A class of nitrogen-containing fused ring STING modulator compounds, preparation methods and uses

Abstract

The present invention discloses a nitrogen-containing fused ring compound represented by general formula I, or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, solvate, polymorph or prodrug thereof, a preparation method thereof and a pharmaceutical application thereof, wherein the definitions of each group are as described in the specification.

Description

Technical Field

[0001] The present invention belongs to the field of medicinal chemistry, and specifically relates to a class of nitrogen-containing fused ring STING modulator compounds, preparation methods and uses. Background Art

[0002] Stimulator of interferon genes (STING) is a transmembrane protein that usually forms a dimer at the 152-173 region (dimerization domain) and is in a self-inhibited state. STING is an important component of the human innate immune system and is the first line of defense against the invasion of external pathogens such as bacteria and viruses. It plays an important role in maintaining the body's dynamic balance, resisting external infections, and preventing tumors and autoimmune diseases.

[0003] Studies have shown that the absence of STING activity can trigger tumors or some specific viral infections. For example, the protease NS2B3 of dengue virus can hinder the production of IFN-α / β by degrading STING. Therefore, STING activators can be used as vaccine adjuvants or immune activators.

[0004] Conversely, the overactivation of STING can cause a variety of severe human autoinflammatory and autoimmune diseases, including rare diseases such as Aicardi-Goutières syndrome (AGS), STING-associated vasculopathy in infancy (SAVI), and systemic lupus erythematosus (SLE). In the case of mitochondrial dysfunction, abnormal cGAS / STING activation will induce more common diseases such as non-alcoholic steatohepatitis (NASH), chronic obstructive pulmonary disease (COPD), age-related macular degeneration (AMD), and Parkinson's disease.

[0005] The important function of STING in the innate immune system and its correlation with various diseases have made it a popular target for drug research and development. Many pharmaceutical companies are conducting research on agonists and antagonists targeting STING. Cyclic dinucleotide (CDN) compounds are the only class of agonists that have been found to directly activate both murine and human STING proteins. Direct injection of CDN compounds into B16 melanoma, CT26 rectal cancer, and 4T1 breast cancer masses not only results in significant inhibition until the tumors disappear, but also induces systemic and persistent antigen-specific T cell immunity, causing the growth of tumors in other parts of the animal that have not been injected with the drug to be inhibited, leading to changes in the microenvironment of various solid tumors, activating effective tumor-induced CD8+ T cells and achieving long-lasting efficacy.

[0006] However, the clinical application of CDN compounds in the treatment of anti-tumor, anti-viral, degenerative diseases or immune diseases is still in a very early stage of research, and is restricted by the administration route and drug-like characteristics. There are undoubtedly many obstacles to overcome before it can become an effective clinical therapy. Therefore, the discovery and search for STING modulators with high activity and good drug-likeness has become a hot field today. Summary of the Invention

[0007] One of the technical problems to be solved by the present invention is to provide a novel STING protein modulator for preparing therapeutic drugs for tumors, immune diseases, anti-viral and degenerative diseases.

[0008] The solutions to the above technical problems are as follows:

[0009] A nitrogen-containing heterocyclic compound represented by the general formula I, or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, solvate, polymorph or prodrug thereof,

[0010]

[0011] In the formula:

[0012] R1 is selected from hydrogen, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl, C3-C6 heterocycloalkyl, 5-10 membered aryl or heteroaryl, acyl, sulfonyl, sulfone, sulfoxide, etc.;

[0013] R2 is selected from amino, amido, C1-C10 alkyl, 5-10 membered aryl or heteroaryl, C3-C6 cycloalkyl or heterocycloalkyl;

[0014] M1, M2, M3, M4, M5, M6, M7 are independently selected from N, CRa, and Ra is independently selected from hydrogen, halogen, hydroxyl, amino, cyano, carbonyl, amido, ester, urea, sulfone, sulfoxide, sulfonyl, sulfinyl, sulfinimide, C1-C6 alkyl, C1-C6 alkoxy, C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl, C3-C6 heterocycloalkyl, 5-10 membered aryl or heteroaryl; or M3 = M4, M5 = M6 are independently selected from heteroatoms such as O, S, etc.;

[0015] And any two adjacent Ras can form a 4-10 membered saturated or partially unsaturated ring system through a carbon chain or a heteroatom; or any one Ra on M6, M7 can form a 3-10 membered saturated or partially unsaturated ring system with any group on R1 through a carbon chain or a heteroatom.

[0016] One or more hydrogen atoms on any of the above groups may be substituted with substituents selected from the group consisting of, including but not limited to, deuterium, halogen, C1-C8 alkyl; wherein, the heteroaryl contains 1-3 heteroatoms selected from the group consisting of N, O, P or S, the heterocycloalkyl contains 1-3 heteroatoms selected from the group consisting of N, O, P or S, the ring system contains saturated or partially unsaturated ring systems such as spiro rings, bridged rings, fused rings, and annelated rings, and the above ring systems may be further substituted with C1-C6 alkyl, hydroxyl, amino, halogen, alkoxy, etc.

[0017] In some embodiments, the compound of general formula (I) is preferably a compound represented by the following general formula (II), or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, solvate, polymorph or prodrug thereof:

[0018]

[0019]

[0020] Wherein, Ra1, Ra2, Ra3, Ra4, Ra5, Ra6, Ra7 are each independently selected from hydrogen, halogen, hydroxyl, amino, cyano, carbonyl, amide group, ester group, urea, sulfone group, sulfoxide group, sulfonyl group, sulfinyl group, sulfinimide group, C1-C6 alkyl, C1-C6 alkoxy, C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl, C3-C6 heterocycloalkyl, 5-10 membered aryl or heteroaryl; and any two adjacent Ra 1-7 may form a 4-10 membered saturated or partially unsaturated ring system through a carbon chain or a heteroatom; or any one Ra 1-7 may form a 3-10 membered saturated or partially unsaturated ring system with any group on R1 through a carbon chain or a heteroatom; R1 and R2 are as defined above.

[0021] In some embodiments, it is preferably a compound represented by the following general formula (III), or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, solvate, polymorph or prodrug thereof:

[0022]

[0023] Among them, R2a, R2b, R2c, Ra4, Ra6, and R1’ are each independently selected from hydrogen, halogen, hydroxyl, amino, cyano, carbonyl, amide, ester, urea, sulfone, sulfoxide, sulfonyl, sulfinyl, sulfinimide, C1-C6 alkyl, C1-C6 alkoxy, C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl, C3-C6 heterocycloalkyl, 5-10-membered aryl or heteroaryl; Ma8 is independently selected from NRa8, O, S(O)p, C(Ra8)q, where Ra8 is selected from hydrogen, halogen, C1-C6 alkyl, p is selected from 0-2, and q is selected from 1-2;

[0024] And any one or more hydrogens on the above groups can be substituted by a group selected from the following group: -ORm, -NRmRn, -NRmCORn, -CO2Rm, -OCORm, -CONRmRn, -SO2NRmRn, -NRmSO2Rn, -SRm, -SORm, -SO2Rm, -OCONRmRn, -NRpCONRmRn; Rm and Rn are each independently selected from C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkylhydroxyl, C1-C6 alkylalkoxy, C1-C6 alkylamino or substituted amino, C1-C6 alkylcycloamino, C3-C6 cycloalkyl or heterocycloalkyl, 5-8-membered aryl or heteroaryl;

[0025] Or Rm and Rn, R2c and R2b can be respectively connected through a carbon atom or a heteroatom to form a 3-12-membered monocyclic or polycyclic alkyl, 3-12-membered monocyclic or polycyclic heterocycloalkyl, 3-12-membered spiro or fused ring alkyl, and 3-12-membered spiro or fused ring heterocycloalkyl;

[0026] Among them, R1, Ra4, and Ra6 are as defined above.

[0027] The structures shown by the above general formula III exclude the following two molecules:

[0028]

[0029] In some embodiments, the compound of formula (I) has a structure shown by the following formula (VI-1) or (VI-2):

[0030]

[0031] Among them, M1, M2, M3, M4, M5, M6, M7, and R2 are as defined above;

[0032] R2’ is selected from amino, amide, C1-C10 alkyl, 5-10-membered aryl or heteroaryl, C3-C6 cycloalkyl or heterocycloalkyl;

[0033] M1’, M2’, M3’, M4’, M5’, M6’, M7’ are independently selected from N, CRa, and Ra is independently selected from hydrogen, halogen, hydroxyl, amino, cyano, carbonyl, amido, ester, urea, sulfone, sulfoxide, sulfonyl, sulfinyl, sulfinimide, C1-C6 alkyl, C1-C6 alkoxy, C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl, C3-C6 heterocycloalkyl, 5-10 membered aryl or heteroaryl; or M3’ = M4’, M5’ = M6’ are independently selected from heteroatoms such as O, S;

[0034] M is selected from O, NRb, S(O) t , CRcRd; Rb is independently preferably selected from hydrogen, C1-C10 alkyl, 3-10 membered cycloalkyl or heterocycloalkyl, acyl or sulfonyl; Rc, Rd are independently selected from hydrogen, deuterium, halogen, C1-C10 alkyl, 3-10 membered cycloalkyl or heterocycloalkyl, or Rc and Rd are connected through a carbon atom or a heteroatom to form a 3-12 membered monocyclic or polycyclic alkyl, 3-12 membered monocyclic or polycyclic heterocycloalkyl, 3-12 membered spiro or fused ring alkyl, and 3-12 membered spiro or fused ring heterocycloalkyl;

[0035] m, n are integers selected from 0-3; t is an integer selected from 0-2.

[0036] In some preferred embodiments, the compounds of the general formulas (I), (II), (III), (IV) and (VI-1), or their pharmaceutically acceptable salts, or their enantiomers, diastereomers, tautomers, solvates, polymorphs or prodrugs, are characterized in that the compounds include but are not limited to the following structures:

[0037]

[0038]

[0039]

[0040]

[0041]

[0042]

[0043] A method for preparing the compound of the above general formula I, characterized in that the method comprises steps a-c:

[0044] a) Reacting the compound of the general formula (A) with an allyl dihalide under basic conditions to obtain the compound of the general formula (B); and

[0045] b) Reacting the compound of general formula B with the amine of general formula R1’R1”NH under basic conditions to obtain the compound of general formula (I); and

[0046] c) Subjecting the compound of general formula (I) to a hydrogenation reduction reaction under the catalysis of a transition metal to obtain the compound of general formula (I’);

[0047]

[0048] Wherein, X is a leaving group such as a halogen, a sulfonate, etc., and the definitions of the other groups are as described above.

[0049] Preferably, steps a), b), and c) are each carried out in a solvent, and the solvent is selected from the group consisting of: water, methanol, ethanol, isopropanol, butanol, ethylene glycol, ethylene glycol methyl ether, N-methylpyrrolidone, dimethyl sulfoxide, tetrahydrofuran, toluene, dichloromethane, 1,2-dichloroethane, acetonitrile, N,N-dimethylformamide, N,N-dimethylacetamide, dioxane, or a combination thereof.

[0050] Preferably, the inorganic base is selected from the group consisting of: sodium hydride, potassium hydroxide, sodium acetate, potassium acetate, potassium tert-butoxide, sodium tert-butoxide, potassium fluoride, cesium fluoride, potassium phosphate, potassium carbonate, potassium bicarbonate, sodium carbonate, sodium bicarbonate, or a combination thereof; the organic base is selected from the group consisting of: pyridine, triethylamine, N,N-diisopropylethylamine, 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU), hexamethyldisilazane lithium, hexamethyldisilazane sodium, dimethylpyridine, or a combination thereof.

[0051] Another object of the present invention is to provide a drug and its composition for treating or preventing tumors, viral infections, autoimmune diseases, and degenerative diseases. The technical solution for achieving the above object is as follows:

[0052] A pharmaceutical composition for treating or preventing the above diseases, which is composed of a nitrogen heterocyclic compound represented by the above general formula (I), or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, solvate, polymorph, or prodrug thereof and a pharmaceutically acceptable carrier.

[0053] Another object of the present invention is to provide a use of the above compound. The technical solution for achieving the above object is as follows:

[0054] The nitrogen-containing heterocyclic compounds represented by the general formula (I), or pharmaceutically acceptable salts thereof, or their enantiomers, diastereomers, tautomers, solvates, polymorphs or prodrugs are used for preparing drugs for treating STING protein-dependent diseases, especially drugs for treating tumors, viral infections, immune diseases and inflammatory diseases, and are a class of therapeutic drugs with a novel mechanism of action. The tumors are each independently selected from: non-small cell lung cancer, small cell lung cancer, lung adenocarcinoma, squamous cell carcinoma of the lung, breast cancer, prostate cancer, liver cancer, skin cancer, gastric cancer, intestinal cancer, cholangiocarcinoma, brain cancer, leukemia, lymphoma, fibroma, sarcoma, basal cell carcinoma, glioma, kidney cancer, melanoma, bone cancer, thyroid cancer, nasopharyngeal carcinoma, pancreatic cancer, etc. The immune diseases and inflammatory diseases are independently selected from rejection of transplanted organs, gout, rhinitis, hair loss, Alzheimer's disease, appendicitis, atherosclerosis, asthma, arthritis, allergic dermatitis, Behcet's disease, bullous skin disease, cholecystitis, chronic idiopathic thrombocytopenic purpura, chronic obstructive pulmonary disease, liver cirrhosis, degenerative joint disease, dermatitis, dermatomyositis, eczema, enteritis, encephalitis, gastritis, nephritis, Hashimoto's thyroiditis, hepatitis, hypophysitis, inflammatory bowel disease, irritable bowel syndrome, Kawasaki disease, meningitis, multiple sclerosis, myocarditis, myasthenia gravis, mycosis fungoides, myositis, nephritis, osteomyelitis, pancreatitis, Parkinson's disease, pericarditis, pernicious anemia, pneumonia, primary biliary sclerosing cholangitis, polyarteritis nodosa, psoriasis, fibrosis, lupus erythematosus, tissue transplant rejection, thyroiditis, type I diabetes, urethritis, uveitis, vasculitis, vitiligo and Waldenström's macroglobulinemia, etc.

[0055] It should be understood that within the scope of the present invention, the above technical features of the present invention and the technical features specifically described below (such as in the examples) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be elaborated one by one here. Detailed implementation manners

[0056] Through long-term and in-depth research, the inventors prepared a class of novel compounds with the structure shown in formula I, and found that they have good STING protein binding activity, and the compounds can specifically bind to STING protein at a relatively low concentration (as low as ≤1 nmol / L), and can regulate the release or inhibition of downstream cytokines such as IFN-β, IL-6, TNF, etc. of the STING pathway, so they can be used to regulate the activity of the STNG pathway and treat related diseases such as tumors, inflammation, antiviral, etc. Based on the above findings, the inventors completed the present invention.

[0057] Term

[0058] Unless otherwise defined, all technical terms used herein have the same meanings as commonly understood by those skilled in the art to which the claimed subject matter pertains. All patents, patent applications, and published materials cited herein in their entirety are incorporated herein by reference unless otherwise indicated.

[0059] It should be understood that the foregoing summary and the following detailed description are exemplary and explanatory only and do not limit the subject matter of the present invention. In this application, the singular form also includes the plural unless specifically stated otherwise. It must be noted that, unless clearly stated otherwise in the text, the singular forms used in this specification and the claims include the plural forms of the indicated items. It should also be noted that, unless otherwise indicated, the terms "or" or "either...or" mean "and / or". In addition, the term "comprising" and other forms such as "comprises", "including", and "contains" are not restrictive.

