Selenocyclic compounds and their applications

Abstract

A class of selenocyclic compounds and their applications. Specifically disclosed are the compounds shown by formula (II) and their pharmaceutically acceptable salts.

Description

[0001] This invention claims priority

[0002] CN202011136978.2, filing date: October 21, 2020;

[0003] CN202110242560.8, filing date: March 4, 2021. Technical Field

[0004] This invention relates to selenocyclic compounds and their applications. Specifically, it relates to the compounds shown in formula (II) and their pharmaceutically acceptable salts. Background Art

[0005] Inflammatory bowel disease (IBD) is an intestinal inflammatory disease with unknown etiology, including Crohn's disease (CD) and ulcerative colitis (UC), which are characterized by inflammation and ulcers in the mucosal layer of the rectum and large intestine. Common symptoms include diarrhea, bloody stools, and abdominal pain. The clinical course proceeds in an intermittent cycle of exacerbation and remission, and patients with ulcerative colitis have an increased risk of developing colorectal cancer (Dennis et al. N Engl J Med, 2011, 365, 1713 - 1725). The excessive inflammatory response in the gastrointestinal tract is mediated by inflammatory cytokines (such as TNFα, IFN-γ, IL-1, IL-6, IL-12, IL-21, and IL-23), and acts on cells of the innate and adaptive immune systems, including T and B lymphocytes, epithelial cells, macrophages, and dendritic cells (Neurath, M.F Nat.Rev.Immunol.2014, 14, 329). The Janus kinase (JAK) family: JAK1, JAK2, JAK3, and Tyk2, are non-receptor tyrosine kinases that play a key role in the transduction responses of many of the above-mentioned cytokines. When a cytokine binds to a receptor, the relevant JAK homo- or heterodimers are phosphorylated and activated, thereby enabling subsequent recruitment, phosphorylation, and activation of the signal transducer and activator of transcription (STAT) family of transcription factors. Phosphorylated STATs (pSTATs) are transported to the nucleus and induce the gene transcription of several chemokines, cytokines, and proteases related to the pathogenesis of IBD.

[0006] Genetic studies of patients with IBD have found that polymorphisms in several proteins related to the JAK / STAT pathway (such as IL-23R, IL-12B, JAK2, Tyk2, and STAT3) are risk factors for the development of IBD. It provides an attractive target for treating the excessive inflammatory response caused by IBD (Boland, B.S. et al., Gastroenterology clinics of North America 2014, 43, 603). Currently, several JAK inhibitors, such as tofacitinib and filgotinib, are in clinical development for IBD. Tofacitinib is approved in the United States for the treatment of rheumatoid arthritis (RA) and UC. Tofacitinib is also in clinical development for CD. Due to the lack of significant efficacy in the 4-week clinical trial of phase II in moderate-to-severe CD patients, further trials for this indication were aborted. Although there is conclusive evidence of target engagement based on biomarker analysis, it is currently unclear whether the efficacy of tofacitinib is related to the clinical study design, the mechanistic differences between UC and CD, or dose-limiting systemic adverse events (AEs) that prevent the drug from being fully exposed to intestinal tissues. Common systemic adverse events (AEs) in phase II and III clinical trials of tofacitinib for IBD include decreased hemoglobin, decreased absolute neutrophil count (ANC), increased total cholesterol (low-density and high-density lipids), and infections (Sandborn, W.J. et al., N. Engl. J. Med. 2012, 367, 616). Such AEs are consistent with those observed in patients with rheumatoid arthritis taking tofacitinib and are consistent with the JAK2-dependent inhibition of EPO and TPO. According to the results of the post-marketing safety trial of tofacitinib, using tofacitinib at a dose of 10 mg twice daily increases the risk of thrombosis and death, and the FDA has issued a black box warning for tofacitinib.

[0007] There are several possible ways to overcome the systemic AEs caused by JAK inhibition. One approach is to develop oral JAK1-selective inhibitors such as filgotinib and upadacitinib, which are currently being used in phase 3 clinical trials for CD and UC. Despite the theoretical safety advantages of JAK1-selective molecules, the recently approved twice-daily dose of 15 mg of upadacitinib for the treatment of rheumatoid arthritis also has a black box warning from the FDA regarding the risk of thrombosis (upadacitinib prescribing information). Another approach is to maximize the intestinal tissue exposure of JAK inhibitors while avoiding potential systemic exposure. Due to the regulatory role of the JAK / STAT pathway in the immune system, systemic exposure to JAK inhibitors may have adverse systemic immunosuppressive effects. Therefore, there is a need to provide new JAK inhibitors that act at the site of the lesion while having no significant systemic effects. Specifically, this has advantages for the treatment of gastrointestinal inflammatory diseases such as UC and CD. SUMMARY OF THE INVENTION

[0008] The present invention provides a compound of formula (II) or a pharmaceutically acceptable salt thereof,

[0009]

[0010] wherein,

[0011] L1 is -(CH2) m -, -C(=O)-, -S(=O)2- or -CH2-C(=O)-;

[0012] T1 is N or CH;

[0013] T2 is N or CH;

[0014] T3 is N or CH;

[0015] R1 is H, CN, C 1-3 alkyl, -NH-C 1-3 alkyl, 4-6 membered heterocycloalkyl or 5-6 membered heteroaryl, wherein the C 1-3 alkyl, -NH-C 1-3 alkyl, 4-6 membered heterocycloalkyl and 5-6 membered heteroaryl are each independently optionally substituted by 1, 2 or 3 R a substituents;

[0016] R2 is R 21 or

[0017] D1 is O, NH or CH2;

[0018] D2 is CH or N;

[0019] E1 and E2 are each independently a single bond or CH2;

[0020] R 21 is H, CN, NH2, C 1-3 alkyl, -NH-C 1-3 alkyl, -N(C 1-3 alkyl)2, -C(=O)-NH-C 1-3 alkyl and -C(=O)-NH-C 3-6 cycloalkyl, wherein the C 1-3 alkyl, -NH-C 1-3 alkyl, -N(C 1-3 alkyl)2, -C(=O)-NH-C 1-3 alkyl and -C(=O)-NH-C 3-6 cycloalkyl are each independently optionally substituted by 1, 2 or 3 R b substituents;

[0021] R3 is H or C 1-3 alkyl;

[0022] R4 is H or C 1-3 alkyl;

[0023] q is 1 or 2;

[0024] m is 0, 1 or 2;

[0025] n is 1 or 2;

[0026] R a and R b are each independently F, Cl, Br, I, OH, NH2, CN or COOH;

[0027] The "4-6 membered heteroalkyl" and "5-6 membered heteroaryl" each independently contain 1, 2 or 3 heteroatoms or heteroatom groups independently being -O-, -NH-, -S-, -Se- or N.

[0028] The present invention provides a compound of formula (I) or a pharmaceutically acceptable salt thereof,

[0029]

[0030] wherein,

[0031] L1 is -(CH2) m -, -C(=O)-, -S(=O)2- or -CH2-C(=O)-;

[0032] T1 is N or CH;

[0033] T2 is N or CH;

[0034] R1 is H, CN, C 1-3 alkyl, -NH-C 1-3 alkyl or 4-6 membered heteroalkyl, wherein said C 1-3 alkyl, -NH-C 1-3 alkyl and 4-6 membered heteroalkyl are optionally substituted by 1, 2 or 3 R a substituents;

[0035] R2 is R 21 or

[0036] D1 is O or CH2;

[0037] R 21 is H, C 1-3 alkyl, -NH-C 1-3 alkyl or -N(C 1-3 alkyl)2, wherein said C 1-3 alkyl, -NH-C 1-3 alkyl and -N(C 1-3 alkyl)2 are optionally substituted by 1, 2 or 3 R b substituents;

[0038] R3 is H or C 1-3 alkyl;

[0039] q is 1 or 2;

[0040] m is 0, 1 or 2;

[0041] R a and R b are each independently F, Cl, Br, I, OH, NH2, CN or COOH.

[0042] Said "4-6 membered heteroalkyl" contains 1, 2 or 3 heteroatoms or heteroatom groups independently being -O-, -NH-, -S-, -Se- or N.

[0043] In some embodiments of the present invention, the above R1 is H, CN, CH3, -NH-CH3, wherein said CH3, -NH-CH3, are each independently optionally substituted by 1, 2 or 3 R a substituents, and R a and other variables are as defined in the present invention.

[0044] In some embodiments of the present invention, the above R1 is H, CN, CH3, -NH-CH3 or wherein said CH3, -NH-CH3 and are optionally substituted by 1, 2 or 3 R a substituents, and R aand other variables are as defined in the present invention.

[0045] In some embodiments of the present invention, the above R1 is H, CN, CF3, -NH-CH3, and other variables are as defined in the present invention.

[0046] In some embodiments of the present invention, the above R1 is H, CN, CF3, -NH-CH3 or and other variables are as defined in the present invention.

[0047] In some embodiments of the present invention, the above structural unit is and other variables are as defined in the present invention.

[0048] In some embodiments of the present invention, the above R 21 is H, CN, NH2, CH3, wherein the CH3, is independently optionally substituted by 1, 2 or 3 R b substituents, and R b and other variables are as defined in the present invention.

[0049] In some embodiments of the present invention, the above R 21 is H, CN, NH2, CH3,

[0050] In some embodiments of the present invention, the above R2 is H, CN, NH2, CH3,

[0051] In some embodiments of the present invention, the above R2 is H, CH3, the CH3, is optionally substituted by 1, 2 or 3 R b substituents, and other variables are as defined in the present invention.

[0052] In some embodiments of the present invention, the above R2 is H, CH3, and other variables are as defined in the present invention.

[0053] In some embodiments of the present invention, the above R3 is H or CH3, and other variables are as defined in the present invention.

[0054] In some embodiments of the present invention, the above R4 is H or CH3, and other variables are as defined in the present invention.

[0055] In some embodiments of the present invention, the above L1 is -CH2-, -(CH2)2-, -C(=O)-, -S(=O)2- or -CH2-C(=O)-, and other variables are as defined in the present invention.

[0056] In some embodiments of the present invention, the above structural unit for Other variables are as defined herein.

[0057] In some embodiments of the present invention, the above structural unit for Other variables are as defined in the present invention.

[0058] In some embodiments of the present invention, the above structural unit for Other variables are as defined herein.

[0059] In some embodiments of the present invention, the above structural unit for Other variables are as defined in the present invention.

[0060] In some embodiments of the present invention, the above structural unit for Other variables are as defined in the present invention.

[0061] Some other solutions of the present invention are obtained by arbitrarily combining the above variables.

[0062] In some embodiments of the present invention, the above compound is

[0063]

[0064] in,

[0065] n, R1, R2, R3, L1, T3, T2 and q are as defined herein.

[0066] In some embodiments of the present invention, the above compound is

[0067]

[0068] in,

[0069] R1, R2, R3, L1, T2 and q are as defined herein.

[0070] The present invention also provides a compound of the following formula or a pharmaceutically acceptable salt thereof:

[0071]

[0072]

[0073]

[0074] The present invention provides the use of the above-mentioned compound or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating JAK-related diseases.

[0075] The present invention provides the use of the above-mentioned compound or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating gut-restricted pan-JAK-related diseases.

[0076] The present invention further provides a method for treating gut-restricted pan-JAK-related diseases in a subject in need thereof, comprising administering to the subject an effective dose of the compound or a pharmaceutically acceptable salt or a pharmaceutical composition thereof as defined in any of the above technical solutions.

[0077] In some technical solutions of the present invention, the gut-restricted pan-JAK-related disease is inflammatory bowel disease.

[0078] Technical effects

[0079] The compounds of the present invention showed good inhibitory activity in in vitro activity tests of two kinase subtypes, JAK1 and JAK2. The compounds of the present invention showed good inhibitory activity in in vitro activity tests of cell (THP1 and HT29) function experiments. The compounds of the present invention showed good drug exposure levels in the small intestine and colon of rats, and the ratios of small intestine / plasma and colon / plasma of the compounds were relatively high, showing good tissue selectivity. The compounds of the present invention showed good drug exposure levels in the small intestine and colon of mice, and the ratios of small intestine / plasma and colon / plasma of the compounds were relatively high, showing good tissue selectivity. In an oxazolone (OXA)-induced murine colitis model, the compounds of the present invention were able to alleviate OXA-induced weight loss, significantly improve the disease activity index (DAI) score and the colon weight / length ratio at the experimental endpoint, and showed good therapeutic effects.

[0080] Definitions and explanations

[0081] Unless otherwise specified, the following terms and phrases used herein are intended to have the following meanings. A particular term or phrase should not be considered indeterminate or unclear without a specific definition, but should be understood in its ordinary meaning. When a trade name appears herein, it is intended to refer to the corresponding product or its active ingredient.

[0082] The term "pharmaceutically acceptable" as used herein refers to those compounds, materials, compositions and / or dosage forms that are within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and animals, without excessive toxicity, irritation, allergic response or other problems or complications, and commensurate with a reasonable benefit / risk ratio.

[0083] The term "pharmaceutically acceptable salts" refers to salts of the compounds of the present invention, which are prepared from the compounds with specific substituents found in the present invention and relatively non-toxic acids or bases. When the compounds of the present invention contain relatively acidic functional groups, the base addition salts can be obtained by contacting such compounds with a sufficient amount of base in a pure solution or a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amine or magnesium salts or similar salts. When the compounds of the present invention contain relatively basic functional groups, the acid addition salts can be obtained by contacting such compounds with a sufficient amount of acid in a pure solution or a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include inorganic acid salts, where the inorganic acids include, for example, hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, bicarbonate, phosphoric acid, monohydrogen phosphate, dihydrogen phosphate, sulfuric acid, hydrogen sulfate, hydroiodic acid, phosphorous acid, etc.; and organic acid salts, where the organic acids include, for example, acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid, methanesulfonic acid and similar acids; also include salts of amino acids (such as arginine, etc.), and salts of organic acids such as glucuronic acid. Certain specific compounds of the present invention contain both basic and acidic functional groups, and thus can be converted into either base or acid addition salts.

[0084] The pharmaceutically acceptable salts of the present invention can be synthesized by conventional chemical methods from parent compounds containing acid radicals or basic groups. Generally, the preparation method of such salts is to react these compounds in the form of free acids or bases with a stoichiometric amount of appropriate bases or acids in water, an organic solvent or a mixture of both.

[0085] The compounds of the present invention can exist in specific geometric or stereoisomeric forms. The present invention contemplates all such compounds, including cis- and trans-isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereoisomers, (D)-isomers, (L)-isomers, and their racemic mixtures and other mixtures, such as enantiomer- or diastereomer-enriched mixtures, and all these mixtures are within the scope of the present invention. Additional asymmetric carbon atoms may be present in substituents such as alkyl groups. All these isomers and their mixtures are included within the scope of the present invention.

[0086] Unless otherwise specified, the term "enantiomer" or "optical isomer" refers to stereoisomers that are mirror images of each other.

[0087] Unless otherwise specified, the terms "cis-trans isomers" or "geometric isomers" are caused by the inability of double bonds or single bonds of ring carbon atoms to rotate freely.

[0088] Unless otherwise indicated, the term "diastereomer" refers to stereoisomers of a molecule that has two or more chiral centers and whose molecules have a non-mirror-image relationship.

[0089] Unless otherwise indicated, "(+)" indicates dextrorotation, "(-)" indicates levorotation, and "(±)" indicates racemization.

[0090] Unless otherwise indicated, the solid wedge bond and the dashed wedge bond represent the absolute configuration of a stereocenter, and the solid straight bond and the dashed straight bond represent the relative configuration of a stereocenter. The wavy line represents the solid wedge bond or the dashed wedge bond or the wavy line represents the solid straight bond and the dashed straight bond

[0091] Unless otherwise indicated, the term "tautomer" or "tautomeric form" refers to different functional group isomers that are in dynamic equilibrium at room temperature and can rapidly interconvert. If tautomers are possible (e.g., in solution), a chemical equilibrium of tautomers can be achieved. For example, proton tautomers (also known as prototropic tautomers) include interconversions that occur through proton migration, such as keto-enol isomerization and imine-enamine isomerization. Valence tautomers include interconversions that occur through the reorganization of some bonding electrons. A specific example of keto-enol tautomerization is the interconversion between the two tautomers of pentane-2,4-dione and 4-hydroxypent-3-en-2-one.

[0092] Unless otherwise indicated, the terms "enriched in one isomer", "isomer-enriched", "enriched in one enantiomer", or "enantiomer-enriched" mean that the content of one isomer or enantiomer is less than 100%, and the content of this isomer or enantiomer is greater than or equal to 60%, or greater than or equal to 70%, or greater than or equal to 80%, or greater than or equal to 90%, or greater than or equal to 95%, or greater than or equal to 96%, or greater than or equal to 97%, or greater than or equal to 98%, or greater than or equal to 99%, or greater than or equal to 99.5%, or greater than or equal to 99.6%, or greater than or equal to 99.7%, or greater than or equal to 99.8%, or greater than or equal to 99.9%.

[0093] Unless otherwise specified, the term "isomer excess" or "enantiomeric excess" refers to the difference between the relative percentages of two isomers or two enantiomers. For example, if the content of one isomer or enantiomer is 90% and the content of the other isomer or enantiomer is 10%, then the isomer or enantiomer excess (ee value) is 80%.

[0094] The optically active (R)- and (S)-isomers as well as D and L isomers can be prepared by chiral synthesis or chiral reagents or other conventional techniques. If one enantiomer of a compound of the present invention is desired, it can be prepared by asymmetric synthesis or derivatization with a chiral auxiliary, where the resulting mixture of diastereomers is separated and the auxiliary group is cleaved to provide the pure desired enantiomer. Alternatively, when the molecule contains a basic functional group (such as an amino group) or an acidic functional group (such as a carboxyl group), it forms a diastereomeric salt with an appropriate optically active acid or base, and then the diastereomers are resolved by conventional methods known in the art, and then the pure enantiomer is recovered. In addition, the separation of enantiomers and diastereomers is usually accomplished by using chromatography, which employs a chiral stationary phase and optionally in combination with chemical derivatization (such as forming a carbamate from an amine).

[0095] The compounds of the present invention may contain non-natural proportions of atomic isotopes on one or more atoms constituting the compound. For example, the compound can be labeled with a radioactive isotope, such as tritium ( 3 H), iodine-125 ( 125 I) or C-14 ( 14 C). Also, for example, deuterium can be used to replace hydrogen to form deuterated drugs. The bond formed by deuterium and carbon is stronger than the bond formed by ordinary hydrogen and carbon. Compared with the non-deuterated drug, the deuterated drug has advantages such as reducing toxic and side effects, increasing drug stability, enhancing efficacy, and prolonging the biological half-life of the drug. All transformations of the isotopic composition of the compounds of the present invention, whether radioactive or not, are included within the scope of the present invention. "Optional" or "optionally" means that the subsequent described event or condition may but does not necessarily occur, and the description includes the case where the described event or condition occurs and the case where the described event or condition does not occur.

[0096] The term "substituted" means that any one or more hydrogen atoms on a specific atom are replaced by a substituent, and the substituent can include deuterium and variants of hydrogen, as long as the valence state of the specific atom is normal and the resulting compound is stable. When the substituent is oxygen (i.e., =O), it means that two hydrogen atoms are replaced. Oxygen substitution does not occur on an aromatic group. The term "optionally substituted" means that it can be substituted or not substituted. Unless otherwise specified, the type and number of substituents can be arbitrary based on what is chemically achievable.

[0097] When any variable (e.g., R) appears more than once in the composition or structure of a compound, its definition in each case is independent. Thus, for example, if a group is substituted with 0 - 2 R's, the group may optionally be substituted with up to two R's, and each R in each case has independent options. In addition, combinations of substituents and / or their variants are only permitted if such combinations result in stable compounds.

[0098] When the number of a linking group is 0, such as -(CRR)0-, it indicates that the linking group is a single bond.

[0099] When one of the variables is selected from a single bond, it indicates that the two groups it connects are directly linked. For example, in A - L - Z, when L represents a single bond, it means the structure is actually A - Z.

[0100] When a substituent is vacant, it means the substituent is absent. For example, in A - X, when X is vacant, it means the structure is actually A. When it is not specified which atom of the listed substituent is connected to the group being substituted, such a substituent can be bonded through any of its atoms. For example, a pyridyl group as a substituent can be connected to the group being substituted through any carbon atom on the pyridine ring.

[0101] When the listed linking group does not specify its connection direction, the connection direction is arbitrary. For example, in which the linking group L is -M - W -, at this time -M - W - can connect ring A and ring B in the same direction as the reading order from left to right to form or can connect ring A and ring B in the opposite direction to the reading order from left to right to form The combinations of the said linking groups, substituents and / or their variants are only permitted if such combinations result in stable compounds.

[0102] Unless otherwise specified, when a group has one or more connectable sites, any one or more of these sites of the group can be connected to other groups through chemical bonds. When the connection mode of the chemical bond is not fixed and there are H atoms at the connectable sites, then when connecting the chemical bonds, the number of H atoms at this site will correspondingly decrease according to the number of connected chemical bonds to become a group with the corresponding valence. The chemical bonds connecting the said site to other groups can be represented by a straight solid line bond a straight dashed line bond or a wavy line For example, the straight solid line bond in -OCH3 indicates connection to other groups through the oxygen atom in this group; the straight dashed line bond in The wavy line in it indicates connection to other groups through the 1- and 2-carbon atoms in the phenyl group; It means that any connectable site on the piperidyl group can be connected to other groups through 1 chemical bond, including at least These 4 connection methods. Even if an H atom is drawn on -N-, but still includes Groups with this connection method. Just when connecting 1 chemical bond, the H at this site will correspondingly decrease by 1 to become the corresponding monovalent piperidyl group.

[0103] Unless otherwise specified, the number of atoms in a ring is usually defined as the ring member count. For example, a "5-7 membered ring" refers to a "ring" composed of 5-7 atoms arranged in a ring.

[0104] Unless otherwise specified, the term "C 1-3 alkyl" is used to represent a straight-chain or branched saturated hydrocarbon group composed of 1 to 3 carbon atoms. The C 1-3 alkyl includes C 1-2 and C 2-3 alkyl, etc.; it can be monovalent (such as methyl), divalent (such as methylene), or polyvalent (such as methine). Examples of C 1-3 alkyl include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), etc.

[0105] Unless otherwise specified, "C 3-6 cycloalkyl" represents a saturated cyclic hydrocarbon group composed of 3 to 6 carbon atoms, which is a monocyclic and bicyclic system. The C 3-6 cycloalkyl includes C 3-5 、C 4-5 and C 5-6 cycloalkyl, etc.; it can be monovalent, divalent, or polyvalent. Examples of C 3-6 cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, etc.

