Menin-MLL protein inhibitor as well as pharmaceutical composition and application thereof

Abstract

The invention provides a compound as shown in a formula I, pharmaceutically acceptable salts, hydrates, isomers, prodrugs or a mixture thereof, a pharmaceutical composition containing the compound, and application of the compound and the pharmaceutically acceptable salts, hydrates, isomers and prodrugs in preparation of drugs for preventing, relieving or treating related diseases caused by interaction with Menin-MLL protein. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to a Menin-MLL protein inhibitor, a pharmaceutical composition containing the same, and their use in the preparation of a drug for preventing or treating related diseases caused by the interaction of Menin-MLL protein. Background Art

[0002] The MLL (Mixed Lineage Leukemia) protein is a histone methyltransferase, also known as KMT2A (Histone-lysine N-methyltransferase 2A). MLL rearrangement (MLL-r) leukemia is caused by the translocation of the chromosomal locus of 11q23 containing the gene encoding KMT2A. Currently, it is known that this chromosomal translocation can produce more than 60 oncogenic fusion proteins - formed by the fusion of the amino terminus of MLL with a variety of different proteins, among which the MLL-AF4 / 9 fusion mutation has the highest malignancy. MLL-r leukemia accounts for 5% - 10% of adult acute leukemia and 70% of infant acute leukemia. Currently, the treatment options are limited, mostly chemotherapy drugs, with a poor prognosis and high recurrence rate.

[0003] Menin is a protein mainly localized in the nucleus, encoded by the Multiple Endocrine Neoplasia Type 1 (MEN1) gene, and is an important cofactor of the oncogenic MLL-r fusion protein, having a high affinity for the MLL-r protein. After Menin binds to the MLL-r protein, it recruits chromatin-modifying enzymes such as Dot1L or the pTEFb complex, resulting in enhanced transcription of genes including HOXA, MEIS1, etc. The abnormal expression of these genes hinders the differentiation of hematopoietic cells and promotes their proliferation. In vitro and in vivo experiments have shown that Menin inhibitors can disrupt the interaction between Menin and MLL-r, and specifically cause growth inhibition and apoptosis of leukemia cells carrying the MLL–r mutation (Cancer Cell 36, 660–673). Studies have also found that Menin inhibitors are effective against leukemia with NPM1 gene mutations, and NPM1 mutations account for about 20 - 30% of patients with acute myeloid leukemia (Science 367, 586–590).

[0004] Currently, several Menin inhibitors are conducting clinical phase 1 / 2 studies for patients with relapsed / refractory acute leukemia carrying MLL-r or NPM1 mutations, such as SNDX-5613 from Syndax Pharmaceuticals, KO-539 from Kura Oncology, and DS-1594b from Daiichi Sankyo Group in Japan. In a clinical trial named AUGMENT-101, among the 60 evaluable patients, 53% of the patients responded to the drug. However, after the second treatment cycle, some patients developed resistance to SNDX-5613. The study found that these drug-resistant patients had MEN1 gene mutations, resulting in amino acid changes in Menin proteins M3271, M327V, G331R, G331D, T349M, and S160C. These amino acid point mutations within the Menin drug-binding pocket interfere with the binding of drug molecules to the target protein, thereby reducing drug affinity. Importantly, the affinity of these mutant Menin proteins for the KMT2A peptide is not significantly affected by the structural changes. The study showed that the proteins with M327I and T349M point mutations had significantly reduced sensitivity to the reported inhibitors, thus gaining a significant selective advantage (Nature 615, 913–919). Therefore, it is particularly important to develop inhibitors that can bind to both wild-type and mutant Menin proteins, especially the proteins with M327I and T349M point mutations, which will bring hope for the cure of these drug-resistant patients.

[0005] In addition, excessively high Menin expression levels can lead to hindrance in the β-cell proliferation process, resulting in relatively insufficient insulin secretion. A Menin-MLL inhibitor has been shown to enhance β-cell proliferation, thus providing the possibility of application in the field of diabetes. Currently, one compound, BMF-219, is conducting a phase 2 clinical study for type 2 diabetes.

[0006] In summary, Menin-MLL interaction inhibitors have good application prospects as drug research and development, and there is a good clinical need to develop Menin-MLL interaction inhibitors. Summary of the Invention

[0007] The present invention provides a compound represented by formula I, its pharmaceutically acceptable salts, hydrates, isomers, prodrugs, or mixtures thereof:

[0008]

[0009] In formula I:

[0010] m = 0, 1 or 2; n = 0, 1, 2 or 3.

[0011] R1 is selected from hydrogen, hydroxyl, halogen, cyano, C1-C3 alkyl, C1-C3 alkoxy or C1-C3 haloalkyl.

[0012] In some embodiments, R1 is selected from hydrogen, fluorine, chlorine, bromine, iodine, cyano, hydroxyl, amino, methyl, ethyl, n-propyl, isopropyl, methoxy, ethoxy, chloromethyl, trifluoromethyl or perfluoroethyl. In some embodiments, R1 is hydrogen or methyl.

[0013] R2 and R3 are each independently selected from hydrogen, C1-C3 alkyl, C1-C3 alkoxy, C1-C3 haloalkyl, R x CO-, sulfonyl or phosphonyl, wherein R x is alkyl or alkenyl.

[0014] In some embodiments, Rx is C1-C3 alkyl; in some specific embodiments, Rx is methyl, ethyl, n-propyl or isopropyl. In some embodiments, Rx is C2-C4 alkenyl, and in some specific examples, Rx is vinyl, 1-propenyl or 2-propenyl.

[0015] In some embodiments, R2 is hydrogen.

[0016] In some embodiments, R3 is hydrogen.

[0017] In some embodiments, R2 is hydrogen and R3 is methyl, ethyl, n-propyl, isopropyl, methoxy, ethoxy, chloromethyl, trifluoromethyl, perfluoroethyl, acetyl, acryloyl, dimethylphosphonyl, methanesulfonyl or ethanesulfonyl.

[0018] R4 and R5 are each independently selected from hydrogen, hydroxyl, halogen, C1-C3 alkyl, C1-C3 alkoxy or C1-C3 haloalkyl.

[0019] In some embodiments, R4 is hydrogen.

[0020] In some embodiments, R5 is hydrogen.

[0021] In some embodiments, R4 is hydrogen and R5 is selected from hydroxyl, halogen, C1-C3 alkyl, C1-C3 alkoxy or C1-C3 haloalkyl.

[0022] In some embodiments, R4 and R5 are independently selected from hydrogen, hydroxyl, fluorine, chlorine, bromine, iodine, cyano, methyl, ethyl, n-propyl, isopropyl, methoxy, ethoxy, chloromethyl, trifluoromethyl or perfluoroethyl.

[0023] R6 is selected from substituted or unsubstituted 5- to 7-membered aryl, 5- to 7-membered heteroaryl, 9- to 16-membered saturated or partially unsaturated cycloalkyl, 9- to 16-membered saturated or partially unsaturated heterocyclic group, 9- to 16-membered fused aryl, 9- to 16-membered fused heteroaryl.

[0024] In some embodiments, R6 is selected from substituted or unsubstituted 5- to 7-membered aryl, 5- to 7-membered heteroaryl, 9- to 12-membered saturated or partially unsaturated cycloalkyl, 9- to 12-membered saturated or partially unsaturated heterocyclic group, 9- to 12-membered fused aryl, 9- to 12-membered fused heteroaryl, wherein the heterocyclic group or heteroaryl optionally contains 1 to 3 heteroatoms selected from N, O, and S in the framework atoms.

[0025] In some embodiments, R6 can be monocyclic or polycyclic.

[0026] In some embodiments, R6 is a 9- to 12-membered fused bicyclic ring.

[0027] In some embodiments, R6 is a 9- to 12-membered fused heterocyclic group or 9- to 12-membered fused heteroaryl.

[0028] In some embodiments, R6 is optionally selected from the following substituted or unsubstituted cyclic groups: furyl, thienyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl; phenyl, pyridyl, pyrimidinyl, pyrazinyl, 1,2,3-triazinyl, 1,2,4-triazinyl, 1,3,5-triazinyl; indolyl, benzimidazolyl, benzofuryl, benzothiazolyl, benzisothiazole, benzoxazolyl, benzisoxazole, benzothienyl, indazolyl, pyrrolo[1,2-a]pyridine, imidazo[1,2-a]pyridine, imidazo[1,5-a]pyridine, pyrazolo[1,5-a]pyridine, isoindolin-1-one group, 3,4-dihydroisoquinolin-1(2H)-one group; naphthyl, quinolinyl, isoquinolinyl, naphthyridinyl, pyridinediazinyl, benzotriazinyl, benzoxatriazinyl, benzopyranyl, benzogamma-pyrone group, tetrahydroisoquinolinyl, tetrahydronaphthyl, 3,4-dihydro-2H-1,4-benzoxazinyl, 2H-1,4-benzoxazin-3(4H)-one group, 1,2,3,5,6,7-hexahydro-S-dipentacyclophenyl.

[0029] In some embodiments, R6 is optionally substituted at any possible position by one or more groups selected from oxygen, hydroxyl, amino, carboxyl, cyano, halogen, phosphonyl, sulfonyl, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, R a CO-, C2-C4 alkenyl, C3-C6 cycloalkyl, 3- to 6-membered heterocycloalkyl, 5- to 7-membered aryl or 5- to 7-membered heteroaryl, wherein R a represents alkyl or alkenyl.

[0030] In some embodiments, R a is a C1-C5 alkyl group; in some embodiments, R a is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, or neopentyl. In some embodiments, R a is a C2-C4 alkenyl group, and in some specific embodiments, R a is vinyl, 1-propenyl, or 2-propenyl.

[0031] In some embodiments, R6 is selected from the following substituted or unsubstituted groups:

[0032]

[0033]

[0034] In some embodiments, R6 may optionally be substituted at any possible position with one or more groups selected from oxygen, hydroxyl, amino, carboxyl, cyano, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, methoxy, ethoxy, chloromethyl, trifluoromethyl, 2,2,2-trifluoroethyl, perfluoroethyl, formyl, acetyl, acryloyl, methylphosphonyl, dimethylphosphonyl, methanesulfonyl, ethanesulfonyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, furyl, thienyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, triazole, tetrazole, phenyl, pyridyl.

[0035] In some embodiments, the present invention provides a compound represented by Formula II, its pharmaceutically acceptable salt, hydrate, isomer, prodrug, or a mixture thereof:

[0036]

[0037] In Formula II: n = 0, 1, 2, or 3.

[0038] The value ranges of R1, R2, R3, R4, R5, and R6 are the same as those described above.

[0039] In some embodiments, the present invention provides a compound represented by Formula III, its pharmaceutically acceptable salt, hydrate, isomer, prodrug, or a mixture thereof:

[0040]

[0041] In Formula III, n = 0, 1, 2, or 3.

[0042] The value ranges of R1, R2, R3, R4, R5, and R6 are the same as those described above.

[0043] The present invention provides the following compounds, their pharmaceutically acceptable salts, hydrates, isomers, prodrugs or mixtures thereof:

[0044]

[0045]

[0046]

[0047]

[0048] The present invention also provides a pharmaceutical composition, which comprises any of the aforementioned compounds, their pharmaceutically acceptable salts, hydrates, isomers, prodrugs or mixtures thereof, and a pharmaceutically acceptable excipient and / or carrier.

[0049] The present invention also provides the use of any of the aforementioned compounds, their pharmaceutically acceptable salts, hydrates, isomers, prodrugs or mixtures thereof or a pharmaceutical composition in the preparation of a drug for preventing, alleviating or treating diseases associated with the interaction of Menin-MLL proteins.