[0060] Definitions of standard chemical terms can be found in reference works (including Carey and Sundberg "ADVANCED ORGANIC CHEMISTRY 4TH ED.", Vols. A (2000) and B (2001), Plenum Press, New York). Unless otherwise indicated, conventional methods within the skill of the art are employed, such as mass spectrometry, NMR, IR, and UV / VIS spectroscopy and pharmacological methods. Unless otherwise specifically defined, the terms used in the relevant descriptions of analytical chemistry, organic synthetic chemistry, and medicinal and pharmaceutical chemistry herein are known in the art. Standard techniques can be used in chemical synthesis, chemical analysis, drug preparation, formulation, and delivery, as well as in the treatment of patients. For example, the instructions provided by the manufacturer for a kit can be utilized, or reactions and purifications can be carried out in a manner known in the art or as described in the present invention. Generally, the above-mentioned techniques and methods can be implemented according to the descriptions in a number of general and more specific documents cited and discussed in this specification, in accordance with conventional methods well-known in the art. In this specification, groups and their substituents can be selected by those skilled in the art to provide stable structural moieties and compounds.

[0061] When a substituent is described by a conventional chemical formula written from left to right, the substituent also includes the chemically equivalent substituent obtained when the structural formula is written from right to left. For example, -CH2O- is equivalent to -OCH2-.

[0062] The section headings used herein are for the purpose of organizing the article only and should not be construed as limiting the subject matter described. All documents or portions of documents cited in this application, including but not limited to patents, patent applications, articles, books, manuals of operations, and theses, are incorporated herein by reference in their entirety.

[0063] The total number of carbon atoms present in certain chemical groups defined herein is represented by a simplified symbol in front of the group. For example, C1-6 alkyl refers to an alkyl group having a total of 1 to 6 carbon atoms as defined below. The total number of carbon atoms in the simplified symbol does not include the carbon in the substituents that may be present in the group.

[0064] Except as otherwise described above, when used in the specification and claims of this application, unless otherwise specifically indicated, the following terms have the meanings shown below.

[0065] In this application, the term "halogen" means fluorine, chlorine, bromine or iodine; "hydroxyl" means the -OH group; "hydroxyalkyl" means an alkyl group as defined below substituted by a hydroxyl (-OH) group; "carbonyl" means the -C(=O)- group; "nitro" means -NO2; "cyano" means -CN; "amino" means -NH2; "substituted amino" means an amino group substituted by one or two alkyl groups, alkylcarbonyl groups, aralkyl groups, heteroaralkyl groups as defined below, for example, monoalkylamino, dialkylamino, alkylcarbonylamino, aralkylamino, heteroaralkylamino; "carboxyl" means -COOH.

[0066] In this application, as a group or as part of other groups (such as in groups like halogen-substituted alkyl), the term "alkyl" means a straight-chain or branched-chain hydrocarbon chain group consisting only of carbon atoms and hydrogen atoms, containing no unsaturated bonds, having, for example, 1 to 12 (preferably 1 to 8, more preferably 1 to 6) carbon atoms and connected to the rest of the molecule by a single bond. Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, 2-methylbutyl, 2,2-dimethylpropyl, n-hexyl, heptyl, 2-methylhexyl, 3-methylhexyl, octyl, nonyl, and decyl, etc.

[0067] In this application, as a group or as part of other groups, the term "alkenyl" means a straight-chain or branched-chain hydrocarbon chain group consisting only of carbon atoms and hydrogen atoms, containing at least one double bond, having, for example, 2 to 14 (preferably 2 to 10, more preferably 2 to 6) carbon atoms and connected to the rest of the molecule by a single bond, such as, but not limited to, vinyl, propenyl, allyl, but-1-enyl, but-2-enyl, pent-1-enyl, pent-1,4-dienyl, etc.

[0068] In the present application, as a group or as part of another group, the term "alkynyl" means a straight-chain or branched-chain hydrocarbon chain group consisting only of carbon and hydrogen atoms, containing at least one triple bond and optionally one or more double bonds, having, for example, 2 to 14 (preferably 2 to 10, more preferably 2 to 6) carbon atoms and being linked to the remainder of the molecule by a single bond. Examples include, but are not limited to, ethynyl, prop-1-ynyl, but-1-ynyl, pent-1-en-4-ynyl, and the like.

[0069] In the present application, as a group or as part of another group, the term "cycloalkyl" means a stable non-aromatic monocyclic or polycyclic hydrocarbon group consisting only of carbon and hydrogen atoms, which may include fused ring systems, bridged ring systems or spiro ring systems, having 3 to 15 carbon atoms, preferably 3 to 10 carbon atoms, more preferably 3 to 8 carbon atoms, and being saturated or unsaturated and being linked to the remainder of the molecule by a single bond through any suitable carbon atom. Unless otherwise specifically indicated in the present specification, the carbon atoms in the cycloalkyl may optionally be oxidized. Examples of cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cyclooctyl, 1H-indenyl, 2,3-dihydroindenyl, 1,2,3,4-tetrahydro-naphthalenyl, 5,6,7,8-tetrahydro-naphthalenyl, 8,9-dihydro-7H-benzocyclohepten-6-yl, 6,7,8,9-tetrahydro-5H-benzocycloheptenyl, 5,6,7,8,9,10-hexahydro-benzocyclooctenyl, fluorenyl, bicyclo[2.2.1]heptyl, 7,7-dimethyl-bicyclo[2.2.1]heptyl, bicyclo[2.2.1]heptenyl, bicyclo[2.2.2]octyl, bicyclo[3.1.1]heptyl, bicyclo[3.2.1]octyl, bicyclo[2.2.2]octenyl, bicyclo[3.2.1]octenyl, adamantyl, octahydro-4,7-methano-1H-indenyl, and octahydro-2,5-methano-s-indacenyl, and the like.

[0070] In the present application, as a group or as part of another group, the term "heterocyclic group" means a stable 3- to 20-membered non-aromatic cyclic group composed of 2 to 14 carbon atoms and 1 to 6 heteroatoms selected from nitrogen, phosphorus, oxygen, and sulfur. Unless otherwise specifically indicated in this specification, the heterocyclic group can be a monocyclic, bicyclic, tricyclic, or more-ring ring system, which can include a fused-ring system, a bridged-ring system, or a spiro-ring system; the nitrogen, carbon, or sulfur atoms in the heterocyclic group can be optionally oxidized; the nitrogen atoms can be optionally quaternized; and the heterocyclic group can be partially or fully saturated. The heterocyclic group can be connected to the rest of the molecule via a carbon atom or a heteroatom by a single bond. In a heterocyclic group containing fused rings, one or more rings can be an aryl or heteroaryl group as defined below, provided that the point of attachment to the rest of the molecule is a non-aromatic ring atom. For the purposes of the present invention, the heterocyclic group is preferably a stable 4- to 11-membered non-aromatic monocyclic, bicyclic, bridged-ring, or spiro-ring group containing 1 to 3 heteroatoms selected from nitrogen, oxygen, and sulfur, and more preferably a stable 4- to 8-membered non-aromatic monocyclic, bicyclic, bridged-ring, or spiro-ring group containing 1 to 3 heteroatoms selected from nitrogen, oxygen, and sulfur. Examples of the heterocyclic group include, but are not limited to: pyrrolidinyl, morpholinyl, piperazinyl, homopiperazinyl, piperidinyl, thiomorpholinyl, 2,7-diaza-spiro[3.5]nonan-7-yl, 2-oxa-6-aza-spiro[3.3]heptan-6-yl, 2,5-diaza-bicyclo[2.2.1]heptan-2-yl, azetidinyl, pyranyl, tetrahydropyranyl, thiopyranyl, tetrahydrofuryl, oxazinyl, dioxolanyl, tetrahydroisoquinolinyl, decahydroisoquinolinyl, imidazolinyl, imidazolidinyl, quinuclidinyl, thiazolidinyl, isothiazolidinyl, isoxazolidinyl, dihydroindolyl, octahydroindolyl, octahydroisoindolyl, pyrrolidinyl, pyrazolidinyl, phthalimido, etc.

[0071] In the present application, as a group or as part of another group, the term "aryl" means a conjugated hydrocarbon ring system group having 6 to 18 carbon atoms (preferably having 6 to 10 carbon atoms). For the purposes of the present invention, the aryl can be a monocyclic, bicyclic, tricyclic, or more-ring ring system, and can also be fused with a cycloalkyl or heterocyclic group as defined above, provided that the aryl is connected to the rest of the molecule via an atom on the aromatic ring by a single bond. Examples of the aryl include, but are not limited to: phenyl, naphthyl, anthracenyl, phenanthrenyl, fluorenyl, 2,3-dihydro-1H-isoindolyl, 2-benzoxazolinone, 2H-1,4-benzoxazin-3(4H)-one-7-yl, etc.

[0072] In the present application, the term "arylalkyl" means an alkyl group as defined above substituted by an aryl group as defined above.

[0073] In the present application, as a group or as part of another group, the term "heteroaryl" means a 5- to 16-membered conjugated ring system group having 1 to 15 carbon atoms (preferably 1 to 10 carbon atoms) and 1 to 6 heteroatoms selected from nitrogen, oxygen, and sulfur within the ring. Unless otherwise specifically indicated in this specification, the heteroaryl can be a monocyclic, bicyclic, tricyclic, or more-ring ring system, and can also be fused with the cycloalkyl or heterocyclic group defined above, provided that the heteroaryl is connected to the rest of the molecule by a single bond through an atom on the aromatic ring. The nitrogen, carbon, or sulfur atoms in the heteroaryl can be optionally oxidized; the nitrogen atoms can be optionally quaternized. For the purposes of the present invention, the heteroaryl is preferably a stable 5- to 12-membered aromatic group containing 1 to 5 heteroatoms selected from nitrogen, oxygen, and sulfur, more preferably a stable 5- to 10-membered aromatic group containing 1 to 4 heteroatoms selected from nitrogen, oxygen, and sulfur or a 5- to 6-membered aromatic group containing 1 to 3 heteroatoms selected from nitrogen, oxygen, and sulfur. Examples of heteroaryl include, but are not limited to, thienyl, imidazolyl, pyrazolyl, thiazolyl, oxazolyl, oxadiazolyl, isoxazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, benzimidazolyl, benzopyrazolyl, indolyl, furyl, pyrrolyl, triazolyl, tetrazolyl, triazinyl, indolizinyl, isoindolyl, indazolyl, isoindazolyl, purinyl, quinolinyl, isoquinolinyl, phthalazinyl, naphthyridinyl, quinoxalinyl, pteridinyl, carbazolyl, carbolinyl, phenanthridinyl, phenanthrolinyl, acridinyl, phenazinyl, isothiazolyl, benzothiazolyl, benzothienyl, oxatriazolyl, cinnolinyl, quinazolinyl, phenylthio, indolizinyl, phenanthrolinyl, isoxazolyl, phenoxazinyl, phenothiazinyl, 4,5,6,7-tetrahydrobenzo[b]thienyl, naphthopyridyl, [1,2,4]triazolo[4,3-b]pyridazine, [1,2,4]triazolo[4,3-a]pyrazine, [1,2,4]triazolo[4,3-c]pyrimidine, [1,2,4]triazolo[4,3-a]pyridine, imidazo[1,2-a]pyridine, imidazo[1,2-b]pyridazine, imidazo[1,2-a]pyrazine, etc.

[0074] In the present application, the term "heteroarylalkyl" means an alkyl group as defined above substituted by a heteroaryl group as defined above.

[0075] In the present application, "optional" or "optionally" means that the subsequent described event or condition may or may not occur, and this description includes both the occurrence and non-occurrence of the event or condition. For example, "optionally substituted aryl" means that the aryl is substituted or unsubstituted, and this description includes both the substituted aryl and the unsubstituted aryl.

[0076] As used herein, the terms "moiety", "structural moiety", "chemical moiety", "group", "chemical group" refer to specific fragments or functional groups in a molecule. A chemical moiety is generally considered to be a chemical entity embedded in or attached to a molecule.

[0077] "Stereoisomers" refer to compounds that are composed of the same atoms, bonded by the same bonds, but have different three-dimensional structures. The present invention will cover various stereoisomers and their mixtures.

[0078] When the compounds of the present invention contain olefinic double bonds, unless otherwise specified, the compounds of the present invention are intended to include E- and Z-geometric isomers.

[0079] "Tautomers" refer to isomers formed by the transfer of a proton from one atom of a molecule to another atom of the same molecule. All tautomeric forms of the compounds of the present invention will also be included within the scope of the present invention.

[0080] The compounds of the present invention or their pharmaceutically acceptable salts may contain one or more chiral carbon atoms and may thus give rise to enantiomers, diastereomers and other stereoisomeric forms. Each chiral carbon atom can be defined as (R)- or (S)- based on stereochemistry. The present invention is intended to include all possible isomers, as well as their racemic and optically pure forms. The preparation of the compounds of the present invention may use racemates, diastereomers or enantiomers as starting materials or intermediates. Optically active isomers can be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques, such as crystallization and chiral chromatography.

[0081] Conventional techniques for the preparation / isolation of individual isomers include chiral synthesis from suitable optically pure precursors or the resolution of racemates (or racemates of salts or derivatives) using, for example, chiral high performance liquid chromatography. See, for example, Gerald Gübitz and Martin G. Schmid (Eds.), Chiral Separations, Methods and Protocols, Methods in Molecular Biology, Vol. 243, 2004; A.M. Stalcup, Chiral Separations, Annu. Rev. Anal. Chem. 3:341-63, 2010; Fumiss et al. (eds.), VOGEL'S ENCYCLOPEDIA OF PRACTICAL ORGANIC CHEMISTRY 5.sup.TH ED., Longman Scientific and Technical Ltd., Essex, 1991, 809-816; Heller, Acc. Chem. Res. 1990, 23, 128.

[0082] In the present application, the term "pharmaceutically acceptable salt" includes pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.

[0083] "Pharmaceutically acceptable acid addition salts" refer to salts formed with inorganic acids or organic acids that are able to retain the biological effectiveness of the free base without other side effects. Inorganic acid salts include, but are not limited to, hydrochloride, hydrobromide, sulfate, nitrate, phosphate, etc.; organic acid salts include, but are not limited to, formate, acetate, 2,2-dichloroacetate, trifluoroacetate, propionate, caproate, caprylate, caprate, undecylenate, glycolate, gluconate, lactate, sebacate, adipate, glutarate, malonate, oxalate, maleate, succinate, fumarate, tartrate, citrate, palmitate, stearate, oleate, cinnamate, laurate, malate, glutamate, pyroglutamate, aspartate, benzoate, methanesulfonate, benzenesulfonate, p-toluenesulfonate, alginate, ascorbate, salicylate, 4-aminosalicylate, naphthalenedisulfonate, etc. These salts can be prepared by methods known in the art.

[0084] "Pharmaceutically acceptable base addition salts" refer to salts formed with inorganic or organic bases that retain the biological effectiveness of the free acid without other adverse effects. Salts derived from inorganic bases include, but are not limited to, sodium salts, potassium salts, lithium salts, ammonium salts, calcium salts, magnesium salts, iron salts, zinc salts, copper salts, manganese salts, aluminum salts, etc. Preferred inorganic salts are ammonium salts, sodium salts, potassium salts, calcium salts and magnesium salts. Salts derived from organic bases include, but are not limited to, the following salts: primary amines, secondary amines and tertiary amines, substituted amines, including natural substituted amines, cyclic amines and basic ion exchange resins, such as ammonia, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, triethanolamine, dimethylethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, choline, betaine, ethylenediamine, glucosamine, methylglucosamine, theobromine, purine, piperazine, piperidine, N-ethylpiperidine, polyamine resins, etc. Preferred organic bases include isopropylamine, diethylamine, ethanolamine, trimethylamine, dicyclohexylamine, choline and caffeine. These salts can be prepared by methods known in the art.

[0085] "Polymorphs" refer to different solid crystalline phases of certain compounds of the present invention in the solid state due to the presence of two or more different molecular arrangements. Some compounds of the present invention can exist in more than one crystal form, and the present invention aims to include all crystal forms and their mixtures.