[0106] Unless otherwise specified, the term "4-6 membered heteroalkyl" alone or in combination with other terms respectively represents a saturated cyclic group composed of 4 to 6 ring atoms, in which 1, 2, 3, or 4 ring atoms are heteroatoms independently selected from O, S, -Se-, and N, and the rest are carbon atoms, where the nitrogen atom is optionally quaternized, and the nitrogen and sulfur heteroatoms can be optionally oxidized (i.e., NO and S(O) p, where p is 1 or 2). It includes monocyclic and bicyclic systems, and the bicyclic system includes spirocycles, fused rings, and bridged rings. In addition, for this "4- to 6-membered heterocycloalkyl", the heteroatom can occupy the connection position between the heterocycloalkyl and the rest of the molecule. The 4- to 6-membered heterocycloalkyl includes 5- to 6-membered, 4-membered, 5-membered, and 6-membered heterocycloalkyls, etc. Examples of the 4- to 6-membered heterocycloalkyl include, but are not limited to, azetidinyl, oxetanyl, thietanyl, pyrrolidinyl, pyrazolidinyl, imidazolidinyl, tetrahydrothienyl (including tetrahydrothiophen-2-yl, tetrahydrothiophen-3-yl, etc.), tetrahydrofuryl (including tetrahydrofuran-2-yl, etc.), tetrahydropyranyl, piperidinyl (including 1-piperidinyl, 2-piperidinyl, 3-piperidinyl, etc.), piperazinyl (including 1-piperazinyl, 2-piperazinyl, etc.), morpholinyl (including 3-morpholinyl, 4-morpholinyl, etc.), dioxolanyl, dithiolanyl, isoxazolidinyl, isothiazolidinyl, 1,2-oxazinyl, 1,2-thiazinyl, hexahydropyridazinyl, homopiperazinyl, or homopiperidinyl, etc.

[0107] Unless otherwise specified, the terms "5- to 6-membered heteroaryl ring" and "5- to 6-membered heteroaryl group" in the present invention can be used interchangeably. The term "5- to 6-membered heteroaryl group" refers to a monocyclic group composed of 5 to 6 ring atoms with a conjugated π-electron system, and 1, 2, 3, or 4 of its ring atoms are heteroatoms independently selected from O, S, and N, and the rest are carbon atoms. Among them, the nitrogen atom is optionally quaternized, and the nitrogen and sulfur heteroatoms can be optionally oxidized (i.e., NO and S(O) p , where p is 1 or 2). The 5- to 6-membered heteroaryl group can be connected to the rest of the molecule through a heteroatom or a carbon atom. The 5- to 6-membered heteroaryl group includes 5-membered and 6-membered heteroaryl groups. Examples of the 5- to 6-membered heteroaryl group include, but are not limited to, pyrrolyl (including N-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl, etc.), pyrazolyl (including 2-pyrazolyl, 3-pyrazolyl, etc.), imidazolyl (including N-imidazolyl, 2-imidazolyl, 4-imidazolyl, 5-imidazolyl, etc.), oxazolyl (including 2-oxazolyl, 4-oxazolyl, 5-oxazolyl, etc.), triazolyl (1H-1,2,3-triazolyl, 2H-1,2,3-triazolyl, 1H-1,2,4-triazolyl, 4H-1,2,4-triazolyl, etc.), tetrazolyl, isoxazolyl (3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl, etc.), thiazolyl (including 2-thiazolyl, 4-thiazolyl, 5-thiazolyl, etc.), furyl (including 2-furyl, 3-furyl, etc.), thienyl (including 2-thienyl, 3-thienyl, etc.), pyridinyl (including 2-pyridinyl, 3-pyridinyl, 4-pyridinyl, etc.), pyrazinyl, or pyrimidinyl (including 2-pyrimidinyl, 4-pyrimidinyl, etc.).

[0108] Unless otherwise specified, C n-n+m or C n -C n+m includes any specific case of n to n + m carbons, for example, C1-12 including C1, C2, C3, C4, C5, C6, C7, C8, C9, C 10 , C 11 , and C 12 , and also includes any range from n to n + m, for example C 1-12 includes C 1-3 , C 1-6 , C 1-9 , C 3-6 , C 3-9 , C 3-12 , C 6-9 , C 6-12 , and C 9-12 etc.; similarly, n-membered to (n + m)-membered means that the number of atoms in the ring is from n to n + m. For example, a 3- to 12-membered ring includes a 3-membered ring, 4-membered ring, 5-membered ring, 6-membered ring, 7-membered ring, 8-membered ring, 9-membered ring, 10-membered ring, 11-membered ring, and 12-membered ring, and also includes any range from n to n + m. For example, a 3- to 12-membered ring includes a 3- to 6-membered ring, 3- to 9-membered ring, 5- to 6-membered ring, 5- to 7-membered ring, 6- to 7-membered ring, 6- to 8-membered ring, and 6- to 10-membered ring etc.

[0109] The term "leaving group" refers to a functional group or atom that can be replaced by another functional group or atom through a substitution reaction (such as a nucleophilic substitution reaction). For example, representative leaving groups include trifluoromethanesulfonate; chlorine, bromine, iodine; sulfonate groups such as methanesulfonate, tosylate, p-bromobenzenesulfonate, p-toluenesulfonate, etc.; acyloxy groups such as acetoxy, trifluoroacetoxy, and so on.

[0110] The term "protecting group" includes but is not limited to "amino protecting group", "hydroxy protecting group" or "mercapto protecting group". The term "amino protecting group" refers to a protecting group suitable for preventing side reactions at the amino nitrogen position. Representative amino protecting groups include but are not limited to: formyl; acyl groups such as alkanoyl groups (such as acetyl, trichloroacetyl or trifluoroacetyl); alkoxycarbonyl groups such as tert-butoxycarbonyl (Boc); arylmethoxycarbonyl groups such as benzyloxycarbonyl (Cbz) and 9-fluorenylmethoxycarbonyl (Fmoc); arylmethyl groups such as benzyl (Bn), trityl (Tr), 1,1-bis-(4'-methoxyphenyl)methyl; silyl groups such as trimethylsilyl (TMS) and tert-butyldimethylsilyl (TBS), and so on. The term "hydroxy protecting group" refers to a protecting group suitable for preventing side reactions of the hydroxyl group. Representative hydroxy protecting groups include but are not limited to: alkyl groups such as methyl, ethyl and tert-butyl; acyl groups such as alkanoyl groups (such as acetyl); arylmethyl groups such as benzyl (Bn), p-methoxybenzyl (PMB), 9-fluorenylmethyl (Fm) and diphenylmethyl (diphenylmethyl, DPM); silyl groups such as trimethylsilyl (TMS) and tert-butyldimethylsilyl (TBS), and so on.

[0111] The compounds of the present invention can be prepared by various synthetic methods well-known to those skilled in the art, including the specific embodiments listed below, the embodiments formed by their combination with other chemical synthesis methods, and the equivalent replacement methods well-known to those skilled in the art. Preferred embodiments include, but are not limited to, the examples of the present invention.

[0112] The structure of the compounds of the present invention can be confirmed by conventional methods well-known to those skilled in the art. If the present invention relates to the absolute configuration of a compound, the absolute configuration can be confirmed by conventional technical means in the art. For example, single crystal X-ray diffraction (SXRD), where the grown single crystal is used to collect diffraction intensity data with a Bruker D8 venture diffractometer, the light source is CuKα radiation, and the scanning mode: After scanning and collecting relevant data, the crystal structure is further analyzed by the direct method (Shelxs97), and the absolute configuration can be confirmed.

[0113] The solvents used in the present invention are commercially available. The following abbreviations are used in the present invention: aq represents water; TFA represents trifluoroacetic acid; ACN represents acetonitrile; DMSO represents dimethyl sulfoxide. Detailed Description of Specific Embodiments

[0114] The present invention will be described in detail below by way of examples, but this does not mean any adverse limitation to the present invention. The present invention has been described in detail herein, and specific embodiments thereof have also been disclosed. It will be obvious to those skilled in the art that various changes and improvements can be made to the specific embodiments of the present invention without departing from the spirit and scope of the present invention.

[0115] Example 1

[0116]

[0117] Step 1: Dissolve compound 1-1 (2 g, 11.56 mmol) in dimethyl sulfoxide (10 mL) at 25 °C, add 1-2 (2.88 g, 12.72 mmol) and N,N-diisopropylethylamine (2.99 g, 23.12 mmol), stir at 100 °C for 16 hours, add 200 mL of water to the reaction solution, and extract with ethyl acetate (500 mL × 3). The combined organic phases are washed successively with 0.2 M hydrochloric acid aqueous solution (100 mL) and saturated brine (200 mL). Finally, the organic phase is dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain crude product 1-3. MS ESI calculated value: C 18 H 23 ClN4O2 [M+H] + 363, found 363.

[0118] Step 1': At 0 °C, dissolve compound 1-4a (10 g, 78.68 mmol) in dichloromethane (100 mL), add N,N-diisopropylethylamine (12.20 g, 94.41 mmol, 16.45 mL) and 1-4b ((15.74 g, 94.41 mmol, 16.71 mL), stir at 25 °C for 16 hours. Add 200 mL of water to the reaction solution, and extract with ethyl acetate (500 mL × 3). The combined organic phases are washed with saturated brine (200 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether:ethyl acetate = 1 / 0 to 30 / 1) to obtain compound 1-4c.

[0119] Step 2': Dissolve compound 1-4c (11 g, 42.74 mmol) in methanol (100 mL), under nitrogen protection, add palladium on carbon (0.1 g, palladium content 10%), then displace with hydrogen three times, and stir at 25 °C for 16 hours. Filter through diatomaceous earth and concentrate under reduced pressure to obtain crude product 1-4. 1H NMR (400 MHz, DMSO-d6) δ -0.05--0.03 (s, 9H), 0.77-0.86 (t, 2H), 1.95-1.97 (s, 3H), 3.47-3.53 (t, 2H), 5.06-5.08 (s, 1H), 5.08-5.11 (s, 2H), 5.16 (s, 2H).

[0120] Step 2: Dissolve compound 1-3 (2.5 g, 6.89 mmol) in dioxane (30 mL), add compound 1-4 (1.64 g, 7.23 mmol), cesium carbonate (4.49 g, 13.78 mmol) and [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] palladium(II) mesylate mesylate (312.28 mg, 344.49 μmol). After displacing with nitrogen three times, heat to 100 °C and stir under nitrogen protection for 16 hours. Add 50 mL of ammonium chloride aqueous solution to the reaction solution, and extract with ethyl acetate (100 mL × 3). The combined organic phases are washed again with saturated brine (100 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 to 5 / 1) to obtain compound 1-5. MS ESI calculated value: C 28 H 43 N7O3Si [M+H] + 554, found 554. 11H NMR (400 MHz, CD3OD) δ -0.09--0.06 (s, 9H), 0.81 - 0.88 (t, 2H), 1.45 - 1.52 (m, 11H), 1.76 - 1.85 (m, 2H), 1.87 - 2.01 (m, 4H), 2.21 - 2.25 (s, 3H), 3.49 - 3.55 (t, 2H), 4.16 - 4.23 (m, 2H), 4.33 - 4.48 (m, 1H), 5.34 (s, 2H), 6.04 - 6.09 (m, 1H), 6.09 - 6.13 (m, 1H), 6.22 - 6.30 (s, 1H).

[0121] Step 3: At 0 - 5 °C, selenium powder (4.39 g, 54.17 mmol) was added to ethanol (30 mL), then sodium borohydride (2.38 g, 62.91 mmol) was slowly added, and the mixture was stirred at room temperature until the solid particles completely disappeared. Pyridine (8.57 g, 108.35 mmol) and compound 1-5 (3 g, 5.42 mmol) were added to the reaction solution, and the temperature was raised to 80 °C and stirred for half an hour. Then 2 M hydrochloric acid aqueous solution (32.50 mL,) was slowly added, and stirring was continued for half an hour. LC-MS showed that the raw materials were completely consumed. 50 mL of ammonium chloride aqueous solution was added to the reaction solution, and it was extracted with ethyl acetate (50 mL × 3). The combined organic phases were washed with saturated brine (50 mL), and finally the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 1 / 1) to obtain compound 1-6. MS ESI calculated value: C 28 H 45 N7O3SeSi [M + H] + 636, found 636. 1 1H NMR (400 MHz, CD3OD) δ -0.02 - 0.01 (s, 9H), 0.90 - 0.96 (t, 2H), 1.52 - 1.55 (m, 11H), 1.85 - 1.93 (m, 2H), 1.96 - 2.08 (m, 4H), 2.24 - 2.30 (s, 3H), 3.58 - 3.64 (t, 2H), 4.22 - 4.31 (m, 2H), 4.37 - 4.53 (m, 1H), 5.36 - 5.43 (s, 2H), 6.22 - 6.25 (m, 1H), 6.32 - 6.35 (s, 1H), 6.36 - 6.39 (m, 1H).

[0122] Step 4: Dissolve compound 1-6 (1.4 g, 2.21 mmol) in ethanol (15 mL) at 25 °C, add 1-7 (204.07 mg, 2.21 mmol), and stir at 80 °C for 1 hour. The reaction solution was directly concentrated under reduced pressure to obtain crude 1-8.

[0123] Step 5: Dissolve compound 1-8 (1.4 g, 2.08 mmol) in ethyl acetate (2 mL), add hydrochloric acid ethyl acetate solution (4 M, 14 mL), and stir at 25 °C for 1 hour. The reaction solution was directly filtered to obtain the filter cake as the crude hydrochloride salt of compound 1-9. MS ESI calculated value C 20 H 25 N7Se[M+H] + 444, found 444. 1 H NMR (400 MHz, CD3OD) δ 1.93 - 2.05 (m, 2H), 2.20 - 2.42 (m, 9H), 2.56 - 2.62 (s, 3H), 4.13 - 4.26 (m, 3H), 5.98 (s, 1H), 6.67 (s, 1H), 6.79 (s, 1H), 8.06 - 8.16 (s, 1H).

[0124] Step 6: Dissolve the crude hydrochloride salt of 1-9 (0.85 g, 1.65 mmol) in methanol (10 mL), add N,N-diisopropylethylamine (639.52 mg, 4.95 mmol, 862 μL), stir the mixture at 25 °C for 10 minutes, then add compound 1-10 (0.290 g, 5.47 mmol, 363 μL), and stir at 25 °C for 16 hours. Add 50 mL of aqueous ammonium chloride solution to the reaction solution, extract with ethyl acetate (50 mL × 3), wash the combined organic phases with saturated brine (50 mL), dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain the crude product. The crude product was separated by preparative high performance liquid chromatography (column: Phenomenex Luna C18 150*40mm*15μm; mobile phase: [water (0.1% TFA)-ACN]; B (ACN)%: 8% - 38%, 11 minutes) to obtain the trifluoroacetate salt of compound 1-11. MS ESI calculated value C 23 H 28 N8Se[M+H] + 497, found 497, 11H NMR (400 MHz, CD3OD) δ 2.10 - 2.20 (m, 2H), 2.31 - 2.50 (m, 9H), 2.51 - 2.56 (s, 3H), 3.10 - 3.17 (t, 2H), 3.48 - 3.59 (t, 2H), 4.17 - 4.31 (m, 3H), 5.90 - 5.95 (s, 1H), 6.68 - 6.74 (m, 2H), 7.96 (m, 1H).

[0125] Example 2

[0126]

[0127] Step 1: At 25 °C, N,N - diisopropylethylamine (21.0 mg, 162.46 μmol) and O-(7 - azabenzotriazol - 1 - yl)-N,N,N′,N′ - tetramethyluronium hexafluorophosphate (34.0 mg, 89.35 μmol) were added to a DMF (0.5 mL) solution of 2 - 1 (10 mg, 81.23 μmol), and the mixture was stirred at 25 °C for 0.5 h. Then, the hydrochloride salt of 1 - 9 (41.86 mg, 81.23 μmol) and a DMF (0.5 mL) solution of N,N - diisopropylethylamine (21.00 mg, 162.46 μmol, 28.30 μL) were added to the reaction system, and the mixture was stirred at 0 - 5 °C for 1 h. 50 mL of aqueous ammonium chloride solution was added to the reaction solution, and the mixture was extracted with ethyl acetate (50 mL × 3). The combined organic phases were washed with saturated brine (50 mL), and finally the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the crude product. The crude product was separated by preparative high - performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA) - ACN]; B (ACN)%: 24% - 54%, 10 minutes) to obtain the trifluoroacetate salt of compound 2 - 2. MS ESI calculated value C 26 H 28 N8OSe [M + H] + 549, found 549, 11H NMR (400 MHz, CD3OD) δ 1.88 - 2.01 (m, 2H), 2.07 - 2.16 (m, 2H), 2.17 - 2.27 (m, 2H), 2.27 - 2.34 (m, 2H), 2.34 - 2.38 (s, 3H), 2.50 - 2.54 (s, 3H), 4.25 - 4.36 (m, 1H), 4.64 - 4.70 (m, 1H), 4.90 - 4.97 (m, 1H), 5.84 - 5.89 (s, 1H), 6.64 - 6.69 (m, 2H), 7.51 - 7.57 (m, 1H), 7.74 - 7.80 (m, 1H), 7.94 - 8.01 (m, 2H), 8.59 - 8.67 (m, 1H).

[0128] Example 3

[0129]

[0130] Step 1: Dissolve the hydrochloride salt of compound 1 - 9 (50 mg, 97.02 μmol) in N - methylpyrrolidone (1 mL) at 20 °C, add triethylamine (49.09 mg, 485.10 μmol, 67.52 μL), stir at 20 °C for 0.5 h. Add N - methylpyrrolidone (0.5 mL) to the reaction solution, cool the reaction solution to 0 °C, stir for more than 0.5 h, slowly add 3 - 1 (21.03 mg, 116.42 μmol), warm the mixture to 20 °C and stir for 3 h. Add 20 mL of saturated ammonium chloride aqueous solution to the reaction solution and extract with ethyl acetate (20 mL × 3). The combined organic phases are washed with saturated brine (5 mL × 3), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain the crude product. The crude product is separated by preparative high - performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA) - ACN]; ACN%: 30% - 60%, 10 minutes) to obtain the trifluoroacetate salt of compound 3 - 2. MS ESI calculated value: C 24 H 29 N9O2SSe [M + H] + 588, found 588, 1 1H NMR (400 MHz, METHANOL - d4) δ 1.81 - 1.92 (m, 2H), 2.02 - 2.10 (m, 2H), 2.18 - 2.27 (m, 4H), 2.35 (s, 3H), 2.52 (s, 3H), 3.66 (m, 1H), 4.02 (m, 2H), 4.09 - 4.18 (m, 3H), 4.30 (m, 2H), 5.87 (s, 1H), 6.67 (m, 2H), 7.97 (s, 1H).

[0131] Example 4

[0132]

[0133] Step 1: At 20 °C, dissolve the hydrochloride salt of Compound 1-9 (60 mg, 116.43 μmol) in DMF (1 mL), add potassium carbonate (32.18 mg, 232.86 μmol) and 4-1 (26.08 mg, 116.43 μmol), and stir the mixture at 50 °C for 16 hours. Add 20 mL of water to the reaction solution and extract with dichloromethane (10 mL × 3). Wash the combined organic phases with saturated brine (5 mL × 3). Finally, dry the organic phase with anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain the crude product. The crude product was separated by preparative high-performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA)-ACN]; ACN%: 16%-46%, 10 minutes) to obtain the trifluoroacetate salt of Compound 4-2. MS ESI calculated value: C 23 H 28 F3NSe[M+H] + 540, found 540, 1 H NMR (400 MHz, METHANOL-d4) δ 2.08 (m, 2H), 2.31 - 2.38 (m, 5H), 2.39 - 2.49 (m, 4H), 2.53 (d, 3H), 2.81 - 2.93 (m, 2H), 3.34 - 3.47 (m, 2H), 4.14 - 4.33 (m, 3H), 5.92 (s, 1H), 6.64 - 6.79 (m, 2H), 7.96 (d, 1H).

[0134] Example 5

[0135]

[0136] Step 1: At 20 °C, dissolve compound 5-1 (33.38 mg, 176.41 μmol) in DMF (1 mL), add hydroxybenzotriazole (47.67 mg, 352.81 μmol) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (67.63 mg, 352.81 μmol), and stir the mixture at 20 °C for 0.5 h. Add a DMF (1 mL) solution of the hydrochloride salt of compound 1-9 (100 mg, 194.05 μmol) and diisopropylethylamine (68.40 mg, 529.22 μmol, 92.18 μL) to the mixture, and stir at 20 °C for 4 h. Add 10 mL of water to the reaction solution, and extract with dichloromethane (20 mL × 3). Wash the combined organic phases with saturated brine (5 mL × 3), and finally dry the organic phase over anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain the crude product 5-2. MS ESI calculated value: C 28 H 38 N9O3Se[M+H] + 615, found 615.

[0137] Step 2: At 20 °C, dissolve compound 5-2 (150 mg, 244.45 μmol) in dichloromethane (3 mL), add trifluoroacetic acid (1 mL), and stir the mixture at 20 °C for 16 h. Concentrate the reaction solution to dryness by rotary evaporation to obtain the crude product. The crude product was separated by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA)-ACN]; ACN%: 13%-43%, 10 min) to obtain the trifluoroacetate salt of compound 5-3. MS ESI calculated value: C 23 H 30 N8OSe[M+H] + 515, found 515, 1 H NMR (400 MHz, METHANOL-d4) δ 1.66 - 1.79 (m, 2H), 2.04 - 2.39 (m, 10H), 2.52 (d, 3H), 2.77 (s, 3H), 4.00 - 4.18 (m, 2H), 4.22 - 4.31 (m, 1H), 4.31 - 4.38 (m, 1H), 4.78 (m, 1H), 5.88 (s, 1H), 6.68 (s, 2H), 7.97 (d, 1H).

[0138] Example 6

[0139]

[0140]

[0141] Step 1: Dissolve compound 1-1 (200 mg, 1.16 mmol) and compound 6-1 (305.64 mg, 1.27 mmol) in DMSO (2 mL) at 25 °C, add N,N-diisopropylethylamine (298.83 mg, 2.31 mmol), stir at 100 °C for 2 hours. Add 10 mL of ethyl acetate to the reaction solution, and wash with saturated brine (10 mL × 3). The organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain crude product 6-2. MS ESI calculated value: C 19 H 25 ClN4O2[M+H] + 377, found 377.

[0142] Step 2: Dissolve compound 6-2 (300 mg, 796.01 μmol), compound 1-4 (217.19 mg, 955.21 mmol), cesium carbonate (518.71 mg, 1.59 mmol) and palladium(II) mesylate of [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] (72.16 mg, 89.60 μmol) in dioxane (3 mL). After purging with nitrogen three times, heat to 100 °C and stir for 16 hours under nitrogen protection. The reaction solution is concentrated under reduced pressure to obtain a crude product, which is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 20 / 1 - 5 / 1) to obtain compound 6-3. MS ESI calculated value: C 29 H 45 N7O3Si[M+H] + 568, found 568.

