4 are each independently selected from hydrogen, C1-C8 alkyl, C1-C8 alkoxy, wherein the C1-C8 alkyl and C1-C8 alkoxy are unsubstituted or substituted by one or more halogens or cyano groups; n is an integer selected from 1 to 3.

3. The compound according to claim 2, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, wherein: L is a single bond or -CH2-; R 1 Selected from cyano, -C(O)NH2, halogenated C1-C6 alkyl, C6-C10 aryl, 5-8 membered heteroaryl containing 1-3 heteroatoms selected from N, O and S, C3-C8 cycloalkyl, 4-8 membered saturated heterocyclic group containing 1-2 heteroatoms selected from N and O, wherein the C6-C10 aryl, 5-8 membered heteroaryl, C3-C8 cycloalkyl, 4-8 membered saturated heterocyclic group are unsubstituted or substituted by 1-3 R x wherein the R x are each independently selected from fluorine, chlorine, bromine, cyano, C1-C4 alkyl, C3-C6 cycloalkyl, 4-6 membered saturated heterocyclic group; wherein the C1-C4 alkyl, C3-C6 cycloalkyl are unsubstituted or substituted by 1-3 substituents selected from fluorine, chlorine, bromine, cyano; R 2 、R 3 、R 4 are each independently selected from hydrogen, C1-C8 alkyl, C1-C8 alkoxy, wherein the C1-C8 alkyl and C1-C8 alkoxy are unsubstituted or substituted by 1-3 halogen atoms or cyano groups.

4. The compound according to claim 2, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, wherein: L is a single bond or -CH2-; R 1 selected from cyano, -C(O)NH2, halo C1-C4 alkyl, phenyl, 5-6 membered heteroaryl containing 1-3 heteroatoms selected from N, O and S, C3-C6 cycloalkyl, 4-6 membered saturated heterocyclic group containing 1 heteroatom selected from N and O, wherein the phenyl, 5-6 membered heteroaryl, C3-C6 cycloalkyl, 4-6 membered saturated heterocyclic group are unsubstituted or substituted by 1-3 R x wherein the R x are each independently selected from fluorine, chlorine, cyano, C1-C4 alkyl, C3-C6 cycloalkyl, 4-6 membered saturated heterocyclic group containing 1 heteroatom selected from N and O; wherein the C1-C4 alkyl, C3-C6 cycloalkyl are unsubstituted or substituted by 1-3 substituents selected from fluorine and cyano; R 2 、R 3 、R 4 are each independently selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, wherein the C1-C6 alkyl and C1-C6 alkoxy are unsubstituted or substituted with 1-3 fluorine, chlorine, bromine or cyano groups.

5. The compound according to claim 2, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, wherein: L is a single bond or -CH2-; R 1 Selected from cyano, -C(O)NH2, C1-C4 alkyl substituted with 1-3 fluorines, 5-membered heteroaryl containing 1-3 heteroatoms selected from N, O and S, 4-6 membered saturated heterocyclic group containing 1 O heteroatom, wherein the 5-membered heteroaryl and 4-6 membered saturated heterocyclic group are unsubstituted or substituted with 1-3 R x wherein the R x each independently selected from fluorine, cyano, methyl, ethyl, C3-C6 cycloalkyl, 4-6 membered saturated heterocyclic group containing 1 O heteroatom; wherein the methyl, ethyl, C3-C6 cycloalkyl are unsubstituted or substituted with 1-3 substituents selected from fluorine and cyano; R 2 、R 3 、R 4 Each independently selected from C1-C4 alkyl and C1-C4 alkoxy, wherein the C1-C4 alkyl is unsubstituted or substituted with 1-3 substituents selected from fluorine and cyano.

6. The compound according to claim 2, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, wherein: L is a single bond or -CH2-; R 1 selected from cyano, -C(O)NH2, R 2 、R 3 、R 4 Each independently selected from methyl, methoxy, monofluoromethyl, difluoromethyl, cyanomethyl.

7. A compound, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, which are selected from:

8. A pharmaceutical composition comprising the compound according to any one of claims 1-7, its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, and a pharmaceutically acceptable carrier, diluent or excipient.

9. Use of the compound according to any one of claims 1-7 or its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts or the pharmaceutical composition according to claim 8 in the preparation of a drug for use as a RIPK1 inhibitor.

10. Use of the compound according to any one of claims 1-7 or its isomers, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts or the pharmaceutical composition according to claim 8 in the preparation of a drug for the treatment or prevention of RIPK1-mediated related diseases or disorders.

11. The use according to claim 10, wherein the related diseases or disorders are selected from Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis, acute neurological diseases, inflammatory bowel disease, Crohn's disease, ulcerative colitis, psoriasis, atopic dermatitis, rheumatoid arthritis, spondyloarthritis, gout, systemic lupus erythematosus, non-alcoholic steatohepatitis, alcoholic steatohepatitis, autoimmune hepatitis, autoimmune hepatobiliary diseases, systemic inflammatory response syndrome, cerebrovascular accident, Huntington's disease, pancreatic cancer, bacterial infections, hematological malignancies, solid organ malignancies.

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