[0086] Generally, crystallization results in solvates of the compounds of the present invention. The term "solvate" as used in the present invention refers to an aggregate containing one or more molecules of a compound of the present invention and one or more solvent molecules. The solvent can be water, in which case the solvate is a hydrate. Alternatively, the solvent can be an organic solvent. Thus, the compounds of the present invention can exist as hydrates, including monohydrates, dihydrates, hemihydrates, sesquihydrates, trihydrates, tetrahydrates, etc., as well as the corresponding solvated forms. The compounds of the present invention can form true solvates, but in some cases, they can also retain only indeterminate water or a mixture of water and some indeterminate solvent. The compounds of the present invention can react in a solvent or precipitate or crystallize out from a solvent. The solvates of the compounds of the present invention are also included within the scope of the present invention.

[0087] The present invention also includes prodrugs of the above compounds. In the present application, the term "prodrug" refers to a compound that can be converted into the bioactive compound of the present invention under physiological conditions or by solvolysis. Therefore, the term "prodrug" refers to a pharmaceutically acceptable metabolic precursor of the compound of the present invention. When administered to an individual in need, the prodrug may be inactive, but is converted into the active compound of the present invention in vivo. Prodrugs are generally rapidly converted in vivo to produce the parent compound of the present invention, for example, by hydrolysis in the blood. Prodrugs generally offer the advantages of solubility, tissue compatibility, or slow release in mammalian organisms. Prodrugs include known amino protecting groups and carboxyl protecting groups. Specific methods for preparing prodrugs can refer to Saulnier, M.G., et al., Bioorg. Med. Chem. Lett. 1994, 4, 1985 - 1990; Greenwald, R.B., et al., J. Med. Chem. 2000, 43, 475.

[0088] In the present application, "pharmaceutical composition" refers to a preparation of the compound of the present invention and a medium generally accepted in the art for delivering a bioactive compound to a mammal (such as a human). This medium includes a pharmaceutically acceptable carrier. The purpose of the pharmaceutical composition is to facilitate the administration to an organism, facilitate the absorption of the active ingredient, and thus exert its biological activity.

[0089] As used herein, the term "pharmaceutically acceptable" refers to a substance (such as a carrier or diluent) that does not affect the biological activity or properties of the compound of the present invention and is relatively non-toxic, that is, the substance can be administered to an individual without causing adverse biological reactions or interacting with any component contained in the composition in an adverse manner.

[0090] In the present application, "pharmaceutically acceptable carrier" includes, but is not limited to, any adjuvant, carrier, excipient, glidant, sweetening agent, diluent, preservative, dye / colorant, flavoring agent, surfactant, wetting agent, dispersing agent, suspending agent, stabilizing agent, isotonic agent, solvent, or emulsifying agent that is permitted by the relevant government regulatory authorities for use in humans or livestock.

[0091] The "tumors", "diseases related to abnormal cell proliferation", etc. described in the present invention include, but are not limited to, diseases such as leukemia, gastrointestinal stromal tumor, histiocytic lymphoma, non-small cell lung cancer, small cell lung cancer, pancreatic cancer, squamous cell carcinoma of the lung, adenocarcinoma of the lung, breast cancer, prostate cancer, liver cancer, skin cancer, epithelial cell carcinoma, cervical cancer, ovarian cancer, intestinal cancer, nasopharyngeal cancer, brain cancer, bone cancer, esophageal cancer, melanoma, kidney cancer, oral cancer, etc.

[0092] As used herein, the terms "preventive", "prevent", and "prevention" include reducing the likelihood of the occurrence or exacerbation of a disease or disorder in a patient.

[0093] As used herein, the terms "treat", "treatment" and other similar synonyms include the following meanings:

[0094] (i) Preventing the occurrence of a disease or disorder in a mammal, particularly when such mammal is predisposed to the disease or disorder but has not been diagnosed as having the disease or disorder;

[0095] (ii) Inhibiting a disease or disorder, i.e., arresting its development;

[0096] (iii) Alleviating a disease or disorder, i.e., causing the state of the disease or disorder to regress; or

[0097] (iv) Relieving the symptoms caused by the disease or disorder.

[0098] As used herein, the terms "effective amount", "therapeutically effective amount" or "pharmaceutically effective amount" refer to an amount of at least one agent or compound that, when administered, is sufficient to alleviate to some extent one or more symptoms of the disease or disorder being treated. The result can be a reduction and / or alleviation of the signs, symptoms or causes, or any other desired change in a biological system. For example, an "effective amount" for treatment is the amount of a composition comprising a compound disclosed herein that provides a significant alleviation of the disorder in a clinical setting. Techniques such as dose escalation trials can be used to determine the effective amount suitable for any individual case.

[0099] As used herein, the terms "administer", "administration", "administered" and the like refer to methods capable of delivering a compound or composition to a desired site of biological action. These methods include, but are not limited to, oral route, duodenal route, parenteral injection (including intravenous, subcutaneous, intraperitoneal, intramuscular, intraarterial injection or infusion), topical administration and rectal administration. Techniques for the administration of the compounds and methods described herein are well known to those skilled in the art, such as those discussed in Goodman and Gilman, The Pharmacological Basis of Therapeutics, current ed.; Pergamon; and Remington's, Pharmaceutical Sciences (current edition), Mack Publishing Co., Easton, Pa. In a preferred embodiment, the compounds and compositions discussed herein are administered orally.

[0100] As used herein, the terms "drug combination", "drug co - administration", "combination therapy", "administering other therapy", "administering other therapeutic agent", etc. refer to a pharmaceutical treatment obtained by mixing or combining more than one active ingredient, which includes fixed and non - fixed combinations of active ingredients. The term "fixed combination" refers to the simultaneous administration to a patient of at least one compound described herein and at least one synergistic agent in the form of a single entity or a single dosage form. The term "non - fixed combination" refers to the simultaneous administration, co - administration or sequential administration at variable intervals to a patient of at least one compound described herein and at least one synergistic agent in the form of separate entities. These also apply to cocktail therapies, such as the administration of three or more active ingredients.

[0101] Those skilled in the art should also understand that in the methods described hereinafter, the functional groups of intermediate compounds may need to be protected by appropriate protecting groups. Such functional groups include hydroxyl, amino, mercapto and carboxylic acid. Suitable hydroxyl protecting groups include trialkylsilyl or diarylalkylsilyl (such as tert - butyldimethylsilyl, tert - butyldiphenylsilyl or trimethylsilyl), tetrahydropyranyl, benzyl, etc. Suitable protecting groups for amino, amidino and guanidine groups include tert - butoxycarbonyl, benzyloxycarbonyl, etc. Suitable mercapto protecting groups include - C(O) - R” (where R” is alkyl, aryl or aralkyl), p - methoxybenzyl, triphenylmethyl, etc. Suitable carboxyl protecting groups include alkyl, aryl or aralkyl esters.

[0102] Protecting groups can be introduced and removed according to standard techniques known to those skilled in the art and as described herein. The use of protecting groups is described in detail in Greene, T.W. and P.G.M. Wuts, Protective Groups in Organic Synthesis, (1999), 4th Ed., Wiley. The protecting group can also be a polymer resin.

[0103] The present invention will be further described below in conjunction with specific embodiments. It should be understood that these embodiments are only used to illustrate the present invention and not to limit the scope of the present invention. The experimental methods without specific conditions noted in the following embodiments are generally carried out under conventional conditions or according to the conditions recommended by the manufacturers. Unless otherwise stated, percentages and parts are weight percentages and weight parts.

[0104] Preparation of Intermediate A

[0105] Intermediate A1: Methyl 3 - (1,3 - dimethyl - 1H - pyrazol - 5 - yl) - 6 - methoxy - 5H - pyrido[4,3 - b]indole - 8 - carboxylate

[0106]

[0107] Step 1: Dissolve methyl 4-iodo-3-methoxybenzoate (3.2 g, 11.0 mmol) and 5-bromo-2-chloropyridin-4-amine (2.5 g, 12.1 mmol) in N,N-dimethylformamide (DMF) (30 mL). Under nitrogen, add palladium acetate (747.6 mg, 3.34 mmol), cesium carbonate (14.5 g, 44.5 mmol), and 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (Xantphos) (341.8 mg, 0.59 mmol). React at 115°C overnight. Cool to room temperature, filter, concentrate under reduced pressure, and purify by column chromatography to obtain methyl 4-((5-bromo-2-chloropyridin-4-yl)amino)-3-methoxybenzoate (2.9 g, white solid). LC-MS: m / z 371.0 / 373.0 [M+H] + .

[0108] Step 2: Dissolve methyl 4-((5-bromo-2-chloropyridin-4-yl)amino)-3-methoxybenzoate (365 mg, 0.99 mmol) in DMF (10 mL). Under nitrogen, add sodium acetate (533.1 mg, 6.50 mmol) and bis(triphenylphosphine)palladium(II) chloride (Pd(PPh3)2Cl2) (140.4 mg, 0.20 mmol). React at 125°C overnight. Cool to room temperature, filter, concentrate under reduced pressure, and purify by column chromatography to obtain methyl 3-chloro-6-methoxy-5H-pyrido[4,3-b]indole-8-carboxylate (160 mg, white solid). 1 H NMR (DMSO-d6, 400MHz): δ 12.42 (s, 1H), 9.30 (s, 1H), 8.57 (s, 1H), 7.61 (d, J = 1.2Hz, 1H), 7.46 (s, 1H), 4.06 (s, 3H), 3.91 (s, 3H). LC-MS:m / z 291.0 / 293.0[M+H] + .

[0109] Step 3: Methyl 3-chloro-6-methoxy-5H-pyrido[4,3-b]indole-8-carboxylate (500 mg, 1.72 mmol) and 1,3-dimethyl-5-pyrazolopinacol borate (453.3 mg, 2.04 mmol) were dissolved in dioxane and water (20 mL / 4 mL). Sodium carbonate (720.8 mg, 6.8 mmol) and tetrakistriphenylphosphine palladium (Pd(PPh3)4) (196.5 mg, 0.17 mmol) were added under nitrogen. The mixture was reacted at 90°C overnight. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, and purified by column chromatography to obtain Intermediate A1 (160 mg, white solid). 11H NMR (DMSO-d6, 400 MHz): δ 12.37 (s, 1H), 9.53 (d, J = 0.8 Hz, 1H), 8.58 (d, J = 1.2 Hz, 1H), 7.68 (d, J = 0.8 Hz, 1H), 7.60 (d, J = 1.2 Hz, 1H), 6.52 (s, 1H), 4.09 (s, 3H), 4.08 (s, 3H), 3.92 (s, 3H), 2.21 (s, 3H). LC-MS: m / z 351.2 [M+H] + 。

[0110] Intermediate A2: Methyl 3-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-6-methoxy-5H-pyrido[4,3-b]indole-8-carboxylate

[0111] Using 1-ethyl 3-methyl-5-pyrazolylboronic acid pinacol ester as the raw material, the target intermediate was prepared by referring to the synthesis method of Intermediate A1. 1 1H NMR (DMSO-d 6, 400 MHz): δ 12.36 (s, 1H), 9.53 (s, 1H), 8.58 (s, 1H), 7.67 (s, 1H), 7.61 (s, 1H), 6.49 (s, 1H), 4.55 - 4.56 (m, 2H), 4.08 (s, 3H), 3.92 (s, 3H), 2.22 (s, 3H), 1.30 - 1.32 (t, J = 7.2 Hz, 3H). LC-MS: m / z 365.2 [M+H] + 。

[0112] Intermediate A3: Methyl 3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-(3-morpholinopropoxy)-5H-pyrido[4,3-b]indole-8-carboxylate

[0113]

[0114] First step: Methyl 3-hydroxy-4-iodobenzoate (9.3 g, 33.5 mmol) and 3-morpholinopropyl methanesulfonate (15.0 g, 67.3 mmol) were dissolved in DMF (100 mL), potassium carbonate (13.8 g, 100.0 mmol) was added, and the reaction was carried out at room temperature overnight. It was washed with ethyl acetate, the filtrate and the washing solution were concentrated under reduced pressure, and purified by column chromatography to obtain methyl 4-iodo-3-(3-morpholinopropoxy)benzoate (12.3 g, white solid). 11H NMR (DMSO-d6, 400 MHz): δ 7.94 (d, J = 8.0 Hz, 1H), 7.40 (d, J = 1.6 Hz, 1H), 7.31 (dd, J = 1.6, 8.0 Hz, 1H), 4.14 (t, J = 6.0 Hz, 2H), 3.86 (s, 3H), 3.55 - 3.58 (m, 4H), 3.30 - 3.35 (m, 2H), 2.48 - 2.51 (m, 4H), 1.90 - 1.93 (m, 2H). LC-MS: m / z 406.1 [M+H] + 。

[0115] Step 2: Dissolve methyl 4-iodo-3-(3-morpholinopropoxy)benzoate (12.3 g, 30.4 mmol) and 5-bromo-2-chloropyridin-4-amine (6.9 g, 33.5 mmol) in DMF (150 mL). Under nitrogen protection, add palladium acetate (680.5 mg, 3.04 mmol), cesium carbonate (29.6 g, 90.8 mmol) and Xantphos (1.75 g, 3.04 mmol), and react at 100 °C overnight. Cool to room temperature, filter, concentrate under reduced pressure, and purify by column chromatography to obtain methyl 4-((5-bromo-2-chloropyridin-4-yl)amino)-3-(3-morpholinopropoxy)benzoate (l2.0 g, pale yellow solid). LC-MS: m / z 484.4 / 486.3 [M+H] + 。

[0116] Step 3: Dissolve methyl 4-((5-bromo-2-chloropyridin-4-yl)amino)-3-(3-morpholinopropoxy)benzoate (12.0 g, 24.84 mmol) in DMF (100 mL). Under nitrogen protection, add sodium acetate (8.15 g, 99.36 mmol) and Pd(PPh3)2Cl2 (1.74 g, 2.48 mmol), and react at 120 °C overnight. Cool to room temperature, filter, concentrate under reduced pressure, and purify by column chromatography to obtain methyl 3-chloro-6-(3-morpholinopropoxy)-5H-pyrido[4,3-b]indole-8-carboxylate (6.1 g, white solid). LC-MS: m / z 404.4 / 406.3 [M+H] + 。

[0117] Step 4: Dissolve methyl 3-chloro-6-(3-morpholinopropoxy)-5H-pyrido[4,3-b]indole-8-carboxylate (220 mg, 0.55 mmol) and 1,3-dimethyl-5-pyrazolylboronic acid pinacol ester (169.7 mg, 0.76 mmol) in dioxane and water (20 mL / 4 mL). Under nitrogen protection, add sodium carbonate (233.8 mg, 2.2 mmol) and Pd(PPh3)4 (63.6 mg, 0.055 mmol), and react at 100 °C overnight. Cool the reaction solution to room temperature, concentrate it under reduced pressure, and purify it by column chromatography to obtain intermediate A3 (120 mg, white solid). 1 H NMR (DMSO-d6, 400 MHz): δ 12.23 (s, 1H), 9.54 (s, 1H), 8.57 (s, 1H), 7.71 (d, J = 0.8 Hz, 1H), 7.59 (d, J = 1.2 Hz, 1H), 6.51 (s, 1H), 4.28 - 4.32 (m, 2H), 4.09 (s, 3H), 3.91 (s, 3H), 3.57 - 3.61 (m, 4H), 2.50 - 2.60 (m, 2H), 2.41 - 2.50 (m, 4H), 2.21 (s, 3H), 1.90 - 2.12 (m, 2H). LC-MS: m / z 464.3 [M+H] + 。

[0118] Using commercial reagents as raw materials, the following intermediates were prepared by referring to the synthesis method of intermediate A3.