[0143] Step 3: At 0 - 5 °C, add selenium powder (285.23 mg, 3.52 mmol) to ethanol (4 mL), then slowly add sodium borohydride (159.91 mg, 4.23 mmol) in portions and stir at room temperature until the solid particles completely disappear. Add pyridine (557.24 mg, 7.04 mmol) and compound 6-3 (200 mg, 352.24 μmol) to the reaction solution, and heat to 80 °C and stir for 0.5 hour. Then slowly add 2 M aqueous hydrochloric acid solution (2.11 mL), continue to stir for 0.5 hour. Add 10 mL of water to the reaction solution, and extract with ethyl acetate (10 mL × 3). The combined organic phases are washed with saturated brine (10 mL × 2) again. Finally, the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product, which is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 10 / 0 - 1 / 1) to obtain compound 6-4. MS ESI calculated value: C 29 H47 N7O3SeSi[M+H] + 649, the measured value is 649.

[0144] Step 4: Dissolve compound 6-4 (140 mg, 159.69 μmol) in ethanol (1 mL) at 25 °C, add compound 6-5 (17.73 mg, 196.62 μmol), and stir at 80 °C for 1 hour. The reaction solution was directly concentrated under reduced pressure to obtain the crude product 6-6. MS ESI calculated value: C 32 H 49 N7O3SeSi[M+H] + 687, the measured value is 687.

[0145] Step 5: Dissolve compound 6-6 (110 mg, 160.16 μmol) in ethyl acetate (1 mL), add hydrochloric acid ethyl acetate solution (4 M, 1.11 mL), and stir at 25 °C for 12 hours. The reaction solution was directly filtered to obtain the filter cake as the crude hydrochloride salt of compound 6-7. MS ESI calculated value C 21 H 27 N7Se[M+H] + 456, the measured value is 456.

[0146] Step 6: Dissolve the crude hydrochloride salt of 6-7 (45 mg, 76.51 μmol) in methanol (1 mL), add N,N-diisopropylethylamine (49.44 mg, 382.53 μmol), stir the mixture at 25 °C for 5 minutes, then add acrylonitrile (1-10) (12.18 mg, 229.52 μmol, 363 μL), and stir at 25 °C for 16 hours. Add 10 mL of aqueous ammonium chloride solution to the reaction solution, extract with ethyl acetate (20 mL × 3), wash the combined organic phases with saturated brine (20 mL), and finally dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain the crude product. The crude product was separated by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA)-ACN]; B (ACN)%: 15%-45%, 10 minutes) to obtain the trifluoroacetate salt of compound 6-8. MS ESI calculated value C 24 H 30 N8Se[M+H] + 510, the measured value is 510, 11H NMR (400 MHz, CD3OD) δ 1.90 - 2.13 (m, 4H), 2.13 - 2.30 (m, 4H), 2.33 (s, 1H), 2.36 (s, 3H), 2.37 - 2.42 (m, 1H), 2.49 (s, 2H), 2.52 (s, 3H), 3.10 (t, J = 7.25 Hz, 2H), 3.79 (t, J = 7.25 Hz, 2H), 3.87 (s, 2H), 4.69 (tt, J = 11.76, 6.07 Hz, 2H), 5.94 (s, 1H), 6.67 (d, J = 1.13 Hz, 1H), 6.69 (s, 1H), 7.94 (s, 1H).

[0147] Example 7

[0148]

[0149] Step 1: Dissolve the crude hydrochloride of compound 6 - 7 (45 mg, 76.51 μmol) in methanol (1 mL), add triethylamine (38.71 mg, 382.53 μmol), then add compound 3 - 1 (16.58 mg, 91.81 μmol) at 0 °C and stir at 25 °C for 16 hours. LC - MS showed that the raw material was completely consumed and the main peak was the product peak. Add 10 mL of aqueous solution to the reaction mixture and extract with ethyl acetate (10 mL × 3). The combined organic phases were washed with saturated brine (10 mL × 2), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain the crude product. The crude product was separated by preparative high - performance liquid chromatography (column: Phenomenex Synergi C18 150*25 mm*10 μm; mobile phase: [water (0.1% TFA) - ACN]; B (ACN)%: 31% - 61%, 10 minutes) to obtain the trifluoroacetate of compound 7 - 1. MS ESI calculated value C 25 H 31 N9O2SSe [M + H] + 601, found 601, 1 1H NMR (400 MHz, CD3OD) δ 1.87 - 1.99 (m, 5H), 2.00 - 2.08 (m, 2H), 2.09 - 2.24 (m, 1H), 2.29 (d, 1H), 2.31 (d, 1H), 2.36 (s, 3H), 2.53 (d, J = 0.88 Hz, 3H), 3.65 (m, 1H), 3.99 (m, 2H), 4.09 - 4.19 (m, 4H), 4.60 - 4.74 (m, 1H), 5.88 (s, 1H), 6.66 (s, 2H), 7.97 (d, J = 0.88 Hz, 1H).

[0150] Example 8

[0151]

[0152] Step 1: Compound 8-1 (15 g, 78.13 mmol) was dissolved in dichloromethane (200 mL). 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (17.97 g, 93.75 mmol) and 1-hydroxybenzotriazole (12.67 g, 93.75 mmol) were added, and the mixture was stirred at 20 °C for 1 hour. Then N,O-dimethylhydroxylamine hydrochloride (11.43 g, 117.19 mmol) and triethylamine (23.72 g, 234.38 mmol, 32.62 mL) were added, and the reaction was carried out at 20 °C for 3 hours. The reaction solution was diluted with 400 mL of water and extracted with 450 mL (150 mL × 3) of ethyl acetate. The combined organic layers were washed with 200 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. It was purified by silica gel column (petroleum ether / ethyl acetate = 10 / 1 to 3 / 1) to obtain compound 8-2. 1 H NMR (400 MHz, CDCl3) δ 3.39 (s, 3H), 3.59 (s, 3H), 7.51 (s, 2H).

[0153] Step 2: Compound 8-2 (14.54 g, 61.85 mmol) was dissolved in tetrahydrofuran (200 mL). Methylmagnesium bromide ether solution (3 M, 24.74 mL) was added dropwise at 0 °C. After the addition, the reaction was carried out at 0 °C for 1 hour. The reaction was quenched with 200 mL of saturated ammonium chloride aqueous solution, diluted with 200 mL of water, and extracted with 400 mL (200 mL × 2) of ethyl acetate. The combined organic layers were washed with 200 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure to obtain crude compound 8-3. 1 H NMR (400 MHz, CDCl3) δ 2.62 (s, 3H), 7.68 (s, 2H).

[0154] Step 3: Compound 8-3 (11.65 g, 61.31 mmol) and ammonium chloride (327.94 mg, 6.13 mmol) were mixed, and dichloromethane (200 mL) was added. Bromine (10.29 g, 64.37 mmol, 3.32 mL) was added dropwise, and the reaction solution was reacted at 30 - 40 °C for 4 hours. 200 mL of saturated sodium bicarbonate aqueous solution was added, then diluted with 200 mL of water, and extracted with 400 mL (200 mL × 2) of dichloromethane. The combined organic layers were washed with 150 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure to obtain crude 8-4. 1 H NMR (400 MHz, CDCl3) δ 4.36 (s, 2H), 7.73 (s, 2H)

[0155] Step 4: Selenium (2.89 g, 35.67 mmol, 2.83 mL) was added to ethanol (30 mL), and sodium borohydride (1.83 g, 48.37 mmol) was added portionwise at 0 - 5 °C. After addition, the reaction was carried out at 20 °C for 1 hour, then 4-morpholinecarbonitrile (2 g, 17.84 mmol) was added slowly, and then pyridine hydrochloride (8.24 g, 71.35 mmol) was added slowly. The mixture was reacted at 20 °C for 16 hours. 50 mL of water was added to the reaction solution, and it was extracted with dichloromethane (30 mL × 3). The combined organic phases were washed with brine (30 mL × 1), dried over anhydrous Na2SO4, filtered and concentrated in vacuo to dryness, and recrystallized and purified in toluene (20 mL) to obtain compound 8-6. 1 1H NMR (400 MHz, DMSO-d6) δ 3.52 - 3.59 (m, 4H), 3.60 - 3.97 (m, 4H), 7.93 (s, 2H).

[0156] Step 5: Compound 8-4 (278.52 mg, 1.04 mmol) was added to methanol (6 mL) and water (2 mL), sodium fluoride (21.74 mg, 517.85 μmol) and 8-6 (0.2 g, 1.04 mmol) were added, and the reaction was carried out at 20 °C for 1 hour. 20 mL of water was added, and a solid precipitated. It was filtered, and the filter cake was dried in vacuo to obtain compound 8-7. 1 1H NMR (400 MHz, CDCl3) δ 3.51 - 3.59 (m, 4H), 3.82 - 3.88 (m, 4H), 7.67 (s, 2H), 7.68 (s, 1H).

[0157] Step 6: Compound 8-7 (1.71 g, 4.71 mmol), 1-2 (1.17 g, 5.18 mmol), [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)]palladium(II) methanesulfonate (213.45 mg, 235.50 μmol) and cesium carbonate (3.07 g, 9.42 mmol) were added to dioxane (35 mL), and the reaction was carried out at 80 °C for 16 hours under a nitrogen atmosphere. The reaction solution was diluted with 120 mL of water and extracted with 150 mL of ethyl acetate (50 mL × 3). The combined organic layers were washed with 50 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column (petroleum ether / ethyl acetate = 10 / 1 to 3 / 1) to obtain compound 8-8. MS ESI calculated value: C 24 H 32 ClN5O3Se [M+H] + 554, found 554.

[0158] Step 7: Compound 8-8 (680 mg, 1.12 mmol) and 8-9 (264.86 mg, 1.34 mmol) were added to toluene (15 mL), followed by the addition of palladium(II) mesylate of [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] (101.44 mg, 111.91 μmol) and cesium carbonate (729.24 mg, 2.24 mmol). The reaction was carried out at 80 °C under a nitrogen atmosphere for 16 hours. The reaction solution was diluted with 60 mL of water and extracted with 60 mL (30 mL × 2) of ethyl acetate. The combined organic layers were washed with 30 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1 to 3 / 1) to obtain Compound 8-10. MS ESI calculated value: C 33 H 46 N8O5Se[M+H]+715, found 715.

[0159] Step 8: Hydrochloric acid ethyl acetate (4 M, 3 mL) was added to Compound 8-10 (160.67 mg, 225.11 μmol), and the reaction was carried out at 20 °C for 1.5 hours. The reaction solution was concentrated under reduced pressure to obtain Compound 8-11. MS ESI calculated value: C 23 H 30 N8OSe[M+H]+515, found 515.

[0160] Step 9: Compound 8-11 (120 mg, 218.20 μmol) was dissolved in methanol (3 mL), and diisopropylethylamine (141.00 mg, 1.09 mmol, 190.03 μL) and acrylonitrile (0.11 g, 2.07 mmol, 137.50 μL) were added. The reaction was carried out at 20 °C for 16 hours. 30 mL of water was added to the reaction solution, and it was extracted with 40 mL (20 mL × 2) of ethyl acetate. The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by high performance liquid chromatography (column model: Phenomenex Synergi C18 150×25mm×10μm; mobile phase: [water (0.1% trifluoroacetic acid)-acetonitrile]; acetonitrile %: 16%-46%, 10 minutes) to obtain the trifluoroacetate of Compound 8-12. MS ESI calculated value: C 26 H 33 N9OSe[M+H]+568, found 568. 11H NMR (400 MHz, CD3OD) δ 2.12 (t, J = 12.0 Hz, 2H), 2.27 - 2.38 (m, 5H), 2.38 - 2.50 (m, 4H), 3.10 (t, J = 7.2 Hz, 2H), 3.42 - 3.62 (m, 6H), 3.77 - 3.84 (m, 4H), 4.15 - 4.23 (m, 1H), 4.24 (s, 2H), 5.83 (d, J = 0.4 Hz, 1H), 6.73 (d, J = 1.2 Hz, 1H), 6.81 (d, J = 1.2 Hz, 1H), 8.10 (s, 1H).

[0161] Example 9

[0162]

[0163] Step 1: At 20 °C, dissolve compound 9 - 1 (15.36 mg, 105.84 μmol) in DMF (0.5 mL), add 1 - hydroxybenzotriazole (28.60 mg, 211.69 μmol) and 1 - (3 - dimethylaminopropyl) - 3 - ethylcarbodiimide hydrochloride (40.58 mg, 211.69 μmol), and stir the mixture at 20 °C for 0.5 h. Add a solution of hydrochloride salt of compound 1 - 9 (60 mg, 116.43 μmol) and diisopropylethylamine (41.04 mg, 317.53 μmol, 55.31 μL) in DMF (0.5 mL) to the mixture, and stir at 20 °C for 16 h. LC - MS shows that the raw materials are consumed completely and the main peak is the product peak. Add 20 mL of water to the reaction solution, extract with ethyl acetate (20 mL × 3), wash the combined organic phases with saturated brine (5 mL × 3), finally dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain the crude product. The crude product is separated by preparative high - performance liquid chromatography (column: Waters Xbridge 150*25mm*5μm; mobile phase: [water (10 mM NH4HCO3) - ACN]; ACN%: 23% - 53%, 8 min) to obtain compound 9 - 2. MS ESI calculated value: C 26 H34N8O2Se [M + H] + 571, found 571, 11H NMR (400 MHz, CD3OD) δ 1.24 - 1.41 (m, 1H), 1.50 - 1.71 (m, 2H), 1.87 - 2.38 (m, 11H), 2.47 (s, 3H), 2.54 (m, 4H), 3.23 (s, 2H), 3.68 - 3.77 (m, 4H), 4.56 (m, 2H), 4.69 (m, 1H), 6.19 - 6.43 (m, 2H), 6.55 (m, 1H), 7.61 - 7.81 (m, 1H).

[0164] Example 10

[0165]

[0166] Step 1: At 20 °C, dissolve compound 1 - 6 (80 mg, 126.03 μmol) in ethanol (1 mL), add 10 - 1 (13.43 mg, 126.03 μmol, 12.79 μL), and stir the mixture at 80 °C for 7 hours. Concentrate the reaction solution under reduced pressure to obtain crude product 10 - 2. MS ESI calculated value: C 32 H 49 N7O3SeSi [M + H] + 688, found 688.

[0167] Step 2: At 20 °C, dissolve compound 10 - 2 (90 mg, 131.04 μmol) in methanol (0.5 mL), add hydrochloric acid / ethyl acetate (4 M, 2 mL), and stir the mixture at 20 °C for 16 hours. Concentrate the reaction solution under reduced pressure to obtain crude product 10 - 3 hydrochloride. MS ESI calculated value: C 21 H 27 N7Se [M + H] + 458, found 458.

[0168] Step 3: At 20 °C, dissolve compound 10-3 hydrochloride (80 mg, 162.30 μmol, HCl) in methanol (1 mL), add diisopropylethylamine (62.93 mg, 486.91 μmol, 84.81 μL) and acrylonitrile (1-10) (120 mg, 2.26 mmol, 150.00 μL). Stir the mixture at 20 °C for 2 hours. Add 20 mL of water to the reaction solution and extract with ethyl acetate (20 mL × 3). Wash the combined organic phases with saturated brine (5 mL × 3). Finally, dry the organic phase over anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain the crude product. The crude product is separated by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA)-ACN]; ACN%: 15%-45%, 10 minutes) to obtain the trifluoroacetate of compound 10-4. MS ESI calculated value: C 24 H 30 N8O8Se[M+H] + 511, found 511, 1 H NMR (400 MHz, CD3OD) δ 1.29 (m, 1H), 2.09 (m, 2H), 2.29 - 2.49 (m, 14H), 2.55 (s, 3H), 3.09 (t, 2H), 3.44 - 3.55 (m, 2H), 4.24 (m, 3H), 5.90 (s, 1H), 6.63 (m, 2H).

[0169] Example 11

[0170]

[0171] Step 1: Add selenium (4.62 g, 57.07 mmol) to ethanol (40 mL). Add sodium borohydride (2.19 g, 57.89 mmol) portionwise at 0 - 5 °C. After addition, react at 20 °C for 1 hour. Then slowly add dimethylcyanamide (11-1) (2 g, 28.53 mmol), and then slowly add pyridine hydrochloride (13.19 g, 114.13 mmol). React the mixture at 20 °C for 16 hours. Add 100 mL of water to the reaction solution and extract with dichloromethane:methanol = 10:1 (50 mL × 4). Dry over anhydrous Na2SO4, filter and concentrate to dryness under vacuum to obtain compound 11-2. 1 H NMR (400 MHz, DMSO-d4) δ 3.12 - 3.45 (m, 6H), 7.62 (d, J = 2.4 Hz, 2H)

[0172] Step 2: Compound 8-4 (1 g, 3.72 mmol) was added to methanol (30 mL), and compound 11-2 (561.76 mg, 3.72 mmol) was added. The reaction was carried out at 20 °C for 1 hour. Ammonia water was added to adjust the pH of the system to 7. The mixture was concentrated to dryness and purified by silica gel column (petroleum ether∶ethyl acetate = 10∶1) to obtain compound 11-3. 1 H NMR (400 MHz, CDCl3) δ 3.17 (s, 6H), 7.58 (s, 1H), 7.69 (s, 2H)

[0173] Step 3: Compound 11-3 (0.8 g, 2.49 mmol), 1-2 (563.91 mg, 2.49 mmol), 2,2-bis(diphenylphosphino)-1,1-binaphthyl (155.15 mg, 249.17 μmol), palladium acetate (55.94 mg, 249.17 μmol) and cesium carbonate (1.62 g, 4.98 mmol) were added to dioxane (20 mL). The reaction was carried out at 90 °C for 5 hours under a nitrogen atmosphere. The reaction solution was diluted with 100 mL of water and extracted with 100 mL of ethyl acetate (50 mL×2). The combined organic layers were washed with 50 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure. Purification by silica gel column (petroleum ether / ethyl acetate = 20 / 1 to 5 / 1) gave compound 11-4. MS ESI calculated value: C 22 H 30 ClN5O2Se [M+H] + 512, found 512.

[0174] Step 4: Compound 11-4 (300 mg, 587.18 μmol) and 8-9 (127.39 mg, 645.90 μmol) were added to dioxane (10 mL). Then, [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] palladium(II) mesylate (53.23 mg, 58.72 μmol) and cesium carbonate (382.63 mg, 1.17 mmol) were added. The reaction was carried out at 90 °C under a nitrogen atmosphere for 5 hours. The reaction solution was diluted with 50 mL of water and extracted with 40 mL (20 mL×2) of ethyl acetate. The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure. Purification by silica gel column (petroleum ether / ethyl acetate = 10 / 1 to 3 / 1) gave compound 11-5. MS ESI calculated value: C 31 H 44 N8O4Se [M+H] + 673, found 673.

[0175] Step 5: Compound 11-5 (150 mg, 223.32 μmol) was dissolved in ethyl acetate (3 mL), hydrochloric acid ethyl acetate (4 M, 3 mL) was added, and the reaction was carried out at 20 °C for 1 hour. The reaction solution was concentrated under reduced pressure to obtain Compound 11-6. MS ESI calculated value: C 21 H 28 N8Se[M+H] + 473, found 473.

[0176] Step 6: Compound 11-6 (150 mg, 295.32 μmol) was dissolved in methanol (5 mL), diisopropylethylamine (190.84 mg, 1.48 mmol, 257.20 μL) and acrylonitrile (0.1 g, 1.88 mmol, 125.00 μL) were added, and the reaction was carried out at 20 °C for 16 hours. 30 mL of water was added to the reaction solution, and it was extracted with 40 mL (20 mL×2) of ethyl acetate. The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by high performance liquid chromatography (column model: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% trifluoroacetic acid)-acetonitrile]; acetonitrile%: 15%-45%, 10 minutes) to obtain the trifluoroacetate salt of Compound 11-7. MS ESI calculated value: C 21 H 28 N8Se[M+H] + 473, found 473. 1 H NMR (400 MHz, MeOD) δ 2.13 (m, 2H), 2.30 - 2.38 (m, 5H), 2.38 - 2.52 (m, 4H), 3.11 (t, J = 7.2 Hz, 2H), 3.16 (s, 6H), 3.51 (s, 2H), 4.12 - 4.23 (m, 1H), 4.25 (s, 2H), 5.83 (s, 1H), 6.72 (s, 1H), 6.81 (d, J = 0.8 Hz, 1H), 7.96 (s, 1H)

[0177] Example 12

[0178]

[0179] Step 1: Compound 12-1 (3 g, 73.08 mmol, 3.85 mL) and selenium powder (12.84 g, 146.16 mmol) were added to DMF (30 mL) and water (3 mL), replaced with carbon monoxide 3 times, filled with carbon monoxide, maintained at a pressure of 50 psi, and reacted at 80 °C for 16 hours. After the reaction was completed, it was dried by an oil pump and purified by a silica gel column (petroleum ether∶ethyl acetate = 1∶2) to obtain Compound 12-2. 11H NMR (400 MHz, DMSO-d6) δ 2.39 (s, 3H), 9.72 - 10.09 (m, 1H), 10.23 (s, 1H)

[0180] Step 2: Compound 8-4 (3 g, 12.16 mmol) was added to methanol (120 mL), and compound 12-2 (1.80 g, 12.27 mmol) was added. The reaction was carried out at 20 °C for 2 hours. Ammonia water was added to adjust the pH of the system to 7, concentrated to dryness, and purified by silica gel column (petroleum ether∶ethyl acetate = 5∶1) to obtain compound 12-3. 1 1H NMR (400 MHz, CDCl3) δ 2.82 (s, 3H), 7.75 (s, 2H), 8.30 (s, 1H)

[0181] Step 3: Compound 12-3 (0.5 g, 1.71 mmol), 1-2 (387.49 mg, 1.71 mmol), 2,2-bis(diphenylphosphino)-1,1-binaphthyl (106.61 mg, 171.22 μmol), palladium acetate (38.44 mg, 171.22 μmol) and cesium carbonate (1.12 g, 3.42 mmol) were added to dioxane (10 mL). The reaction was carried out at 90 °C for 5 hours under a nitrogen atmosphere. The reaction solution was diluted with 100 mL of water, extracted with 100 mL of ethyl acetate (50 mL × 2), the combined organic layers were washed with 50 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column (petroleum ether / ethyl acetate = 20 / 1 to 5 / 1) to obtain compound 12-4. MS ESI calculated value: C 21 H 27 ClN4O2Se [M+H] + 483, found 483.

[0182] Step 4: Compound 12-4 (590 mg, 1.22 mmol) and 8-9 (241.49 mg, 1.22 mmol) were added to dioxane (15 mL), and [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] palladium(II) methanesulfonate (120.99 mg, 122.44 μmol) and cesium carbonate (797.85 mg, 2.45 mmol) were added. The reaction was carried out at 90 °C under a nitrogen atmosphere for 5 hours. The reaction solution was diluted with 50 mL of water and extracted with 40 mL (20 mL × 2) of ethyl acetate. The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column (petroleum ether / ethyl acetate = 10 / 1 to 3 / 1) to obtain compound 12-5. MS ESI calculated value: C 30 H41 N7O4Se[M+H] + 644, the measured value is 644.