[0050] In the present invention, the diseases associated with the interaction of Menin-MLL proteins include malignant tumors, diabetes or complications associated with said diseases. Among them, malignant tumors include hematological tumors, lymphomas, solid tumors.

[0051] Hematological tumors include leukemia and myeloma, including but not limited to acute lymphoblastic leukemia, chronic lymphocytic leukemia, acute myeloid leukemia, chronic myeloid leukemia, acute monocytic leukemia, chronic monocytic leukemia, childhood leukemia, acute granulocytic leukemia, chronic granulocytic leukemia, mixed lineage leukemia, hairy cell leukemia, precursor T-cell lymphoblastic leukemia, large granular lymphocytic leukemia, meningeal leukemia, myelodysplastic syndrome, myeloproliferative diseases, myeloproliferative neoplasia, plasmacytoma, multiple myeloma.

[0052] Lymphomas include but not limited to cutaneous T-cell lymphoma, lymphoid neoplasm, AIDS-related lymphoma, Hodgkin lymphoma, non-Hodgkin lymphoma or malignant lymphoma.

[0053] Solid tumors include but not limited to pancreatic cancer, colon cancer, rectal cancer, liver cancer, gastric cancer, glioblastoma multiforme, lung cancer, breast cancer, prostate cancer.

[0054] Related complications include but not limited to leukemic meningitis.

[0055] Experimental results have shown that the Menin-MLL protein inhibitor of the present invention has excellent in vitro enzyme inhibition activity and cell proliferation inhibition activity. In particular, compared with existing compounds, it has excellent activity in inhibiting the interaction between Menin mutant protein and MLL protein, and thus has good prospects for resistance to drug resistance.

[0056] This invention is funded by the Sichuan Provincial Science and Technology Plan. Detailed implementation manners

[0057] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Apparently, the described embodiments are only a part of the embodiments of the present invention, rather than all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention. Definitions and general descriptions:

[0058] Indicates the connection position of this group to other structures.

[0059] Unless otherwise specified, "substituted" means that a hydrogen atom in a molecule is replaced by other different atoms or groups.

[0060] "Oxygen substitution" means that an oxygen atom forms a "C=O" structure with a directly adjacent carbon atom.

[0061] "Alkyl" refers to a saturated hydrocarbon group composed only of carbon atoms and hydrogen atoms, with single bonds between carbon-carbon and carbon-hydrogen atoms. The alkyl group can be straight-chain or branched-chain. Representative branched-chain alkyl groups have one, two or three branches. Exemplary alkyl groups include but are not limited to methyl, ethyl, propyl (n-propyl and isopropyl), butyl (n-butyl, isobutyl and tert-butyl), pentyl (n-pentyl, isopentyl and neopentyl) and hexyl.

[0062] "Heteroatom" refers to a non-carbon atom in a carbon chain or backbone atom. Typical heteroatoms include but are not limited to N, O and S, etc.

[0063] "Member" represents the number of backbone atoms constituting a ring. Typical 5-membered rings include, for example, cyclopentyl, pyrrole, pyrrolidine, imidazole, thiazole, furan, tetrahydrofuran, thiophene, etc.; typical 6-membered rings include, for example, cyclohexyl, piperidine, piperazine, pyridine, pyran, pyrazine, thiopyran, pyridazine, pyrimidine, benzene, etc. A ring containing a heteroatom in the backbone atoms is a heterocycle, an aromatic group containing a heteroatom is a heteroaryl; a non-aromatic group containing a heteroatom is a heterocycloalkyl.

[0064] "Cycloalkyl" refers to a non-aromatic cyclic group in which all the skeletal atoms are carbon atoms, including saturated or unsaturated monocyclic, bicyclic or polycyclic systems, such as fused 2, 3, 4 rings or spiro rings, etc., specifically such as cyclized alkyl and alkenyl groups. Typical examples of cycloalkyl include but are not limited to cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclopentenyl, cyclohexenyl, etc. The definition of "cycloalkyl" also includes cyclic groups formed by the fusion of a non-aromatic ring and an aromatic ring, such as benzo- or pyrido-derivative groups of cyclopentane, cyclopentene, cyclohexane, etc., and specific examples include tetrahydronaphthalene.

[0065] "Heterocycloalkyl" and "heterocyclic group" have the same meaning, referring to a saturated or unsaturated monocyclic, bicyclic or polycyclic system (such as 2, 3 or 4 fused rings or spiro rings, etc.) in which one or more of the skeletal atoms are heteroatoms. The heteroatoms are selected from N, O and S, and the remaining ring atoms are carbon. Examples of heterocycloalkyl include but are not limited to: azetidinyl, oxetanyl, pyrrolinyl, pyrrolidinyl, tetrahydrofuranyl, tetrahydrothienyl, piperidinyl, piperazinyl, tetrahydropyranyl or morpholinyl. The definition of "heterocycloalkyl" also includes cyclic groups formed by the fusion of the aforementioned cycloalkyl ring or heterocycloalkyl ring with an aromatic ring, where the aromatic ring may or may not include the aforementioned heteroatoms, and specific examples include but are not limited to tetrahydroquinolinyl, tetrahydroisoquinolinyl or benzogamma-pyrone group.

[0066] "Aryl" refers to a monocyclic or polycyclic (such as having 2, 3 or 4 fused rings) aromatic hydrocarbon group. Typical aryls include but are not limited to phenyl, naphthyl, anthracenyl, phenanthryl, etc.

[0067] "Heteroaryl" refers to a monocyclic or polycyclic (such as having 2, 3 or 4 fused rings) aromatic heterocyclic moiety having one or more ring members that are heteroatoms selected from N, S and O. Typical heteroaryls include but are not limited to pyridyl, indolyl, quinoxalinyl, quinolinyl, isoquinolinyl, benzothienyl, benzofuranyl, benzothiophenyl, benzopyranyl, benzothiopyranyl, furyl, pyrrolyl, thiazolyl, oxazolyl, isoxazolyl, triazolyl, tetrazolyl, pyrazolyl, imidazolyl, thienyl, oxadiazolyl, benzimidazolyl, benzothiazolyl, benzoxazolyl, etc.

[0068] "Alkenyl" refers to an unsaturated hydrocarbon group having one or more -C=C- (carbon-carbon double bonds). Examples of alkenyl include but are not limited to vinyl, 1-propenyl, 2-propenyl, 1-butenyl, 2-butenyl, 3-butenyl, 2-methyl-1-propenyl, etc.

[0069] "Halogen" refers to fluorine, chlorine, bromine or iodine.

[0070] "Cyano" refers to -CN.

[0071] "Halogenated alkyl" means an alkyl group substituted by one or more halogen atoms, where the alkyl has the meaning as described in the present invention. Halogenated alkyl includes but is not limited to mono-halogenated alkyl, di-halogenated alkyl, tri-halogenated alkyl, per-halogenated alkyl, etc., such as chloromethyl, dichloromethyl, difluoromethyl, dibromomethyl, trifluoromethyl, 2,2,2-trifluoroethyl, perfluoroethyl, 2,2,2-trifluoro-1,1-dichloroethyl, etc.

[0072] "Alkoxy" refers to -O-alkyl, where the alkyl has the meaning as described in the present invention.

[0073] "Amino", used alone or in combination with other terms, refers to the group of formula -NH2.

[0074] "Hydroxy" refers to -OH.

[0075] "Sulfonyl" refers to the group of formula where Rb represents an alkyl or aryl as defined in the present invention. Exemplary sulfonyl groups include but are not limited to methylsulfonyl, ethylsulfonyl, benzenesulfonyl, p-toluenesulfonyl.

[0076] "Phosphonyl" refers to the group of formula where Rc represents a hydroxy group or an alkyl or aryl as defined in the invention, and Rd represents an alkyl or aryl as defined in the present invention. Exemplary phosphonyl groups include but are not limited to methylphosphonyl, dimethylphosphonyl, diethylphosphonyl.

[0077] In this article, unless otherwise specified, the term "Ca-Cb" means that the moiety modified thereby has a - b carbon atoms (b > a, and both are integers). For example, C 1- C3 means that the moiety modified thereby has 1 - 3 carbon atoms, such as 1 carbon atom, 2 carbon atoms, or 3 carbon atoms.

[0078] "p - q membered (hetero)cycloalkyl" refers to the moiety modified by this term having p - q carbon atoms (q > p, and both are integers) and the number of heteroatoms participating in the ring formation. For example, 3 - 6 membered hetero cycloalkyl means that the ring - forming structure modified thereby has 3, 4, 5, or 6 atoms, including carbon atoms and at least one heteroatom.

[0079] "y - z membered (hetero)aryl" refers to the moiety modified by this term having y - z carbon atoms (z > y, and both are integers) and the total number of heteroatoms participating in the ring formation. For example, 5 - 7 membered aryl means that the aryl structure modified thereby has 5, 6, or 7 carbon atoms; for example, 5 - 7 membered heteroaryl means that the heteroaryl structure modified thereby has 5, 6, or 7 atoms, including carbon atoms and at least one heteroatom.

[0080] "Optionally" means that the subsequently described events or circumstances can be freely chosen or not chosen.

[0081] "Hydrate" refers to an aggregate of one or more water molecules contained in the compounds of the present invention, including hemihydrate, monohydrate, dihydrate, trihydrate, etc.

[0082] "Isomer" means that when the compounds of the present invention contain one or more asymmetric centers, they can exist in the form of racemates and racemic mixtures, single enantiomers, diastereoisomer mixtures and single diastereoisomers. The compounds of the present invention may have asymmetric centers, which may result in the existence of two optical isomers. The scope of the present invention includes all possible optical isomers and their mixtures. If the compounds of the present invention contain olefin double bonds, unless otherwise specified, the scope of the present invention includes cis isomers and trans isomers. The compounds of the present invention may exist in the form of tautomers (a kind of functional group isomers), which have different hydrogen attachment points through the displacement of one or more double bonds. For example, a ketone and its enol form are keto-enol tautomers. Each tautomer and its mixture are within the scope of the present invention. All enantiomers, diastereoisomers, racemates, meso forms, cis-trans isomers, tautomers, geometric isomers, epimers and their mixtures of the compounds are within the scope of the present invention.

[0083] "Prodrug" refers to a derivative compound that can directly or indirectly provide the compound of the present invention after being administered to an individual. Particularly preferred derivative compounds or prodrugs are compounds that can improve the bioavailability of the compound of the present invention (e.g., more easily absorbed into the blood) when administered to an individual, or compounds that promote the delivery of the parent compound to the site of action (e.g., the lymphatic system). Unless otherwise indicated, all prodrug forms of the compounds of the present invention are within the scope of the present invention, and various prodrug forms are known in the art. For example, see T. Higuchi, V. Stella, Pro-drugs as Novel Drug Delivery Systems[J], American Chemical Society, Vol. 14, 1975. In addition, the present invention also encompasses compounds of the present invention containing protecting groups. During any process for preparing the compounds of the present invention, it may be necessary and / or desirable to protect sensitive groups or reactive groups on any relevant molecules, thereby forming chemically protected forms of the compounds of the present invention. This can be achieved by conventional protecting groups, such as those described in T.W. Greene, P.G.M. Wuts, Protective Groups in Organic Synthesis[M], John Wiley & Sons, 2006. These protecting groups can be removed at appropriate subsequent stages using methods known in the art.