[0119]

[0120]

[0121] Preparation of Intermediate B

[0122] Intermediate B1: Methyl 2-(1,3-dimethyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxylate

[0123]

[0124] Step 1: Methyl 4-amino-3-methoxybenzoate (6.0 g, 33.15 mmol) and 2,4-dichloro-5-iodopyrimidine (11.8 g, 43.10 mmol) were dissolved in 2-pentanol (100 mL), and N,N-diisopropylethylamine (DIEA) (12.8 g, 99.22 mmol) was added. The reaction was carried out under reflux at 130 °C overnight. The reaction mixture was cooled to room temperature, filtered, and the solid was purified by trituration with methanol to obtain methyl 4-((2-chloro-5-iodopyrimidin-4-yl)amino)-3-methoxybenzoate (9.4 g, white solid). LC-MS: m / z 420.0 / 422.0 [M+H] + .

[0125] Step 2: Methyl 4-((2-chloro-5-iodopyrimidin-4-yl)amino)-3-methoxybenzoate (9.4 g, 22.4 mmol) was dissolved in DMF (100 mL). Under nitrogen protection, sodium acetate (7.4 g, 90.2 mmol) and bis(triphenylphosphine)palladium(II) chloride (Pd(PPh3)2Cl2) (1.57 g, 2.24 mmol) were added, and the reaction was carried out at 120 °C overnight. The reaction mixture was cooled to room temperature, poured into water to precipitate a solid, filtered, and the filter cake was purified by trituration with methanol to obtain methyl 2-chloro-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxylate (5.6 g, white solid). LC-MS: m / z 292.1 / 294.1 [M+H] + .

[0126] Step 3: Methyl 2-chloro-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxylate (200 mg, 0.69 mmol) and 1,3-dimethyl-5-pyrazolylborate pinacol ester (233.3 mg, 1.05 mmol) were dissolved in dioxane and water (20 mL / 4 mL). Under nitrogen protection, sodium carbonate (300.8 mg, 2.84 mmol) and Pd(PPh3)4 (196.5 mg, 0.17 mmol) were added, and the reaction was carried out at 100 °C overnight. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, and purified by column chromatography to obtain intermediate B1 (160 mg, white solid). 1 1H-NMR (DMSO-d6, 400 MHz): δ 12.94 (s, 1H), 9.67 (s, 1H), 8.57 (d, J = 1.2 Hz, 1H), 7.63 (d, J = 1.2 Hz, 1H), 6.81 (s, 1H), 4.27 (s, 3H), 4.06 (s, 3H), 3.92 (s, 3H), 2.22 (s, 3H). LC-MS: m / z 352.2 [M+H] + .

[0127] Intermediate B2: Methyl 2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxylate

[0128] Using 1-ethyl-3-methyl-5-pyrazolylboronic acid pinacol ester as the raw material, the target intermediate was prepared by referring to the synthesis method of Intermediate B1. 1 H NMR (DMSO-d6, 400 MHz): δ 12.92 (s, 1H), 9.66 (s, 1H), 8.57 (d, J = 0.8 Hz, 1H), 7.63 (d, J = 1.2 Hz, 1H), 6.81 (s, 1H), 4.75 - 4.79 (m, 2H), 4.06 (s, 3H), 3.92 (s, 3H), 2.23 (s, 3H), 1.39 (t, J = 7.2 Hz, 3H). LC-MS: m / z 366.2 [M + H] + 。

[0129] Intermediate B3: Methyl 2-(1,3-dimethyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxylate

[0130]

[0131] First step: Dissolve methyl 4-amino-3-hydroxybenzoate (80 mg, 0.48 mmol) and 3-morpholinopropan-1-ol (73 mg, 0.50 mmol) in tetrahydrofuran (THF) (25 mL), add diisopropyl azodicarboxylate (DIAD) (194 mg, 0.96 mmol) and triphenylphosphine (PPh3) (180 mg, 0.70 mmol), and react overnight at room temperature. The reaction solution was concentrated under reduced pressure and purified by column chromatography to obtain methyl 4-amino-3-(3-morpholinopropoxy)benzoate (70 mg, white solid). LC-MS: m / z 295.2 [M + H] + 。

[0132] The subsequent operations were carried out by referring to the synthesis method of Intermediate B1 to obtain Intermediate B3. 1 H NMR (DMSO-d6, 400 MHz): δ 12.89 (s, 1H), 9.65 (s, 1H), 8.54 (s, 1H), 7.60 (s, 1H), 6.81 (s, 1H), 4.28 - 4.31 (m, 5H), 3.91 (s, 3H), 3.59 - 3.62 (m, 4H), 2.43 - 2.60 (m, 9H), 1.90 - 2.10 (m, 2H). LC-MS: m / z 465.2 [M + H] + 。

[0133] Intermediate B4: Methyl 2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-hydroxypropoxy)-9H-pyrimido[4,5-b]indole-6-carboxylate

[0134] Intermediate B4 was prepared by the same method as Intermediate B3, LC-MS: m / z 410.2 [M+H] + 。

[0135] Intermediate B5: Methyl 2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-((4-methoxybenzyl)oxy)-9H-pyrimido[4,5-b]indole-6-carboxylate

[0136]

[0137] First step: Methyl 3-hydroxy-4-nitrobenzoate (33.0 g, 167.5 mmol) was dissolved in DMF (300 mL), potassium carbonate (69.4 g, 502.5 mmol) was added, the temperature was cooled to zero degree, and p-methoxybenzyl chloride (PMB-Cl) (31.5 g, 201.0 mmol) was slowly added dropwise. The reaction was carried out at 80 °C for 1 h. LC-MS showed that the reaction was complete. After filtration, the solvent was evaporated under reduced pressure. Water (300 mL) and ethyl acetate (150 mL) were added for extraction twice. After evaporation of the solvent, the residue was triturated with ethyl acetate (100 mL) to obtain methyl 3-((4-methoxybenzyl)oxy)-4-nitrobenzoate (yellow solid, 49.2 g). LC-MS [M+H]+: m / z 316.2

[0138] Second step: The above yellow solid (2.0 g, 6.3 mmol) was dissolved in methanol and water (20 mL / 2 mL), iron powder (1.76 g, 31.5 mmol) and ammonium chloride (1.70 g, 31.5 mmol) were added, and the reaction was carried out at 75 °C overnight. LC-MS showed that the reaction was complete. After filtration through diatomaceous earth twice, the solvent was evaporated under reduced pressure to obtain a white solid, which was triturated with water (20 mL) to obtain methyl 4-amino-3-((4-methoxybenzyl)oxy)benzoate (white solid, 1.6 g).

[0139] Third step: The above white solid compound (3.0 g, 10.2 mmol) and 2,4-dichloro-5-iodopyrimidine (5.58 g, 20.4 mmol) were dissolved in 2-pentanol (10 mL), and then DIEA (5.22 g, 40.8 mmol) was added. The reaction was carried out by sealed tube at 130 °C overnight. After filtration, the solid was triturated with methanol to obtain methyl 4-((2-chloro-5-iodopyrimidin-4-yl)amino)-3-((4-methoxybenzyl)oxy)benzoate (3.5 g).

[0140] Step 4: Dissolve the compound obtained in the previous step (2.0 g, 3.8 mmol) in DMF (30 mL), add sodium acetate (2.06 g, 15.2 mmol), and then add PdCl2(PPh3)2Cl2 (266 mg, 0.38 mmol). Under nitrogen protection, react at 120 °C overnight. Add water (60 mL) and ethyl acetate (60 mL), and perform column chromatography to obtain methyl 2-chloro-8-((4-methoxybenzyl)oxy)-9H-pyrimido[4,5-b]indole-6-carboxylate (1.3 g). LC-MS [M+H]+: m / z 398.4.

[0141] Step 5: Dissolve the compound obtained in the previous step (500 mg, 1.25 mmol) and 1-ethyl-3-methyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (450.0 mg, 1.9 mmol) in dioxane and water (24 mL / 4 mL), add sodium carbonate (530.0 mg, 5.0 mmol), and then add Pd(PPh3)4 (144.0 mg, 0.125 mmol). Under nitrogen protection, react at 85 °C overnight. Concentrate, and separate by column chromatography (EA / PE = 1 / 1) to obtain Intermediate B5 (white solid, 320 mg). LC-MS [M+H]+: m / z 472.2. 1H NMR (400 MHz, DMSO-d6): δ 9.29 (s, 1H), 8.35 (s, 1H), 7.87 - 7.92 (m, 1H), 7.49 - 7.62 (m, 2H), 6.73 - 7.16 (m, 4H), 5.32 (s, 2H), 4.82 (s, 2H), 3.77 (s, 3H), 1.99 (s, 3H), 1.17 - 1.35 (m, 3H).

[0142] Intermediate C: Methyl 2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrido[2,3-b]indole-6-carboxylate

[0143]

[0144] Step 1: Methyl 4-amino-3-iodo-5-methoxybenzoate (1.0 g, 3.3 mmol) and (2,6-dichloropyridin-3-yl)boronic acid (1.26 g, 6.6 mmol) were added to DMF (50 mL). Under nitrogen protection, palladium acetate (Pd(OAc)2) (74 mg, 0.33 mmol), triphenylphosphine (PPh3) (86 mg, 0.33 mmol) and triethylamine (TEA) (1.0 g, 9.9 mmol) were added. The mixture was heated to 85 °C and reacted for 5 h. The reaction solution was cooled to room temperature, concentrated under reduced pressure, and purified by column chromatography to obtain methyl 4-amino-3-(2,6-dichloropyridin-3-yl)-5-methoxybenzoate (0.7 g, pale yellow solid). 1 1H NMR (DMSO-d6, 400 MHz): δ 7.82 (d, J = 8.0 Hz, 1H), 7.61 (d, J = 8.0 Hz, 1H), 7.35 (s, 1H), 7.24 (s, 1H), 5.53 (s, 2H), 3.87 (s, 3H), 3.76 (s, 3H). LC-MS: m / z 327.0 / 329.0 [M+H] + 。

[0145] Step two: Methyl 4-amino-3-(2,6-dichloropyridin-3-yl)-5-methoxybenzoate (420 mg, 1.29 mmol) was dissolved in DMF (20 mL), 18-crown-6 (680 mg, 2.57 mmol) and sodium hydride (NaH) (60% in mineral oil, 206 mg, 5.15 mmol) were added. Under nitrogen protection, the temperature was raised to 100 °C and reacted for 2 h. The reaction solution was cooled to room temperature, quenched with water (50 mL), extracted three times with ethyl acetate (50 mL), the organic phases were combined, dried, filtered, concentrated under reduced pressure, and purified by column chromatography to obtain methyl 2-chloro-8-methoxy-9H-pyrido[2,3-b]indole-6-carboxylate (142 mg, white solid). 1 1H NMR (DMSO-d6, 400 MHz): δ 12.62 (brs, 1H), 8.72 (d, J = 8.0 Hz, 1H), 8.53 (s, 1H), 7.59 (s, 1H), 7.34 (d, J = 8.0 Hz, 1H), 4.05 (s, 3H), 3.91 (s, 3H). LC-MS: m / z 291.1 / 293.1 [M+H] + 。

[0146] Step 3: Dissolve methyl 2-chloro-8-methoxy-9H-pyrido[2,3-b]indole-6-carboxylate (273 mg, 0.94 mmol) and 1,3-dimethyl- pyrazole-5-boronic acid pinacol ester (278 mg, 1.18 mmol) in dioxane and water (30 mL / 5 mL). Under nitrogen protection, add sodium carbonate (398 mg, 3.75 mmol) and tetrakis(triphenylphosphine)palladium (Pd(PPh3)4) (100 mg, 0.09 mmol), and react at 85 °C overnight. Cool the reaction solution to room temperature, concentrate under reduced pressure, and purify by column chromatography to obtain intermediate C (160 mg, white solid). 1 H NMR (DMSO-d6, 400 MHz): δ 12.48 (brs, 1H), 8.73 (d, J = 8.4 Hz, 1H), 8.54 (s, 1H), 7.60 - 7.64 (m, 2H) 6.63 (s, 1H), 4.67 - 4.70 (m, 2H), 4.08 (s, 3H), 3.93 (s, 3H), 2.23 (s, 3H), 1.38 (t, J = 7.2 Hz, 3H). LC-MS: m / z 365.4 [M+H] + 。

[0147] Using commercial reagents as raw materials, the following intermediates were prepared by referring to the synthesis method of intermediate C1.

[0148]

[0149] Intermediate D: (S)-2-(1-ethyl-3-methyl-1H-pyrazole-5-carbamoyl)-3-(3-hydroxypropoxy)-3,4-dihydro-5-oxa-1,2a-diazacenaphthylene-7-carboxamide

[0150]

[0151] Intermediate D was prepared by the same method as in Example 10 of Document WO2020006432A1, LC-MS [M+H] + : m / z 413.3. 1 H NMR (400 MHz, DMSO): δ 12.72 (s, 1H), 7.93 (s, 1H), 7.59 (s, 1H), 7.34 (s, 2H), 6.64 (s, 1H), 4.08 - 4.66 (m, 7H), 3.41 - 3.43 (m, 2H), 1.83 (s, 3H), 1.34 - 1.59 (m, 7H).

[0152] Intermediate E: (E)-2-(1-Ethyl-3-methyl-1H-pyrazole-5-carbamoyl)-3-(4-hydroxybut-2-en-1-yl)-3H-imidazo[4,5-b]pyridine-6-carboxamide

[0153]

[0154] First step: Dissolve the compound (E)-2-(4-bromobut-2-en-1-yl)isoindoline-1,3-dione (17 g, 60.93 mmol) in DMF (200 ml), then add potassium acetate (11.9 g, 121.86 mmol), under nitrogen protection, heat to 80 °C and react for 16 h. Cool to room temperature, pour into 600 mL of water, and then extract with ethyl acetate (200 mL × 2). The organic layer is washed successively with saturated ammonium chloride (500 mL) and saturated brine (500 mL), dried, and purified by column chromatography to obtain (E)-4-(1,3-dioxoisoindolin-2-yl)but-2-en-1-yl acetate (white solid, 13.6 g). 1 1H-NMR (400 MHz, DMSO-d6): δ 8.01 - 7.77 (m, 4H), 5.88 - 5.75 (m, 1H), 5.75 - 5.58 (m, 1H), 4.49 (dd, 2H), 4.20 (dd, J = 5.2, 2H), 2.08–1.90 (m, 3H).

[0155] Second step: Dissolve the product from the previous step (13.6 g, 52.5 mmol) in methanol (1.3 L), then add sodium methoxide (0.28 g, 5.25 mmol), stir at room temperature for 16 h, then add 14 mL of 1N HCl to quench the reaction, evaporate to dryness, and purify by column chromatography to obtain (E)-2-(4-hydroxybut-2-en-1-yl)isoindoline-1,3-dione (white solid, 10.6 g). 1 1H-NMR (400 MHz, DMSO-d6): δ 8.29 - 7.61 (m, 4H), 5.66 (q, 2H), 4.71 (t, 1H), 4.30 - 4.00 (m, 2H), 3.89 (dd, 2H).

[0156] Step 3: Dissolve the product of the previous step (10.6 g, 48.85 mmol) in dichloromethane (150 mL), then add imidazole (6.65 g, 97.7 mmol) and N,N-dimethylaminopyridine (DMAP) (1.2 g, 9.77 mmol), stir for 5 min, add tert-butyldiphenylchlorosilane (14 mL, 53.37 mmol) at room temperature. After addition, stir for 4 h, add 200 mL of water, separate the layers, dry the organic layer, concentrate by rotary evaporation, and purify by column chromatography to obtain (E)-2-(4-((tert-butyldiphenylsilyl)oxy)but-2-en-1-yl)isoindoline-1,3-dione (white solid, 21 g). 1 1H-NMR (400 MHz, DMSO-d6): δ 7.99 - 7.79 (m, 4H), 7.58 (dd, 4H), 7.45 - 7.28 (m, 6H), 5.78 (dt, 1H), 5.69 (dt, 1H), 4.19 (d, 2H), 4.18 - 4.13 (m, 2H), 0.97 (s, 9H).