[0183] Step 5: Compound 12-5 (150 mg, 210.07 μmol) was dissolved in ethyl acetate (3 mL), hydrochloric acid ethyl acetate (4 M, 2.70 mL) was added, and the reaction was carried out at 20 °C for 1 hour. The reaction solution was concentrated under reduced pressure to obtain Compound 12-6. MS ESI calculated value: C 20 H 25 N7Se[M+H] + 444, the measured value is 444.

[0184] Step 6: Compound 12-6 (100 mg, 226.03 μmol) was dissolved in methanol (3 mL), diisopropylethylamine (146.06 mg, 1.13 mmol, 196.85 μL) and acrylonitrile (0.12 g, 2.26 mmol, 150.00 μL) were added, and the reaction was carried out at 15 °C for 2 hours. 30 mL of water was added to the reaction solution, and it was extracted with 40 mL (20 mL×2) of ethyl acetate. The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by high performance liquid chromatography (column model: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% trifluoroacetic acid) - acetonitrile]; acetonitrile %: 13% - 43%, 10 minutes) to obtain the trifluoroacetate of Compound 12-7. MS ESI calculated value: C 23 H 28 N8Se[M+H]+497, the measured value is 497. 1 H NMR (400 MHz, CD3OD) δ 2.13 (t, J = 12.4 Hz, 2H), 2.35 (s, 3H), 2.35 - 2.50 (m, 6H), 2.81 (s, 3H), 3.10 (t, J = 7.2 Hz, 2H), 3.51 (s, 2H), 4.14 - 4.31 (m, 3H), 5.85 (s, 1H), 6.77 (d, J = 1.2 Hz, 1H), 6.84 (d, J = 1.2 Hz, 1H), 8.77 (s, 1H).

[0185] Example 13

[0186]

[0187] Step 1: Dissolve the crude product of hydrochloride (55 mg, 96.41 μmol) of 6 - 7 in DMF (1 mL), add N,N - diisopropylethylamine (37.38 mg, 289.24 μmol), stir at 0 °C for 0.5 h, then add a DMF (1 mL) solution of compound 5 - 1 (16.58 mg, 91.81 μmol), 1 - hydroxybenzotriazole (23.69 mg, 175.29 μmol) and 1 - (3 - dimethylaminopropyl) - 3 - ethylcarbodiimide hydrochloride (36.96 mg, 192.82 μmol) at 0 °C, and stir at 25 °C for 15.5 h. LC - MS shows that the raw materials are basically consumed completely. Add 10 mL of ethyl acetate to dilute the reaction solution, and wash it with saturated brine (10 mL × 2). The organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product is separated by preparative chromatography plate (SiO2, PE∶EA = 0∶1) to obtain compound 13 - 1. MS ESI calculated value C 29 H 40 N8O3Se[M + H] + 628, measured value 628.

[0188] Step 2: Dissolve 13 - 1 (20 mg, 31.87 μmol) in dichloromethane (1 mL), add trifluoroacetic acid (616 mg, 5.4 mmol), stir at 0 °C for 16 h. LC - MS shows that the raw materials are basically consumed, and the main peak is the product peak. The reaction solution is concentrated under reduced pressure to obtain a crude product. The crude product is separated by preparative high - performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA) - ACN]; B (ACN)%: 17% - 47%, 10 min) to obtain the trifluoroacetate of compound 13 - 2. MS ESI calculated value C 24 H 32 N8OSe[M + H] + 528, measured value 528, 1 H NMR (400 MHz, CD3OD) δ1.29 (m, 2H), 1.71 - 1.81 (m, 1H), 1.86 (m, 1H), 1.91 (m, 1H), 1.94 (m, 2H), 1.98 - 2.04 (m, 2H), 2.27 (m, 1H), 2.35 (s, 3H), 2.37 - 2.46 (m, 2H), 2.53 (s, 3H), 2.77 (s, 3H), 4.11 (s, 2H), 4.69 - 4.76 (m, 1H), 5.90 (s, 1H), 6.67 (s, 1H), 6.68 (s, 1H), 7.97 (s, 1H).

[0189] Example 14

[0190]

[0191] Step 1: Dissolve the crude hydrochloride of 6 - 7 (55 mg, 96.41 μmol) in DMF (2 mL), add potassium carbonate (53.30 mg, 385.65 μmol) and compound 4 - 1 (43.19 mg, 192.82 μmol), and stir at 120 °C for 3 hours. Add 10 mL of water to dilute the reaction solution, and extract with ethyl acetate (10 mL × 3). Combine the organic phases, dry over anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain the crude product. The crude product is separated by preparative high - performance liquid chromatography (column: Phenomenex Gemini - NXC18 75*30mm*3μm; mobile phase: [water (0.1% TFA) - ACN]; B(ACN)%: 40% - 70%, 8 minutes), and then (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA) - ACN]; B(ACN)%: 15% - 45%, 10 minutes) to obtain the trifluoroacetate of compound 14 - 1. MS ESI calculated value C 24 H 30 F3N7Se[M + H] + 553, found 553, 1 H NMR (400 MHz, CD3OD) δ 1.95 - 2.01 (m, 1H), 2.03 - 2.12 (m, 1H), 2.12 - 2.28 (m, 5H), 2.29 - 2.35 (m, 2H), 2.35 (s, 3H), 2.45 - 2.51 (m, 1H), 2.52 (s, 3H), 2.85 (m, 2H), 3.65 - 3.72 (m, 2H), 3.87 (s, 2H), 4.65 - 4.80 (m, 1H), 5.97 (s, 1H), 6.66 (s, 1H), 6.69 (s, 1H), 7.94 (s, 1H).

[0192] Example 15

[0193]

[0194]

[0195] Step 1: Compound 15-1 (2 g, 13.23 mmol) and diisopropylethylamine (2.56 g, 19.84 mmol) were added to tetrahydrofuran (30 mL). Cyanogen bromide (1.78 g, 16.80 mmol, 1.24 mL) was added at 0 °C, and the reaction was carried out at 20 °C for 16 h. The reaction solution was diluted with 100 mL of water, extracted with 120 mL of ethyl acetate (40 mL × 3), and the combined organic layers were washed with 50 mL of brine and dried over sodium sulfate to obtain Compound 15-2. 1 H NMR (400 MHz, CDCl3) δ 2.68 (s, 3H), 3.74 (s, 3H), 4.01 (s, 2H), 6.84 (d, J = 8.8 Hz, 2H), 7.14 - 7.23 (m, 2H).

[0196] Step 2: Selenium (2.10 g, 25.99 mmol) was added to ethanol (50 mL). Sodium borohydride (983.24 mg, 25.99 mmol) was added in portions at 0 - 5 °C. After addition, the reaction was carried out at 20 °C for 1 h, then Compound 15-2 (2.29 g, 13.00 mmol) was added slowly, and then pyridine hydrochloride (6.01 g, 51.98 mmol) was added slowly. The mixture was reacted at 20 °C for 16 h. 100 mL of water was added to the reaction solution, and it was extracted with dichloromethane∶methanol = 10∶1 (50 mL × 3), dried over anhydrous Na2SO4, filtered and concentrated to dryness under vacuum, and purified by silica gel column (petroleum ether∶ethyl acetate = 10∶1 to 2∶1) to obtain Compound 15-3. 1 H NMR (400 MHz, DMSO-d6) δ 2.58 - 3.04 (m, 3H), 3.49 - 3.84 (m, 3H), 4.50 - 5.49 (m, 2H), 6.67 - 7.00 (m, 2H), 7.03 - 7.43 (m, 2H), 7.43 - 8.17 (m, 2H).

[0197] Step 3: Compound 8-4 (0.6 g, 2.23 mmol) was added to methanol (40 mL), and Compound 15-3 (631.21 mg, 2.45 mmol) was added. The reaction was carried out at 20 °C for 1 h. After the reaction was completed, it was filtered, and the filter cake was dried under vacuum to obtain Compound 15-4. MS ESI calculated value: C 17 H 15 Cl2N3OSe [M+H] + 428, found 428.

[0198] Step 4: Compound 15-4 (560 mg, 1.31 mmol), 1-2 (296.68 mg, 1.31 mmol), 2,2-bis(diphenylphosphino)-1,1-binaphthyl (29.43 mg, 131.09 μmol), palladium(II) acetate (29.43 mg, 131.09 μmol) and cesium carbonate (854.24 mg, 2.62 mmol) were added to dioxane (15 mL), and the reaction was carried out at 90 °C for 16 h under a nitrogen atmosphere. The reaction mixture was diluted with 100 mL of water and extracted with 100 mL of ethyl acetate (50 mL × 2). The combined organic layers were washed with 50 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column chromatography (petroleum ether / ethyl acetate = 20 / 1 to 5 / 1) to obtain Compound 15-5. MS ESI calculated value: C 29 H 36 ClN5O3Se [M+H] + 618, found 618.

[0199] Step 5: Compound 15-5 (440 mg, 648.91 μmol) and 8-9 (127.99 mg, 648.91 μmol) were added to dioxane (15 mL), and then [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] palladium(II) mesylate (58.82 mg, 64.89 μmol) and cesium carbonate (422.85 mg, 1.30 mmol) were added. The reaction was carried out at 90 °C for 5 h under a nitrogen atmosphere. The reaction mixture was diluted with 50 mL of water and extracted with 40 mL of ethyl acetate (20 mL × 2). The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1 to 3 / 1) to obtain Compound 15-6. MS ESI calculated value: C 38 H 50 N8O5Se [M+H] + 779, found 779.

[0200] Step 6: Compound 15-6 (150 mg, 223.32 μmol) was dissolved in trifluoroacetic acid (7.70 g, 67.53 mmol, 5 mL), and the reaction was carried out at 70 °C for 20 h. The reaction mixture was concentrated under reduced pressure to obtain Compound 15-7. MS ESI calculated value: C 20 H 26 N8Se [M+H]+459, found 459.

[0201] Step 7: Compound 15-7 (90 mg, 157.49 μmol) was dissolved in methanol (3 mL), and diisopropylethylamine (101.77 mg, 787.46 μmol, 137.16 μL) and acrylonitrile (0.15 g, 2.07 mmol, 137.50 μL) were added. The reaction was carried out at 15 °C for 2 hours. 30 mL of water was added to the reaction solution, and it was extracted with 40 mL (20 mL × 2) of ethyl acetate. The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by high performance liquid chromatography (column model: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% trifluoroacetic acid)-acetonitrile]; acetonitrile %: 10%-40%, 10 minutes) to obtain the trifluoroacetate salt of compound 15-8. MS ESI calculated value: C 23 H 29 N9Se[M+H] + 512, found 512. 1 H NMR (400 MHz, CD3OD) δ 7.85 (s, 1H), 6.69 (s, 1H), 6.64 (d, J = 0.8 Hz, 1H), 5.85 (s, 1H), 4.24 (s, 2H), 4.14 - 4.22 (m, 1H), 3.51 (s, 2H), 3.10 (t, J = 7.2 Hz, 2H), 3.03 (s, 3H), 2.38 - 2.48 (m, 4H), 2.28 - 2.38 (m, 5H), 2.12 (t, J = 12.4 Hz, 2H).

[0202] Example 16

[0203]

[0204] Step 1: At 20 °C, dissolve compound 16-1 (11.88 mg, 139.71 μmol) in DMF (0.5 mL), add O-(7-azabenzotriazol-1-yl)-N,N,N,N-tetramethyluronium hexafluorophosphate (88.54 mg, 232.86 μmol, 34.62 μL), and stir the mixture at 20 °C for 0.5 h. Add a solution of compound 1-9 hydrochloride (60 mg, 116.43 μmol) and diisopropylethylamine (45.14 mg, 349.28 μmol, 60.84 μL) in DMF (0.5 mL) to the mixture and stir at 20 °C for 16 h. Add 10 mL of water to the reaction solution and extract with ethyl acetate (20 mL × 3). Wash the combined organic phases with saturated brine (5 mL × 3), and finally dry the organic phase over anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain the crude product. The crude product was separated by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA)-ACN]; ACN%: 23%-53%, 10 min) to obtain the trifluoroacetate of compound 16-2. MS ESI calculated value: C 23 H 26 N8OSe[M+H] + 511, found 511, 1 H NMR (400 MHz, CDCl3) δ 1.73-1.84 (m, 2H), 1.85-1.98 (m, 2H), 1.98-2.06 (m, 2H), 2.13-2.29 (m, 4H), 2.34 (s, 3H), 2.55 (s, 3H), 3.45-3.56 (m, 2H), 4.01 (m, 1H), 4.26-4.33 (m, 1H), 4.76-4.84 (m, 1H), 5.76 (s, 1H), 6.47 (s, 1H), 6.64 (s, 1H), 7.63 (s, 1H), 8.27 (s, 1H).

[0205] Example 17

[0206]

[0207] Step 1: Dissolve compound 1-3 (2 g, 5.51 mmol) in tetrahydrofuran (20 mL) at 0 °C, add sodium hydride (330.71 mg, 8.27 mmol, 60% purity), and stir at 0 °C for 0.5 h. Add methyl iodide (0.17 g, 8.27 mmol, 514.70 μL) to the reaction solution and stir at 20 °C for 1 h. Add 30 mL of saturated aqueous sodium bicarbonate to the reaction solution, extract with ethyl acetate (30 mL × 3), wash the combined organic phases with saturated brine (5 mL × 3), finally dry the organic phase over anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 50 / 1 - 20 / 1) to obtain compound 17-1. MS ESI calculated value: C 19 H 25 ClN4O2[M+H] + 377, found 377.

[0208] Step 2: Dissolve compound 17-1 (1.86 g, 4.94 mmol) in dioxane (20 mL), add compound 1-4 (1.18 g, 5.18 mmol), cesium carbonate (3.22 g, 9.87 mmol) and [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)]palladium(II) mesylate (223.69 mg, 246.76 μmol). After purging with nitrogen three times, heat to 100 °C and stir under nitrogen protection for 16 h. Add 30 mL of water to the reaction solution, extract with ethyl acetate (40 mL × 3), wash the combined organic phases with saturated brine (5 mL × 3) again, finally dry the organic phase over anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether:ethyl acetate = 10 / 1 - 5 / 1) to obtain compound 17-2. MS ESI calculated value: C 29 H 45 N7O3Si[M+H] + 568, found 568.

[0209] Step 3: At 0 - 5 °C, selenium powder (885.68 mg, 10.94 mmol) was added to ethanol (20 mL), and then sodium borohydride (413.75 mg, 10.94 mmol) was slowly added. After stirring at room temperature for half an hour until the solid particles completely disappeared, pyridine hydrochloride (1.69 g, 14.58 mmol) and compound 17 - 2 (2.07 g, 3.65 mmol) were added to the reaction solution, and the temperature was raised to 80 °C and stirred for 2 hours. 40 mL of water was added to the reaction solution, and it was extracted with ethyl acetate (30 mL × 3). The combined organic phases were washed with saturated brine (15 mL), and finally the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 8 / 1 - 2 / 1) to obtain compound 17 - 3. MS ESI calculated value: C 29 H 47 N7O3SeSi[M + H] + 650, found 650.

[0210] Step 4: At 25 °C, compound 17 - 3 (300 mg, 462.41 μmol) was dissolved in ethanol (3 mL), 6 - 5 (300 mg, 3.24 mmol) was added, and stirred at 80 °C for 2 hours. The reaction solution was directly concentrated under reduced pressure to obtain the crude product 17 - 4. MS ESI calculated value C 32 H 49 N7O3SeSi[M + H] + 688, found 688.

[0211] Step 5: Compound 17 - 4 (200 mg, 359.35 μmol) was dissolved in methanol (0.5 mL), hydrochloric acid ethyl acetate solution (4 M, 3 mL) was added, and stirred at 20 °C for 16 hours. The reaction solution was directly rotary evaporated to obtain the crude hydrochloride of compound 17 - 5. MS ESI calculated value C 21 H 27 N7Se[M + H] + 458, found 458.

[0212] Step 6: Dissolve the crude product of 17-5 hydrochloride (160 mg, 324.61 μmol) in methanol (1 mL), add N,N-diisopropylethylamine (125.86 mg, 973.82 μmol, 169.62 μL), then add acrylonitrile (1-10) (120 mg, 2.26 mmol, 150.00 μL), and stir at 20 °C for 2 hours. Add 20 mL of water to the reaction solution, extract with dichloromethane (20 mL × 3), wash the combined organic phases with saturated brine (5 mL × 3), finally dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain the crude product. The crude product is separated by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA)-ACN]; ACN%: 16%-46%, 10 minutes) to obtain the trifluoroacetate salt of compound 17-6. MS ESI calculated value C 24 H 30 N8Se[M+H] + 511, found 511, 1 H NMR (400 MHz, CDCl3) δ 1.26 (s, 1H), 1.91 - 2.00 (m, 2H), 2.30 - 2.40 (m, 7H), 2.45 - 2.62 (m, 6H), 2.97 (s, 3H), 3.14 (m, 2H), 3.30 (m, 2H), 4.16 (m, 2H), 4.86 - 5.04 (m, 1H), 6.07 (s, 1H), 6.58 (s, 1H), 6.94 (s, 1H), 7.63 (s, 1H), 10.60 - 10.77 (m, 1H).

[0213] Example 18

[0214]

[0215] Step 1: At 25 °C, dissolve compound 18-1 (0.4 g, 2.3 mmol) in dimethyl sulfoxide (2 mL), add 1-2 (520.30 mg, 2.30 mmol) and N,N-diisopropylethylamine (594.3 mg, 4.60 mmol, 800.89 μL), stir at 25 °C for 1 hour. Add 100 mL of water to the reaction solution, and extract with ethyl acetate (100 mL × 3). The combined organic phases are washed with saturated brine (100 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 4 / 1) to obtain compound 18-2. 1H NMR (400 MHz, CDCl3) δ 1.46 - 1.49 (m, 9H), 1.61 - 1.69 (m, 4H), 1.77 - 1.87 (m, 2H), 1.97 - 2.04 (m, 2H), 4.27 - 4.32 (m, 2H), 4.44 - 4.69 (m, 1H), 5.78 - 5.94 (s, 1H), 6.53 - 6.69 (s, 1H).

[0216] Step 2: Dissolve compound 18-2 (0.6 g, 1.65 mmol) in dioxane (10 mL), add 1-4 (412.46 mg, 1.81 mmol), cesium carbonate (1.07 g, 3.30 mmol) and [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)]palladium(II) mesylate mesylate (74.74 mg, 82.45 μmol). After purging with nitrogen three times, heat to 100 °C and stir for 16 hours under nitrogen protection. Add 50 mL of ammonium chloride aqueous solution to the reaction solution, and extract with ethyl acetate (100 mL × 3). The combined organic phases are washed with saturated brine (100 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 5 / 1) to obtain compound 18-3. MS ESI calculated value: C 27 H 42 N8O3Si [M+H] + 555, found 555. 11H NMR (400 MHz, CD3OD) δ 0.00 - 0.01 (m, 9H), 0.91 - 0.97 (m, 2H), 1.25 - 1.29 (m, 2H), 1.48 - 1.49 (m, 9H), 1.60 - 1.73 (m, 4H), 1.77 - 1.83 (m, 2H), 1.94 - 2.03 (m, 2H), 2.26 - 2.29 (s, 3H), 4.25 - 4.37 (m, 2H), 4.88 - 5.03 (m, 1H), 5.42 (s, 2H), 6.17 - 6.25 (s, 1H), 6.34 - 6.42 (s, 1H), 7.40 - 7.51 (s, 1H).

[0217] Step 3: At 25 °C, dissolve compound 18 - 3 (0.35 g, 630.91 μmol) in DMF (10 mL), sequentially add water (0.5 mL), triethylamine (363.50 mg, 3.59 mmol, 0.5 mL) and selenium powder (150 mg, 1.85 mmol). After displacing with carbon monoxide three times, react at 90 °C under 15 psi pressure for 1.5 h. Add 50 mL of ammonium chloride aqueous solution to the reaction solution, and extract with dichloromethane (50 mL × 3). The combined organic phases are washed with saturated brine (50 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain crude product 18 - 4.

[0218] Step 4: At 25 °C, dissolve compound 18 - 4 (0.4 g, 629.19 μmol) in ethanol (4 mL), add 6 - 5 (58.22 mg, 629.19 μmol), and stir at 80 °C for 1 h. The reaction solution is directly concentrated under reduced pressure to obtain crude product 18 - 5. MS ESI calculated value C 30 H 46 N8O3SeSi [M + H] + 675, found 675.

[0219] Step 5: Dissolve compound 18 - 5 (350 mg, 519.46 μmol) in hydrochloric acid ethyl acetate solution (4 M, 5 mL), and stir at 25 °C for 2 h. The reaction solution is directly filtered to obtain the filter cake as the crude hydrochloride salt of compound 18 - 6. MS ESI calculated value C 19 H 24 N8Se [M + H] + 445, found 445.

[0220] Step 6: Dissolve the crude product of 18 - 6 hydrochloride (100 mg, 193.68 μmol) in methanol (10 mL), add N,N - diisopropylethylamine (75.1 mg, 581.0 μmol, 101.20 μL), stir the mixture at 25 °C for 10 minutes, then add acrylonitrile (1 - 10) (20.55 mg, 387.35 μmol, 25.69 μL), and stir at 25 °C for 16 hours. Add 50 mL of ammonium chloride aqueous solution to the reaction solution, extract with ethyl acetate (50 mL × 3), wash the combined organic phases with saturated brine (50 mL), finally dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain the crude product. The crude product is separated by preparative high - performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA) - ACN]; B (ACN)%: 14% - 44%, 10 minutes) to obtain the trifluoroacetate of compound 18 - 7. MS ESI calculated value C 22 H 27 N9Se[M + H] + 498, found 498, 1 H NMR (400 MHz, CD3OD) δ 2.04 - 2.13 (m, 2H), 2.21 - 2.28 (m, 2H), 2.31 - 2.47 (m, 7H), 2.57 - 2.61 (s, 3H), 3.07 - 3.12 (m, 2H), 3.45 - 3.53 (m, 2H), 4.18 - 4.28 (m, 2H), 4.56 - 4.73 (m, 1H), 5.87 - 6.04 (s, 1H), 6.66 - 6.88 (s, 1H), 8.07 - 8.13 (s, 1H).