[0084] "Pharmaceutical composition" refers to a preparation of the compound of the present invention and a medium generally accepted in the art for delivering bioactive compounds to mammals (such as humans). This medium includes pharmaceutically acceptable carriers. The purpose of the pharmaceutical composition is to facilitate the administration to an organism, promote the absorption of the active ingredient, and thereby exert its biological activity.

[0085] "Pharmaceutically acceptable" refers to substances such as carriers, diluents, or excipients that do not affect the biological activity or properties of the compounds of the present invention and are relatively non-toxic, i.e., the substance can be administered to an individual without causing adverse biological reactions or interacting in an adverse manner with any component contained in the composition. For example, "excipients" include, but are not limited to, any adjuvants, carriers, excipients, glidants, sweeteners, diluents, preservatives, dyes / colorants, flavoring agents, surfactants, wetting agents, dispersants, suspending agents, stabilizers, isotonic agents, solvents, or emulsifiers that are permitted by the relevant government regulatory authorities for use in humans or livestock.

[0086] "Pharmaceutically acceptable salts" include pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.

[0087] The present invention also provides a method for synthesizing the above compound. The synthesis method of the present invention mainly conducts relevant syntheses starting from the preparation methods reported in chemical literature or using commercially available chemical reagents as starting materials.

[0088] Abbreviation Explanation

[0089] Boc represents tert-butyl carbonate;

[0090] TsOH represents 4-methylbenzenesulfonic acid;

[0091] IPA represents isopropanol;

[0092] TEA represents triethylamine;

[0093] DCM represents dichloromethane;

[0094] DBU represents 1,8-diazabicyclo[5.4.0]undec-7-ene;

[0095] THF represents tetrahydrofuran;

[0096] TMEDA represents N,N,N',N'-tetramethylethylenediamine;

[0097] TMSCl represents trimethylchlorosilane;

[0098] EDCl represents 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride;

[0099] HOBT represents 1-hydroxybenzotriazole;

[0100] DIPEA represents N,N-diisopropylethylamine;

[0101] XPhos represents 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl;

[0102] XPhosPdG3 represents methanesulfonic acid (2-dicyclohexylphosphino-2',4',6'-tri-isopropyl-1,1'-biphenyl)(2'-amino-1,1'-biphenyl-2-yl)palladium(II);

[0103] KOAc represents potassium acetate;

[0104] dppf represents 1,1'-bis(diphenylphosphino)ferrocene;

[0105] Fmoc represents 9-fluorenylmethyl formate;

[0106] EA represents ethyl acetate;

[0107] MeI represents methyl iodide;

[0108] DCC represents N,N'-dicyclohexylcarbodiimide;

[0109] PPh3 represents triphenylphosphine. Specific examples

[0111] Preparation Example 1

[0112] 7-Bromo-1-(2,2,2-trifluoroethyl)-1H-indole-2-carbonitrile

[0113] Synthesis route:

[0114]

[0115] Step 1: Preparation of 7-bromo-1H-indole-2-carboxamide

[0116] 7-Bromo-1H-indole-2-carboxylic acid (3 g) was dissolved in 40 mL of tetrahydrofuran, N,N'-carbonyldiimidazole (3.04 g) was added, and after reacting at room temperature for 1.5 h, ammonia water (5.78 mL) was added and the reaction continued for 2 h. The reaction was monitored by TLC and LCMS until completion, concentrated under reduced pressure, and purified by column chromatography to obtain the title compound (3.0 g).

[0117]

[0118] MS(ESI) m / z 239.0 (M+H) +

[0119] Step 2: Preparation of 7-bromo-1H-indole-2-carbonitrile

[0120] 7-Bromo-1H-indole-2-carboxamide (3 g) was dissolved in 40 mL of toluene, phosphorus oxychloride (5.86 mL) was added, the mixture was purged with nitrogen three times, and reacted at 120 °C for 2 h. The reaction was monitored by TLC until completion, concentrated under reduced pressure, and purified by column chromatography to obtain the title compound (2.63 g).

[0121]

[0122] 1 1H NMR (400 MHz, DMSO-d6) δ 12.62 (s, 1H), 7.71 (d, J = 8.0 Hz, 1H), 7.59 (d, J = 7.5 Hz, 1H), 7.51 (d, J = 2.0 Hz, 1H), 7.11 (t, J = 7.8 Hz, 1H).

[0123] Step 3: Preparation of 7-bromo-1-(2,2,2-trifluoroethyl)-1H-indole-2-carbonitrile

[0124] 7-Bromo-1H-indole-2-carbonitrile (600 mg) was dissolved in 15 mL of N,N-dimethylformamide, potassium carbonate (1.13 g) was added, and 2,2,2-trifluoroethyl trifluoromethanesulfonate (0.79 mL) was reacted at 80 °C for 10 h. The reaction was monitored by TLC, quenched with water, extracted twice with ethyl acetate, washed with saturated brine, and purified by column chromatography to obtain the title compound (800 mg).

[0125]

[0126] 1 H NMR (400 MHz, DMSO-d6) δ 7.82 (dd, J=8.0, 1.0 Hz, 1H), 7.78 (s, 1H), 7.74 (dd, J=7.6, 1.1 Hz, 1H), 7.21 (t, J=7.8 Hz, 1H), 5.65 (q, J=8.6 Hz, 2H).

[0127] Using 7-bromo-1H-indole-2-carbonitrile (the title compound in Step 2 of Preparation Example 1) as the substrate, reacting with different commercially available halogenating reagents, referring to Step 3 in the preparation example, different intermediates were obtained, as shown in Table 1 specifically.

[0128] Table 1 Intermediates

[0129]

[0130] Preparation Example 2: Preparation of Methyl (S)-3-((tert-Butoxycarbonyl)amino)-4-iodobutyrate

[0131]

[0132] Step 1: Preparation of 1-(2,5-Dioxopyrrolidin-1-yl) 4-Methyl (tert-Butoxycarbonyl)-L-Aspartic Acid

[0133] (S)-2-((tert-Butoxycarbonyl)amino)-4-methoxy-4-oxobutyric acid was dissolved in ethyl acetate (400 mL), 1-hydroxypyrrolidine-2,5-dione (15.4 g) was added under an ice-water bath, and a solution of N,N'-dicyclohexylcarbodiimide (26.3 g) in ethyl acetate (100 mL) was added dropwise. After the addition, it was transferred to room temperature and left overnight, and the reaction was monitored by TLC until completion. It was filtered, and the filtrate was washed successively with saturated sodium bicarbonate solution and saturated brine, dried over anhydrous sodium sulfate, and concentrated to obtain the crude title compound (41.8 g).

[0134]

[0135] MS (ESI) m / z (M+H) + =345.1。

[0136] Step 2: Preparation of methyl (S)-3-((tert-butoxycarbonyl)amino)-4-hydroxybutyrate.

[0137] Sodium borohydride (7.4 g) was added to a mixed solvent of tetrahydrofuran (300 mL) and water (40 mL) under an ice-water bath. A solution of 1-(2,5-dioxopyrrolidin-1-yl)-4-methyl (tert-butoxycarbonyl)-L-aspartic acid (41.8 g) in tetrahydrofuran (100 mL) was added dropwise at this temperature. After the addition was completed in 40 minutes, the reaction was continued for half an hour under an ice-water bath. When the raw materials disappeared as monitored by TLC, the reaction was quenched with saturated ammonium chloride solution, and extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the crude title compound (17.3 g).

[0138]

[0139] MS(ESI) m / z(M+Na) + = 256.1

[0140] Step 3: Preparation of methyl (S)-3-((tert-butoxycarbonyl)amino)-4-iodobutyrate

[0141] Triphenylphosphine (19.5 g), imidazole (5.1 g), and iodine (18.9 g) were mixed in dry dichloromethane (300 mL). Under nitrogen protection and an ice-water bath, a solution of methyl (S)-3-((tert-butoxycarbonyl)amino)-4-hydroxybutyrate (17.3 g) in dichloromethane (100 mL) was added dropwise. The reaction mixture was warmed to room temperature and reacted for 3 h. When the raw materials were consumed as monitored by TLC, part of triphenylphosphine oxide was removed by filtration. After adding water to the filtrate, the system was quenched with sodium thiosulfate solution until it became colorless, and then extracted with dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (15.9 g).

[0142]

[0143] MS(ESI) m / z(M+Na) + = 366.1.

[0144] Some intermediates used in the present invention are shown in Table 2, and all the intermediates in this table are commercially available.

[0145] Table 2 Intermediates

[0146]

[0147]

[0148] Example 1

[0149] (R)-2-((5-(5-(3-Amino-4-phenylbutanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0150] Synthesis route:

[0151]

[0152] Step 1: Preparation of tert-butyl 3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate 4-methylbenzenesulfonate

[0153] Dissolve di-tert-butyl 4,6-dihydropyrrolo[3,4-c]pyrrole-2,5(1H,3H)-dicarboxylate (1.7 g) in isopropanol (30 mL), add 4-methylbenzenesulfonic acid (1.04 g), and react at 55 °C overnight. Monitor the reaction by LCMS until completion. Cool to room temperature, filter, wash the filter cake with ethyl acetate, and dry to obtain the title compound (1.65 g).

[0154]

[0155] MS(ESI) m / z(M+H) + = 211.1

[0156] Step 2: Preparation of tert-butyl 5-(3,6-dichloro-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate

[0157] Add tert-butyl 3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate 4-methylbenzenesulfonate (1.64 g) to dichloromethane (30 mL), dropwise add triethylamine (1.30 g), and then add trichloro-1,2,4-triazine (0.95 g) in portions. React at room temperature for 2 h. Monitor the reaction by LCMS until completion, quench with water, extract with dichloromethane three times, combine the organic phases, dry over anhydrous sodium sulfate, concentrate, and purify by column chromatography to obtain the title compound (1.5 g).

[0158]

[0159] MS(ESI) m / z(M+H) + = 358.1

[0160] Step 3: Preparation of tert-butyl 5-(3-chloro-6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate

[0161] tert-Butyl 5-(3,6-dichloro-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate (1.5 g) was added to tetrahydrofuran (20 mL), followed by the addition of DBU (0.96 g), N-ethyl-5-fluoro-2-hydroxy-N-isopropylbenzamide (1.04 g), and the reaction was carried out at 55 °C overnight. The reaction was monitored by LCMS until completion, then the reaction was stopped, cooled to room temperature, quenched with water, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (1.75 g).

[0162]

[0163] MS(ESI) m / z(M+H) + = 547.1

[0164] Step 4: Preparation of tert-Butyl 5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate

[0165] tert-Butyl 5-(3-chloro-6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate (1.75 g) was added to tetrahydrofuran (30 mL), sodium borohydride (0.61 g) was added portionwise, followed by the addition of tetramethylethylenediamine (0.74 g) and 1,1'-bis(diphenylphosphino)ferrocene palladium(II) dichloride (0.23 g). The reaction was carried out under nitrogen protection at room temperature overnight. The reaction was monitored by LCMS until completion, quenched with water, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (1.27 g).

[0166]

[0167] MS(ESI) m / z(M+H) + = 513.1

[0168] Step 5: Preparation of N-Ethyl-5-fluoro-N-isopropyl-2-((5-(3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrole-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)benzamide

[0169] tert-Butyl 5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate (1.27 g) was added to trifluoroethanol (20 mL), and trimethylchlorosilane (0.8 mL) was added. The reaction was carried out at room temperature for 2 h. The reaction was monitored by LCMS until completion, the solvent was evaporated under reduced pressure, the residue was adjusted to alkaline with saturated sodium bicarbonate solution, extracted three times with a dichloromethane / methanol mixed solvent, the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to dryness to obtain the title compound (1 g).