[0157] Step 4: Dissolve the product of the previous step (21 g, 46.15 mmol) in ethanol (500 mL), add hydrazine hydrate (80% content, 5.3 mL, 184.6 mmol), then reflux for 16 h, cool, filter, concentrate the filtrate by rotary evaporation, add water (200 mL) and ethyl acetate (200 mL), separate the layers, wash the organic layer with water, dry, and concentrate by rotary evaporation to obtain (E)-4-((tert-butyldiphenylsilyl)oxy)but-2-en-1-amine (colorless transparent liquid, 15 g). 1 1H-NMR (400 MHz, DMSO-d6): δ 7.64 - 7.60 (m, 4H), 7.50 - 7.34 (m, 6H), 5.78 (dt, 1H), 5.70 - 5.52 (m, 1H), 4.17 (dd, 2H), 3.15 (dd, 2H), 1.54 - 1.30 (m, 2H), 1.00 (s, 9H).

[0158] Step 5: Dissolve the product of the previous step (3.5 g, 16.4 mmol) and methyl 6-chloro-5-nicotinate (5.9 g, 18.03 mmol) in dioxane (60 mL), add DIEA (8.1 mL, 49.2 mmol) at room temperature, stir at room temperature for 3 h, pour the reaction mixture into water (200 mL), extract with ethyl acetate (300 mL), wash the organic layer with saturated brine, dry, concentrate by rotary evaporation, and purify by column chromatography to obtain methyl (E)-6-((4-((tert-butyldiphenylsilyl)oxy)but-2-en-1-yl)amino)-5-nicotinate (yellow solid, 6.3 g). 1H-NMR(400MHz, DMSO-d6): δ 9.09 (t, 1H), 8.86 (dd, 1H), 8.74 (dd, 1H), 7.59 (dd, 4H), 7.47 - 7.28 (m, 6H), 5.88 - 5.83 (m, 1H), 5.76 - 5.62 (m, 1H), 4.26 (t, 2H), 4.17 (t, 2H), 3.88 (d, 3H), 1.00 (s, 9H).

[0159] Step 6: Dissolve the product of the previous step (1.5 g, 0.97 mmol) in methanol / tetrahydrofuran / water (25 mL / 25 mL / 25 mL), add ammonium chloride (1.6 g, 29.7 mmol) and zinc powder (1.93 g, 29.7 mmol), stir at room temperature for 3 h, filter, pour the filtrate into water, extract with ethyl acetate, then wash the organic layer with saturated brine, dry, rotary evaporate to obtain methyl (E)-5-amino-6-((4-((tert-butyldiphenylsilyl)oxy)but-2-en-1-yl)amino)nicotinate (reddish-brown viscous substance, 1.4 g). LC-MS [M+H] + : m / z 476.2. 1 H-NMR(400MHz, DMSO-d6): δ 8.03 (t, 1H), 7.61 (dd, 4H), 7.50 - 7.32 (m, 6H), 7.17 (d, 1H), 6.53 (t, 1H), 5.87 (dt, 1H), 5.72 (dd, 1H), 5.02 (s, 2H), 4.19 (d, 2H), 4.14 - 4.01 (m, 2H), 3.74 (s, 3H), 0.99 (s, 9H).

[0160] Step 7: Dissolve the product of the previous step (200 mg, 0.42 mmol) in dichloromethane / anhydrous methanol (3 mL / 3 mL), add cyanogen bromide (108 mg, 1.0 mmol) at room temperature, then stir at room temperature for 16 h. Add saturated sodium bicarbonate solution (50 mL) to the reaction solution, stir for 1 h, extract with dichloromethane (50 mL), wash the organic layer with saturated brine, dry, rotary evaporate, and purify by TLC plate to obtain methyl (E)-2-amino-3-(4-((tert-butyldiphenylsilyl)oxy)but-2-en-1-yl)-3H-imidazo[4,5-b]pyridine-6-carboxylate (white solid, 50 mg). LC-MS [M+H] + : m / z 501.2. 11H-NMR (400 MHz, DMSO-d6): δ 8.51 (d, 1H), 7.85 (d, 1H), 7.55 (dd, 4H), 7.45 - 7.39 (m, 2H), 7.35 (dd, 4H), 7.17 (s, 2H), 5.88 (m, 1H), 5.64 (m, 1H), 4.75 (d, 2H), 4.15 (d, 2H), 3.81 (m, 3H), 0.96 (s, 9H).

[0161] Example

[0162] Example 9: (E)-9,9'-(but-2-ene-1,4-diyl)bis(2-(1,3-dimethyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide)

[0163]

[0164] First step: Dissolve intermediate B1 (160 mg, 0.46 mmol) in methanol (10 mL), add 2N NaOH (5 mL), and react at 100 °C overnight. Cool the reaction solution to room temperature, concentrate under reduced pressure to remove most of the organic solvents, adjust the pH to 3 with 1M dilute hydrochloric acid, precipitate the solid, filter, and dry to obtain 2-(1,3-dimethyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxylic acid (120 mg, white solid). LC-MS: m / z 338.3 [M+H] + . Second step: Dissolve 2-(1,3-dimethyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxylic acid (120 mg, 0.36 mmol) in DMF (15 mL), add NH4Cl (199 mg, 3.8 mmol), DIEA (140 mg, 1.08 mmol), and HATU (410.4 mg, 1.08 mmol). React at room temperature for 2 hours. Concentrate the reaction solution under reduced pressure and purify by column chromatography to obtain 2-(1,3-dimethyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide (60 mg, white solid). 1 1H NMR (DMSO-d6, 400 MHz): δ 12.77 (s, 1H), 9.53 (s, 1H), 8.44 (s, 1H), 8.15 (brs, 1H), 7.66 (s, 1H), 7.33 (brs, 1H), 6.81 (s, 1H), 4.28 (s, 3H), 4.05 (s, 3H), 2.22 (s, 3H). LC-MS: m / z 337.4 [M+H]+ .

[0166] Step 3: Dissolve 2-(1,3-dimethyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide (60 mg, 0.18 mmol) and (E)-1,4-dibromobut-2-ene (19.3 mg, 0.09 mmol) in DMF (10 mL), add cesium carbonate (120 mg, 0.37 mmol) and sodium iodide (60 mg, 0.40 mmol), and react at room temperature for 1 hour. Concentrate the reaction solution under reduced pressure and purify by column chromatography to obtain (E)-9,9'-(but-2-ene-1,4-diyl)bis(2-(1,3-dimethyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide (1 mg, white solid). 1 1H NMR (TFA-d, 400 MHz): δ 11.67 (s, 2H), 10.60 (d, J = 0.8 Hz, 2H), 9.80 (s, 2H), 9.37 (s, 2H), 8.11 (brs, 2H), 7.53 (brs, 4H), 6.46 (s, 6H), 5.88 (s, 6H), 4.55 (s, 6H). LC-MS: m / z 725.0 [M+H] + .

[0167] Using intermediate B as the raw material, the following example compounds were prepared with reference to the synthesis method of Example 9.

[0168]

[0169] Example 12: (E)-5-(4-(8-carbamoyl-3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-(3-morpholinopropoxy)-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-methoxy-5H-pyrido[4,3-b]indole-8-carboxamide

[0170]

[0171] Step 1: Dissolve intermediate A1 (500 mg, 1.43 mmol) and (E)-1,4-dibromobut-2-ene (606 mg, 2.86 mmol) in DMF (20 mL), add cesium carbonate (1.4 g, 4.29 mmol) and sodium iodide (429 mg, 2.86 mmol), and react at room temperature overnight. The reaction solution was concentrated under reduced pressure and purified by column chromatography to obtain methyl (E)-5-(4-bromobut-2-en-1-yl)-3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-methoxy-5H-pyrido[4,3-b]indole-8-carboxylate (275 mg, white solid). 1 1H NMR (DMSO-d6, 400 MHz): δ 9.56 (s, 1H), 8.60 (s, 1H), 8.03 (s, 1H), 7.63 (s, 1H), 6.66 (s, 1H), 5.95 - 6.23 (m, 2H), 5.37 - 5.38 (m, 2H), 4.05 - 4.16 (m, 8H), 3.93 (s, 3H), 2.22 (s, 3H). LC-MS: m / z 483.1 [M+H] + 。

[0172] Step 2: Dissolve methyl (E)-5-(4-bromobut-2-en-1-yl)-3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-methoxy-5H-pyrido[4,3-b]indole-8-carboxylate (275 mg, 0.57 mmol) and intermediate A3 (264 mg, 0.57 mmol) in DMF (15 mL), add cesium carbonate (372 mg, 1.14 mmol) and sodium iodide (86 mg, 0.57 mmol), and react at room temperature overnight. The reaction solution was concentrated under reduced pressure and purified by column chromatography to obtain methyl (E)-3-(1,3-dimethyl-1H-pyrazol-5-yl)-5-(4-(3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-methoxy-8-(methoxycarbonyl)-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-6-(3-morpholinopropoxy)-5H-pyrido[4,3-b]indole-8-carboxylate (120 mg, white solid). LC-MS: m / z 866.4 [M+H] + 。

[0173] Step 3: Dissolve methyl (E)-3-(1,3-dimethyl-1H-pyrazol-5-yl)-5-(4-(3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-methoxy-8-(methoxycarbonyl)-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-6-(3-morpholinopropoxy)-5H-pyrido[4,3-b]indole-8-carboxylate (120 mg, 0.14 mmol) in methanol (10 mL), add 2N NaOH (5 mL), and react at 100 °C overnight. Cool the reaction solution to room temperature, concentrate under reduced pressure to remove most of the organic solvents, adjust the pH to 3 with 1M dilute hydrochloric acid, precipitate the solid, filter, and dry to obtain (E)-5-(4-(8-carboxy-3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-(3-morpholinopropoxy)-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-methoxy-5H-pyrido[4,3-b]indole-8-carboxylic acid (90 mg, white solid). LC-MS: m / z 838.3 [M+H] + 。

[0174] Step 4: Dissolve (E)-5-(4-(8-carboxy-3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-(3-morpholinopropoxy)-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-methoxy-5H-pyrido[4,3-b]indole-8-carboxylic acid (90 mg, 0.11 mmol) in DMF (15 mL), add NH4Cl (58 mg, 1.1 mmol), DIEA (206 mg, 1.6 mmol) and HATU (84 mg, 0.22 mmol). React at room temperature overnight. Concentrate the reaction solution under reduced pressure and purify by preparative chromatography to obtain (E)-5-(4-(8-carbamoyl-3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-(3-morpholinopropoxy)-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-methoxy-5H-pyrido[4,3-b]indole-8-carboxamide (17 mg, white solid). 11H NMR (CD3OD, 400 MHz): δ 9.58 (s, 1H), 9.57 (s, 1H), 8.56 (d, J = 1.2 Hz, 1H), 8.53 (d, J = 1.2 Hz, 1H), 8.06 (s, 1H), 8.05 (s, 1H), 7.74 (s, 1H), 7.69 (d, J = 1.2 Hz, 1H), 6.58 (s, 1H), 6.54 (s, 1H), 5.73 - 5.85 (m, 2H), 5.39 - 5.44 (m, 4H), 4.05 - 4.20 (m, 4H), 3.71 - 3.81 (m, 11H), 3.24 - 3.44 (m, 6H), 2.34 (s, 3H), 2.33 (s, 3H), 2.10 - 2.15 (m, 2H). LC-MS: m / z 836.0 [M + H] + 。

[0175] Example 13: (E)-5-(4-(8-Carbamoyl-3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-methoxy-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-3-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-6-methoxy-5H-pyrido[4,3-b]indole-8-carboxamide

[0176]

[0177] First step: Dissolve 3-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-6-methoxypyrido[4,3-b]indole-8-carboxamide (100 mg, 0.29 mmol) and (E)-1,4-dibromobut-2-ene (318 mg, 1.5 mmol) in DMF (20 mL), add cesium carbonate (391 mg, 1.2 mmol) and sodium iodide (50 mg, 0.3 mmol), and stir at room temperature for 1 hour. Add water (40 mL) and ethyl acetate (40 mL) to the reaction solution for liquid separation. Extract the aqueous phase with ethyl acetate (40 mL) once, combine the organic phases, concentrate under reduced pressure, grind the residue with ethyl acetate (20 mL), filter, and dry to obtain (E)-9-(4-bromobut-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrido[2,3-b]indole-6-carboxamide (100 mg, yellow solid). LC-MS: m / z 482.3 [M + H] + 。

[0178] Step 2: Dissolve (E)-9-(4-bromobut-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrido[2,3-b]indole-6-carboxamide (50 mg, 0.1 mmol) and 3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-methoxy-5H-pyrido[4,3-b]indole-8-carboxamide (34 mg, 0.1 mmol) in DMF (20 mL), add cesium carbonate (131 mg, 0.4 mmol) and sodium iodide (16 mg, 0.1 mmol), and stir at room temperature for 1 hour. Concentrate the reaction solution under reduced pressure and purify by preparative chromatography to obtain (E)-5-(4-(8-carbamoyl-3-(1,3-dimethyl-1H-pyrazol-5-yl)-6-methoxy-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-3-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-6-methoxy-5H-pyrido[4,3-b]indole-8-carboxamide (5.6 mg, white solid). 1 H NMR(CD3OD,400MHz):δ9.45(s,1H),9.43(s,1H),8.46(d,J=1.2Hz,1H),8.44(d,J=1.2Hz,1H),7.87(s,1H),7.81(s,1H),7.63(s,1H),7.61(s,1H),6.30(s,1H),6.24(s,1H),5.93(br s,2H),5.38(brs,4H),4.24-4.26(m,2H),3.81-3.83(m,9H),2.29(s,3H),2.28(s,3H),1.23(t,J=7.2Hz,3H). LC-MS:m / z 737.8[M+H] + 。

[0179] Using intermediates A and B as raw materials, the following example compounds were prepared with reference to the synthesis methods of Example 12 and Example 13.

[0180]

[0181]

[0182] Example 19: (S)-3-(3-(6-carbamoyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indol-9-yl)propyl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-carboxamide)-3,4-dihydro-5-oxa-1,2a-diazacenaphthylen-7-carboxamide

[0183]

[0184] Prepared by the same method as in Example 12 to obtain the target compound (white solid, 3.1 mg), MS (ESI): m / z = 745.4 [M+H]. 1H NMR (400 MHz, MeOD-d4): (9.13 (s, 1H), 8.21 (s, 1H), 7.44 (s, 1H), 7.36 (s, 1H), 7.23 (s, 1H), 6.78 (s, 1H), 6.31 (s, 1H), 5.34 (s, 2H), 4.54 - 4.80 (m, 6H), 4.09 - 4.11 (m, 1H), 3.79 (s, 3H), 2.27 (s, 3H), 2.12 (s, 3H), 1.20 - 1.59 (m, 10H).

[0185] Example 25: (E)-9-(4-(6-Carboxamido-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-8-(3-(difluoromethoxy)propoxy)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0186]

[0187] First step: Dissolve 3-((4-methoxybenzyl)oxy)propyl-1-ol (1.0 g, 5.10 mmol) in acetonitrile (40 mL), then add CuI (193.8 mg, 1.02 mmol), heat to 70 °C and slowly add 2,2-difluoro-2-(fluorosulfonylfluoro)acetic acid (1.81 g, 10.2 mmol), and react at room temperature for 2 hours. Wash with ethyl acetate / water (40 / 40 mL) 3 times, spin dry and mix the sample, and purify layer by layer with petroleum ether / ethyl acetate (volume ratio PE / EA = 10 / 1) to obtain the crude oil 1-((3-(difluoromethoxy)propoxy)methyl)-4-methoxybenzene (300 mg). 1 H-NMR (400 MHz, MeOD-d4): δ 7.24 (d, 2H), 6.88 (d, 2H), 6.12 (t, 1H), 4.42 (s, 2H), 3.92 (d, 2H), 3.78 (s, 3H), 3.31 (t, 2H), 1.86 - 1.90 (m, 2H).