[0221] Example 19

[0222]

[0223] Step 1: Dissolve the crude product of 18 - 6 hydrochloride (30 mg, 44.68 μmol) in DMF (0.5 mL), add triethylamine (22.61 mg, 223.40 μmol, 31.09 μL), stir the mixture for 0.5 h, then add DMF (0.5 mL) to the reaction solution and cool to 0 °C, stir for more than 0.5 h, slowly add 3 - 1 (9.68 mg, 53.62 μmol), warm the mixture to 20 °C and stir for 3 h. LC - MS shows that the raw materials are consumed completely and the main peak is the product peak. Add saturated ammonium chloride aqueous solution (20 mL) to the reaction solution, extract with ethyl acetate (20 mL × 3), wash the combined organic phases with saturated brine (5 mL × 3), finally dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain the crude product. The crude product is separated by preparative high - performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA) - ACN]; ACN%: 28% - 58%, 10 minutes) to obtain the trifluoroacetate of compound 19 - 1. MS ESI calculated value C 23 H 28 N 10 O2SSe[M + H] + 589, found 589, 1 H NMR (400 MHz, CDCl3) δ 1.24 - 1.36 (m, 1H), 1.82 (m, 2H), 1.87 - 1.94 (m, 2H), 2.10 - 2.17 (m, 2H), 2.22 - 2.27 (m, 2H), 2.40 (s, 3H), 2.57 (s, 3H), 3.39 - 3.48 (m, 1H), 3.87 - 4.00 (m, 1H), 4.10 - 4.18 (m, 4H), 4.32 (m, 2H), 5.86 (s, 1H), 6.80 (s, 1H), 7.74 (d, 1H), 7.96 (m, 1H), 9.01 - 9.31 (m, 1H).

[0224] Example 20

[0225]

[0226] Step 1: At 20 °C, diisopropylethylamine (2.50 g, 19.32 mmol) and compound 20-1 (2 g, 9.66 mmol) were added to tetrahydrofuran (40 mL). 1-2 (2.19 g, 9.66 mmol) was dissolved in tetrahydrofuran (10 mL) and added dropwise to the former system at 0 °C, and the reaction was carried out at 20 °C for 18 hours. The reaction solution was diluted with 150 mL of water, extracted with 120 mL of ethyl acetate (40 mL × 3), the combined organic layers were washed with 40 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column (petroleum ether / ethyl acetate = 10 / 1 to 1 / 1) to obtain compound 20-2.

[0227] Step 2: Compound 20-2 (2.15 g, 5.42 mmol) was dissolved in methanol (20 mL) and water (6 mL), lithium hydroxide (454.63 mg, 10.83 mmol) was added, and the reaction was carried out at 20 °C for 2 hours. The pH of the reaction solution was adjusted to 4 with 0.5 M aqueous hydrochloric acid, the reaction solution was diluted with 100 mL of water, extracted with 120 mL of ethyl acetate (40 mL × 3), the combined organic layers were washed with 40 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure to obtain compound 20-3. MS ESI calculated value: C 17 H 23 ClN4O4[M+H]+383, found 383.

[0228] Step 3: Compound 20-3 (1.9 g, 4.96 mmol) was dissolved in dichloromethane (40 mL), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (1.14 g, 5.96 mmol) and 1-hydroxybenzotriazole (804.70 mg, 5.96 mmol) were added, and the reaction was carried out at 20 °C for 0.5 hour. Then methoxymethylamine hydrochloride (726.12 mg, 7.44 mmol) and triethylamine (2.01 g, 19.85 mmol) were added, and the reaction solution was reacted at 20 °C for 16 hours. The reaction solution was diluted with 100 mL of water, extracted with 100 mL of ethyl acetate (50 mL × 2), the combined organic layers were washed with 50 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column (petroleum ether / ethyl acetate = 10 / 1 to 3 / 1) to obtain compound 20-4. MS ESI calculated value: C 19 H 28 ClN5O4[M+H]+426, found 426.

[0229] Step 4: Compound 20-4 (1.80 g, 4.24 mmol) was dissolved in tetrahydrofuran (30 mL), and a solution of methylmagnesium bromide in diethyl ether (3 M, 3.11 mL) was added dropwise at 0 °C. The reaction was carried out at 0 °C for 1 hour. The reaction was quenched by adding 20 mL of saturated ammonium chloride solution. The reaction mixture was diluted with 60 mL of water and extracted with 60 mL of ethyl acetate (30 mL×2). The combined organic layers were washed with 30 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. Purification by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1 to 4 / 1) gave Compound 20-5. 1 H NMR (400 MHz, CDCl3) δ 1.47 (s, 9H), 1.52 - 1.93 (m, 4H), 2.05 (s, 4H), 2.61 (s, 3H), 4.14 - 4.37 (m, 2H), 4.37 - 4.73 (m, 1H), 5.37 - 5.74 (m, 1H), 6.82 (s, 1H)

[0230] Step 5: Compound 20-5 (660 mg, 1.73 mmol) was dissolved in dichloromethane (20 mL). Triethylamine (1.05 g, 10.40 mmol) was added, and then tert-butyldimethylsilyl trifluoromethanesulfonate (1.37 g, 5.20 mmol) was added dropwise. The reaction was carried out at 15 °C for 1 hour. The reaction mixture was diluted with 60 mL of dichloromethane and washed with 60 mL of saturated aqueous sodium bicarbonate solution (30 mL×2), 20 mL of water, and 30 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure to give Compound 20-6. MS ESI calculated value: C 30 H 53 ClN4O3Si [M+H]+ 609, found 609.

[0231] Step 6: Compound 20-6 (1.14 g, 1.87 mmol) was dissolved in tetrahydrofuran (30 mL) and water (5 mL). N-Bromosuccinimide (332.95 mg, 1.87 mmol) was added, and the reaction was carried out at 15 °C for 2 hours. The reaction mixture was diluted with 100 mL of water and extracted with 100 mL of ethyl acetate (50 mL×2). The combined organic layers were washed with 50 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure to give Compound 20-7. MS ESI calculated value: C 18 H 24 BrClN4O3 [M+H] + 459, found 459.

[0232] Step 7: Compound 20-7 (650 mg, 1.13 mmol) was dissolved in methanol (25 mL). Compound 12-2 (138.18 mg, 1.13 mmol) was added at 0 - 5 °C, and the reaction was carried out at 15 °C for 0.5 h. Then Boc anhydride (296.56 mg, 1.36 mmol), 2,6-dimethylpyridine (13.83 mg, 113.24 μmol) and triethylamine (458.33 mg, 4.53 mmol) were added, and the reaction was carried out at 15 °C for 16 h. The reaction solution was diluted with 100 mL of water, extracted with 90 mL of ethyl acetate (30 mL×3), the combined organic layers were washed with 50 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column (petroleum ether / ethyl acetate = 5 / 1 to 3 / 1) to obtain Compound 20-8. MS ESI calculated value: C 20 H 26 ClN5O2Se[M+H] + 484, found 484.

[0233] Step 8: Compound 20-8 (100 mg, 205.03 μmol) and 8-9 (48.53 mg, 246.03 μmol) were added to dioxane (3 mL), and then [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] palladium(II) mesylate (18.59 mg, 20.50 μmol) and cesium carbonate (133.60 mg, 410.05 μmol) were added. The reaction was carried out at 80 °C under a nitrogen atmosphere for 3 h. The reaction solution was diluted with 50 mL of water and extracted with 40 mL (20 mL×2) of ethyl acetate. The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column (petroleum ether / ethyl acetate = 5 / 1 to 1 / 1) to obtain Compound 20-9. MS ESI calculated value: C 29 H 40 N8O4Se[M+H]+645, found 645.

[0234] Step 9: Compound 20-9 (65 mg, 100.99 μmol) was dissolved in methanol (1 mL), and hydrochloric acid ethyl acetate (4 M, 2 mL) was added. The reaction was carried out at 50 °C for 2 h. The reaction solution was concentrated under reduced pressure to obtain Compound 20-10. MS ESI calculated value: C 19 H 24 N8Se[M+H] + 444, found 444.

[0235] Step 10: Compound 20-10 (50 mg, 104.20 μmol) was dissolved in methanol (2 mL), and diisopropylethylamine (67.33 mg, 520.98 μmol) and acrylonitrile (0.22 g, 4.15 mmol) were added. The reaction was carried out at 15 °C for 2 hours. 5 mL of water and 5 mL of methanol were added to the reaction solution, and it was stirred for 15 minutes. The resulting precipitate was filtered, and the filter cake was slurried with 5 mL of methanol and then filtered to obtain Compound 20-11. Calculated value by MSESI: C 22 H 27 N9Se[M+H]+ 498, found 498. 1 H NMR (400 MHz, DMSO-d6) δ 1.52 (d, J = 10.4 Hz, 2H), 1.71 (d, J = 6.8 Hz, 2H), 1.80 (s, 2H), 1.92 (d, J = 2.8 Hz, 2H), 2.20 (s, 3H), 2.60 (m, 4H), 2.73 (s, 3H), 3.25 - 3.32 (m, 2H), 4.13 - 4.43 (m, 1H), 6.60 (s, 2H), 6.94 - 7.24 (m, 1H), 8.36 - 9.11 (m, 2H), 11.72 (s, 1H).

[0236] Example 21

[0237]

[0238] Step 1: Compound 20-7 (600 mg, 1.05 mmol) was dissolved in methanol (25 mL), and Compound 8-6 (201.84 mg, 1.05 mmol) was added at 0 - 5 °C. The reaction was carried out at 15 °C for 0.5 hour. Then, (Boc)2O (273.75 mg, 1.25 mmol), 2,6-dimethylpyridine (12.77 mg, 104.53 μmol) and triethylamine (423.07 mg, 4.18 mmol) were added, and the reaction was carried out at 15 °C for 16 hours. The reaction solution was diluted with 100 mL of water and extracted with 90 mL of ethyl acetate (30 mL × 3). The combined organic layers were washed with 50 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column chromatography (petroleum ether / ethyl acetate = 5 / 1 to 3 / 1) to obtain Compound 21-1. Calculated value by MS ESI: C 23 H 31 ClN6O3Se[M+H] + 555, found 555.

[0239] Step 2: Compound 21-1 (250 mg, 451.31 μmol), 8-9 (106.82 mg, 541.57 μmol) were added to dioxane (8 mL), and then [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] palladium(II) methanesulfonate (40.91 mg, 45.13 μmol) and cesium carbonate (294.09 mg, 902.62 μmol) were added. The reaction was carried out at 80 °C under a nitrogen atmosphere for 3 hours. The reaction solution was diluted with 50 mL of water and extracted with 40 mL (20 mL×2) of ethyl acetate. The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column (petroleum ether / ethyl acetate = 5 / 1 to 1 / 1) to obtain Compound 21-2. MS ESI calculated value: C 32 H 45 N9O5Se[M+H] + 716, found 716.

[0240] Step 3: Compound 21-2 (220 mg, 270.88 μmol) was dissolved in methanol (2 mL), and hydrochloric acid ethyl acetate (4 M, 6 mL) was added. The reaction was carried out at 40 °C for 2 hours. The reaction solution was concentrated under reduced pressure to obtain Compound 21-3. MS ESI calculated value: C 22 H 29 N9Se[M+H] + 516, found 516.

[0241] Step 4: Compound 21-3 (150 mg, 272.26 μmol) was dissolved in methanol (5 mL), and diisopropylethylamine (175.93 mg, 1.36 mmol) and acrylonitrile (28.89 mg, 544.52 μmol) were added. The reaction was carried out at 15 °C for 2 hours. 10 mL of water and 10 mL of methanol were added to the reaction solution, and it was stirred for 15 minutes. The filtrate was obtained by filtration. The filter cake was slurried with 10 mL of methanol and filtered to obtain Compound 21-4. MS ESI calculated value: C 25 H 32 N 10 OSe[M+H] + 569, found 569. 11H NMR (400 MHz, DMSO-d6) δ 1.40 - 1.56 (m, 2H), 1.70 (d, J = 7.2 Hz, 2H), 1.77 (s, 2H), 1.86 - 1.97 (m, 2H), 2.20 (s, 3H), 2.59 (m, 4H), 3.29 (s, 2H), 3.43 (s, 4H), 3.72 (d, J = 4.40 Hz, 4H), 4.10 - 4.37 (m, 1H), 6.49 (d, J = 14.4 Hz, 2H), 7.03 (d, J = 2.4 Hz, 1H), 7.97 (s, 1H), 8.55 (s, 1H), 11.70 (s, 1H)

[0242] Example 22

[0243]

[0244] Step 1: Dissolve 22-1 (1.2 g, 2.48 mmol) in ethanol (50 mL), add 22-2 (580.83 mg, 2.98 mmol), and stir at 80 °C for 1 hour. The reaction solution was concentrated under reduced pressure to obtain a crude product, diluted with 50 mL of tetrahydrofuran, add triethylamine (502.31 mg, 4.96 mmol) and di-tert-butyl dicarbonate (541.70 mg, 2.48 mmol), and stir at 80 °C for 1 hour. Add 100 mL of water to the reaction solution for dilution, and extract with ethyl acetate (50 mL × 2), wash with saturated brine (50 mL × 2), finally dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain a crude product. The crude product was purified by column chromatography (SiO2, petroleum ether: ethyl acetate = 20 / 1 - 3 / 1) to obtain compound 22-3. MS ESI calculated value C 23 H 29 ClN4O4Se [M+H] + 540, found 540.

[0245] Step 2: Dissolve 22-3 (280 mg, 518.60 μmol) in tetrahydrofuran (6 mL), add a solution of lithium hydroxide monohydrate (65.29 mg, 1.56 mmol) in water (2 mL), and stir at 35 °C for 16 hours. Adjust the pH of the reaction solution to 4 - 5 with 1 M dilute hydrochloric acid, concentrate to obtain a crude product, soak the crude product with tetrahydrofuran (10 mL) and methanol (5 mL), filter, and concentrate the filtrate to obtain compound 22-4. MS ESI calculated value C 21 H 25 C1N4O4Se [M+H] + 512, found 512.

[0246] Step 3: Dissolve 22-4 (160 mg, 312.59 μmol) in acetonitrile (4 mL), add a solution of 22-5 (35.69 mg, 625.17 μmol) and N,N-diisopropylethylamine (161.59 mg, 1.25 mmol) in DMF (2 mL), then add 2-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (130.74 mg, 343.84 μmol), and stir at 30 °C for 16 h. The reaction solution was diluted with water (10 mL), extracted with ethyl acetate (10 mL × 2), washed with saturated brine (20 mL × 2), the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product, and the crude product was purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 5 / 1~1 / 1) to obtain compound 22-6. MS ESI calculated value C 24 H 30 ClN5O3Se[M+H] + 551, found 551.

[0247] Step 4: Dissolve compound 22-6 (150 mg, 272.26 μmol), 8-9 (64.44 mg, 326.71 μmol), cesium carbonate (177.42 mg, 544.52 μmol) and palladium(II) mesylate of [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] (24.68 mg, 27.23 μmol) in dioxane (5 mL), after purging with nitrogen three times, heat to 100 °C, and stir under nitrogen protection for 16 h. The reaction solution was concentrated under reduced pressure to obtain a crude product, and the crude product was purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 4 / 1~1 / 2) to obtain compound 22-7. MS ESI calculated value: C 33 H 44 N8O5Se[M+H] + 712, found 712.

[0248] Step 5: Dissolve compound 22-7 (100 mg, 140.51 μmol) in ethyl acetate (2 mL), add hydrochloric acid ethyl acetate solution (4 M, 2 mL), and stir at 30 °C for 16 h. The reaction solution was directly filtered to obtain a filter cake which was the crude product of the hydrochloride salt of compound 22-8. MS ESI calculated value C 23 H 28 N8OSe[M+H] + 512, found 512.

[0249] Step 6: Dissolve the crude product of 22-8 hydrochloride (70 mg, 136.86 μmol, HCl) in methanol (5 mL), add N,N-diisopropylethylamine (88.44 mg, 684.29 μmol), and acrylonitrile (1-10) (21.79 mg, 410.57 μmol), and stir at 25 °C for 16 hours. LC-MS showed that the raw materials were completely consumed and the main peak was the product peak. Add 10 mL of ammonium chloride aqueous solution to the reaction solution, extract with ethyl acetate (20 mL × 3), wash the combined organic phases with saturated brine (20 mL), dry over anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain the crude product. The crude product was separated by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA)-ACN]; B (ACN)%: 12%-42%, 10 minutes), adjust the pH = 8 with ammonia water, extract with ethyl acetate (10 mL × 3), dry the combined organic phases over anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain compound 22-9. MS ESI calculated value C 26 H 31 N9OSe[M+H] + 565, found 565, 1 H NMR (400 MHz, DMSO-d6) δ 0.61-0.67 (m, 2H), 0.69-0.75 (m, 2H), 1.50 (m, 2H), 1.73 (m, 2H), 1.76-1.85 (m, 2H), 1.87-1.97 (m, 2H), 2.19 (s, 3H), 2.52 (m, 1H), 2.56-2.65 (m, 4H), 2.86 (m, 1H), 3.31 (s, 3H), 4.08-4.29 (m, 1H), 6.30 (s, 1H), 6.33-6.49 (m, 1H), 6.50-6.67 (m, 1H), 8.20 (s, 1H), 8.85 (s, 1H), 11.56-11.88 (m, 1H).

[0250] Example 23

[0251]

[0252] Step 1: Dissolve 22-3 (80 mg, 156.29 μmol) in acetonitrile (4 mL), add a solution of 23-1 (32.50 mg, 312.59 μmol) and N,N-diisopropylethylamine (80.80 mg, 625.17 μmol) in DMF (1 mL), then add 2-(7-azabenzotriazol)-N,N,N,N-tetramethyluronium hexafluorophosphate (65.37 mg, 171.92 μmol), and stir at 50 °C for 16 h. Dilute the reaction solution with water (10 mL), extract with ethyl acetate (10 mL × 2), wash with saturated brine (20 mL × 2), dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain the crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 3 / 1 - 1 / 1) to obtain compound 23-2. MS ESI calculated value for C 22 H 28 ClN5O3Se[M+H] + 525, found 525.

[0253] Step 2: Dissolve compound 23-2 (60 mg, 114.31 μmol), 8-9 (27.05 mg, 137.17 μmol), cesium carbonate (74.49 mg, 228.61 μmol) and [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] palladium(II) mesylate mesylate (20.72 mg, 22.86 μmol) in dioxane (5 mL), displace with nitrogen three times and then heat to 100 °C, and stir under nitrogen protection for 16 h. Concentrate the reaction solution under reduced pressure to obtain the crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 4 / 1 - 1 / 1) to obtain compound 23-3. MS ESI calculated value: C 31 H 42 N8O5Se[M+H] + 686, found 686.

[0254] Step 3: Dissolve compound 23-3 (40 mg, 58.34 μmol) in ethyl acetate (2 mL), add hydrochloric acid ethyl acetate solution (4 M, 2 mL), and stir at 30 °C for 3 h. Filter the reaction solution directly to obtain the filter cake as the crude hydrochloride of compound 23-4. MS ESI calculated value for C 21 H 26 N8OSe[M+H] + 485, found 485.

[0255] Step 4: Dissolve the crude product of 23-4 hydrochloride (60 mg, 123.60 μmol) in methanol (5 mL), add N,N-diisopropylethylamine (79.87 mg, 617.99 μmol), and acrylonitrile (1-10) (19.68 mg, 370.80 μmol), and stir at 25 °C for 1 hour. Add 10 mL of aqueous ammonium chloride solution to the reaction solution, extract with ethyl acetate (20 mL × 3), wash the combined organic phases with saturated brine (20 mL), dry the organic phase over anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain the crude product. The crude product is separated by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA)-ACN]; B (ACN)%: 10%-40%, 10 minutes), adjust the pH = 8 with ammonia water, extract with ethyl acetate (10 mL × 3), dry the combined organic phases over anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain compound 23-5. MS ESI calculated value C 24 H 29 N9OSe[M+H] + 539, found 539, 1 H NMR (400 MHz, DMSO-d6) δ 1.51 (m, 2H), 1.73 (m, 2H), 1.79 (s, 2H), 1.91 (m, 2H), 2.19 (s, 3H), 2.55 - 2.65 (m, 5H), 2.81 (d, J = 4.75 Hz, 3H), 3.31 (s, 3H), 4.07 - 4.31 (m, 1H), 6.29 (s, 1H), 6.37 (s, 1H), 6.60 (s, 1H), 8.23 (s, 1H), 8.84 (s, 1H), 11.58 - 11.86 (s, 1H)

[0256] Example 24

[0257]

[0258] Step 1: Dissolve 22-3 (200 mg, 370.43 μmol) in tetrahydrofuran (5 mL), add a toluene solution of diisobutylaluminum hydride (1 M, 1.18 mL) at -65 °C, and stir at 0 °C for 2 hours. At 0 °C, add saturated potassium sodium tartrate (5 mL) to the reaction solution, stir for 0.5 hour to quench, then add saturated brine (10 mL) for dilution, extract with ethyl acetate (20 mL × 2), wash the organic phase with saturated brine (20 mL × 2), dry over anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain compound 24-1. MS ESI calculated value C 21 H 27 ClN4O3Se[M+H] +498, measured value 498.

[0259] Step 2: Dissolve 24-1 (180 mg, 361.54 μmol) in dichloromethane (5 mL), add Dess-Martin periodinane (230.01 mg, 542.30 μmol 1), and stir at 20 °C for 1 hour. Add saturated sodium sulfite (5 mL) and saturated sodium bicarbonate (5 mL) to the reaction solution, stir at 20 °C for 10 minutes to quench, then add water (10 mL) for dilution, extract with dichloromethane (20 mL × 2), wash the organic phase with saturated brine (20 mL × 2), dry over anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain the crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 10 / 1 - 3 / 1) to obtain compound 24-2. MS ESI calculated value C 21 H 25 ClN4O3Se [M+H] + 496, measured value 496.

[0260] Step 3: Dissolve 24-2 (170 mg, 310.20 μmol) in tetrahydrofuran (5 mL), add elemental iodine (393.66 mg, 1.55 mmol) and ammonia water (434.85 mg, 3.10 mmol, 25%), and stir at 30 °C for 16 hours. Add saturated sodium sulfite (5 mL) and saturated sodium bicarbonate (5 mL) to the reaction solution, stir at 20 °C for 10 minutes to quench, then add water (10 mL) for dilution, extract with dichloromethane (20 mL × 2), wash the organic phase with saturated brine (20 mL × 2), dry over anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain the crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 10 / 1 - 3 / 1) to obtain compound 24-3. MS ESI calculated value C 21 H 24 ClN5O2Se [M+H] + 496, measured value 493.

[0261] Step 4: Dissolve compound 24-3 (100 mg, 198.84 μmol), 8-9 (43.14 mg, 218.72 μmol), cesium carbonate (194.36 mg, 596.52 μmol) and palladium(II) mesylate of [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] (18.02 mg, 19.88 μmol) in dioxane (4 mL). After purging with nitrogen three times, heat the solution to 90 °C and stir for 2 hours under nitrogen protection. Concentrate the reaction solution under reduced pressure to obtain a crude product, which is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 10 / 1 - 3 / 1) to obtain compound 24-4. MSESI calculated value: C 30 H 38 N8O4Se[M+H] + 654, found 654.

[0262] Step 5: Dissolve compound 24-4 (100 mg, 151.00 μmol) in ethyl acetate (2 mL), add hydrochloric acid in ethyl acetate solution (4 M, 2 mL), and stir at 45 °C for 2 hours. Concentrate the reaction solution directly to obtain the crude hydrochloride of compound 24-5. MS ESI calculated value C 20 H 22 N8Se[M+H] + 453, found 453.