[0170]

[0171] MS(ESI)[M+H] + = 413.1.

[0172] Step 6: Preparation of tert-Butyl (R)-(4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxo-1-phenylbutan-2-yl)carbamate

[0173] (R)-3-((tert-Butoxycarbonyl)amino)-4-phenylbutanoic acid (36 mg) was added to N,N-dimethylformamide (2 mL), and N,N-diisopropylethylamine (45 mg), 1-hydroxybenzotriazole (30 mg), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (40 mg) were added successively. After reacting at room temperature for half an hour, N-ethyl-5-fluoro-N-isopropyl-2-((5-(3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)benzamide (50 mg) was added, and the reaction was carried out overnight at room temperature. The reaction was monitored by LCMS until completion, quenched with water, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (50 mg).

[0174]

[0175] MS(ESI)[M+H] + = 674.3.

[0176] Step 6: Preparation of (R)-2-((5-(5-(3-Amino-4-phenylbutanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0177] (R)-(4-(5-(6-(2-(Ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxo-1-phenylbutan-2-yl)carbamic acid tert-butyl ester (50 mg) was added to trifluoroethanol (3 mL), and trimethylchlorosilane (0.1 mL) was added. After reacting at room temperature for 2 hours, the reaction was monitored by LCMS to be complete. The solvent was distilled off under reduced pressure. The residue was adjusted to weak alkalinity with saturated sodium bicarbonate solution, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by pre-HPLC (preparative high performance liquid chromatography, the same below) to obtain the title compound (23.73 mg).

[0178]

[0179] MS(ESI)[M+H] + = 574.3.

[0180] 1 H NMR(400MHz,DMSO-d6)δ8.56(d,J = 2.0Hz,1H),7.49 - 7.44(td,J = 8.9,4.7Hz,1H),7.40 - 7.34(m,2H),7.31 - 7.27(t,J = 7.5Hz,2H),7.22 - 7.18(m,3H),4.78(m,2H),4.37 - 4.25(m,4H),4.19 - 4.10(m,2H),3.70 - 3.63(p,J = 6.6Hz,1H),3.34(m,1H),3.07 - 3.00(m,1H),2.72 - 2.67(dd,J = 13.1,5.9Hz,1H),2.61 - 2.54(m,1H),2.24 - 2.21(m,2H),1.56(br,2H),1.09 - 1.03(m,3H),1.03 - 0.89(m,4H),0.72(m,3H).

[0181] Using N-ethyl-5-fluoro-N-isopropyl-2-((5-(3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)benzamide (i.e., the title compound in step 5 of Example 1) as the starting material, and using the intermediates shown in Table 2 and reaction reagents available through commercial channels, the compounds of Examples 2 - 21 were obtained by referring to the synthetic routes in steps 6 - 7 of Example 1. The specific structures and characterizations of the compounds are shown in Table 3.

[0182] Table 3 Examples 2 - 21

[0183]

[0184]

[0185]

[0186]

[0187]

[0188]

[0189]

[0190] Example 22

[0191] (R)-3-(2-Amino-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutyl)benzoic acid

[0192] The synthetic route is as follows:

[0193]

[0194] Step 1: Preparation of methyl (R)-3-(2-((tert-butoxycarbonyl)amino)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutyl)benzoate

[0195] Add (R)-(1-(3-bromophenyl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamic acid tert-butyl ester (80 mg, synthesized according to Example 18) into methanol (2 mL), successively add triethylamine (30 μL), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium dichloromethane complex (9 mg). After the system is replaced with carbon monoxide, react at 110 °C for two days. Monitor the reaction by LCMS until it is complete. Distill off the solvent under reduced pressure, concentrate, and purify by column chromatography to obtain the title compound (30 mg).

[0196]

[0197] MS(ESI) m / z [M+H] + = 732.3

[0198] Step 2: Preparation of Methyl (R)-3-(2-Amino-4-(5-(6-(2-(Ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutyl)benzoate

[0199] Methyl (R)-3-(2-((tert-Butoxycarbonyl)amino)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutyl)benzoate (30 mg) was added to trifluoroethanol (3 mL), and trimethylchlorosilane (0.1 mL) was added. After reacting at room temperature for 2 hours, the reaction was monitored by LCMS to be complete. The solvent was distilled off under reduced pressure. The residue was adjusted to weak alkalinity with saturated sodium bicarbonate solution, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (15 mg).

[0200]

[0201] MS(ESI) m / z [M+H] + = 632.3

[0202] Step 3: Preparation of (R)-3-(2-Amino-4-(5-(6-(2-(Ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutyl)benzoic Acid

[0203] Methyl (R)-3-(2-amino-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutyl)benzoate (15 mg) was added to a mixed solvent of tetrahydrofuran (1 mL), methanol (0.3 mL) and water (0.3 mL), and lithium hydroxide monohydrate (6 mg) was added. After reacting at room temperature for 2 hours, the reaction was monitored by LCMS to be complete. The pH of the system was adjusted to 2 - 3 with 2M dilute hydrochloric acid, the solvent was distilled off under reduced pressure, and the residue was purified by pre-HPLC to obtain the title compound (2.41 mg).

[0204]

[0205] MS(ESI) m / z [M+H] + = 618.3

[0206] 1 1H NMR (400 MHz, DMSO-d6) δ 8.56 (s, 1H), 7.80 (s, 1H), 7.77 - 7.76 (d, J = 6.7 Hz, 1H), 7.49 - 7.44 (m, 1H), 7.40 - 7.33 (m, 4H), 4.77 (m, 2H), 4.36 - 4.28 (m, 4H), 4.17 - 4.12 (m, 2H), 3.70 - 3.63 (m, 2H), 3.06 (m, 1H), 2.83 - 2.73 (m, 2H), 2.39 - 2.37 (m, 2H), 1.09 - 1.03 (m, 4H), 0.99 - 0.89 (m, 3H), 0.72 (m, 3H)

[0207] Example 23

[0208] (R)-2-(2-Amino-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutyl)benzoic acid

[0209] Synthetic route:

[0210]

[0211] Step 1: Preparation of (R)-2-(2-((tert-butoxycarbonyl)amino)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutyl)benzoic acid

[0212] Methyl (R)-2-(2-((tert-butoxycarbonyl)amino)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutyl)benzoate (80 mg, synthesized starting from (R)-4-(2-bromophenyl)-3-((tert-butoxycarbonyl)amino)butyric acid in Example 19 and following the method of Step 1 in Example 22) was added to a mixed solvent of tetrahydrofuran (1 mL), methanol (0.3 mL) and water (0.3 mL). Lithium hydroxide monohydrate (14 mg) was added, and the reaction was carried out at room temperature for 2 hours. After monitoring by LCMS that the raw material disappeared, the pH of the system was adjusted to 2 - 3 with 2 M dilute hydrochloric acid, and then extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and the residue was purified by pre-TLC to obtain the title compound (15 mg).

[0213]

[0214] MS(ESI) m / z [M+H] + = 718.3

[0215] Step 2: Preparation of (R)-2-(2-amino-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutyl)benzoic acid

[0216] Add (R)-2-(2-((tert-butoxycarbonyl)amino)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrol[3,4-c]pyrrol-2(1H)-yl)-4-oxobutyl)benzoic acid (15 mg) to trifluoroethanol (3 mL), add trimethylchlorosilane (0.1 mL), and react at room temperature for 2 hours. After monitoring the reaction to completion by LCMS, evaporate the solvent under reduced pressure. Adjust the residue to weak alkalinity with saturated sodium bicarbonate solution, then evaporate the solvent under reduced pressure. The residue is purified by pre-HPLC to obtain the title compound (1.77 mg).

[0217]

[0218] MS(ESI) m / z [M+H] + = 618.3

[0219] 1 1H NMR (400 MHz, DMSO-d6) δ 8.98 (br, 1H), 8.57 (s, 1H), 7.51 - 7.45 (m, 2H), 7.41 - 7.35 (m, 2H), 7.29 - 7.18 (m, 3H), 4.82 (m, 2H), 4.41 - 4.36 (m, 4H), 4.21 - 4.16 (m, 2H), 3.70 - 3.61 (m, 2H), 3.06 - 2.94 (m, 3H), 2.82 - 2.77 (dd, J = 17.1, 4.6 Hz, 1H), 2.68 - 2.60 (m, 1H), 1.10 - 1.03 (m, 4H), 1.00 - 0.87 (m, 3H), 0.79 (m, 3H).

[0220] Example 24

[0221] (R)-2-((5-(5-(3-Amino-4-(2-cyanophenyl)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0222] Synthetic route:

[0223]

[0224] Step 1: Preparation of tert-butyl (R)-(1-(2-cyanophenyl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamate

[0225] tert-Butyl (R)-(1-(2-bromophenyl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamate (120 mg, referring to Example 19), potassium ferrocyanide trihydrate (48 mg), 2-(dicyclohexylphosphino)-2',4',6'-tri-isopropyl-1,1'-biphenyl (8 mg), methanesulfonic acid (2-dicyclohexylphosphino-2',4',6'-tri-isopropyl-1,1'-biphenyl)(2'-amino-1,1'-biphenyl-2-yl)palladium(II) (7 mg), potassium acetate (32 mg) were placed in a mixed solvent of 1,4-dioxane (1.5 mL) and water (0.4 mL). Under nitrogen protection, the reaction was sealed at 120 °C for 3 h, and the formation of the product was monitored by LCMS. After cooling to room temperature, water was added, and the mixture was extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (30 mg).

[0226]

[0227] MS(ESI) m / z [M+H] + = 699.3.

[0228] Step 2: Preparation of (R)-2-((5-(5-(3-Amino-4-(2-cyanophenyl)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0229] (R)-(1-(2-Cyanophenyl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamic acid tert-butyl ester (30 mg) was added to trifluoroethanol (3 mL), and trimethylchlorosilane (0.1 mL) was added. After reacting at room temperature for 2 hours, the reaction was monitored by LCMS to completion. The solvent was evaporated under reduced pressure. The residue was adjusted to weakly basic with saturated sodium bicarbonate solution, and then the solvent was evaporated under reduced pressure. The residue was purified by pre-HPLC to obtain the title compound (4.04 mg).

[0230]

[0231] MS(ESI) m / z [M+H] + = 599.3

[0232] 1 1H NMR (400 MHz, DMSO-d6) δ 8.56 (s, 1H), 7.78 - 7.76 (d, J = 7.7 Hz, 1H), 7.66 - 7.62 (t, J = 7.6 Hz, 1H), 7.52 - 7.45 (m, 2H), 7.43 - 7.34 (m, 3H), 4.79 (s, 2H), 4.38 - 4.29 (m, 4H), 4.20 - 4.13 (m, 2H), 3.70 - 3.63 (p, J = 6.6 Hz, 1H), 3.42 - 3.38 (m, 1H), 3.07 - 3.02 (m, 1H), 2.95 - 2.90 (dd, J = 13.4, 5.7 Hz, 1H), 2.85 - 2.76 (ddd, J = 13.1, 7.8, 3.4 Hz, 1H), 2.36 - 2.34 (d, J = 6.8 Hz, 2H), 1.72 (br, 2H), 1.10 - 1.03 (m, 4H), 0.99 - 0.90 (m, 3H), 0.72 (m, 3H).