[0188] Second step: Dissolve the crude product from the previous step (10 mg, 0.04 mmol) in methanol (MeOH) (5 mL), add palladium on carbon (2 mg), and react overnight at room temperature under hydrogen. Filter and separate to obtain the oily product 3-(difluoromethoxy)propyl-1-ol (4.0 mg).

[0189] Step 3: Dissolve the crude product from the previous step (4 mg, 0.03 mmol) in DCM (10 mL), add TEA (9.1 mg, 0.09 mmol), cool down to 0 °C, and slowly add methanesulfonyl chloride (5.2 mg, 0.045 mmol). React at room temperature for 2 hours. Wash with saturated sodium carbonate (10 mL) three times, rotary evaporate to obtain the crude oil 3-(difluoromethoxy)propyl methanesulfonate (4 mg), which is directly used for the next reaction.

[0190] Step 4: Dissolve the crude product from the previous step (5 mg, 0.006 mmol) and (E)-9-(4-(6-amide-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-hydroxy-pyrimido[4,5-b]indole-6-carboxamide (2.5 mg, 0.012 mmol) in DMF (5 mL), add cesium carbonate (5.9 mg, 0.018 mmol), react at room temperature overnight, extract with dichloromethane, wash with saturated sodium bicarbonate solution and water, concentrate, and prepare and isolate Example Compound 25 (white solid, 0.6 mg). LC-MS [M+H] + : m / z 960.4.

[0191] Example 26: (E)-9-(4-(6-amide-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-8-(3-(4,4-difluoropiperidin-1-yl)propoxy)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0192]

[0193] Example 26 was prepared by the same method as Example 25 (white solid, 6.3 mg). LC-MS [M+H]+: m / z 1013.4. 1H-NMR (400 MHz, MeOD-d4): δ 9.29 (s, 2H), 8.27 (s, 2H), 7.32 (s, 2H), 6.82 (s, 2H), 5.63 (s, 2H), 5.17 (s, 4H), 4.60 - 4.66 (m, 6H), 3.68 - 3.70 (m, 4H), 3.54 - 3.57 (m, 4H), 2.27 - 2.32 (m, 4H), 2.25 (s, 6H), 2.20 - 2.23 (m, 6H), 1.83 - 1.87 (m, 4H), 1.46 - 1.49 (m, 4H), 1.37 (t, 6H).

[0194] Example 27: (E)-9-(4-(6-Carboxamido-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-8-(3-(3,3-difluoroazetidin-1-yl)propoxy)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0195]

[0196] Example 27 (white solid, 5.5 mg) was prepared by the same method as in Example 25. LC-MS [M+H] + : m / z 985.4. 1 H-NMR (400 MHz, MeOD-d4): δ 9.28 (s, 2H), 8.27 (s, 2H), 7.36 (d, 2H), 6.79 (d, 2H), 5.64 (s, 2H), 5.19 (s, 4H), 4.60 - 4.64 (m, 6H), 3.74 - 3.78 (m, 4H), 3.56 - 3.59 (m, 4H), 4.41 - 4.47 (m, 4H), 2.33 - 2.47 (m, 2H), 2.26 - 2.27 (m, 6H), 2.22 - 2.24 (m, 4H), 1.34 - 1.38 (m, 4H), 1.30 - 1.34 (m, 6H).

[0197] Example 28: (E)-9-(4-(6-Carboxamido-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrido[2,3-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0198]

[0199] Step 1: Dissolve 2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrido[2,3-b]indole-6-carboxamide (100 mg, 0.3 mmol) and (E)-1,4-dibromobut-2-ene (190.0 mg, 0.9 mmol) in DMF (25 mL), add cesium carbonate (200 mg, 0.6 mmol), and react at room temperature for 1 h. Add water (40 mL) and ethyl acetate (40 mL) for extraction once, then slurry with ethyl acetate and petroleum ether (5 mL, volume ratio 1 / 1), filter, and dry to obtain (E)-9-(4-bromobut-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrido[2,3-b]indole-6-carboxamide (75 mg). LC-MS [M+H]+: m / z 484.4.

[0200] Step 2: Dissolve the crude product from the previous step (75 mg, 0.16 mmol) and 2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide (74 mg, 0.16 mmol) in DMF (5 mL), add cesium carbonate (104.3 mg, 0.32 mmol), and react at room temperature overnight. Separate by column chromatography (volume ratio DCM / MeOH = 10 / 1) to obtain (E)-9-(4-(6-carboxamide-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrido[2,3-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide (white solid, 50 mg). LC-MS [M+H]+: m / z 865.6.

[0201] Step 3: Dissolve the crude product from the previous step (20 mg, .02 mmol) in DCM (10 mL), then add boron tribromide (BBr3) (5 mL), and react at 50 °C overnight. Quench with methanol, rotary evaporate to obtain the crude product (E)-9-(4-(6-carboxamide-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-hydroxy-9H-pyrido[2,3-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide (20 mg). LC-MS [M+H] + : m / z 851.4.

[0202] Step 4: Dissolve the compound from the previous step (20 mg, 0.023 mmol) and 3-morpholinopropyl-1-ol (13.29 mg, 0.046 mmol) in DMF (5 mL), add cesium carbonate (22.49 mg, 0.069 mmol), and react overnight at room temperature. Extract with dichloromethane, wash with saturated sodium bicarbonate solution and water, concentrate, and prepare and isolate Example 28 (white solid, 3.7 mg). LC-MS [M+H] + : m / z 978.5. 1 H-NMR (400 MHz, MeOD-d4): δ 9.32 (s, 1H), 8.40 (d, 1H), 8.30 (d, 2H), 7.57 (d, 1H), 7.39 (d, 2H), 6.84 (s, 1H), 6.59 (s, 1H), 5.60 - 5.61 (m, 1H), 5.45 - 5.49 (m, 1H), 5.21 - 4.26 (m, 2H), 5.27 - 5.38 (m, 2H), 4.40 - 4.62 (m, 2H), 4.38 - 4.40 (m, 2H), 3.78 - 3.79 (m, 12H), 3.08 - 3.28 (m, 12H), 2.31 (d, 6H), 1.94 - 1.96 (m, 2H), 1.81 - 1.84 (m, 2H), 1.34 - 1.8. (m, 3H), 1.26 - 1.36 (m, 3H).

[0203] Example 29: (E)-9-(4-(6-Amidino-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrido[2,3-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0204]

[0205] Step 1: Dissolve 2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrido[2,3-b]indole-6-carboxamide (50 mg, 0.14 mmol) and (E)-1,4-dibromobut-2-ene (148.4 mg, 0.7 mmol) in DMF (20 mL), add cesium carbonate (136.9 mg, 0.42 mmol), and react at room temperature for 1 h. Dilute with water (40 mL) and EA (40 mL), separate the organic phase, spin-dry, triturate with ethyl acetate (20 mL), filter, and dry to obtain (E)-9-(4-bromobut-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrido[2,3-b]indole-6-carboxamide (gray solid, 20 mg). LC-MS [M+H] + : m / z 484.2.

[0206] Step 2: Dissolve the compound obtained in the previous step (20 mg, 0.045 mmol) and 2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide (19 mg, 0.041 mmol) in DMF (20 mL), add cesium carbonate (58.7 mg, 0.18 mmol), and react at room temperature for 1 h. Extract with dichloromethane, wash with saturated sodium bicarbonate solution and water, concentrate, and prepare and separate to obtain the compound of Example 29 (white solid, 35.3 mg). LC-MS [M+H] + : m / z 865.5. 1 1H-NMR (400 MHz, DMSO-d6): δ 9.48 (s, 1H), 8.51 - 8.53 (m, 1H), 8.41 (s, 2H), 8.01 - 8.05 (m, 2H), 7.35 - 7.60 (m, 5H), 6.78 (s, 1H), 6.58 (s, 1H), 5.72 - 5.77 (m, 2H), 5.22 (s, 4H), 4.57 - 4.59 (m, 2H), 4.22 - 4.25 (m, 2H), 3.93 (s, 2H), 3.77 (s, 3H), 3.45 (s, 4H), 2.11 - 2.21 (m, 12H), 1.54 (s, 2H), 1.02 - 1.23 (m, 6H).

[0207] Example 30: (E)-9-(4-(8-carboxamido-3-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-6-(3-morpholinopropoxy)-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide

[0208]

[0209] Step 1: Dissolve 2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide (50 mg, 0.14 mmol) and A (148.4 mg, 0.7 mmol) in DMF (20 mL), add cesium carbonate (136.9 mg, 0.42 mmol), and react at room temperature for 1 h. Add water (40 mL) and ethyl acetate (EA) (40 mL), extract with ethyl acetate (40 mL), evaporate to dryness, triturate with ethyl acetate (20 mL), filter, and dry to obtain a gray solid (40 mg). LC-MS [M+H] + : m / z 485.1.

[0210] Step 2: Dissolve the gray solid from the previous step (40 mg, 0.09 mmol) and 3-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-6-(3-morpholinopropoxy)-5H-pyrido[4,3-b]indole-8-carboxamide (38 mg, 0.81 mmol) in DMF (20 mL), add cesium carbonate (117.4 mg, 0.36 mmol), and react at room temperature for 1 h. Extract with dichloromethane, wash with saturated sodium bicarbonate solution and water, concentrate, and prepare for separation to obtain Example 30 (white solid, 35.3 mg). LC-MS [M+H] + : m / z 865.5. 1 1H-NMR (400 MHz, CD3OD): δ 9.42 (s, 1H), 9.31 (s, 1H), 8.44 (s, 1H), 8.33 (s, 1H), 7.87 (s, 1H), 7.59 (s, 1H), 7.54 (s, 1H), 6.73 (s, 1H), 6.29 (s, 1H), 5.79 (s, 2H), 5.35 (s, 2H), 5.28 (s, 2H), 4.49 - 4.51 (m, 2H), 4.02 - 4.20 (m, 5H), 3.76 (s, 4H), 3.21 - 3.30 (m, 8H), 2.05 - 2.30 (m, 8H), 1.19 - 1.25 (m, 6H).

[0211] Example 31: (E)-9,9'-(but-2-ene-1,4-diyl)bis(2-(1-ethyl-3-methyl-IH-pyrazol-5-yl)-8-((4-methoxybenzyl)oxy)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0212]

[0213] Dissolve the intermediate B5 compound (250 mg, 0.54 mmol) and 1,4-dibromobutene (58.0 mg, 0.3 mmol) in DMF (25 mL), add cesium carbonate (540 mg, 3.0 mmol), and react at room temperature for 1 h. Add water (40 mL) to precipitate the solid, slurry it with methanol (10 mL) and DMF (10 mL), filter, and dry to obtain Example 31 (gray solid, 72 mg). LC-MS [M+H]+: m / z 965.5. 1H-NMR (400 MHz, TFA-d6): δ 11.58 (s, 2H), 10.47 (s, 2H), 9.72 (s, 2H), 9.38 (s, 2H), 8.14 (s, 2H), 7.51 (s, 4H), 6.93 (s, 4H), 4.55 (s, 6H), 3.63 (s, 6H).

[0214] Example 32: (E)-9,9'-(But-2-ene-1,4-diyl)bis(2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-hydroxy-9H-pyrimido[4,5-b]indole-6-carboxamide)

[0215]

[0216] Dissolve Example 31 (72 mg, 0.08 mmol) in DCM (10 mL), then add trifluoroacetic acid (TFA) (5 mL), and heat the reaction at 50 °C overnight. Evaporate to dryness, slurry it with methanol (10 mL), and filter to obtain Example 32 (white solid, 42 mg). 1H-NMR (400 MHz, TFA-d6): δ 11.15 (s, 2H), 10.03 (s, 2H), 9.28 (s, 2H), 8.94 (s, 2H), 7.70 (s, 2H), 7.07 (s, 4H), 6.50 (s, 4H), 4.11 (s, 6H), 3.19 (s, 6H).

[0217] Example 33: (E)-9-(4-(6-Carbamoyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-((4-methoxybenzyl)oxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide

[0218]

[0219] Dissolve compound 3a (520.0 mg, 0.88 mmol) and compound 2d (309.5 mg, 0.88 mmol) in DMF (20 mL), add cesium carbonate (860.6 mg, 2.64 mmol), and react at room temperature for 2 h. Add water (40 mL) and ethyl acetate (40 mL) for extraction once, then slurry with ethyl acetate and petroleum ether (5 mL, v:v = 1:1), filter, and dry to obtain Example 33 (white solid, 580.0 mg). LC-MS [M+H] + : m / z 859.6. 1 H NMR (400 MHz, DMSO-d6): δ 9.48 (s, 2H), 8.45 (s, 2H), 8.05 (s, 2H), 7.69 (s, 1H), 7.57 (s, 1H), 7.40 (s, 1H), 6.41 - 7.57 (m, 2H), 7.08 - 7.11 (m, 2H), 6.75 - 6.99 (m, 2H), 6.53 - 6.63 (m, 2H), 5.69 - 5.74 (m, 2H), 5.55 - 5.60 (m, 4H), 5.10 (s, 2H), 4.47 - 4.57 (m, 4H), 3.59 - 3.80 (m, 6H), 2.20 (s, 6H), 1.30 (t, 3H), 1.20 (t, J = 6.4 Hz, 3H).

[0220] Example 34: (E)-9-(4-(6-Carbamoyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-hydroxy-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide

[0221]

[0222] Dissolve the compound of Example 33 (60.0 mg, 0.07 mmol) in DCM (10 mL), add TFA (5 mL), and react at room temperature overnight. Slurry with methanol and dichloromethane (5 mL) = 1 / 1, filter, and dry to obtain Example 34 (white solid, 50.0 mg). LC-MS [M+H] + : m / z 739.5. 1 H NMR (400 MHz, TFA-d): δ 11.64 (s, 2H), 10.57 (s, 2H), 9.78 (s, 2H), 9.40 (s, 2H), 8.14 (s, 2H), 7.54 (s, 4H), 6.92 - 6.95 (m, 4H), 5.89 (s, 3H), 4.57 (s, 6H), 3.60 - 3.67 (m, 6H).

[0223] (E)-9-(4-(6-Carbamoyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-methoxypropoxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide

[0224]

[0225] Dissolve the compound of Example 34 (25.0 mg, 0.03 mmol) and the bromide raw material (9.3 mg, 0.06 mmol) in DMF (10 mL), add cesium carbonate (29.1 mg, 0.09 mmol), and react at room temperature overnight. After preparative chromatography separation, Example 35 (white solid, 2.4 mg) was obtained. LC-MS [M+H] + : m / z 811.6. 1 H NMR (400 MHz, DMSO-d6): δ 9.47 (s, 1H), 9.46 (s, 1H), 8.43 (s, 1H), 8.42 (s, 1H), 8.06 (s, 2H), 7.59 (s, 1H), 7.55 (s, 1H), 7.38 - 7.41 (m, 2H), 6.75 (s, 1H), 6.72 (s, 1H), 5.79 (s, 2H), 5.20 - 5.23 (m, 4H), 4.49 - 4.58 (m, 4H), 4.00 - 4.04 (m, 2H), 3.77 (s, 3H), 3.25 - 3.28 (m, 2H), 3.11 (s, 3H), 2.19 (s, 3H), 2.16 (s, 3H), 1.71 - 1.74 (m, 2H), 1.20 (t, 3H), 1.15 (t, 3H).

[0226] (E)-9-(4-(6-Carbamoyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-((4-(trifluoromethyl)benzyl)oxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide

[0227]

[0228] Example compound 34 (25.0 mg, 0.03 mmol) and 4-(trifluoromethyl)benzyl chloride (11.6 mg, 0.06 mmol) were dissolved in DMF (10 mL), cesium carbonate (29.3 mg, 0.09 mmol) was added, and the reaction was carried out overnight at room temperature. After preparative chromatography, example compound 36 (white solid, 17.1 mg) was obtained. LC-MS [M+H] + : m / z 897.5. 1 1H-NMR (400 MHz, DMSO-d6): δ 9.50 (s, 1H), 9.44 (s, 1H), 8.46 (s, 1H), 8.43 (s, 1H), 8.06 (s, 2H), 7.69 (s, 1H), 7.55 (s, 1H), 7.40 - 7.49 (m, 6H), 6.77 (s, 1H), 6.69 (s, 1H), 5.84 - 5.90 (m, 1H), 5.50 - 5.63 (m, 1H), 5.21 - 5.23 (m, 2H), 5.12 - 5.15 (m, 4H), 4.57 - 4.59 (m, 2H), 4.48 - 4.49 (m, 2H), 3.64 (s, 3H), 2.19 (s, 3H), 2.16 (s, 3H), 1.21 (t, 3H), 1.11 (t, 3H).