[0263] Step 6: Dissolve the crude hydrochloride of 24-5 (70 mg, 154.39 μmol) in methanol (2 mL), add N,N-diisopropylethylamine (99.77 mg, 771.94 μmol), and acrylonitrile (1-10) (24.58 mg, 463.17 μmol), and stir at 20 °C for 1 hour. LC-MS shows that the raw material is consumed and the main peak is the product peak. Add 10 mL of ammonium chloride aqueous solution to the reaction solution and extract with ethyl acetate (20 mL × 3). Wash the combined organic phases with saturated brine (20 mL), and finally dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain a crude product. The crude product is separated by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA)-ACN]; B (ACN)%: 13% - 43%, 10 minutes) to obtain the trifluoroacetate of compound 24-6. MS ESI calculated value C 23 H 25 N9Se[M+H] + 506, found 506, 11H NMR (400 MHz, CD3OD) δ 2.05 (m, 3H), 2.33 (m, 2H), 2.36 (s, 3H), 2.38 (m, 1H), 2.42 (m, 2H), 3.07 - 3.11 (m, 2H), 3.50 (m, 2H), 4.22 (s, 2H), 4.32 (m, 1H), 6.00 (s, 1H), 6.67 (s, 1H), 6.72 (s, 1H), 9.27 (s, 1H).

[0264] Example 25

[0265]

[0266] Step 1: Add 25-1 (10 g, 109.73 mmol) to anhydrous ethanol (30 mL), then add methyl iodide (18.38 g, 129.48 mmol). Stir at 80 °C in a sealed flask for 4 hours. The solid precipitates and is directly filtered to obtain the hydroiodide salt of compound 25-2. 1 1H NMR (400 MHz, D2O) δ 2.60 (s, 3H).

[0267] Step 2: At 25 °C, add selenium powder (6.95 g, 85.81 mmol) to ethanol (140 mL), then add sodium borohydride (3.57 g, 94.39 mmol) in batches. After the black selenium powder completely disappears, add sodium carbonate (4.55 g, 42.90 mmol) and the hydroiodide salt of 25-2 (10 g, 42.90 mmol), and warm up to 25 °C and stir for 16 hours. Add 20 mL of glacial acetic acid to quench the reaction solution, concentrate under reduced pressure to obtain the crude product, and the crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 2 / 1 - 0 / 1 then dichloromethane∶methanol = 20 / 1 - 10 / 1) to obtain compound 25-3. 1 1H NMR (400 MHz, DMSO-d6) δ 3.80 (s, 2H), 8.26 (s, 1H), 8.57 (s, 1H), 9.67 (s, 1H).

[0268] Step 3: Add 25-3 (4 g, 28.98 mmol) and 1-1 (4.01 g, 23.18 mmol) to trifluoroacetic acid (50 mL), and heat to 80 °C and stir for 1 hour. Concentrate the reaction solution under reduced pressure to obtain a crude product. Dilute the crude product with ethyl acetate (100 mL), adjust the pH to 7-8 with saturated aqueous sodium bicarbonate solution, extract with ethyl acetate (50 mL × 3), wash the organic phase with saturated brine (50 mL × 2), dry over anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether: ethyl acetate = 4 / 1 - 3 / 2) to obtain compound 25-4. MS ESI calculated value C7H4Cl2N4Se [M+H] + 294, found 294.

[0269] Step 4: Add 25-4 (1 g, 2.99 mmol) to DMF (20 mL), then add triethylamine (605.76 mg, 5.99 mmol), 4-dimethylaminopyridine (73.13 mg, 598.64 μmol) and di-tert-butyl dicarbonate (979.88 mg, 4.49 mmol), and stir at 25 °C for 1 hour. Add water (50 mL) to dilute, extract with ethyl acetate (30 mL × 3), wash the organic phase with saturated brine (50 mL × 2), dry over anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 20 / 1 - 5 / 1) to obtain compound 25-5. MS ESI calculated value C 12 H 12 Cl2N4O2Se [M+H] + 394, found 394.

[0270] Step 5: Add compound 25-5 (1.2 g, 3.04 mmol), 1-2 (826.90 mg, 3.65 mmol), tris(dibenzylideneacetone)dipalladium(0) (278.82 mg, 304.48 μmol), 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (352.35 mg, 608.96 μmol) and cesium carbonate (1.98 g, 6.09 mmol) to dioxane (40 mL). After purging with nitrogen three times, heat to 100 °C and stir under nitrogen protection for 16 hours. Concentrate the reaction solution under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether: ethyl acetate = 10 / 1 - 3 / 1) and then by preparative separation using high performance liquid chromatography (column: Waters Xbridge C18 150*50mm*10μm; mobile phase: [water (10 mM ammonium bicarbonate)-ACN]; B (ACN) %: 56% - 86%) to obtain compound 25-6. MS ESI calculated value: C 24 H33 ClN6O4Se[M+H] + 584, the measured value is 584.

[0271] Step 6: Add compound 25-6 (120 mg, 182.89 μmol), 8-9 (43.29 mg, 219.46 μmol), cesium carbonate (178.76 mg, 548.66 μmol), and [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] palladium(II) mesylate mesylate (16.58 mg, 18.29 μmol) to dioxane (5 mL). After purging with nitrogen three times, the temperature is raised to 90 °C, and the mixture is stirred under nitrogen protection for 3 hours. The reaction solution is concentrated under reduced pressure to obtain a crude product, which is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 10 / 1 - 2 / 1) to obtain compound 25-7. MSESI calculated value: C 33 H 47 N9O6Se[M+H] + 745, the measured value is 745.

[0272] Step 7: Dissolve compound 25-7 (80 mg, 107.42 μmol) in ethyl acetate (2 mL), add hydrochloric acid ethyl acetate solution (4 M, 2 mL), and stir at 45 °C for 2 hours. The reaction solution is directly concentrated to obtain a crude product of the hydrochloride salt of compound 25-8. MSESI calculated value C 18 H 23 N9Se[M+H] + 444, the measured value is 444.

[0273] Step 8: Dissolve the crude hydrochloride salt of 25-8 (50 mg, 103.98 μmol) in methanol (2 mL), add N,N-diisopropylethylamine (67.19 mg, 519.91 μmol), and acrylonitrile (1-10) (16.55 mg, 311.94 μmol), and stir at 20 °C for 1 hour. Add 10 mL of aqueous ammonium chloride solution to the reaction solution, and extract with ethyl acetate (20 mL × 3). The combined organic phases are washed with saturated brine (20 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product is separated by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA)-ACN]; B (ACN)%: 9% - 39%, 10 minutes) to obtain the trifluoroacetate salt of compound 25-9. MSESI calculated value C 21 H 26 N 10Se[M+H] + 497, measured value 497, 1 H NMR(400 MHz, CD3OD) δ 2.10(m, 3H), 2.33(m 1H), 2.36(s, 3H), 2.39 - 2.40(m, 1H), 2.43(m, 3H), 3.10(m, 3H), 3.51(m, 3H), 4.24(m, 3H), 5.92(s, 1H), 6.53(s, 1H), 6.56(s, 1H).

[0274] Example 26

[0275]

[0276] Step 1: At 20 °C, dissolve the hydrochloride salt of compound 18 - 6 (60 mg, 89.36 μmol) in DMF (0.5 mL), add potassium carbonate (24.70 mg, 178.72 μmol) and 4 - 1 (24.02 mg, 107.23 μmol, 12.57 μL), and stir the mixture at 50 °C for 16 hours. The reaction solution was directly concentrated under reduced pressure to obtain a crude product. The crude product was separated by preparative high - performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA) - ACN]; ACN%: 16% - 46%, 10 minutes) to obtain the trifluoroacetate salt of compound 26 - 1. MS ESI calculated value: C 22 H 27 F3N8Se[M+H] + 541, measured value 541, 1 H NMR(400 MHz, DMSO - d6) δ 1.83(m, 2H), 2.07 - 2.11(m, 2H), 2.23(s, 3H), 2.30 - 2.34(m, 2H), 2.44(s, 3H), 2.81 - 2.99(m, 2H), 3.17 - 3.30(m, 2H), 4.14(m, 3H), 4.25 - 4.49(m, 2H), 6.33(m, 1H), 6.70(s, 1H), 7.62 - 7.84(m, 1H), 7.96(s, 1H), 9.46 - 9.65(m, 1H), 9.69(m, 1H).

[0277] Example 27

[0278]

[0279] Step 1: Dissolve compound 27-1 (1.8 g, 12.28 mmol) in tetrahydrofuran (10 mL) and saturated sodium bicarbonate solution (10 mL) at 0 °C. Add CbzCl (2.30 g, 13.50 mmol, 1.92 mL), and stir at 20 °C for 16 hours. Add 10 mL of water to the reaction solution, and extract with dichloromethane (20 mL × 3). The combined organic phases are washed successively with 0.5 M aqueous hydrochloric acid solution (5 mL × 3) and saturated brine (5 mL × 3). Finally, the organic phase is dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 8 / 1 - 3 / 1) to obtain compound 27-2. 1 H NMR (400 MHz, CDCl3) δ 1.77 - 1.96 (m, 4H), 2.84 (m, 1H), 3.46 (m, 2H), 3.73 (m, 2H), 5.14 (s, 2H), 7.30 - 7.44 (m, 5H).

[0280] Step 2: Dissolve compound 27-2 (950 mg, 3.89 mmol) and selenium (629.80 mg, 7.78 mmol, 617.45 μL) in N,N-DMF (10 mL) and water (1 mL). Evacuate the mixture under vacuum and replace the gas with carbon monoxide three times. The reaction mixture is stirred at 80 °C for 16 hours under carbon monoxide gas (50 psi). The reaction solution is directly concentrated under reduced pressure to remove the solvent. Add 20 mL of water, and extract with ethyl acetate (20 mL × 3). The combined organic phases are washed again with saturated brine (20 mL × 3). Finally, the organic phase is dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether:ethyl acetate = 4 / 1 - 1 / 2) to obtain compound 27-3. 1 H NMR (400 MHz, CDCl3) δ 1.67 - 1.83 (m, 3H), 1.94 (m, 2H), 2.79 - 2.87 (m, 2H), 4.21 - 4.41 (m, 2H), 5.13 (m, 2H), 7.32 - 7.40 (m, 5H), 7.77 (m, 1H), 8.51 (m, 1H).

[0281] Step 3: Dissolve compound 27-3 (1.35 g, 4.15 mmol) in methanol (15 mL). Add 8-4 (858.58 mg, 3.19 mmol), and stir at 20 °C for 16 hours. The reaction solution is directly concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether:ethyl acetate = 4 / 1 - 3 / 1) to obtain compound 27-4. MS ESI calculated value C 21 H 19Cl2N3O2Se[M+H] + 496, the measured value is 496.

[0282] Step 4: Dissolve compound 27-4 (500 mg, 1.01 mmol), 1-2 (228.58 mg, 1.01 mmol), (±)-2,2-bis(diphenylphosphino)-1,1'-binaphthalene (62.89 mg, 101.00 μmol), palladium acetate (22.68 mg, 101.00 μmol) and cesium carbonate (658.15 mg, 2.02 mmol) in dioxane (12 mL). The mixture is purged with nitrogen three times and stirred at 90 °C for 16 hours. Add 30 mL of water to the reaction solution and extract with ethyl acetate (30 mL × 3). The combined organic phases are washed with saturated brine (5 mL × 3). Finally, the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 8 / 1 to 1 / 1) to obtain compound 27-5. MS ESI calculated value C 33 H 40 ClN5O4Se[M+H] + 686, the measured value is 686.

[0283] Step 5: Dissolve compound 27-5 (500 mg, 729.81 μmol), 8-9 (158.34 mg, 802.79 μmol), methanesulfonic acid (2-dicyclohexylphosphino)-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)(2′-amino-1,1′-biphenyl-2-yl)palladium(II) (BrettPhos Pd G3) (66.16 mg, 72.98 μmol) and cesium carbonate (475.57 mg, 1.46 mmol) in dioxane (12 mL). The mixture is purged with nitrogen three times and stirred at 90 °C for 16 hours. Add 40 mL of water to the reaction solution and extract with dichloromethane (30 mL × 3). The combined organic phases are washed with saturated brine (5 mL × 3). Finally, the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether: ethyl acetate = 5 / 1 to 1 / 1) to obtain compound 27-6. MS ESI calculated value C 42 H 54 N s O6Se[M+H] + 847, the measured value is 847.

[0284] Step 6: Dissolve compound 27-6 (380 mg, 449.23 μmol) in methanol (1 mL), add hydrochloric acid ethyl acetate solution (4 M, 4 mL), and stir at 20 °C for 16 h. The reaction solution was directly concentrated in vacuo to obtain the crude hydrochloride salt of compound 27-7. Calculated value of MSESI C 32 H 38 N8O2Se[M+H] + 647, found 647.

[0285] Step 7: Dissolve the crude hydrochloride salt of 27-7 (300 mg, 417.49 μmol) in methanol (3 mL), add N,N-diisopropylethylamine (161.87 mg, 1.25 mmol, 218.16 μL), then add compound acrylonitrile (1-10) (33.23 mg, 626.24 μmol, 41.54 μL), and stir at 20 °C for 2 h. LC-MS showed that the raw material was consumed completely and the main peak was the product peak. The reaction solution was directly concentrated under reduced pressure to obtain the crude product, which was separated by preparative high performance liquid chromatography (column: Phenomenex luna C18 150*40 mm*15 μm; mobile phase: [water (0.1% TFA)-ACN]; ACN%: 18%-48%, 11 min) to obtain the trifluoroacetate salt of compound 27-8. Calculated value of MS ESI C 35 H 41 N9O2Se[M+H] + 700, found 700.

[0286] Step 8: Dissolve the trifluoroacetate salt of 27-8 (90 mg, 128.81 μmol) in hydrobromic acid (0.3 mL), and stir at 15 °C for 16 h. The reaction solution was filtered to obtain the crude product, which was separated by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150*25 mm*10 μm; mobile phase: [water (0.1% TFA)-ACN]; ACN%: 8%-38%, 10 min) to obtain the trifluoroacetate salt of compound 27-9. Calculated value of MS ESI C 27 H 35 N9Se[M+H] + 566, found 566. 11H NMR (400 MHz, DMSO-d6) δ 1.84 - 1.95 (m, 4H), 2.14 (m, 2H), 2.19 - 2.24 (m, 2H), 2.25 (m, 3H), 2.28 (m, 2H), 3.06 - 3.13 (m, 4H), 3.40 (m, 6H), 4.03 - 4.25 (m, 4H), 6.02 (s, 1H), 6.57 (s, 1H), 6.78 (s, 1H), 8.46 - 8.64 (m, 1H), 8.74 (m, 1H), 8.84 (m, 1H), 9.62 - 10.48 (m, 2H).

[0287] Example 28

[0288]

[0289] Step 1: At 25 °C, dissolve compound 18-1 (300 mg, 1.72 mmol) in acetonitrile (5 mL), add 6-1 (414.41 mg, 1.72 mmol) and N,N-diisopropylethylamine (445.70 mg, 3.45 mmol, 600.67 μL), stir at 0 °C for 1 hour. TLC shows that the raw materials are consumed. Add 100 mL of saturated ammonium chloride solution to the reaction solution, and extract with ethyl acetate (100 mL × 3). The combined organic phases are washed with saturated brine (100 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 4 / 1) to obtain compound 28-1.

[0290] Step 2: Dissolve compound 28-1 (0.6 g, 1.59 mmol) in dioxane (10 mL), add 1-4 (397.15 mg, 1.75 mmol), cesium carbonate (1.03 g, 3.18 mmol) and [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] palladium(II) mesylate mesylate (143.94 mg, 158.79 μmol). After purging with nitrogen three times, heat to 100 °C and stir under nitrogen protection for 4 hours. Add 50 mL of aqueous ammonium chloride solution to the reaction solution, and extract with ethyl acetate (100 mL × 3). The combined organic phases are washed with saturated brine (100 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 5 / 1) to obtain compound 28-2. MS ESI calculated value C 28 H 44 N8O3Si [M + H] +569, the measured value is 569. 1 1H NMR (400 MHz, MeOD) δ 1.49 - 1.51 (s, 9H), 1.58 - 1.61 (s, 9H), 1.62 - 1.65 (s, 2H), 1.88 - 2.11 (m, 8H), 2.38 - 2.42 (s, 3H), 2.46 - 2.54 (m, 4H), 4.24 - 4.28 (m, 2H), 4.39 - 4.53 (m, 1H), 5.67 (m, 1H), 6.35 - 6.44 (m, 1H), 6.65 - 6.71 (m, 1H), 7.49 - 7.57 (s, 1H).

[0291] Step 3: At 25 °C, dissolve compound 28 - 2 (500 mg, 879.07 μmol) in DMF (10 mL), successively add water (0.5 mL), triethylamine (363.50 mg, 3.59 mmol, 0.5 mL) and selenium powder (213.55 mg, 2.64 mmol), displace with carbon monoxide three times and then react at 90 °C under 15 psi pressure for 1.5 hours. Add 50 mL of ammonium chloride aqueous solution to the reaction solution, extract with dichloromethane (50 mL × 3), wash the combined organic phases with saturated brine (50 mL), and finally dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain crude product 28 - 3.

[0292] Step 4: At 25 °C, dissolve compound 28 - 3 (600 mg, 923.42 μmol) in ethanol (5 mL), add 6 - 5 (85.44 mg, 923.42 μmol), and stir at 80 °C for 1 hour. The reaction solution is directly concentrated under reduced pressure to obtain crude product 28 - 4. MS ESI calculated value C 31 H 48 N8O3SeSi [M + H] + 689, the measured value is 689.

[0293] Step 5: Dissolve compound 28 - 4 (600 mg, 872.34 μmol) in hydrochloric acid ethyl acetate solution (4 M, 5 mL), and stir at 25 °C for 2 hours. The reaction solution is directly filtered to obtain the filter cake as the crude hydrochloride salt of compound 28 - 5. MS ESI calculated value C 20 H 26 N8Se [M + H] + 459, the measured value is 459.

[0294] Step 6: The crude hydrochloride of 28-5 (100 mg, 202.47 μmol) was dissolved in methanol (2 mL), N,N-diisopropylethylamine (52.34 mg, 404.95 μmol, 70.53 μL) was added, and the mixture was stirred at 25 °C for 10 minutes, and then compound acrylonitrile (1-10) (10.74 mg, 202.47 μmol, 13.43 μL) was added and stirred at 25 °C for 16 hours. 50 mL of ammonium chloride aqueous solution was added to the reaction solution, and extracted with ethyl acetate (50 mL×3). The combined organic phase was washed with saturated brine (50 mL), and finally the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product was separated by high performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA)-ACN]; B (ACN) %: 17%-47%, 10 minutes) to obtain the trifluoroacetate salt of compound 28-6. MS ESI calculated value C 23 H 29 N9Se[M+H] + 512, measured value 512, 1 H NMR (400MHz, CD3OD) δ1.84-2.09(m, 4H), 2.11-2.29(m, 4H), 2.32-2.52(m, 5H), 2.56-2.60(s, 3H), 3.08-3.13(m, 2H), 3.77-3.82(m, 2H), 3.85-3.89(m, 2H), 5.08-5.21(m, 1H), 5.88-5.99(s, 1H), 6.78(s, 1H), 8.03-8.18(s, 1H).

[0295] Embodiment 29

[0296]

[0297] Step 1: Compound 29-1 (0.5 g, 4.50 mmol) was dissolved in DMF (10 mL) at 25 °C, and water (1 mL) and selenium powder (728.58 mg, 9.00 mmol) were added in sequence. After replacement with carbon monoxide three times, the mixture was reacted at 80 °C under a pressure of 50 psi for 16 hours. After the raw materials were consumed as monitored by TLC, 50 mL of aqueous ammonium chloride solution was added to the reaction solution, and extracted with dichloromethane (50 mL×3). The combined organic phase was washed with saturated brine (50 mL), and the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography (SiO2, petroleum ether: ethyl acetate = 1 / 0 to 1 / 1) to obtain compound 29-2. 11H NMR (400 MHz, DMSO-d6) δ 1.69 - 1.84 (m, 5H), 3.81 - 3.93 (m, 4H).

[0298] Step 2: Compound 29-2 (0.4 g, 1.49 mmol) was added to methanol (10 mL), and compound 8-4 (285.76 mg, 1.49 mmol) was added. The reaction was carried out at 20 °C for 1 h. TLC showed that the raw materials were completely consumed. 50 mL of ammonium chloride aqueous solution was added to the reaction solution, and it was extracted with ethyl acetate (100 mL × 3). The combined organic phases were washed with saturated brine (100 mL), and finally the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 5 / 1) to obtain compound 29-3. MS ESI calculated value C 13 H 12 Cl2N2OSe [M+H] + 363, found 363.

[0299] Step 3: Compound 29-3 (350 mg, 966.55 μmol), compound 1-2 (262.49 mg, 1.16 mmol), (±)-2,2-bis(diphenylphosphino)-1,1-binaphthalene (120.37 mg, 193.31 μmol), palladium acetate (21.70 mg, 96.65 μmol), and cesium carbonate (629.84 mg, 1.93 mmol) were added to dioxane (5 mL). The reaction was carried out at 80 °C for 3 h under a nitrogen atmosphere. 50 mL of ammonium chloride aqueous solution was added to the reaction solution, and it was extracted with ethyl acetate (100 mL × 3). The combined organic phases were washed with saturated brine (100 mL), and finally the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 2 / 1) to obtain compound 29-4. MS ESI calculated value: C 25 H 33 ClN4O3Se [M+H] + 553, found 553.

[0300] Step 4: Dissolve compound 29-4 (250 mg, 452.93 μmol) in dioxane (5 mL), add compound 8-9 (98.27 mg, 498.22 μmol), cesium carbonate (295.14 mg, 905.85 μmol), and palladium(II) mesylate of [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] (41.06 mg, 45.29 μmol). After purging with nitrogen three times, heat the mixture to 90 °C and stir for 4 hours under nitrogen protection. Add 50 mL of aqueous ammonium chloride solution to the reaction mixture, and extract with ethyl acetate (100 mL × 3). Wash the combined organic phases with saturated brine (100 mL), dry the organic phase over anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain the crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 10 / 1) to obtain compound 29-5. MS ESI calculated value for C 34 H 47 N7O5Se[M+H] + 714, found 714.

[0301] Step 5: Dissolve compound 29-5 (200 mg, 280.61 μmol) in hydrochloric acid ethyl acetate solution (4 M, 2 mL), and stir at 25 °C for 2 hours. Filter the reaction mixture directly to obtain the crude product of the hydrochloride salt of compound 29-6. MS ESI calculated value for C 24 H 31 N7OSe[M+H] + 514, found 514.