[0233] Example 25

[0234] (R)-2-((5-(5-(3-Amino-4-(3-(dimethylphosphoryl)phenyl)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0235] Synthetic route:

[0236]

[0237] Step 1: Preparation of Methyl (R)-4-(3-bromophenyl)-3-((tert-butoxycarbonyl)amino)butyrate

[0238] Dissolve (R)-4-(3-bromophenyl)-3-((tert-butoxycarbonyl)amino)butyric acid (200 mg) in N,N-dimethylformamide. Add potassium bicarbonate (84 mg) and methyl iodide (55 μL) at room temperature and react for 4 h at room temperature. Monitor the reaction by LCMS until completion. Quench the reaction with water, extract with ethyl acetate, combine the organic phases, wash with saturated brine, dry over anhydrous sodium sulfate, and concentrate to obtain the title compound (220 mg).

[0239]

[0240] MS(ESI) m / z [M+H] + = 372.1

[0241] Step 2: Preparation of (R)-3-((tert-butoxycarbonyl)amino)-4-(3-(dimethylphosphoryl)phenyl)butyric acid

[0242] Mix methyl (R)-4-(3-bromophenyl)-3-((tert-butoxycarbonyl)amino)butyrate (130 mg), dimethylphosphine oxide (82 mg), palladium acetate (8 mg), 1,1'-bis(diphenylphosphino)ferrocene (40 mg), and N,N-diisopropylethylamine (0.15 mL) in DMF (2 mL), protect with nitrogen, and react at 120 °C for 8 h. Monitor the reaction by LCMS until completion. Cool to room temperature, quench the reaction with water, extract with ethyl acetate, combine the organic phases, dry over anhydrous sodium sulfate, and concentrate to obtain the crude product of methyl (R)-3-((tert-butoxycarbonyl)amino)-4-(3-(dimethylphosphoryl)phenyl)butyrate.

[0243] Add the crude product to a mixed solvent of tetrahydrofuran (2 mL), methanol (0.6 mL), and water (0.6 mL), add lithium hydroxide monohydrate (40 mg), and react at room temperature for 2 h. Monitor by LCMS until the raw materials disappear. Extract the system three times with ethyl acetate, discard the organic phase, adjust the pH of the aqueous phase to 2 - 3 with 2 M dilute hydrochloric acid, then extract with a dichloromethane / methanol mixed solvent, combine the organic phases, wash with saturated brine, dry over anhydrous sodium sulfate, and concentrate to obtain the crude title compound (80 mg).

[0244]

[0245] MS(ESI) m / z [M-H] -- = 354.1

[0246] Step 3: Preparation of tert-butyl (R)-(1-(3-(dimethylphosphoryl)phenyl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamate

[0247] (R)-3-((tert-Butoxycarbonyl)amino)-4-(3-(dimethylphosphoryl)phenyl)butyric acid (80 mg) was added to N,N-dimethylformamide (2 mL). N,N-Diisopropylethylamine (0.07 mL), 1-hydroxybenzotriazole (35 mg), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (50 mg) were added successively. After reacting at room temperature for half an hour, N-ethyl-5-fluoro-N-isopropyl-2-((5-(3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)benzamide (70 mg) was added, and the reaction was carried out overnight at room temperature. The reaction was monitored by LCMS until completion, quenched with water, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (45 mg).

[0248]

[0249] MS(ESI) m / z [M+H] + = 750.3.

[0250] Step 4: Preparation of (R)-2-((5-(5-(3-amino-4-(3-(dimethylphosphoryl)phenyl)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0251] Tert-butyl (R)-(1-(3-(dimethylphosphoryl)phenyl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamate (45 mg) was added to trifluoroethanol (3 mL), and trimethylchlorosilane (0.1 mL) was added. After reacting at room temperature for 2 hours, the reaction was monitored by LCMS until completion. The solvent was evaporated under reduced pressure. The residue was adjusted to weakly basic with saturated sodium bicarbonate solution, and then the solvent was evaporated under reduced pressure. The residue was purified by pre-HPLC to obtain the title compound (26.94 mg).

[0252]

[0253] MS(ESI) m / z [M+H] + = 650.3

[0254] 1 H NMR (400 MHz, DMSO-d6) δ 8.56 (s, 1H), 7.63 - 7.57 (m, 2H), 7.49 - 7.34 (m, 5H), 4.79 (m, 2H), 4.38 - 4.30 (m, 4H), 4.19 - 4.14 (m, 2H), 3.70 - 3.63 (p, J = 6.7 Hz, 1H), 3.09 - 3.01 (m, 2H), 2.81 - 2.76 (dd, J = 13.3, 5.8 Hz, 1H), 2.69 - 2.63 (q, J = 6.6 Hz, 1H), 2.38 - 2.25 (m, 2H), 1.65 (s, 3H), 1.62 (s, 3H), 1.09 - 0.99 (m, 4H), 0.97 - 0.89 (m, 3H), 0.74 - 0.70 (m, 3H).

[0255] Using the intermediate (R)-4-(2-bromophenyl)-3-((tert-butoxycarbonyl)amino)butyric acid as the starting material and reaction reagents available through commercial channels, the compound of Example 26 was obtained with reference to the synthetic route of Example 25, as shown in Table 4 specifically.

[0256] Table 4 Example 26

[0257]

[0258] Example 27

[0259] (R)-2-((5-(5-(3-Amino-4-(1-methyl-1H-indol-3-yl)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0260] Synthetic route:

[0261]

[0262] Step 1: Preparation of tert-butyl (R)-(4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1-(1-methyl-1H-indol-3-yl)-4-oxobutan-2-yl)carbamate

[0263] (R)-(4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1-(1H-indol-3-yl)-4-oxobutan-2-yl)carbamic acid tert-butyl ester (40 mg, refer to Example 6) was added to tetrahydrofuran (2 mL). Sodium hydroxide (34 mg) was added under an ice-water bath, and then methyl iodide (16 mg) was added later. The temperature was raised to room temperature and the reaction was carried out overnight. TLC was used to monitor the disappearance of the raw materials. Water was added to quench the reaction, and the mixture was extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by pre-TLC to obtain the title compound (15 mg).

[0264]

[0265] MS(ESI) m / z [M+H] + = 727.3.

[0266] Step 2: Preparation of (R)-2-((5-(5-(3-amino-4-(1-methyl-1H-indol-3-yl)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0267] (R)-(4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1-(1-methyl-1H-indol-3-yl)-4-oxobutan-2-yl)carbamic acid tert-butyl ester (15 mg) was added to trifluoroethanol (3 mL). Trimethylchlorosilane (0.1 mL) was added, and the reaction was carried out at room temperature for 2 hours. LCMS was used to monitor the completion of the reaction. The solvent was evaporated under reduced pressure. The residue was adjusted to weakly basic with saturated sodium bicarbonate solution, and then extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by pre-HPLC to obtain the title compound (4.79 mg).

[0268]

[0269] MS(ESI) m / z [M+H] + = 627.3.

[0270] 11H NMR (400 MHz, DMSO-d6) δ 8.56 (s, 1H), 7.58 - 7.56 (d, J = 7.9 Hz, 1H), 7.49 - 7.44 (dd, J = 8.8, 4.7 Hz, 1H), 7.40 - 7.34 (m, 3H), 7.14 - 7.10 (m, 2H), 7.02 - 6.98 (t, J = 7.4 Hz, 1H), 4.78 (m, 2H), 4.37 - 4.30 (m, 3H), 4.21 - 4.13 (m, 3H), 3.74 (s, 3H), 3.70 - 3.63 (p, J = 6.6 Hz, 1H), 3.41 - 3.37 (m, 1H), 3.05 (m, 1H), 2.82 - 2.78 (dd, J = 14.0, 5.8 Hz, 1H), 2.69 - 2.63 (m, 1H), 2.40 - 2.35 (dd, J = 15.5, 4.5 Hz, 1H), 2.28 - 2.23 (dd, J = 15.5, 8.0 Hz, 1H), 1.65 (br, 2H), 1.09 - 1.03 (m, 4H), 0.99 - 0.89 (m, 3H), 0.72 (m, 3H).

[0271] Example 28

[0272] (R)-2-((5-(5-(3-Amino-4-(1-cyclopropyl-1H-indol-3-yl)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0273] Synthetic route:

[0274]

[0275] Step 1: Preparation of tert-butyl (R)-(1-(1-cyclopropyl-1H-indol-3-yl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamate

[0276] (R)-(4-(5-(6-(2-(Ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1-(1H-indol-3-yl)-4-oxobutan-2-yl)carbamic acid tert-butyl ester (80 mg, Refer to Example 6), copper acetate (44 mg), cyclopropylboronic acid (19 mg), 4-dimethylaminopyridine (27 mg) were mixed in toluene and reacted at 110 °C for 3 h under nitrogen protection. LCMS monitoring showed that the main product was formed with some starting materials remaining. Copper acetate (44 mg) and cyclopropylboronic acid (19 mg) were added and the reaction was continued overnight. LCMS monitoring showed that the reaction was complete. The reaction mixture was cooled to room temperature, filtered through diatomaceous earth, and the filtrate was concentrated and purified by column chromatography to obtain the title compound (35 mg).

[0277]

[0278] MS(ESI)m / z[M+H] + = 753.3.

[0279] Step 2: Preparation of (R)-2-((5-(5-(3-amino-4-(1-cyclopropyl-1H-indol-3-yl)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0280] tert-Butyl (R)-(1-(1-cyclopropyl-1H-indol-3-yl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamate (35 mg) was added to trifluoroethanol (3 mL), and trimethylchlorosilane (0.1 mL) was added. After reacting at room temperature for 2 h, LCMS monitoring showed that the reaction was complete. The solvent was evaporated under reduced pressure, and the residue was adjusted to weak alkalinity with saturated sodium bicarbonate solution and then extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by pre-HPLC to obtain the title compound (6.32 mg).

[0281]

[0282] MS(ESI)m / z[M+H] + = 653.3.

[0283] 11H NMR (400 MHz, DMSO-d6) δ 8.56 (s, 1H), 7.58 - 7.56 (d, J = 7.9 Hz, 1H), 7.51 - 7.45 (m, 2H), 7.40 - 7.34 (m, 2H), 7.16 - 7.12 (m, 2H), 7.04 - 7.01 (t, J = 7.4 Hz, 1H), 4.78 (m, 2H), 4.37 - 4.06 (m, 7H), 3.70 - 3.63 (p, J = 6.6 Hz, 1H), 3.06 - 3.01 (s, 1H), 2.80 - 2.75 (dd, J = 14.0, 5.8 Hz, 1H), 2.67 - 2.60 (m, 1H), 2.40 - 2.35 (dd, J = 15.5, 4.6 Hz, 1H), 2.28 - 2.22 (dd, J = 15.5, 7.9 Hz, 1H), 1.75 (br, 2H), 1.10 - 0.89 (m, 12H), 0.72 (m, 3H).

[0284] Example 29

[0285] (R)-2-((5-(5-(4-(Benzo[b]thiophen-3-yl)-3-(methylamino)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0286] Synthetic route:

[0287]

[0288] Step 1: Preparation of tert-butyl (R)-(1-(benzo[b]thiophen-3-yl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)(methyl)carbamate

[0289] tert-Butyl (R)-(1-(benzo[b]thiophen-3-yl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamate (60 mg, refer to Example 3) was added to dry tetrahydrofuran (2 mL). Sodium hydride (6 mg, Purity 60%) was added under an ice-water bath. Subsequently, methyl iodide (24 mg) was added. The reaction mixture was warmed to room temperature and stirred for 2 h. The reaction was monitored by LCMS until completion. The reaction was quenched with saturated ammonium chloride solution, and then extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (40 mg).