[0229] Example 37: (E)-9,9'-(but-2-ene-1,4-diyl)bis(2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-((4-(trifluoromethyl)benzyl)oxy)-9H-pyrimido[4,5-b]indole-6-carboxamide)

[0230]

[0231] Example compound 32 (30.0 mg, 0.04 mmol) and 4-(trifluoromethyl)benzyl chloride (23.3 mg, 0.12 mmol) were dissolved in DMF (10 mL), cesium carbonate (39.1 mg, 0.12 mmol) was added, and the reaction was carried out overnight at room temperature. After preparative chromatography, example 37 (white solid, 19.8 mg) was obtained. LC-MS [M+H] + : m / z 1041.2. 1H-NMR(400MHz, DMSO-d6): δ 9.48 (s, 2H), 8.51 (s, 2H), 8.09 (s, 2H), 7.67 (s, 2H), 7.47 (s, 2H), 7.34 - 7.37 (m, 4H), 7.29 - 7.31 (m, 4H), 6.75 (s, 2H), 5.51 (s, 2H), 5.14 (s, 4H), 5.01 (s, 4H), 4.54 - 4.56 (m, 4H), 2.19 (s, 6H), 1.19 (t, 6H).

[0232] Example 38: (E)-9,9'-(But-2-ene-1,4-diyl)bis(2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-methoxypropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide)

[0233]

[0234] Dissolve the compound of Example 32 (30. 0 mg, 0.04 mmol) and the bromide raw material (18.2 mg, 0.12 mmol) in DMF (10 mL), add cesium carbonate (39.1 mg, 0.12 mmol), react at room temperature overnight, and prepare Example 38 (white solid, 20.2 mg) by preparative chromatography. LC-MS [M+H] + : m / z 869.6. 1 H NMR(400MHz, DMSO-d6): δ 9.47 (s, 2H), 8.41 (s, 2H), 8.05 (s, 2H), 7.54 (s, 2H), 7.38 (s, 2H), 6.75 (s, 2H), 5.76 (s, 2H), 5.21 (s, 4H), 4.53 - 4.58 (m, 4H), 3.96 - 3.99 (m, 4H), 3.21 - 3.24 (m, 4H), 3.10 (s, 6H), 2.20 (s, 6H), 1.64 - 1.68 (m, 4H), 1.20 (t, 6H).

[0235] Example 39: (E)-9,9'-(But-2-ene-1,4-diyl)bis(8-(2-(benzyloxy)ethoxy)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-9H-pyrimido[4,5-b]indole-6-carboxamide)

[0236] Using 2-(benzyloxy)ethyl bromide and the compound of Example 32 as raw materials, Example 39 (white solid, 12.0 mg) was synthesized according to the method of Example 38 by reference. LC-MS [M+H] + : m / z 993.7. 11H-NMR (400 MHz, DMSO-d6): δ 9.48 (s, 2H), 8.44 (s, 2H), 8.05 (s, 2H), 7.59 (s, 2H), 7.41 (s, 2H), 7.09 - 7.17 (m, 10H), 6.75 (s, 2H), 5.65 (s, 2H), 5.10 (s, 4H), 4.52 - 4.55 (m, 4H), 4.23 (s, 4H), 4.10 (s, 4H), 3.41 - 3.45 (m, 4H), 2.19 (s, 6H), 1.18 (t, 6H).

[0237] Example 40: (E)-8-(2-(Benzyloxy)ethoxy)-9-(4-(6-acyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0238] Using Example 34 and 2-benzyloxybromoethane as raw materials, Example 40 (white solid, 13.3 mg) was synthesized by referring to the method of Example 38. LC-MS [M + H] + : m / z 873.5. 1 1H-NMR (400 MHz, DMSO-d6): δ 9.46 (s, 2H), 8.43 (s, 2H), 8.04 - 8.07 (m, 2H), 7.58 - 7.60 (m, 2H), 7.40 - 7.41 (m, 2H), 7.15 - 7.21 (m, 5H), 6.72 (s, 2H), 5.70 - 5.78 (m, 2H), 5.12 - 5.18 (m, 4H), 4.46 - 4.58 (m, 4H), 4.30 (s, 2H), 4.17 (s, 2H), 3.74 (s, 3H), 3.56 - 3.66 (m, 2H), 2.17 (d, 6H), 1.16 (t, 6H).

[0239] Example 41: (E)-9,9'-(But-2-ene-1,4-diyl)bis(2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(2-hydroxyethoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide)

[0240]

[0241] Step 1: Dissolve the compound of Example 39 (10.0 mg, 0.01 mmol) in TFA (10 mL), and react at 90 °C overnight. The reaction was monitored by LC-MS and found to be complete. The solvent was evaporated to dryness to obtain the crude product (10.0 mg), which was directly used in the next step. LC-MS [M+H] + : m / z 1005.5.

[0242] Step 2: Dissolve the crude product compound from the previous step (10.0 mg, 0.01 mmol) in MeOH / H2O (10 mL / 1 mL), add potassium carbonate (4.2 mg, 0.03 mmol), and react at room temperature for 4 h. The reaction was monitored by LCMS and found to be complete. The product was purified by preparative chromatography to obtain Example 41 (white solid, 1.5 mg). LC-MS [M+H] + : m / z 813.5. 1 1H-NMR (400 MHz, DMSO-d6): δ 9.48 (s, 2H), 8.42 (s, 2H), 8.06 (s, 2H), 7.57 (s, 2H), 7.41 (s, 2H), 6.72 (s, 2H), 5.84 (s, 2H), 5.23 - 5.25 (m, 4H), 4.83 - 4.88 (m, 1H), 4.49 - 4.54 (m, 4H), 3.98 - 4.01 (m, 4H), 3.37 - 3.54 (m, 5H), 2.18 (s, 6H), 1.18 (t, 6H).

[0243] Example 42: (E)-9-(4-(6-Acyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(2-hydroxyethoxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide

[0244] Using the compound of Example 40 as the starting material, Example 42 (white solid, 2.5 mg) was synthesized according to the method of Example 41. LC-MS [M+H] + : m / z 783.5. 1 1H-NMR (400 MHz, DMSO-d6): δ 9.46 (s, 2H), 8.41 (s, 2H), 8.05 (s, 2H), 7.59 (s, 2H), 7.41 (s, 2H), 6.72 (s, 2H), 5.84 (s, 2H), 5.18 - 5.28 (m, 4H), 4.47 - 4.53 (m, 4H), 4.03 - 4.05 (m, 2H), 3.78 (s, 3H), 3.50 - 3.57 (m, 3H), 2.18 (s, 6H), 1.16 (t, 6H).

[0245] Example 43: (E)-9-(4-(6-Acyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-8-(3-(diethylamino)propoxy)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0246]

[0247] Example 43 (white solid, 6.1 mg) was prepared in the same manner as in Example 25. LC-MS [M+H] + : m / z 965.6. 1 H-NMR (400 MHz, MeOD-d4): δ 9.28 (s, 2H), 8.27 (s, 1H), 8.26 (s, 1H), 7.48 (s, 1H), 7.39 (s, 1H), 6.84 (s, 1H), 6.78 (s, 1H), 5.64 (s, 2H), 5.16 (s, 4H), 4.61 - 4.66 (m, 6H), 3.68 - 3.73 (m, 4H), 3.54 - 3.57 (m, 4H), 2.36 - 2.38 (m, 4H), 2.26 - 2.27 (m, 6H), 2.18 - 2.22 (m, 6H), 1.47 - 1.49 (m, 4H), 1.32 - 1.38 (m, 6H), 0.92 (t, 6H).

[0248] Example 44: (E)-8-Butoxy-9-(4-(6-acyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0249] Example 44 (white solid, 2.2 mg) was prepared in the same manner as in Example 25. LC-MS [M+H] + : m / z 908.6. 1H-NMR(400MHz,MeOD-d4):δ9.28 - 9.31(s,2H),8.35(s,1H),8.27(s,1H),7.48(s,1H),7.39(s,1H),6.84(s,1H),6.78(s,1H),5.57 - 5.71(m,2H),5.05 - 5.25(m,4H),4.58 - 4.74(m,4H),3.74 - 3.95(m,8H),3.14 - 3.29(m,4H),2.26 - 2.28(m,6H),1.92 - 1.96(m,2H),1.22 - 1.39(m,10H),0.80(t,3H).

[0250] Example 45: (E)-9-(4-(6-Acyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-8-(cyclopropylmethoxy)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0251] Example 45 was prepared by the same method as in Example 25 (white solid, 12.0 mg). LC-MS [M + H] + : m / z 906.6. 1 H-NMR(400MHz,MeOD-d6):δ9.26(s,1H),8.31(s,1H),8.24(s,1H),7.47(s,1H),7.37(s,1H),6.79(s,1H),6.74(s,1H),5.73 - 5.79(m,2H),5.25 - 5.67(m,5H),4.50 - 4.70(m,3H),3.98 - 4.02(m,4H),3.65 - 3.67(m,4H),3.20 - 3.23(m,4H),2.25 - 2.29(m,6H),1.98 - 2.01(m,2H),1.26 - 1.33(m,6H),0.84 - 0.95(m,1H),0.42 - 0.45(m,2H),0.38 - 040(m,2H).

[0252] Example 46: (E)-9-(4-(6-Carbamoyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-hydroxypropoxy)-9H-pyrido[2,3-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0253] Example 46 (white solid, 19.1 mg) was prepared in the same manner as in Example 25. LC-MS [M+H] + : m / z 909.4. 1 H NMR (400 MHz, DMSO-d6): δ 9.83 (s, 1H), 9.72 (s, 1H), 8.50 - 8.55 (d, 1H), 8.46 (s, 1H), 8.40 (s, 1H), 8.04 - 8.08 (m, 2H), 7.61 - 7.63 (d, 1H), 7.53 - 7.57 (m, 2H), 7.38 - 7.46 (m, 2H), 6.76 (s, 1H), 6.61 (s, 1H), 5.66 (s, 2H), 5.18 - 5.23 (m, 4H), 4.52 - 4.53 (m, 2H), 4.25 - 4.29 (m, 2H), 3.98 - 4.07 (m, 8H), 3.58 - 3.61 (m, 2H), 3.40 - 3.43 (m, 2H), 2.94 - 3.17 (m, 4H), 2.51 - 2.86 (m, 2H), 2.20 (s, 6H), 1.76 - 1.78 (m, 2H), 1.60 - 1.64 (m, 2H), 1.19 (t, 3H), 1.08 (t, J = 5.6 Hz, 3H).

[0254] Example 47: (E)-9-(4-(6-carbamoyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-hydroxypropoxy)-9H-pyrido[2,3-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide

[0255] Example 47 (white solid, 17.5 mg) was prepared in the same manner as in Example 25. LC-MS [M+H] + : m / z 796.4. 11H NMR (400 MHz, DMSO-d6): δ 9.47 (s, 1H), 8.51 (d, 1H), 8.42 (s, 1H), 8.37 (s, 1H), 8.01 - 8.08 (m, 2H), 7.53 - 7.59 (m, 3H), 7.33 - 7.42 (m, 2H), 6.73 (s, 1H), 6.58 (s, 1H), 5.68 - 5.87 (m, 2H), 5.18 - 5.27 (m, 4H), 4.43 - 4.48 (m, 2H), 4.24 - 4.29 (m, 2H), 4.07 - 4.10 (m, 2H), 3.98 (s, 3H), 3.45 - 3.48 (m, 2H), 2.18 (s, 6H), 1.64 - 1.77 (m, 2H), 1.04 - 1.12 (m, 6H).

[0256] Example 48: (E)-9-(4-(6-Carbamoyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-methoxypropoxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0257] Example 48 white solid (44.1 mg) was prepared by the same method as in Example 25. LC-MS [M + H] + : m / z 924.8. 1 1H NMR (400 MHz, DMSO-d6): δ 9.50 (d, 2H), 8.47 (d, 2H), 8.44 (d, 2H), 8.08 (d, 2H), 7.56 (d, 2H), 6.76 (d, 2H), 5.62 - 5.69 (m, 2H), 5.18 - 5.19 (m, 4H), 4.52 - 4.61 (m, 4H), 3.87 - 4.01 (m, 8H), 3.38 - 3.41 (m, 4H), 3.17 - 3.22 (m, 5H), 2.89 - 2.92 (m, 2H), 2.20 (d, 6H), 1.59 - 1.61 (m, 2H), 1.22 - 1.25 (m, 2H), 1.19 - 1.21 (m, 6H).

[0258] (E)-9-(4-(6-carbamoyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(2-hydroxyethoxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide Example 49 was prepared as a white solid (30.1 mg) using the same method as in Example 25. LC-MS [M+H] + : m / z 896.3. 1 H NMR (400 MHz, DMSO-d6): δ 9.48 (d, 2H), 8.45 (d, 2H), 8.08 (d, 2H), 7.58 (d, 2H), 7.44 (d, 2H), 6.73 (d, 2H), 5.62 - 5.89 (m, 2H), 5.19 - 5.25 (m, 4H), 4.50 - 4.55 (m, 4H), 3.89 - 3.99 (m, 4H), 4.00 - 4.02 (m, 2H), 3.54 - 3.61 (m, 5H), 3.28 - 3.30 (m, 2H), 3.13 - 3.19 (m, 2H), 2.91 - 2.94 (m, 2H), 2.17 (d, 6H), 1.15 - 1.22 (m, 2H), 1.10 - 1.15 (m, 6H).

[0259] (E)-9-(4-(6-carbamoyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrido[2,3-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide

[0260] Example 50 was prepared as a white solid (6.2 mg) using the same method as in Example 25. LC-MS [M+H] + : m / z 865.2.

[0261] (E)-9-(4-(6-carbamoyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrido[2,3-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-hydroxypropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0262] Example 51 was prepared in the same manner as in Example 25 (white solid, 5.6 mg). LC-MS [M+H] + : m / z 909.3.

[0263] Example 52: (E)-9-(4-(8-carbamoyl-3-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-6-(3-hydroxypropoxy)-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0264] Example 52 was prepared in the same manner as in Example 25 (white solid, 18.0 mg). LC-MS [M+H] + : m / z 909.5.

[0265] Example 53: (E)-9-(4-(8-carbamoyl-3-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-6-(3-hydroxypropoxy)-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide

[0266] Example 53 was prepared in the same manner as in Example 25 (white solid, 18.0 mg). LC-MS [M+H] + : m / z 769.3.

[0267] Example 54: (E)-9-(4-(8-carbamoyl-3-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-6-(pyrimidin-2-ylmethoxy)-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-methoxy-9H-pyrimido[4,5-b]indole-6-carboxamide

[0268] Example 54 was prepared in the same manner as in Example 25 (white solid, 5.1 mg). LC-MS [M+H] + : m / z 830.2.