[0302] Step 6: Dissolve the crude hydrochloride salt of 29-6 (150 mg, 273.24 μmol) in methanol (1 mL), add N,N-diisopropylethylamine (70.63 mg, 546.48 μmol, 95.19 μL). Stir the mixture at 25 °C for 10 minutes, then add compound acrylonitrile (1-10) (29.00 mg, 546.48 μmol, 36.25 μL), and stir at 25 °C for 16 hours. Add 50 mL of aqueous ammonium chloride solution to the reaction mixture, and extract with ethyl acetate (50 mL × 3). Wash the combined organic phases with saturated brine (50 mL), dry the organic phase over anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain the crude product. The crude product is separated by preparative high performance liquid chromatography (column: Phenomenex Luna C18 75*30mm*3μm; mobile phase: [water (0.1% TFA)-ACN]; B (ACN)%: 20% - 40%, 7 minutes) to obtain the trifluoroacetate salt of compound 29-7. MS ESI calculated value for C 27 H34 N8OSe[M+H] + 567, the measured value is 567. 1 H NMR (400 MHz, CD3OD) δ 1.85 - 1.96 (m, 2H), 2.07 - 2.18 (m, 4H), 2.26 - 2.50 (m, 9H), 3.08 - 3.13 (t, 2H), 3.33 - 3.39 (m, 1H), 3.47 - 3.56 (t, 2H), 3.58 - 3.65 (t, 2H), 4.02 - 4.08 (d, 2H), 4.18 - 4.29 (m, 3H), 5.81 - 5.89 (s, 1H), 6.79 - 6.83 (s, 1H), 6.89 - 6.92 (s, 1H), 8.84 (s, 1H).

[0303] Example 30

[0304]

[0305] Step 1: At 0 °C, compound 30 - 1 (0.5 g, 8.76 mmol, 591.02 μL) was added to dichloromethane (5 mL), and then triethylamine (2.66 g, 26.27 mmol, 3.66 mL) and 30 - 2 (1.11 g, 10.51 mmol, 773.00 μL) were added successively. The reaction was carried out at 20 °C for 2 hours. TLC showed that the raw materials were completely consumed. 50 mL of ammonium chloride aqueous solution was added to the reaction solution, and it was extracted with ethyl acetate (100 mL × 3). The combined organic phases were washed with saturated brine (100 mL), and finally the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain compound 30 - 3.

[0306] Step 2: Selenium (1.58 g, 19.49 mmol) was added to ethanol (30 mL). Sodium borohydride (737.26 mg, 19.49 mmol) was added in portions at 0 - 5 °C. After addition, the reaction was carried out at 20 °C for 1 hour. 30 - 3 (0.8 g, 9.74 mmol) was added, and then pyridine hydrochloride (4.50 g, 38.98 mmol) was added slowly. The mixture was reacted at 80 °C for 1 hour. 50 mL of ammonium chloride aqueous solution was added to the reaction solution, and it was extracted with dichloromethane (50 mL × 3). The combined organic phases were washed with saturated brine (50 mL), and finally the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 1 to dichloromethane∶methanol = 10∶1) to obtain compound 30 - 4. 11H NMR (400 MHz, DMSO-d6) δ 1.96 - 2.21 (m, 2H), 3.73 - 4.07 (m, 4H), 7.26 - 7.84 (m, 2H).

[0307] Step 3: Compound 30-4 (0.45 g, 1.67 mmol) was added to methanol (2 mL), and compound 8-4 (272.89 mg, 1.67 mmol) and sodium fluoride (35.13 mg, 836.67 μmol) were added. The reaction was carried out at 20 °C for 0.5 h. 50 mL of ammonium chloride aqueous solution was added to the reaction solution, and it was extracted with ethyl acetate (100 mL × 3). The combined organic phases were washed with saturated brine (100 mL), and finally the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 5 / 1) to obtain compound 30-5. MS ESI calculated value C 11 H9Cl2N3Se [M+H] + 334, found 334.

[0308] Step 4: Compound 30-5 (600 mg, 1.80 mmol), compound 1-2 (407.68 mg, 1.80 mmol), (±)-2,2-bis(diphenylphosphino)-1,1-binaphthalene (224.34 mg, 360.28 μmol), palladium acetate (40.44 mg, 180.14 μmol), and cesium carbonate (1.17 g, 3.60 mmol) were added to dioxane (4 mL). The reaction was carried out at 80 °C for 3 h under a nitrogen atmosphere. 50 mL of ammonium chloride aqueous solution was added to the reaction solution, and it was extracted with ethyl acetate (100 mL × 3). The combined organic phases were washed with saturated brine (100 mL), and finally the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 5 / 1) to obtain compound 30-6. MS ESI calculated value C 23 H 30 ClN5O2Se [M+H] + 524, found 524.

[0309] Step 5: Dissolve compound 30-6 (400 mg, 764.92 μmol) in dioxane (4 mL), add compound 8-9 (165.96 mg, 841.42 μmol), cesium carbonate (498.45 mg, 1.53 mmol) and palladium(II) mesylate of [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] (69.34 mg, 76.49 μmol). After displacing nitrogen three times, heat the mixture to 100 °C and stir for 4 hours under nitrogen protection. Add 50 mL of ammonium chloride aqueous solution to the reaction mixture, and extract with ethyl acetate (100 mL × 3). Wash the combined organic phases with saturated brine (100 mL), dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 5 / 1 - 1 / 1) to obtain compound 30-7. MS ESI calculated value: C 32 H 44 N8O4Se[M+H] + 685, found 685.

[0310] Step 6: Dissolve compound 30-7 (350 mg, 397.31 μmol) in hydrochloric acid ethyl acetate solution (4 M, 4 mL), and stir at 25 °C for 16 hours. The reaction mixture is directly concentrated under reduced pressure to obtain the crude hydrochloride salt of compound 30-8. MS ESI calculated value C 22 H 28 N8Se[M+H] + 485, found 485.

[0311] Step 7: Dissolve the crude hydrochloride salt of 30-8 (250 mg, 480.83 μmol) in methanol (5 mL), add N,N-diisopropylethylamine (186.43 mg, 1.44 mmol, 251.26 μL). Stir the mixture at 25 °C for 10 minutes, then add compound acrylonitrile (1-10) (51.03 mg, 961.66 μmol, 63.79 μL), and stir at 25 °C for 16 hours. Add 50 mL of ammonium chloride aqueous solution to the reaction mixture, and extract with ethyl acetate (50 mL × 3). Wash the combined organic phases with saturated brine (50 mL), dry the organic phase with anhydrous sodium sulfate, filter and concentrate under reduced pressure to obtain a crude product. The crude product is separated by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150*25mm*10μm; mobile phase: [water (0.1% TFA)-ACN]; B(ACN)%: 15% - 45%, 10 minutes) to obtain the trifluoroacetate salt of compound 30-9. MS ESI calculated value C 25H 31 N9Se[M+H] + 538, the measured value is 538, 1 H NMR (400 MHz, CD3OD) δ 2.08 - 2.17 (s, 2H), 2.30 - 2.48 (m, 9H), 2.50 - 2.58 (m, 2H), 3.09 - 3.13 (s, 2H), 3.46 - 3.56 (m, 2H), 4.14 - 4.26 (m, 7H), 5.84 (s, 1H), 6.66 (s, 1H), 6.71 (s, 1H), 7.96 (s, 1H).

[0312] Example 31

[0313]

[0314] Step 1: At 0 °C, dissolve compound 18-1 (360 mg, 2.07 mmol) in acetonitrile (5 mL), add 6-1 (497.3 mg, 2.07 mmol) and N,N-diisopropylethylamine (534.8 mg, 4.14 mmol, 720.80 μL), stir at 0 °C for 1 hour, add 100 mL of saturated ammonium chloride solution to the reaction solution, and extract with ethyl acetate (100 mL × 3). The combined organic phases are washed with saturated brine (100 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 2 / 1) to obtain compound 31-1.

[0315] Step 2: At 0 °C, dissolve compound 33-1 (0.7 g, 1.85 mmol) in tetrahydrofuran (10 mL), add sodium hydride (110 mg, 2.75 mmol, 60% purity), stir at 0 °C for 0.5 hour, then slowly add methyl iodide (525.8 mg, 3.71 mmol, 230.6 μL) to the system, and then stir the whole system at 35 °C for 3 hours. Add 100 mL of saturated ammonium chloride solution to the reaction solution, and extract with ethyl acetate (100 mL × 3). The combined organic phases are washed with saturated brine (100 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 1 / 1) to obtain compound 31-2. 11H NMR (400 MHz, CDCl3) δ 0.76 - 0.84 (m, 2H), 1.14 - 1.25 (m, 2H), 1.41 - 1.45 (s, 9H), 1.56 - 1.90 (m, 6H), 2.68 - 2.84 (s, 3H), 4.25 - 4.35 (m, 1H), 4.37 - 4.47 (m, 1H), 5.75 - 6.07 (m, 1H), 6.46 - 6.69 (m, 1H).

[0316] Step 3: Dissolve compound 31 - 2 (600 mg, 1.53 mmol) in dioxane (5 mL), add compound 1 - 4 (382.9 mg, 1.68 mmol), cesium carbonate (997.6 mg, 3.06 mmol) and [(2 - di - cyclohexylphosphino - 3,6 - dimethoxy - 2′,4′,6′ - triisopropyl - 1,1′ - biphenyl) - 2 - (2′ - amino - 1,1′ - biphenyl)] palladium(II) mesylate mesylate (138.8 mg, 153.10 μmol). After displacing with nitrogen three times, heat the mixture to 100 °C and stir for 4 hours under nitrogen protection. Add 50 mL of ammonium chloride aqueous solution to the reaction solution, and extract with ethyl acetate (100 mL × 3). The combined organic phases are washed with saturated brine (100 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product is purified by column chromatography (SiO2, petroleum ether∶ethyl acetate = 1 / 0 - 1 / 1) to obtain compound 31 - 3. MS ESI calculated value C 29 H 46 N8O3Si [M + H] + 583, found 583.

[0317] Step 4: At 25 °C, dissolve compound 31 - 3 (250 mg, 428.9 μmol) in DMF (5 mL), successively add water (0.3 mL), triethylamine (218.1 mg, 2.16 mmol, 0.3 mL) and selenium powder (104.2 mg, 1.29 mmol). After displacing with carbon monoxide three times, react at 90 °C under 15 psi pressure for 2 hours. Add 50 mL of ammonium chloride aqueous solution to the reaction solution, and extract with dichloromethane (50 mL × 3). The combined organic phases are washed with saturated brine (50 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain crude product 31 - 4.

[0318] Step 5: At 25 °C, dissolve compound 31 - 4 (300 mg, 451.95 μmol) in ethanol (5 mL), add 6 - 5 (50.2 mg, 542.34 μmol), and stir at 80 °C for 3 hours. The reaction solution is directly concentrated under reduced pressure to obtain crude product 31 - 5. MS ESI calculated value C32 H 50 N8O3SeSi[M+H] + 703, measured value 703.

[0319] Step 6: Dissolve compound 31-5 (0.3 g, 427.45 μmol) in hydrochloric acid methanol solution (4 M, 4 mL), and stir at 30 °C for 1 hour. LCMS shows that the raw material has basically reacted completely, and the main peak is the product peak. The reaction solution is directly concentrated under reduced pressure to obtain the crude hydrochloride of compound 31-6. MS ESI calculated value C 21 H 28 N8Se[M+H] + 473, measured value 473.

[0320] Step 7: Dissolve the crude hydrochloride of 31-6 (200 mg, 393.76 μmol) in methanol (2 mL), add N,N-diisopropylethylamine (152.7 mg, 1.18 mmol, 205.8 μL), stir the mixture at 25 °C for 10 minutes, then add acrylonitrile (1-10) (41.8 mg, 787.52 μmol, 52.2 μL), and stir at 25 °C for 16 hours. Add 50 mL of ammonium chloride aqueous solution to the reaction solution, and extract with ethyl acetate (50 mL × 3). The combined organic phases are washed with saturated brine (50 mL), and finally the organic phase is dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain the crude product. The crude product is separated by preparative high performance liquid chromatography (column: Phenomenex Luna C18 75*30mm*3μm; mobile phase: [water (0.1% TFA)-ACN]; B(ACN)%: 15%-45%, 7 minutes) to obtain the trifluoroacetate of compound 31-7. MS ESI calculated value C 24 H 31 N9Se[M+H] + 526, measured value 526, 1 H NMR (400 MHz, CD3OD) δ 1.87-2.15 (m, 6H), 2.26-2.46 (m, 5H), 2.46-2.62 (m, 5H), 3.10-3.15 (t, 2H), 3.16-3.23 (s, 3H), 3.78-3.84 (t, 2H), 3.87-3.94 (m, 2H), 5.97-6.01 (s, 1H), 6.02-6.21 (m, 1H), 6.99-7.10 (s, 1H), 8.12 (s, 1H).

[0321] Example 32

[0322]

[0323] Step 1: At 20 °C, compound 18-1 (0.6 g, 3.45 mmol) was dissolved in tetrahydrofuran (15 mL), and diisopropylethylamine (891.37 mg, 6.90 mmol) was added. Then, 1-2 (780.45 mg, 3.45 mmol) was added dropwise at 0 °C, and the reaction was carried out at 0 °C for 2 hours and then at 20 °C for 16 hours. The reaction solution was diluted with 15 mL of water and extracted with 120 mL (40 mL × 3) of ethyl acetate. The combined organic layers were washed with 40 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. It was purified by silica gel column (petroleum ether∶ethyl acetate = 10 / 1 to 1 / 1) to obtain compound 32-2. MS ESI calculated value: C 17 H 22 ClN5O2[M+H] + 364, found 364.

[0324] Step 2: Compound 32-2 (740 mg, 1.98 mmol) was dissolved in DMF (15 mL), and sodium hydride (103.00 mg, 2.58 mmol) was added portionwise at 0 °C. After addition, the reaction was carried out at this temperature for 30 minutes, and then iodomethane (309.29 mg, 2.18 mmol) was added dropwise at 0 °C, and the reaction was carried out at 15 °C for 2 hours. 10 mL of saturated ammonium chloride aqueous solution was added, then 50 mL of water was added, and it was extracted with 40 mL (20 mL × 2) of ethyl acetate. The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. It was triturated with petroleum ether:ethyl acetate = 10 mL:10 mL at room temperature to obtain compound 32-3. MS ESI calculated value: C 18 H 24 ClN5O2[M+H] + 378, found 378.

[0325] Step 3: Compound 32-3 (0.6 g, 1.59 mmol) and 8-9 (344.50 mg, 1.75 mmol) were added to dioxane (15 mL), and [(2-di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] palladium(II) mesylate (143.94 mg, 158.79 μmol) and cesium carbonate (1.03 g, 3.18 mmol) were added. The reaction was carried out at 80 °C under a nitrogen atmosphere for 3 hours. The reaction solution was diluted with 50 mL of water and extracted with 40 mL (20 mL × 2) of ethyl acetate. The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. It was purified by silica gel column (petroleum ether / ethyl acetate = 5 / 1 to 1 / 1) to obtain compound 32-4. MS ESI calculated value: C 27 H 38N8O4[M+H] + 539, the measured value is 539.

[0326] Step 4: DMF (5 mL) and water (0.5 mL) were added to a single-necked flask, followed by compound 32-4 (350 mg, 630.29 μmol), selenium powder (153.12 mg, 1.89 mmol), and triethylamine (352.59 mg, 3.48 mmol). The mixture was purged with carbon monoxide three times and then reacted at 90 °C for 1 hour under a carbon monoxide atmosphere (15 psi). The reaction mixture was diluted with 50 mL of water and extracted with 120 mL (40 mL × 3) of ethyl acetate. The combined organic layers were washed with 50 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure to obtain compound 32-5. MS ESI calculated value: C 27 H 40 N8O4Se[M+H] + 621, the measured value is 621.

[0327] Step 5: Compound 32-5 (400 mg, 645.56 μmol) was dissolved in ethanol (10 mL), and chloroacetone (0.26 g, 2.81 mmol) was added. The mixture was reacted at 80 °C for 2 hours. The reaction mixture was concentrated to dryness, and then Boc anhydride (169.07 mg, 774.67 μmol), triethylamine (195.97 mg, 1.94 mmol), and tetrahydrofuran (6 mL) were added. The mixture was heated to 50 °C and reacted for 3 hours, and then concentrated to dryness under reduced pressure. It was purified by silica gel plate (petroleum ether∶ethyl acetate = 1∶2) to obtain compound 32-6. MS ESI calculated value: C 30 H 42 N8O4Se[M+H] + 659, the measured value is 659.

[0328] Step 6: Compound 32-6 (50 mg, 76.03 μmol) was added to a single-necked flask, and hydrochloric acid ethyl acetate (4 M, 3 mL) was added. The mixture was reacted at 50 °C for 1 hour. The reaction mixture was concentrated under reduced pressure to obtain compound 32-7 hydrochloride. MS ESI calculated value: C 20 H 26 N8Se[M+H]+459, the measured value is 459.

[0329] Step 7: Compound 32-7 hydrochloride (50 mg, 101.24 μmol) was dissolved in methanol (2 mL), and diisopropylethylamine (65.42 mg, 506.20 μmol) and acrylonitrile (1-10) (10.74 mg, 202.48 μmol) were added. The reaction was carried out at 15 °C for 2 hours. 30 mL of water was added to the reaction solution, and the mixture was extracted with ethyl acetate (40 mL, 20 mL×2). The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by high performance liquid chromatography (column model: Phenomenex Luna C18 75×30 mm×3 μm; mobile phase: [water (0.1% trifluoroacetic acid)-acetonitrile]; acetonitrile%: 15%-45%, 7 minutes) to obtain the trifluoroacetate of compound 32-8. MS ESI calculated value: C 23 H 29 N9Se[M+H] + 512, found 512. 1 H NMR (400 MHz, CD3OD) δ 2.01 (d, J = 12.0 Hz, 2H), 2.26 - 2.37 (m, 5H), 2.37 - 2.51 (m, 4H), 2.58 (s, 3H), 3.13 (t, J = 7.2 Hz, 2H), 3.18 (s, 3H), 3.49 (t, J = 6.8 Hz, 2H), 4.27 (s, 2H), 5.24 - 5.81 (m, 1H), 6.03 (s, 1H), 7.04 (s, 1H), 8.09 (s, 1H).

[0330] Example 33

[0331]

[0332] Step 1: Hydrochloride of 32-7 (60 mg, 104.99 μmol) was dissolved in N-methylpyrrolidone (1 mL), and triethylamine (53.1 mg, 524.9 μmol, 73.0 μL) was added. The mixture was stirred at 20 °C for 30 minutes, then the reaction system was cooled to 0 °C, and compound 3-1 (20.86 mg, 115.49 μmol) was added. The mixture was stirred at 20 °C for 2 hours. 50 mL of ammonium chloride aqueous solution was added to the reaction solution, and the mixture was extracted with ethyl acetate (50 mL×3). The combined organic phases were washed with saturated brine (50 mL), and finally the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain the crude product. The crude product was separated by preparative high performance liquid chromatography (column: Phenomenex Luna C18 75*30 mm*3 μm; mobile phase: [water (0.1% TFA)-ACN]; B (ACN)%: 30%-60%, 7 minutes) to obtain the trifluoroacetate of compound 33-1. MS ESI calculated value C 24H 30 N 10 O2SSe[M+H] + 603, the measured value is 603, 1 H NMR (400 MHz, CD3OD) δ 1.75 - 1.86 (m, 2H), 1.96 - 2.10 (m, 2H), 2.11 - 2.25 (m, 4H), 2.36 (s, 3H), 2.61 (s, 3H), 3.15 - 3.24 (s, 3H), 3.64 - 3.74 (m, 1H), 4.03 - 4.11 (t, 2H), 4.15 - 4.24 (t, 2H), 4.28 - 4.38 (m, 2H), 5.45 - 5.70 (m, 1H), 5.87 - 6.04 (s, 1H), 6.92 - 7.12 (s, 1H), 8.15 (s, 1H).

[0333] Example 34

[0334]

[0335] Step 1: At 20 °C, diisopropylethylamine (6.24 g, 48.31 mmol) and compound 20 - 1 (5 g, 24.15 mmol) were added to tetrahydrofuran (80 mL). Compound 1 - 2 (5.47 g, 24.15 mmol) was dissolved in tetrahydrofuran (20 mL) and added dropwise to the former system at 0 °C. The reaction was carried out at 20 °C for 16 hours. The reaction solution was diluted with 200 mL of water and extracted with 200 mL of ethyl acetate (100 mL × 2). The combined organic layers were washed with 100 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure. Purification by silica gel column (petroleum ether / ethyl acetate = 10 / 1 to 1 / 1) gave compound 34 - 1. MS ESI calculated value: C 18 H 25 ClN4O4[M+H] + 397, the measured value is 397.

[0336] Step 2: Compound 34 - 1 (5.23 g, 13.18 mmol) was dissolved in tetrahydrofuran (80 mL). Sodium hydride (632.49 mg, 15.81 mmol) was added portionwise at 0 °C. After addition, the reaction was carried out at 0 °C for 30 minutes. Iodomethane (2.06 g, 14.50 mmol) was dissolved in tetrahydrofuran (20 mL) and added dropwise to the above reaction system at 0 °C, and then the reaction was carried out at 15 °C for 16 hours. The reaction solution was quenched with 20 mL of ammonium chloride solution, diluted with 150 mL of water, and extracted with 160 mL of ethyl acetate (80 mL × 2). The combined organic layers were washed with 80 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure. Purification by silica gel column (petroleum ether / ethyl acetate = 4 / 1 to 3 / 1) gave compound 34 - 2. MS ESI calculated value: C19 H 27 ClN4O4[M+H] + 411, measured value 411.

[0337] Step 3: Compound 34-2 (2.4 g, 5.84 mmol) was dissolved in tetrahydrofuran (50 mL), and a solution of methylmagnesium bromide in diethyl ether (3 M, 3.11 mL) was added dropwise at 0 °C. The reaction was carried out at -78 °C for 1 hour. The reaction was quenched by adding 5 mL of saturated ammonium chloride solution. The reaction mixture was diluted with 60 mL of water and extracted with 60 mL of ethyl acetate (30 mL × 2). The combined organic layers were washed with 30 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. It was purified by silica gel column (petroleum ether / ethyl acetate = 3:1) to obtain Compound 34-3. 1 HNMR (400 MHz, CDCl3) δ 1.50 (s, 9H), 1.61 (dd, J = 12.4, 3.2 Hz, 2H), 1.85 (d, J = 6.8 Hz, 4H), 2.05 (d, J = 6.4 Hz, 2H), 2.62 (s, 3H), 2.86 (s, 3H), 4.32 (s, 2H), 5.30 (s, 1H), 6.92 (s, 1H)

[0338] Step 4: Compound 34-3 (800 mg, 2.03 mmol) was dissolved in dichloromethane (20 mL), triethylamine (1.23 g, 12.16 mmol) was added, and then tert-butyldimethylsilyl trifluoromethanesulfonate (1.61 g, 6.08 mmol) was added dropwise. The reaction was carried out at 15 °C for 16 hours. The reaction mixture was diluted with 60 mL of dichloromethane and washed with 60 mL of saturated aqueous sodium bicarbonate solution (30 mL × 2), 20 mL of water, and 30 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure to obtain Compound 34-4. MS ESI calculated value: C 25 H 41 ClN4O3Si[M+H] + 509, measured value 509.