[0290]

[0291] MS(ESI) m / z [M+H] + = 743.3。

[0292] Step 2: Preparation of (R)-2-((5-(5-(4-(benzo[b]thiophen-3-yl)-3-(methylamino)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0293] tert-Butyl (R)-(1-(benzo[b]thiophen-3-yl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)(methyl)carbamate (40 mg) was added to trifluoroethanol (3 mL). Trimethylchlorosilane (0.1 mL) was added. After reacting at room temperature for 2 h, the reaction was monitored by LCMS until completion. The solvent was evaporated under reduced pressure. The residue was adjusted to weakly basic with saturated sodium bicarbonate solution, and then extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by pre-HPLC to obtain the title compound (17.44 mg).

[0294]

[0295] MS(ESI) m / z [M+H] + = 644.3。

[0296] 11H NMR (400 MHz, DMSO-d6) δ 8.56 (s, 1H), 7.97 - 7.91 (m, 2H), 7.50 - 7.44 (m, 2H), 7.42 - 7.33 (m, 4H), 4.78 - 4.74 (m, 2H), 4.36 - 4.33 (m, 2H), 4.26 - 4.12 (m, 4H), 3.70 - 3.63 (dt, J = 13.1, 6.4 Hz, 1H), 3.26 - 3.19 (t, J = 6.3 Hz, 1H), 3.07 - 3.05 (m, 1H), 3.01 - 2.89 (m, 2H), 2.48 - 2.44 (m, 1H), 2.31 (s, 3H), 2.28 - 2.33 (dd, J = 15.6, 6.2 Hz, 1H), 1.71 (br, 1H), 1.09 - 1.03 (m, 4H), 0.99 - 0.90 (m, 3H), 0.72 (m, 3H).

[0297] Example 30

[0298] (R)-2-((5-(3-Amino-4-(4-hydroxyphenyl)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0299] Synthetic route:

[0300]

[0301] Step 1: Preparation of (9H-fluoren-9-yl)methyl (R)-(1-(4-(tert-butoxy)phenyl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamate

[0302] (R)-3-(((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-4-(4-(tert-butoxy)phenyl)butanoic acid (60 mg) was added to N,N-dimethylformamide (2 mL), followed by N,N-diisopropylethylamine (45 mg), 1-hydroxybenzotriazole (30 mg), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (40 mg). After reacting at room temperature for half an hour, N-ethyl-5-fluoro-N-isopropyl-2-((5-(3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)benzamide (50 mg) was added, and the reaction was carried out overnight at room temperature. The reaction was monitored by LCMS until completion, quenched with water, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (50 mg).

[0303]

[0304] MS(ESI) m / z [M+H] + = 868.3。

[0305] Step 2: Preparation of (9H-Fluoren-9-yl)methyl (R)-(4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1-(4-hydroxyphenyl)-4-oxobutan-2-yl)carbamate

[0306] (9H-Fluoren-9-yl)methyl (R)-(1-(4-(tert-butoxy)phenyl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamate (50 mg) was added to ethyl acetate (3 mL), 4M hydrochloric acid (0.5 mL) was added, and the reaction was carried out overnight at room temperature. The reaction was monitored by LCMS until completion, adjusted to about 4 with saturated sodium bicarbonate solution, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to obtain the crude product (50 mg).

[0307]

[0308] MS(ESI) m / z [M+H] + = 812.3。

[0309] Preparation of (R)-2-((5-(5-(3-amino-4-(4-hydroxyphenyl)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0310] (9H-Fluoren-9-yl)methyl (R)-(4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1-(4-hydroxyphenyl)-4-oxobutan-2-yl)carbamate (50 mg crude) was added to dichloromethane (3 mL), piperidine (0.5 mL) was added, and after reacting at room temperature for 4 hours. The reaction was monitored by LCMS to be complete, water was added, and the mixture was extracted with dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by pre-HPLC to obtain the title compound (29.54 mg).

[0311]

[0312] MS(ESI) m / z [M+H] + = 590.3

[0313] 1 1H NMR (400 MHz, DMSO-d6) δ 9.10 (br, 1H), 8.49 (s, 1H), 7.42 - 7.38 (mz, 1H), 7.33 - 7.27 (m, 2H), 6.93 - 6.91 (d, J = 7.9 Hz, 2H), 6.62 - 6.60 (d, J = 7.9 Hz, 2H), 4.71 (m, 2H), 4.30 - 4.06 (m, 6H), 3.63 - 3.56 (p, J = 6.6 Hz, 1H), 3.17 (m, 1H), 3.00 - 2.94 (m, 1H), 2.51 - 2.44 (m, 1H), 2.42 - 2.38 (m, 1H), 2.27 - 2.09 (m, 2H), 1.51 (br, 2H), 1.03 - 0.96 (m, 4H), 0.92 - 0.82 (m, 3H), 0.65 (m, 3H).

[0314] Using N-ethyl-5-fluoro-N-isopropyl-2-((5-(3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)benzamide (i.e., the title compound in Step 5 of Example 1) as the starting material, the intermediates shown in Table 2 and reaction reagents available through commercial channels were used, and the compounds of Examples 31 - 32 were obtained by referring to the synthetic route of Example 30, as specifically shown in Table 5.

[0315] Table 5 Examples 31 - 32

[0316]

[0317]

[0318] Example 33

[0319] (R)-2-((5-(5-(4-(3-(1H-tetrazol-5-yl)phenyl)-3-aminobutanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0320] Synthesis route:

[0321]

[0322] Step 1: Preparation of methyl (R)-4-(3-cyanophenyl)-3-((tert-butoxycarbonyl)amino)butyrate

[0323] Dissolve (R)-4-(3-cyanophenyl)-3-((tert-butoxycarbonyl)amino)butyric acid (200 mg) in N,N-dimethylformamide, add potassium bicarbonate (84 mg) and methyl iodide (55 μL) at room temperature, react at room temperature for 4 h, monitor the reaction by LCMS until completion, quench with water, extract with ethyl acetate, combine the organic phases, wash with saturated brine, dry over anhydrous sodium sulfate, and concentrate to obtain the title compound (220 mg).

[0324]

[0325] MS(ESI) m / z [M + H] + = 319.1

[0326] Step 2: Preparation of methyl (R)-4-(3-(1H-tetrazol-5-yl)phenyl)-3-((tert-butoxycarbonyl)amino)butyrate

[0327] Add methyl (R)-4-(3-cyanophenyl)-3-((tert-butoxycarbonyl)amino)butyrate (220 mg) to N,N-dimethylformamide (3 mL), add ammonium chloride (85 mg) and sodium azide (82 mg) at room temperature, react at 115 °C for two days, monitor the reaction by LCMS, with a small amount of raw material remaining, mainly the product. Cool to room temperature, quench with sodium hypochlorite solution, evaporate the solvent under reduced pressure, and purify the residue by column chromatography to obtain the title compound (220 mg).

[0328]

[0329] MS(ESI) m / z [M+H] + = 362.1。

[0330] Step 3: Preparation of (R)-4-(3-(1H-tetrazol-5-yl)phenyl)-3-((tert-butoxycarbonyl)amino)butyric acid

[0331] Methyl (R)-4-(3-(1H-tetrazol-5-yl)phenyl)-3-((tert-butoxycarbonyl)amino)butyrate (100 mg) was added to a mixed solvent of tetrahydrofuran (2 mL), methanol (0.6 mL) and water (0.6 mL). Lithium hydroxide monohydrate (28 mg) was added. After reacting at room temperature for 2 hours, LCMS monitored the disappearance of the raw material. The pH of the system was adjusted to 7 with 2M dilute hydrochloric acid, and the solvent was distilled off under reduced pressure and concentrated to obtain the crude title compound.

[0332]

[0333] MS(ESI) m / z [M-H] - = 348.1。

[0334] Step 4: Preparation of tert-butyl (R)-(1-(3-(1H-tetrazol-5-yl)phenyl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamate

[0335] The crude (R)-4-(3-(1H-tetrazol-5-yl)phenyl)-3-((tert-butoxycarbonyl)amino)butyric acid obtained in Step 3 was added to N,N-dimethylformamide (2 mL). N,N-Diisopropylethylamine (0.06 mL), 1-hydroxybenzotriazole (30 mg), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (40 mg) were added in sequence. After reacting at room temperature for half an hour, N-ethyl-5-fluoro-N-isopropyl-2-((5-(3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)benzamide (50 mg) was added, and the reaction was carried out overnight at room temperature. LCMS monitored the completion of the reaction, quenched with water, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (25 mg).

[0336]

[0337] MS(ESI) m / z [M+H] + = 742.3。。

[0338] Preparation of (R)-2-((5-(5-(4-(3-(1H-tetrazol-5-yl)phenyl)-3-aminobutanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0339] tert-Butyl (R)-(1-(3-(1H-tetrazol-5-yl)phenyl)-4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-oxobutan-2-yl)carbamate (25 mg) was added to trifluoroethanol (3 mL), and trimethylchlorosilane (0.1 mL) was added. After reacting at room temperature for 2 hours, the reaction was monitored by LCMS until completion. The solvent was evaporated under reduced pressure. The residue was adjusted to weak alkalinity with saturated sodium bicarbonate solution, and then the solvent was evaporated under reduced pressure. The residue was purified by pre-HPLC to obtain the title compound (4.5 mg).

[0340]

[0341] MS(ESI) m / z [M+H] + = 642.3..

[0342] 1 1H NMR (400 MHz, DMSO-d6) δ 8.56 (s, 1H), 7.90 - 7.87 (m, 2H), 7.52 - 7.46 (m, 1H), 7.40 - 7.33 (m, 3H), 7.15 - 7.13 (d, J = 7.5 Hz, 1H), 4.76 - 4.72 (m, 2H), 4.35 (m, 2H), 4.25 - 4.15 (m, 4H), 3.81 - 3.74 (q, J = 6.7 Hz, 1H), 3.69 - 3.63 (p, J = 6.7 Hz, 1H), 3.04 - 2.99 (m, 3H), 2.92 - 2.87 (m, 1H), 2.67 - 2.54 (m, 1H), 1.09 - 1.02 (m, 4H), 0.99 - 0.85 (m, 3H), 0.71 (m, 3H).

[0343] Example 34

[0344] (R)-2-((5-(5-(3-amino-4-(2-methyl-1-oxoisoindolin-4-yl)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0345] Synthetic route:

[0346]

[0347] Step 1: Preparation of Methyl (R)-3-((tert-butoxycarbonyl)amino)-4-(2-methyl-1-oxoisoindolin-4-yl)butyrate

[0348] Zinc powder (171 mg) was added to dry N,N-dimethylformamide (4 mL) under nitrogen protection. Iodine (66 mg) was added at room temperature. After the color of iodine disappeared after five minutes, a DMF solution (0.5 mL) of trimethylchlorosilane (50 μL) was added. The reaction was carried out at room temperature for half an hour. A solution of methyl (S)-3-((tert-butoxycarbonyl)amino)-4-iodobutyrate (300 mg) in N,N-dimethylformamide (1 mL) was added dropwise. The reaction continued at room temperature for five minutes. 4-Bromo-2-methylisoindolin-1-one (236 mg), 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (36 mg), and tris(dibenzylideneacetone)dipalladium (II) (79 mg) were added. The temperature was raised to 70 °C and the reaction was carried out for 4 h. LCMS monitored the formation of the product and the raw materials were consumed completely. The reaction was quenched with saturated sodium bicarbonate solution at room temperature, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (250 mg).