[0269] (E)-9-(4-(8-Carbamoyl-3-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-6-(3-morpholinopropoxy)-5H-pyrido[4,3-b]indol-5-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0270] Example 55 was prepared as a pale yellow solid (5.0 mg) by the same method as in Example 25. LC-MS [M+H] + : m / z 978.4. 1 H NMR (400 MHz, MeOD-d4): δ 9.50 (s, 1H), 9.36 (s, 1H), 8.48 (s, 1H), 8.37 (s, 1H), 7.94 (s, 1H), 7.50 - 7.62 (m, 2H), 6.85 (s, 1H), 6.32 (s, 1H), 5.72 - 5.42 (m, 2H), 5.15 - 5.25 (m, 4H), 4.50 - 4.71 (m, 2H), 4.17 - 4.21 (m, 2H), 3.99 - 4.13 (m, 8H), 3.74 - 3.81 (m, 4H), 3.40 - 3.42 (m, 4H), 3.01 - 3.21 (m, 8H), 2.09 - 2.28 (m, 6H), 1.85 - 1.91 (m, 2H), 1.80 - 1.95 (m, 2H), 1.29 - 1.30 (m, 6H).

[0271] Example 56: (E)-9-(4-(6-Carbamoyl-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-hydroxypropoxy)-9H-pyrimido[4,5-b]indol-9-yl)but-2-en-1-yl)-2-(1-ethyl-3-methyl-1H-pyrazol-5-yl)-8-(3-morpholinopropoxy)-9H-pyrimido[4,5-b]indole-6-carboxamide

[0272] Example 56 was prepared as a white solid (9.0 mg) by the same method as in Example 25. LC-MS [M+H] + : m / z 910.3. 11H NMR (400 MHz, MeOD-d4): δ 10.13 - 12.02 (m, 2H), 8.16 - 8.19 (m, 2H), 7.38 - 7.41 (m, 2H), 6.61 - 6.60 (m, 2H), 5.59 - 5.64 (m, 2H), 5.12 - 5.33 (m, 4H), 4.44 - 4.48 (m, 4H), 4.02 - 4.12 (m, 2H), 3.95 - 4.01 (m, 4H), 3.93 - 3.95 (m, 2H), 3.76 - 3.78 (m, 2H), 3.57 - 3.58 (m, 2H), 3.30 - 3.34 (m, 2H), 3.05 - 3.19 (m, 2H), 1.98 - 2.20 (m, 6H), 1.73 - 1.98 (m, 2H), 1.70 - 1.72 (m, 2H), 1.19 - 1.28 (m, 6H).

[0273] Comparative Compound 1: Prepared according to the synthesis method of Example 4 on page 85 of the reference patent WO2019069275A. LC-MS [M + H] + : 850.5 m / z. 1 1H NMR (400 MHz, CD3OD): δ 7.59 (s, 2H), 7.27 (s, 1H), 7.25 (s, 1H), 6.61 (s, 1H), 6.59 (s, 1H), 5.80 (br s, 2H), 5.01 (br s, 4H), 4.57 - 4.61 (m, 4H), 3.97 (br s, 4H), 3.71 (br s, 5H), 3.12 - 3.31 (m, 6H), 2.18 (s, 6H), 1.95 - 1.98 (m, 2H), 1.31 - 1.39 (m, 6H).

[0274] Comparative Compound 2: Prepared according to the synthesis method of Example 1 in the reference patent WO2020028565A1. 1 1H NMR (TFA-d, 400 MHz): δ 11.66 (s, 2H), 10.58 (d, J = 0.8 Hz, 2H), 9.78 (s, 2H), 9.36 (s, 2H), 8.09 (br s, 2H), 7.51 (br s, 4H), 6.87 - 6.89 (m, 4H), 5.84 (s, 6H), 4.54 (s, 6H), 3.58 (t, J = 7.2 Hz, 6H). MS (ESI): m / z = 753.0 [M + H].

[0275] Comparative Compound 3: The comparative compound 3 was prepared according to the synthesis method of Example 2 in Patent WO2020028565A1. LC-MS [M+H] + : m / z 841.4. 1 H NMR (400 MHz, DMSO-d6): δ 9.46 (s, 2H), 8.40 (s, 2H), 8.06 (s, 2H), 7.56 (s, 2H), 7.39 (s, 2H), 6.73 (s, 2H), 5.77 (s, 4H), 5.21 (s, 4H), 4.50 - 4.54 (m, 6H), 4.01 - 4.05 (m, 4H), 3.37 - 3.39 (m, 4H), 2.17 (s, 6H), 1.59 - 1.63 (m, 4H), 1.14 - 1.18 (m, 6H).

[0276] Test Example 1: Test for the binding activity of the compounds in the examples to the WT hSTING protein

[0277] Operate according to the test method of the WT hSTING kit provided by Cisbio: In a 96-well screening plate, the STING protein reacts with the test compound (starting concentration 10 μM, ten-fold serial dilution in triplicate) and buffer at 37 °C for 30 minutes, and then co-incubates with 20 μM fluorescent substrate at 37 °C for 30 minutes. Use a Thermo Scientific Verioskan Flash multi-functional reader to read the light intensity at 455 nm with 358 nm as the excitation light. Calculate the competitive binding activity of the sample to the protein based on the measured fluorescence value compared to the value of the blank well. The EC 50 value was calculated by fitting with Prism software from Graphpad company.

[0278] The results showed that most of the compounds in the examples of the present invention could bind well to the WT hSTING protein, and their EC 50 value was less than 100 nM, and the EC 50 value of most of the examples was less than 10 nM, and the EC50 value of some of the compounds in the examples was even less than 1 nM, reaching the activity level of pM. (The specific data is shown in Table 1, A < 0.3 nM, 0.3 nM ≤ B < 1 nM, 1 nM ≤ C < 10 nM, 10 nM ≤ D < 100 nM, E ≥ 100 nM).

[0279] Table 1. Binding activity of the compounds in the examples to the WT hSTING protein

[0280] Number <![CDATA[EC 50 > Number <![CDATA[EC 50 > Number <![CDATA[EC 50 > 1 E 2 E 3 E 4 D 5 C 6 B 7 E 8 E 9 E 10 A 11 A 12 E 13 C 14 E 15 C 16 A 17 C 18 A 19 A 20 B 21 B 22 B 23 B 24 B 25 C 26 C 27 C 28 C 29 A 30 A 31 B 32 B 33 B 34 C 35 B 36 A 37 A 38 B 39 A 40 A 41 A 42 B 43 C 44 C 45 C 46 A 47 A 48 A 49 A 50 A 51 A 52 A 53 A 54 A 55 A 56 A Comparative Compound 1 B Comparative Compound 2 B Comparative Compound 3 C

[0281] Test Example 2: Test for the release activity of the compounds in the examples to stimulate THP1 cells to release IFNβ by the Elisa method

[0282] Test procedure: 1. THP-1 cells are cultured in RPMI1640 + 10% FBS + 0.05 mM β-ME, and the density is maintained at 2×10 5 ~1×10 6 viable cells / mL; collect the cells, after discarding the old medium, wash the cells once with serum-free medium, centrifuge to remove the supernatant, and resuspend the cells with serum-free medium. After trypan blue staining and counting, when the cell viability is greater than 95%, proceed with the subsequent experiment; adjust the cell density to 1.1×10 6 viable cells / mL with serum-free medium; add 180 μL of the cell suspension to the wells of a 96-well cell culture plate, and the cell density is 2×10 5 viable cells / well. 2: Dilute the positive control and the test compound with the solvent to form a stock solution and perform serial dilutions to obtain 10-fold concentration solutions; add 20 μL of the 10-fold concentration compound solution to each well, with two replicates for each concentration. The highest concentration of the positive control compound cGAMP is 100 μM, and it is diluted 3-fold for a total of 5 concentrations. The highest concentration of the test compound is 10 μM, and it is diluted 5-fold for a total of 5 concentrations; 3. Place the cell culture plate in an incubator and incubate for 24 hrs; centrifuge the cell culture plate at 2000 g for 5 minutes, transfer the supernatant directly for ELISA detection; calculate the IFN-β concentration in the ELISA test samples according to the IFN-β standard curve respectively. The IFN-β concentration in the cell supernatant is equal to the IFN-β concentration in the ELISA sample multiplied by the dilution factor; 4. Analyze the data using GraphPad Prism 5.0 software, fit the data with a non-linear S curve regression to obtain the dose-effect curve, and calculate the EC 50 value.

[0283] Results: For the compounds in most of the examples of the present invention, the EC 50 value of the IFNβ release activity stimulated in THP1 cells is less than 1 μM, and for some examples, the EC 50 value is even less than 100 nM, and for some examples, the EC 50 value is even less than 1 nM, such as Examples 16, 29, 39, 56, etc. (specific EC 50 data are shown in Table 2, A < 1 nM, 1 nM ≤ B < 10 nM, 10 nM ≤ C < 100 nM, D ≥ 100 nM).

[0284] Table 2. IFNβ release activity stimulated by the compounds of the examples in THP1 cells

[0285]

[0286] Test Example 3: Detection of the inhibitory ability (antagonism) of the compounds in the examples on the secretion of IFN-β by THP-1 stimulated by 2'3'-cGAMP using the Elisa method

[0287] Test method: 1. THP-1 cells were cultured using RPMI1640 + 10% FBS + 0.05 mM β-ME, and maintained at a density of 2×10 5 ~1×10 6 viable cells / mL; the cells were collected, and after discarding the old medium, the cells were washed once with serum-free medium. After centrifugation to remove the supernatant, the cells were resuspended with serum-free medium. After trypan blue staining, the cells were counted. When the cell viability was greater than 95%, the subsequent experiments were carried out. The cell density was adjusted to 1.1×10 6 viable cells / mL using serum-free medium; 160 μL of the cell suspension was added to the wells of a 96-well cell culture plate, and the cell density was 2×10 5 viable cells / well. 2. The positive control and the test compound were diluted with the solvent to form a stock solution and then serially diluted to obtain a 10-fold concentration solution. 20 μL of the 10-fold concentration compound solution was added to each well, and the cells were incubated in an incubator for 2 hrs. Then, 20 μL of cGAMP was added to each well, and the final concentration of cGAMP was 30 μM. The highest concentration of the test compound was 30 μM, and it was diluted 5-fold for a total of 5 concentrations. One replicate well was set for each concentration. 3. The cell culture plate was placed in an incubator and incubated for 24 hrs, centrifuged at 2000 g for 5 minutes, and the supernatant was transferred directly for ELISA detection. 4. The ELISA detection process was carried out with reference to the instructions of R&D Systems (Cat# DY814-05). The concentrations of IFN-β in the ELISA test samples were calculated according to the IFN-β standard curve respectively. The concentration of IFN-β in the cell supernatant was equal to the concentration of IFN-β in the ELISA sample multiplied by the dilution factor. The data was analyzed using GraphPad Prism 5.0 software, and the dose-effect curve was obtained by fitting the data with a non-linear S curve regression to calculate the EC 50 value.

[0288] Results: During the long-term research process, the inventors of the present invention for the first time found that some of the compounds of the general formula (I) in the present invention have good activity in antagonizing the STING function, and the EC 50 is less than 20 uM, such as Examples 11, 18, 30, 36, 55; the inhibitory ability of some of the compounds in the examples on the secretion of IFN-β by THP-1 stimulated by 2'3'-cGAMP, the EC 50 is less than 1 uM, such as Examples 11 and 16.

[0289] Test Example 4: Inhibitory activity of the compounds in the examples on different enzymes

[0290] (1) Prepare 1×Kinase buffer; (2) Preparation of compound concentration gradient: The test concentration of the test compound starts from 10 μM, and is diluted 10 times at a 3-fold dilution, with duplicate wells for testing. In a 96-well plate, it is gradient-diluted into 10 different concentrations of solutions with a final concentration of 100 times. Then, each concentration of the compound is further diluted with 1×Kinase buffer into an intermediate dilution solution with a final concentration of 5 times; (3) Take 5 μL of each prepared compound solution and add it to the compound wells of a 384-well plate for single-well testing of each concentration; add 5 μL of 5% DMSO to the negative control well and the positive control well respectively; (4) Prepare a kinase solution with a final concentration of 2.5 times with 1×Kinase buffer; (5) Add 10 μL of the kinase solution with a final concentration of 2.5 times to the compound wells and the positive control well respectively; add 10 μL of 1×Kinase buffer to the negative control well; (6) Centrifuge at 1000 rpm for 30 seconds, incubate at room temperature for 10 minutes after shaking well; (7) Prepare a mixed solution of ATP and Kinase substrate22 with a final concentration of 2.5 times with 1×Kinase buffer; (8) Add 10 μL of the mixed solution of ATP and substrate with a final concentration of 2.5 times to start the reaction; (9) Centrifuge the 384-well plate at 1000 rpm for 30 seconds, incubate at 28 °C for the corresponding time after shaking well; (10) Add 30 μL of the termination detection solution to stop the kinase reaction, centrifuge at 1000 rpm for 30 seconds, and shake well; (11) Read the conversion rate with Caliper EZ ReaderⅡ. Use the log value of the concentration as the X-axis and the percentage inhibition rate as the Y-axis, and use the log(inhibitor) vs. response-Variable slope of the analysis software GraphPad Prism 5 to fit the dose-effect curve, so as to obtain the IC 50 value of each compound on enzyme activity. Using the above test method, the compounds of the examples are respectively interacted with kinases such as GSK3, TBK, CDK, IKK, AMPK, ULK1, etc., and the inhibitory activities of the compounds on the above enzymes are tested.

[0291] Results: Compounds 11, 16, 18, 19, 39, and 56 of the examples of the present invention have weak inhibitory activities on kinases such as GSK3, TBK, CDK, IKK, AMPK, ULK1, etc., and the IC 50 > 500 nM, showing relatively high selectivity for binding to STING protein.

[0292] All documents mentioned in this invention are cited herein as references, as if each document was individually cited as a reference. In addition, it should be understood that after reading the above teachings of this invention, those skilled in the art can make various changes or modifications to this invention, and these equivalent forms also fall within the scope defined by the appended claims of this application.

Claims

1. A class of nitrogen-containing fused-ring compounds, or pharmaceutically acceptable salts thereof, which are the following compounds:

2. Use of the compound or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, For the preparation of drugs for the prevention or treatment of STING protein-dependent diseases.

3. The use according to claim 2, characterized in that, The diseases are selected from the following group: tumors, autoimmune diseases, viral infections, degenerative diseases.

4. The use according to claim 2, wherein, The diseases are selected from the following group: non-small cell lung cancer, small cell lung cancer, breast cancer, prostate cancer, liver cancer, skin cancer, gastric cancer, intestinal cancer, cholangiocarcinoma, brain cancer, leukemia, lymphoma, fibroma, sarcoma, kidney cancer, bone cancer, thyroid cancer, nasopharyngeal cancer, pancreatic cancer, rejection of transplanted organs, gout, rhinitis, hair loss, Alzheimer's disease, appendicitis, atherosclerosis, asthma, arthritis, atopic dermatitis, Behcet's disease, bullous skin disease, cholecystitis, chronic idiopathic thrombocytopenic purpura, chronic obstructive pulmonary disease, cirrhosis, degenerative joint disease, dermatitis, dermatomyositis, eczema, enteritis, encephalitis, gastritis, nephritis, Hashimoto's thyroiditis, hepatitis, hypophysitis, inflammatory bowel disease, irritable bowel syndrome, Kawasaki disease, meningitis, multiple sclerosis, myocarditis, myasthenia gravis, mycosis fungoides, myositis, nephritis, osteomyelitis, pancreatitis, Parkinson's disease, pericarditis, pernicious anemia, pneumonia, primary biliary sclerosing cholangitis, polyarteritis nodosa, psoriasis, fibrosis, lupus erythematosus, tissue transplant rejection, thyroiditis, type I diabetes, urethritis, uveitis, vasculitis, vitiligo, Waldenström's macroglobulinemia.

5. The use according to claim 2, characterized in that, The diseases are selected from the following group: lung adenocarcinoma, squamous cell carcinoma of the lung, basal cell carcinoma, glioma, melanoma.

6. A pharmaceutical composition comprising the compound as described in claim 1 or a pharmaceutically acceptable salt thereof, characterized in that, The pharmaceutical composition comprises: (i) An effective amount of the compound, or a pharmaceutically acceptable salt thereof; and (ii) A pharmaceutically acceptable carrier.

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