[0339] Step 5: Compound 34-4 (1.15 g, 2.03 mmol) was dissolved in tetrahydrofuran (30 mL) and water (5 mL), and N-bromosuccinimide (721.57 mg, 4.05 mmol) was added. The reaction was carried out at 15 °C for 2 hours. The reaction mixture was diluted with 100 mL of water and extracted with 100 mL of ethyl acetate (50 mL × 2). The combined organic layers were washed with 50 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure to obtain Compound 34-5. MS ESI calculated value: C 19 H 26 BrClN4O3[M+H] + 473, measured value 473.

[0340] Step 6: Compound 34-5 (800 mg, 2.14 mmol) was dissolved in methanol (25 mL). Methylselenamide (12-2) (261.25 mg, 2.14 mmol) was added at 0-5 °C, and the reaction was carried out at 15 °C for 2 hours. Then, BOC anhydride (560.70 mg, 2.57 mmol), 2,6-dimethylpyridine (26.16 mg, 214.09 μmol), and triethylamine (866.55 mg, 8.56 mmol) were added, and the reaction was carried out at 15 °C for 2 hours. The reaction solution was diluted with 100 mL of water and extracted with 90 mL of ethyl acetate (30 mL×3). The combined organic layers were washed with 50 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. It was purified by silica gel column (petroleum ether / ethyl acetate = 5 / 1 to 3 / 1) to obtain Compound 34-6. MS ESI calculated value: C 21 H 28 ClN5O2Se [M+H] + 498, found 498.

[0341] Step 7: Compound 34-6 (120 mg, 241.50 μmol) and tert-butyl 5-amino-3-methyl-1H-pyrazole-1-carboxylate (57.16 mg, 289.80 μmol) were added to dioxane (4 mL). [(2-Di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl)-2-(2′-amino-1,1′-biphenyl)] palladium(II) methanesulfonate (21.89 mg, 24.15 μmol) and cesium carbonate (157.37 mg, 483.00 μmol) were added, and the reaction was carried out at 80 °C under a nitrogen atmosphere for 3 hours. The reaction solution was diluted with 50 mL of water and extracted with 40 mL of ethyl acetate (20 mL×2). The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. It was purified by silica gel column (petroleum ether / ethyl acetate = 5 / 1 to 1 / 1) to obtain Compound 34-7. MS ESI calculated value: C 30 H 42 N8O4Se [M+H] + 659, found 659.

[0342] Step 8: Compound 34-7 (60 mg, 91.23 μmol) was added to hydrochloric acid in methanol (4 M, 3 mL), and the reaction was carried out at 40 °C for 1 hour. The reaction solution was concentrated under reduced pressure to obtain the hydrochloride salt of Compound 34-8. MS ESI calculated value: C 20 H 26 N8Se [M+H] + 459, found 459.

[0343] Step 9: The hydrochloride salt of compound 34-8 (55 mg, 120.24 μmol) was dissolved in methanol (2 mL), and diisopropylethylamine (62.16 mg, 480.94 μmol) and acrylonitrile (1-10) (12.76 mg, 240.47 μmol) were added. The reaction was carried out at 15 °C for 2 hours. 30 mL of water was added to the reaction solution, and it was extracted with 40 mL (20 mL × 2) of ethyl acetate. The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by high performance liquid chromatography (column model: 3-100C18 ultra 150*50mm*3μm; mobile phase: [water (0.225% formic acid)-acetonitrile]; acetonitrile %: 5%-35%, 10 minutes) to obtain the formate salt of compound 34-9. MS ESI calculated value: C 23 H 29 N9Se[M+H]+512, found 512. 1 HNMR (400 MHz, CD3OD) δ 1.48-1.65 (m, 2H), 1.83-1.91 (m, 2H), 1.94-2.03 (m, 2H), 2.05-2.14 (m, 2H), 2.29 (s, 3H), 2.61-2.69 (m, 2H), 2.72-2.77 (m, 2H), 2.81 (s, 3H), 2.99 (s, 3H), 3.46 (s, 2H), 5.00-5.60 (m, 1H), 6.22 (s, 1H), 6.81 (s, 1H), 8.81 (s, 1H).

[0344] Example 35

[0345]

[0346] Step 1: The hydrochloride salt of compound 34-8 (110 mg, 222.72 μmol) was dissolved in tetrahydrofuran (2 mL), and triethylamine (112.68 mg, 1.11 mmol) and compound 3-1 (40.23 mg, 222.72 μmol) were added. The reaction was carried out at 15 °C for 1 hour. 30 mL of water was added to the reaction solution, and it was extracted with 40 mL (20 mL × 2) of ethyl acetate. The combined organic layers were washed with 20 mL of brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and purified by high performance liquid chromatography (column model: Phenomenex luna C18 150*25mm*10μm; mobile phase: [water (0.225% formic acid)-acetonitrile]; acetonitrile %: 16%-46%, 10 minutes) to obtain the formate salt of compound 35-1. MS ESI calculated value: C 24 H 30 N 10 O2SSe[M+H] +603, measured value 603. 1 HNMR (400 MHz, CD3OD) δ 1.68 (d, J = 10.0 Hz, 2H), 1.93 (d, J = 7.6 Hz, 2H), 1.99 - 2.15 (m, 4H), 2.28 (s, 3H), 2.80 (s, 3H), 2.94 (s, 3H), 3.65 (s, 1H), 4.01 (t, J = 7.2 Hz, 2H), 4.09 - 4.17 (m, 2H), 4.22 (s, 2H), 5.18 - 5.66 (m, 1H), 6.14 (s, 1H), 6.77 (s, 1H), 8.82 (s, 1H).

[0347] Example 36

[0348]

[0349] Step 1: At 20 °C, dissolve compound 5-1 (7.69 mg, 40.62 μmol) in DMF (0.3 mL), add 1-hydroxybenzotriazole (10.98 mg, 81.24 μmol) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (15.57 mg, 81.24 μmol), and stir the mixture at 20 °C for 0.5 h. Add a solution of the hydrochloride salt of compound 18-6 (30 mg, 44.68 μmol) and diisopropylethylamine (15.75 mg, 121.85 μmol) in dimethylformamide (0.3 mL) to the mixture and stir at 20 °C for 16 h. LC-MS shows that 33% of the starting material remains. Add 5-1 (3.84 mg, 20.31 μmol) to the reaction solution and stir at 20 °C for another 2 h. Add 10 mL of water to the reaction solution and extract with ethyl acetate (20 mL × 3). Wash the combined organic phases with saturated brine (5 mL × 3). Finally, dry the organic phase over anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain crude product 36-1. MS ESI calculated value: C 27 H 37 N9O3Se [M+H] + 616, measured value 616.

[0350] Step 2: At 20 °C, dissolve compound 36-1 (60 mg, 97.62 μmol) in dichloromethane (3 mL), add trifluoroacetic acid (1 mL), and stir the mixture at 20 °C for 1 hour. LC-MS showed that the raw material was consumed completely and the main peak was the product peak. Concentrate the reaction solution by rotary evaporation to obtain the crude product. The crude product was separated by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150*25 mm*10 μm; mobile phase: [water (0.1% TFA)-ACN]; ACN%: 13%-43%, 10 minutes) to obtain the trifluoroacetate of compound 36-2. MS ESI calculated value: C 22 H 29 N9OSe[M+H] + 516, found 516. 1 H NMR (400 MHz, DMSO-d6) δ 1.36-1.52 (m, 2H), 1.78-1.88 (m, 2H), 1.91 (m, 2H), 2.07 (m, 2H), 2.21 (s, 3H), 2.30 (s, 3H), 2.41 (s, 3H), 3.16-3.27 (m, 2H), 3.39 m, 1H), 4.31 (m, 1H), 4.53 (m, 2H), 6.32-6.49 (m, 1H), 6.51 (s, 1H), 7.24 (m, 1H), 7.86 (s, 1H), 8.88-9.18 (m, 1H), 11.59-12.04 (m, 1H).

[0351] Biological test

[0352] Experimental example 1:

[0353] Main reagents and consumables

[0354]

[0355] Experimental method

[0356] In this test, JAK1, 2, 3 and TYK2 were used Ultra method for activity detection. In the detection plate, mix the enzyme, ULight-labeled polypeptide substrate, ATP and the test compound, and incubate the reaction. After the reaction, add EDTA to terminate the reaction and simultaneously add Eu-labeled antibody. In Ultra kinase detection, the Eu-labeled anti-phosphorylated matrix antibody binds to the phosphorylated ULight-labeled matrix, which can bring the donor and acceptor molecules closer. After irradiation with light of 320 nm wavelength, the kinase reacts, and the energy of the Eu donor will be transferred to the ULight acceptor dye, and light with a wavelength of 665 nm is generated. The emission intensity of the light is proportional to the phosphorylation level of the ULight matrix.

[0357] Final test concentration of the compound: The final test concentration of the test compound ranged from 1 μM to 0.017 nM, with a 3-fold serial dilution and 11 concentrations. The content of DMSO in the detection reaction was 1%.

[0358] Kinase assay: Preparation of the buffer. The buffer included: 50 mM HEPES (pH 7.5), 0.01% Brij-35, 10 mM MgCl2, 1 mM EDTA, 1 mM DTT.

[0359] JAK1 enzyme reaction:

[0360] In the buffer, 2 nM JAK1 and 50 nM substrate were pre-incubated with different concentrations of the pre-diluted compound for 15 minutes. The reaction was started by adding 38 μM ATP and incubated at room temperature for 90 minutes. After the reaction, an antibody was added for detection, incubated at room temperature for 60 minutes, and then detected by Evnvision to collect data.

[0361] JAK2 enzyme reaction:

[0362] In the buffer, 0.03 nM JAK2 and 50 nM substrate were pre-incubated with different concentrations of the pre-diluted compound for 15 minutes. The reaction was started by adding 12 μM ATP and incubated at room temperature for 90 minutes. After the reaction, an antibody was added for detection, incubated at room temperature for 60 minutes, and then detected by Evnvision to collect data.

[0363] Data analysis:

[0364] According to % inhibition vs. log[compound concentration], data analysis and graph fitting were performed using XLfit5 software mode205 to obtain the IC 50 data, and the results are summarized in Table 1.

[0365] Table 1: Summary of the kinase activity of the test compound

[0366]

[0367]

[0368] Conclusion: The test compound of the present invention showed good inhibitory effects on two kinase subtypes, JAK1 and JAK2, in the in vitro activity tests.

[0369] Experimental Example 2

[0370] Main reagents and instruments

[0371] Main reagent consumables

[0372]

[0373]

[0374] Instrument

[0375] Flow cytometer: Brand: BD; Model: Fortessa

[0376] Reagent preparation:

[0377] Complete culture medium: RPMI 1640 medium + 10% fetal bovine serum + 1% penicillin / streptomycin (percentages are all by volume)

[0378] Experimental procedure

[0379] a) Drug treatment and induction

[0380] (1) Discard the culture medium of HT29 cells, add 10 mM ethylenediaminetetraacetic acid, and incubate at 37 °C for 5 min to digest the cells.

[0381] (2) Centrifuge the HT29 cells and THP1 cells together at 320 g for 3 min.

[0382] (3) Count the cells, and then adjust the cell concentration to 7.5×10 5 / mL with complete culture medium; inoculate 200 μL / well into a 96-well round bottom plate.

[0383] (4) Add drugs to the cells at final concentrations of 5000.00 nM, 1000.00 nM, 200.00 nM, 40.00 nM, 8.00 nM, 1.60 nM, 0.32 nM, 0.06 nM. Incubate at 37 °C for 30 min (the negative and positive control groups do not add drugs, but add the same concentration of dimethyl sulfoxide).

[0384] (5) Add IL-13 (final concentration 6 ng / mL) to the HT29 cells and incubate at 37 °C for 30 min; add IL-6 (final concentration 30 ng / mL) to the THP1 cells and incubate at 37 °C for 15 min (the negative control group is not stimulated with cytokines, and the positive control group is added with the same concentration of cytokines).

[0385] b) Cell staining and flow cytometry detection

[0386] (1) Centrifuge the above HT29 and THP1 cells at 320 g for 3 min.

[0387] (2) Wash the cells twice with staining buffer.

[0388] (3) Add 100 μL of cell fixative to each well and fix at 4 °C for 15 min.

[0389] (4) Wash the cells twice with the staining buffer.

[0390] (5) Add 100 μL of cell permeabilization solution to each well and permeabilize the cells at 4 °C for 30 min.

[0391] (6) Add the pSTAT3 antibody or pSTAT6 antibody to the staining buffer at a ratio of 2:48, and add 50 μL / well to THP1 or HT29 cells respectively, and stain at 4 °C for 30 min.

[0392] (7) Wash the cells twice with the staining buffer.

[0393] (8) Resuspend the cells with 150 μL of staining buffer and detect the mean fluorescence intensity (MFI) of the PE channel (pSTAT) using a flow cytometer.

[0394] c) Data processing

[0395] (1) The MFI corresponding to the sample is obtained by flow cytometer detection.

[0396] (2) Response rate (Response%) = 100 × (MFI of sample - MFI of negative control group) / (MFI of positive control group - MFI of negative control group)

[0397] (3) Substitute the response rate into the graphpad prism8 software and fit the curve using the log(inhibitor) vs. response--Variable slope (four parameters) method to obtain the half inhibitory concentration (IC 50 ).

[0398] The results of the JAK inhibitory activity test of THP1 and HT29 cells are shown in Table 2.

[0399] Table 2: Summary of the cell activities of the test compounds

[0400]

[0401]

[0402] Conclusion: The test compounds of the present invention showed good inhibitory effects in the in vitro activity test of the cell (THP1 and HT29) function experiment.

[0403] Experimental example 3

[0404] Test animals:

[0405] SD rats, 2 rats per group, male, Beijing Vital River Laboratory Animal Technology Co., Ltd.

[0406] Drug administration:

[0407] Two male SD rats were used at each time point, and there were 4 time points in total, with 8 rats in total. After fasting overnight, the rats were administered the drug p.o. at a dose of 5 mg / kg and a volume of 5 mL / kg.

[0408] Sample collection:

[0409] After the rats were administered the drug, they were sacrificed by CO2 at 1, 3, 6, and 12 hours. Approximately 200 μL of jugular vein blood was collected and placed in an EDTA-K2 tube. The plasma was separated by centrifugation at 3,200 g for 10 min at 4℃ and stored at -70 ± 10℃. At each time point, the small intestine and colon were taken. After squeezing out the contents, the colon and small intestine were rinsed with normal saline and then weighed and homogenized (the homogenate was methanol∶15 mM PBS = 1∶2). The homogenization ratio was 1∶4 (1 g of tissue with 4 mL of homogenate), and it was stored at -70 ± 10℃.

[0410] Sample treatment:

[0411] Plasma sample treatment:

[0412] Protein precipitation: 200 μL of acetonitrile containing internal standard was added to 20 μL of plasma sample for precipitation. After mixing, it was centrifuged at 12,000 g at 4℃. 50 μL of the supernatant after treatment was taken and added to a 96-well plate. After centrifugation at 3,220 g at 4℃, the supernatant was directly subjected to LC-MS / MS analysis.

[0413] Small intestine homogenate sample treatment:

[0414] 400 μL of acetonitrile containing internal standard was added to 40 μL of small intestine homogenate sample for precipitation. After mixing, it was centrifuged at 12,000 g at 4℃. 50 μL of the supernatant after treatment was taken and added to a 96-well plate. After centrifugation at 3220 g at 4℃, the supernatant was directly subjected to LC-MS / MS analysis.

[0415] Colon homogenate sample treatment:

[0416] 400 μL of acetonitrile containing internal standard was added to 40 μL of colon homogenate sample for precipitation. After mixing, it was centrifuged at 12,000 g at 4℃. 50 μL of the supernatant after treatment was taken and added to a 96-well plate. After centrifugation at 3,220 g at 4℃, the supernatant was directly subjected to LC-MS / MS analysis.

[0417] The experimental results are shown in the following table:

[0418]

[0419]

[0420] Conclusion: The compound of the present invention shows good drug exposure levels in the small intestine and colon of rats, and the ratios of small intestine / plasma and colon / plasma of the compound are relatively high, showing good tissue selectivity.

[0421] Experimental Example 4

[0422] Test animals:

[0423] C57BL / 6J mice, 2 mice per group, male, Lingchang Biotechnology Co., Ltd.

[0424] Drug administration:

[0425] 2 male C57BL / 6J mice at each time point, 4 time points, a total of 8 mice; after fasting overnight, they were administered p.o. at a dose of 3 mg / kg and a volume of 3 mL / kg.

[0426] Sample collection:

[0427] After the mice were administered the drug, they were sacrificed by CO2 at 0.5, 1, 5, and 12 hours. Approximately 30 μL of blood was collected by saphenous vein or cardiac puncture and placed in an EDTA-K2 tube. After centrifugation at 3,200 g for 10 min at 4°C, the plasma was obtained and stored at -60°C or below; at each time point, the small intestine and colon were taken. After squeezing out the contents, the colon and small intestine were rinsed with saline and weighed and homogenized (the homogenate was methanol∶15 mM PBS = 1∶2), and the homogenization ratio was 1∶4 (1 g of tissue to 4 mL of homogenate), and stored at -60°C or below.

[0428] Sample treatment:

[0429] Plasma sample treatment:

[0430] Protein precipitation: 60 μL of acetonitrile containing internal standard was added to 3 μL of plasma sample for precipitation. After mixing, it was centrifuged at 12,000 g at 4°C. 50 μL of the treated supernatant was taken and added to a 96-well plate. After centrifugation at 3,220 g at 4°C, the supernatant was directly subjected to LC-MS / MS analysis.

[0431] Small intestine homogenate sample treatment:

[0432] 400 μL of acetonitrile containing internal standard was added to 20 μL of small intestine homogenate sample for precipitation. After mixing, it was centrifuged at 3,220 g at 4°C. 50 μL of the treated supernatant was taken and added to a 96-well plate. After centrifugation at 3,220 g at 4°C, the supernatant was directly subjected to LC-MS / MS analysis.

[0433] Colon homogenate sample treatment:

[0434] 400 μL of acetonitrile containing internal standard was added to 20 μL of colon homogenate sample for precipitation. After mixing, it was centrifuged at 3220 g at 4°C. 50 μL of the treated supernatant was taken and added to a 96-well plate. After centrifugation at 3220 g at 4°C, the supernatant was directly subjected to LC-MS / MS analysis.

[0435] The experimental results are shown in the following table:

[0436]

[0437]

[0438] Note: ND, below the detection limit

[0439] Conclusion: The compound of the present invention exhibits good drug exposure levels in the small intestine and colon of mice, and the compound has high ratios of small intestine / plasma and colon / plasma, showing good tissue selectivity.

[0440] Experimental Example 5

[0441] Oxazolone (OXA)-induced murine colitis model

[0442] Test animals:

[0443] 45 Balb / c mice, 8 - 10 weeks old, 18 - 20 g, female

[0444] Experimental procedures:

[0445] Randomly divided into 9 groups, a blank control group with 5 mice, sensitized on the back with 100 μL of acetone + olive oil (4:1) for 5 days, and starting from Day 0, rectal perfusion with 50% ethanol; the remaining 8 groups are model groups, with 5 mice in each group, sensitized on the back with 100 μL of 2% Oxazolone (diluted with acetone + olive oil) for 5 days, and starting from Day 0, rectal perfusion with 150 μL of 1% Oxazolone solution. The test compound is administered from Day - 1 to the end of Day 3, and Day 4 is the experimental end point. The specific grouping is shown in Table 3

[0446] Table 3 Animal grouping and dosing regimens

[0447]

[0448] Note: The solvent is 0.5% CMC - Na

[0449] Data processing

[0450] During the experiment (In - life)

[0451] Record the body weight and disease activity index (DAI) score of the animals every day to evaluate the disease onset of each group of animals and the effect of the test compound on the disease. The DAI score consists of 3 parts, and the specific criteria refer to Table 4 below

[0452] Table 4 DAI scoring criteria

[0453] Score Percentage of weight loss Stool consistency Occult blood or bloody stool 0 0 Normal Negative occult blood 1 1～5 Soft stool Weakly positive occult blood 2 6～10 Loose stool Positive occult blood 3 11～20 Watery stool Small amount of blood 4 ＞20 Large amount of blood ​

[0454] Method description of fecal occult blood determination

[0455] When fresh blood is seen in the feces or at the anus, occult blood is no longer measured. The feces of the mice without obvious bloody stools were collected and subjected to occult blood measurement. If the color of the feces becomes darker and darker within 1 - 2 minutes, 2 points are given. If there is no obvious color or the color is very weak within 1 - 2 minutes, and then the color appears, but the color depth is significantly lower than that of the feces of the mice with 2 points, 1 point is given.

[0456] Experimental results

[0457] The data obtained were expressed as mean ± standard error, and statistical differences were analyzed. The results are shown in Tables 5, 6 and 7.

[0458]

[0459] Table 6 Changes in DAI score

[0460]

[0461] Table 7 Changes in colon length and weight

[0462]

[0463] Conclusion: In the oxazolone (OXA)-induced mouse colitis model, the compound of the present invention can relieve the OXA-induced weight loss, significantly improve the disease activity index (DAI) score and the colon weight-length ratio at the experimental end point, showing good therapeutic effects.

Claims

1. A compound of formula (II) or a pharmaceutically acceptable salt thereof, wherein, L1 is -(CH2) m - or -S(=O)2-; T1 is CH; T2 is CH; T3 is CH; R1 is CN or a 4- to 6-membered heterocycloalkyl group, wherein the 4- to 6-membered heterocycloalkyl group is substituted with 1, 2 or 3 R a substituents; R2 is C 1-3 alkyl; R3 is C 1-3 alkyl; R4 is H or C 1-3 alkyl; q is 1 or 2; m is 1 or 2; n is 1; R a is CN; the "4-6 membered heterocycloalkyl" contains 1, 2 or 3 heteroatoms which are N.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein, R1 is CN or wherein said optionally substituted by 1, 2 or 3 R a substituents.

3. The compound according to claim 2 or a pharmaceutically acceptable salt thereof, wherein, R1 is CN or 4. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein, R2 is CH3.

5. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein, R3 is CH3.

6. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein, R4 is H or CH3.

7. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein, Structural unit is 8. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein, Structural unit is 9. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein the compound is wherein, n, R1, R2, R3, L1, T3, T2 and q are as defined in claim 1.

10. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein the compound is wherein, R1, R2, R3, L1, T2 and q are as defined in claim 1.

11. A compound of the following formula or a pharmaceutically acceptable salt thereof, 12. Use of the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 11 in the preparation of a medicament for treating JAK-related diseases.

13. Use of the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 11 in the preparation of a medicament for treating gut-restricted JAK-related diseases.

14. The use according to claim 13, wherein the gut-restricted JAK-related disease refers to inflammatory bowel disease.

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