[0349]

[0350] MS(ESI) m / z (M+Na) + = 385.1

[0351] Step 2: Preparation of (R)-3-((tert-butoxycarbonyl)amino)-4-(2-methyl-1-oxoisoindolin-4-yl)butyric acid

[0352] Methyl (R)-3-((tert-butoxycarbonyl)amino)-4-(2-methyl-1-oxoisoindolin-4-yl)butyrate (250 mg) was added to a mixed solvent of tetrahydrofuran (3 mL), methanol (3 mL), and water (1 mL). Lithium hydroxide monohydrate (60 mg) was added at room temperature. After 2 h, LCMS monitored that the reaction was complete. Water was added to the system, and it was extracted three times with ethyl acetate. The organic phases were discarded. The pH of the aqueous phase was adjusted to 2 with 2 M dilute hydrochloric acid, and it was extracted with a dichloromethane / methanol mixed solvent. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to obtain the title compound (120 mg crude product).

[0353]

[0354] MS(ESI) m / z [M+H] + = 349.1

[0355] Step 3: Preparation of tert-butyl (R)-(4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1-(2-methyl-1-oxoisoindol-4-yl)-4-oxobutan-2-yl)carbamate

[0356] Crude (R)-3-((tert-butoxycarbonyl)amino)-4-(2-methyl-1-oxoisoindolin-4-yl)butanoic acid (120 mg) was added to N,N-dimethylformamide (2 mL). Subsequently, N,N-diisopropylethylamine (0.1 mL), 1-hydroxybenzotriazole (80 mg), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (100 mg) were added successively. After reacting at room temperature for half an hour, N-ethyl-5-fluoro-N-isopropyl-2-((5-(3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)benzamide (90 mg) was added, and the reaction was carried out overnight at room temperature. The reaction was monitored by LCMS until completion, quenched with water, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (110 mg).

[0357]

[0358] MS(ESI) m / z [M+H] + = 743.3

[0359] Step 4: Preparation of (R)-2-((5-(5-(3-amino-4-(2-methyl-1-oxoisoindolin-4-yl)butanoyl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0360] tert-Butyl (R)-(4-(5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)-3,4,5,6-tetrahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1-(2-methyl-1-oxoisoindol-4-yl)-4-oxobutan-2-yl)carbamate (110 mg) was added to trifluoroethanol (3 mL), and trimethylchlorosilane (0.2 mL) was added. After reacting at room temperature for 2 hours, the reaction was monitored by LCMS until completion. The solvent was evaporated under reduced pressure, the residue was adjusted to weakly basic with saturated sodium bicarbonate solution, concentrated, and purified by pre-HPLC to obtain the title compound (56.13 mg).

[0361]

[0362] MS(ESI) m / z [M+H] + = 643.3

[0363] 1 H NMR (400 MHz, DMSO-d6) δ 8.56 (s, 1H), 7.52 - 7.35 (m, 6H), 4.79 (m, 2H), 4.48 (s, 2H), 4.37 - 4.32 (m, 4H), 4.19 - 4.13 (m, 2H), 3.70 - 3.63 (p, J = 6.7 Hz, 1H), 3.39 (m, 1H), 3.08 (s, 3H), 3.01 (m, 1H), 2.82 - 2.77 (dd, J = 13.4, 5.4 Hz, 1H), 2.65 - 2.59 (m, 1H), 2.41 - 2.29 (m, 2H), 1.67 (br, 2H), 1.10 - 1.03 (m, 4H), 0.99 - 0.90 (m, 3H), 0.74 - 0.70 (m, 3H).

[0364] Using methyl (S)-3-((tert-butoxycarbonyl)amino)-4-iodobutyrate as the starting material, and using halogenating reagents that are commercially available or prepared in the preparation examples as reactants, the compounds of Examples 35 - 64 were obtained by referring to the synthetic route of Example 34, as specifically shown in Table 6 below.

[0365] Table 6 Examples 35 - 64

[0366]

[0367]

[0368]

[0369]

[0370] [[ID=3P1]]

[0371]

[0372]

[0373]

[0374]

[0375]

[0376] It should be noted that there seems to be a mislabeling in your original text where " " is likely a typo as you have "3P1" in the translation. I've left it as is based on the instructions but you may want to double-check the original.

[0377] Example 65

[0378] N-Ethyl-2-((5-((((1r,4r)-4-(ethylsulfonamido)cyclohexyl)methyl)hexahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-5-fluoro-N-isopropylbenzamide

[0379]

[0380] Step 1: Preparation of tert-Butyl 5-(3,6-dichloro-1,2,4-triazin-5-yl)hexahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate

[0381] Add tert-butyl hexahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate (5 g) to dichloromethane (50 mL), dropwise add triethylamine (7.24 g), and then add trichloro-1,2,4-triazine (4.4 g) in batches. React at room temperature for 1 h. Monitor the reaction by LCMS until completion, quench with water, extract with dichloromethane three times, combine the organic phases, dry over anhydrous sodium sulfate, concentrate, and purify by column chromatography to obtain the title compound (7.5 g).

[0382]

[0383] MS(ESI) m / z(M+H) + = 360.1.

[0384] Step 2: Preparation of tert-Butyl 5-(3-chloro-6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)hexahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate

[0385] Add tert-butyl 5-(3,6-dichloro-1,2,4-triazin-5-yl)hexahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate (7.5 g) to tetrahydrofuran (50 mL), add DBU (9.5 g), N-ethyl-5-fluoro-2-hydroxy-N-isopropylbenzamide (4.7 g), and react at 50 °C overnight. Monitor the reaction by LCMS until completion, stop the reaction, cool to room temperature, quench with water, extract with ethyl acetate, combine the organic phases, dry over anhydrous sodium sulfate, concentrate, and purify by column chromatography to obtain the title compound (8.74 g).

[0386]

[0387] MS(ESI) m / z(M+H) + = 549.2.

[0388] Step 3: Preparation of tert-butyl 5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)hexahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate

[0389] tert-Butyl 5-(3-chloro-6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)hexahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate (8.74 g) was added to tetrahydrofuran (80 mL). Sodium borohydride (3.02 g) was added portionwise, followed by tetramethylethylenediamine (3.70 g) and 1,1'-bis(diphenylphosphino)ferrocene palladium(II) dichloride (0.58 g). The reaction was carried out overnight at room temperature under nitrogen protection. The reaction was monitored by LCMS until completion, quenched with water, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (5.12 g).

[0390]

[0391] MS(ESI) m / z(M+H) + = 515.2

[0392] Step 4: Preparation of N-ethyl-5-fluoro-2-((5-(hexahydropyrrolo[3,4-c]pyrrole-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-isopropylbenzamide

[0393] tert-Butyl 5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)hexahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylate (5.12 g) was added to trifluoroethanol (20 mL). Trimethylchlorosilane (3.8 mL) was added, and the reaction was carried out at room temperature for 2 h. The reaction was monitored by LCMS until completion, the solvent was evaporated under reduced pressure, the residue was adjusted to alkaline with saturated sodium bicarbonate solution, and after lyophilizing the solvent, it was purified by column chromatography to obtain the title compound (4.0 g).

[0394]

[0395] MS(ESI) [M+H] + = 415.3

[0396] Step 5: Preparation of tert-butyl ((1r,4r)-4-((5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)hexahydropyrrolo[3,4-c]pyrrole-2(1H)-yl)methyl)cyclohexyl)carbamate

[0397] N-Ethyl-5-fluoro-2-((5-(hexahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-isopropylbenzamide (0.27 g) was added to methanol (5 mL), and tert-butyl ((1r,4r)-4-formylcyclohexyl)carbamate (0.1 g), sodium cyanoborohydride (0.08 g) were added. The reaction was carried out at room temperature for 3 h under nitrogen protection. The reaction was monitored by LCMS until completion. After evaporating the methanol solvent under reduced pressure, water was added to the reaction system, and the mixture was extracted with dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (190 mg).

[0398]

[0399] MS(ESI)[M+H] + = 626.3.

[0400] Step 6: Preparation of 2-((5-(5-((1r,4r)-4-aminocyclohexyl)methyl)hexahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide

[0401] Tert-butyl ((1r,4r)-4-((5-(6-(2-(ethyl(isopropyl)carbamoyl)-4-fluorophenoxy)-1,2,4-triazin-5-yl)hexahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)methyl)cyclohexyl)carbamate (190 mg) was added to trifluoroethanol (5 mL), and trimethylchlorosilane (0.2 mL) was added. After reacting at room temperature for 2 h, the reaction was monitored by LCMS until completion. The solvent was evaporated under reduced pressure, and the residue was adjusted to weak alkalinity with saturated sodium bicarbonate solution, then extracted with dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain the title compound (157 mg).

[0402]

[0403] MS(ESI)[M+H] + = 526.3.

[0404] Step 7: Preparation of N-ethyl-2-((5-((((1r,4r)-4-(ethylsulfonamido)cyclohexyl)methyl)hexahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-5-fluoro-N-isopropylbenzamide

[0405] 2-((5-(5-((1R,4R)-4-aminocyclohexyl)methyl)hexahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-1,2,4-triazin-6-yl)oxy)-N-ethyl-5-fluoro-N-isopropylbenzamide (157 mg) was added to dichloromethane (5 mL), triethylamine (90 mg) was added dropwise, and then ethylsulfonyl chloride (38 mg) was added. The reaction was carried out at room temperature for 1 h. The reaction was monitored by LCMS until completion, quenched with water, extracted three times with dichloromethane, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by pre-HPLC to obtain the title compound (60.77 mg).

[0406]

[0407] MS(ESI) m / z (M+H) + = 618.3.

[0408] 1 H NMR (400 MHz, DMSO-d6) δ 7.45 - 7.39 (m, 1H), 7.38 - 7.34 (m, 1H), 7.33 (dt, J = 4.6, 1.5 Hz, 1H), 6.99 (d, J = 7.7 Hz, 1H), 4.14 (s, 1H), 2.95 (q, J = 7.4 Hz, 4H), 2.89 - 2.81 (m, 2H), 2.54 (s, 1H), 2.42 (d, J = 6.7 Hz, 2H), 2.15 (d, J = 7.2 Hz, 2H), 1.86 - 1.80 (m, 2H), 1.74 (d, J = 13.0 Hz, 2H), 1.34 - 1.02 (m, 12H), 1.02 - 0.65 (m, 9H).

[0409] Experimental Example 1: Test for the inhibitory activity of Menin-MLL protein interaction

[0410] (1) Test for the inhibitory activity of Menin-MLL protein interaction

[0411] The IC value of the test compound for inhibiting the Menin-MLL protein interaction was detected by the method of Fluorescence Polarization. 50 value.

[0412] The specific steps are as follows: The compound stock solution (10 mM, prepared with DMSO) was serially diluted three-fold with DMSO to ten concentrations of 10000.00, 3333.33, 1111.11, 370.37, 123.46, 41.15, 13.72, 4.57, 1.52, and 0.51 nM. Using an ECHO665 Series Acoustic Liquid Handler (BECKMAN Inc.), 50 nL of the test compounds at different concentrations (10 gradient concentrations) and DMSO solution without the compound (negative control wells) were respectively taken and transferred to a 384-well plate, and centrifuged at 1000 rpm for standby. Using an I.DOT (DISPENDIX Inc.), 5 μL of Menin (ICE Inc., Cat No. E2208F-H15H) was aspirated and added to each assay well, and incubated at 25 °C for 10 minutes. Using an I.DOT, 5 μL of FITC-MLL4-43 (Genscrip Inc.) was aspirated and added to each assay well, centrifuged at 1000 rpm and incubated for 60 minutes. The FP signal was measured using a PHERAstar FSX multimode microplate reader (BMGLabtech Inc.), and the data was processed.

[0413] The IC50 of the compound was fitted with a non-linear regression equation: Inhibition% = (negative control well signal - compound well signal) / (negative control well signal - background signal) * 100% (the background signal was the signal value detected in the assay well containing only 10 μM SNDX-5613). Using the log value of the compound concentration as the X-axis and the percentage inhibition rate (Inhibition%) as the Y-axis, a dose-effect curve was fitted to obtain the IC50 value of each compound for inhibiting the Menin-MLL protein interaction. The experimental results are shown in Table 7.

[0414] Table 7 IC of the compounds of the present invention for inhibiting the Menin-MLL protein interaction 50 value

[0415]

[0416]

[0417] Note: In Table 7, "-" indicates not yet detected. The numbers under "Compound Number" represent the compounds in the corresponding examples. SNDX5613 refers to N-ethyl-2-((4-(7-(((1r,4r)-4-(ethylsulfonylamino)cyclohexyl)methyl)-2,7-diazaspiro[3.5]nonan-2-yl)pyrimidin-5-yl)oxy)-5-fluoro-N-isopropylbenzamide (the same below), purchased from Shanghai LouLan Biotechnology Co., Ltd., and the structural formula is as follows:

[0418]

[0419] The above experiments show that the compounds of the present invention have excellent activity in inhibiting the interaction between Menin and MLL proteins.

[0420] (2) Test for the inhibitory activity of the interaction between mutant Menin-M327I, Menin-T349M and MLL proteins

[0421] The method of Fluorescence Polarization was used to detect the IC 50 value of the test compound in inhibiting the interaction between Menin-M327I, Menin-T349M and MLL proteins.

[0422] The specific steps are as follows: The compound stock solution (10 mM, prepared with DMSO) was serially diluted three-fold with DMSO to ten concentrations of 10000.00, 3333.33, 1111.11, 370.37, 123.46, 41.15, 13.72, 4.57, 1.52, 0.51 nM. 50 nL of different concentrations of the test compound (10 gradient concentrations) and DMSO solution without compound (negative control well) were respectively taken by ECHO665 Series Acoustic Liquid Handlers (BECKMAN Inc.) and transferred to a 384-well plate, and centrifuged at 1000 rpm for standby. 5 μL of Menin-M327I (ICE Inc. CatNo.A130412011) and Menin-T349M (ICE Inc. Cat No.A130413011) were respectively aspirated into each assay well using I.DOT (DISPENDIX Inc.) and incubated at 25 °C for 10 minutes. 5 μL of FITC-MLL4-43 (Genscrip Inc.) was aspirated into each assay well using I.DOT, centrifuged at 1000 rpm and incubated for 60 minutes. The FP signal was measured using a PHERAstar FSX multimode microplate reader (BMG Labtech Inc.), and the data was processed.

[0423] The IC of the compound was fitted with a non-linear regression equation 50 : Inhibition% = (signal of negative control well - signal of compound well) / (signal of negative control well - background signal) * 100% (the background signal is the signal value detected in the assay well containing only 100 μM SNDX-5613). Taking the log value of the compound concentration as the X-axis and the percentage inhibition rate (Inhibition%) as the Y-axis, a dose-effect curve was fitted to obtain the IC of each compound in inhibiting the interaction between Menin-M327I, Menin-T349M and MLL proteins50 value. The experimental results are shown in Table 8.

[0424] Table 8 IC50 values of the compounds of the present invention for inhibiting the interaction between Menin mutant protein and MLL protein

[0425]

[0426] Note: In Table 8, the numbers under "Compound Number" represent the compounds in the corresponding examples.

[0427] The above tests show that the compounds of the present invention have excellent activity in inhibiting the interaction between Menin mutant protein and MLL protein, and thus have good prospects for resisting drug resistance.

[0428] Experimental Example 2: Cell proliferation inhibition experiment

[0429] (1) Cell seeding:

[0430] Take out the cells from the incubator and place them on the operating table. Gently pipette and mix well, and count with CounterStar.

[0431] Dilute the cells to the required density with fresh complete medium. MV-4-11 (cell source: Nanjing Kebai, product number: CBP60522), medium: RPMI1640 (containing HEPES) (BOSTER, product number: PYG0122) + 10% FBS (GIBCO, product number: 10099-141C) + 1% P / S (HyClone, product number: SV30010), seeding density: 1×10^4 cells / well, 100 μL / well.

[0432] Use an electric multi-channel pipette to aspirate 100 μL of each of the above cell suspensions into a 96-well plate.

[0433] (2) Preparation of compounds:

[0434] Dilute the stock solution of the compound from 10 mM to 4000, 1200, 400, 120, 40, 12, 4, 1.2, 0 μM with DMSO, where 0 μM is used as the control well.

[0435] After thorough mixing, use an electric multi-channel pipette to take out 1.3 μL and add it to 258.7 μL of medium. At this time, it is diluted 100 times, and the DMSO content is 0.5%.

[0436] After thoroughly mixing using the multi-gun mixing mode, take out the cells plated above. Set two replicate wells for each concentration of each compound, and add 100 μL of the compound diluted in Step 2 to each of the two replicate wells. The final concentrations of the compounds are: 10, 3, 1, 0.3, 0.1, 0.03, 0.01, 0.003, 0 μM. At this time, there is 200 μL of culture medium in each well of the cell culture plate, and the DMSO content is 0.25%. The well containing only 0.25% DMSO (compound concentration is 0) is the control well.

[0437] Put the cells back into the incubator at 37 °C and 5% CO2 and continue culturing. After adding the compounds and treating for 3 days, conduct the detection.

[0438] (3) CTG detection:

[0439] When the culturing reaches the specified time, take out the cells, aspirate a part of the culture medium so that 50 μL of culture medium remains in each well, and add 50 μL / well of CTG reagent (cellcounting-Lite2.0, Vazyme, DD1101-02) using a multi-gun.

[0440] Incubate on a shaker at room temperature for 15 min and let it stand at room temperature to equilibrate for 15 min.

[0441] Use a multi-functional microplate reader for detection.

[0442] (4) Data analysis:

[0443] Calculate the cell viability, Cell viability% = As / Ac × 100%. As: test well (culture medium containing cells, CTG, and the compound to be tested), Ac: control well (culture medium containing cells, CTG, without the compound to be tested).

[0444] Use the log value of the compound concentration as the X-axis and the cell viability (Cell viability%) as the Y-axis to fit the dose-response curve to obtain the IC 50 value. The specific results are shown in Table 9.

[0445] Table 9 Proliferation inhibition activity of the compounds of the present invention against MV-4-11 cells

[0446]

[0447]

[0448] In Table 9, "-" indicates no content. The numbers under "Compound Number" represent the compounds in the corresponding examples. The test results show that the compounds of the present application have excellent inhibitory activity against MV-4-11 cells.

Claims

1. A compound represented by formula I, its pharmaceutically acceptable salt, hydrate, isomer, prodrug or a mixture thereof, In formula I: m = 0, 1 or 2; n = 0, 1, 2 or 3; R1 is selected from hydrogen, hydroxy, halogen, cyano, C1-C3 alkyl, C1-C3 alkoxy or C1-C3 haloalkyl; R2 and R3 are each independently selected from hydrogen, C1-C3 alkyl, C1-C3 alkoxy, C1-C3 haloalkyl, RxCO-, sulfonyl or phosphonyl, where Rx is alkyl or alkenyl; R4 and R5 are each independently selected from hydrogen, hydroxy, halogen, C1-C3 alkyl, C1-C3 alkoxy or C1-C3 haloalkyl; R6 is selected from a substituted or unsubstituted 5-7-membered aryl, 5-7-membered heteroaryl, 9-16-membered saturated or partially unsaturated cycloalkyl, 9-16-membered saturated or partially unsaturated heterocycloalkyl, 9-16-membered fused aryl, 9-16-membered fused heteroaryl.

2. The compound, pharmaceutically acceptable salt, hydrate, isomer, prodrug or mixture thereof according to claim 1, characterized in that, R1 is selected from fluorine, chlorine, bromine, iodine, cyano, hydroxy, amino, methyl, ethyl, n-propyl, isopropyl, methoxy, ethoxy, chloromethyl, trifluoromethyl or perfluoroethyl.

3. The compound, pharmaceutically acceptable salt, hydrate, isomer, prodrug or mixture thereof according to any one of claims 1 or 2, characterized in that, Rx is C1-C3 alkyl or C2-C4 alkenyl.

4. The compound, pharmaceutically acceptable salt, hydrate, isomer, prodrug or mixture thereof according to claim 3, characterized in that, R2 is hydrogen, and R3 is selected from hydrogen, methyl, ethyl, n-propyl, isopropyl, methoxy, ethoxy, chloromethyl, trifluoromethyl, perfluoroethyl, acetyl, acryloyl, dimethylphosphonyl, methanesulfonyl or ethanesulfonyl.

5. The compound, pharmaceutically acceptable salt, hydrate, isomer, prodrug or mixture thereof according to claim 1, characterized in that, R4 is hydrogen, and R5 is selected from hydrogen, hydroxy, fluorine, chlorine, bromine, iodine, cyano, methyl, ethyl, n-propyl, isopropyl, methoxy, ethoxy, chloromethyl, trifluoromethyl or perfluoroethyl.

6. The compound, pharmaceutically acceptable salt, hydrate, isomer, prodrug or mixture thereof according to claim 1, characterized in that, R6 is selected from a substituted or unsubstituted 5-7-membered aryl, 5-7-membered heteroaryl, 9-12-membered saturated or partially unsaturated cycloalkyl, 9-12-membered saturated or partially unsaturated heterocycloalkyl, 9-12-membered fused aryl, 9-12-membered fused heteroaryl, where the backbone atoms of the heterocycloalkyl or heteroaryl may optionally contain 1-3 heteroatoms selected from N, O, S.

7. The compound, pharmaceutically acceptable salt, hydrate, isomer, prodrug or mixture thereof according to claim 6, characterized in that, R6 may be selected from the following substituted or unsubstituted cyclic groups: furyl, thienyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl; phenyl, pyridyl, pyrimidinyl, pyrazinyl, 1,2,3-triazinyl, 1,2,4-triazinyl, 1,3,5-triazinyl; indolyl, benzimidazolyl, benzofuryl, benzothiazolyl, benzisothiazole, benzoxazolyl, benzisoxazole, benzothienyl, indazolyl, pyrrolo[1,2-a]pyridine, imidazo[1,2-a]pyridine, imidazo[1,5-a]pyridine, pyrazolo[1,5-a]pyridine, isoindolin-1-one, 3,4-dihydroisoquinolin-1(2H)-one; naphthyl, quinolinyl, isoquinolinyl, naphthyridinyl, pyridinediazinyl, benzotriazinyl, benzoxatriazinyl, benzopyranyl, benzogamma-pyrone, tetrahydroisoquinolinyl, tetrahydronaphthyl, 3,4-dihydro-2H-1,4-benzoxazinyl, 2H-1,4-benzoxazin-3(4H)-one or 1,2,3,5,6,7-hexahydro-S-dipentacyclophenyl.

8. The compound, pharmaceutically acceptable salt, hydrate, isomer, prodrug or mixture thereof according to claim 6 or 7, characterized in that, R6 is optionally substituted at any possible position by one or more groups selected from oxygen, hydroxyl, amino, carboxyl, cyano, halogen, phosphonyl, sulfonyl, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, RaCO-, C2-C4 alkenyl, C3-C6 cycloalkyl, 3-6 membered heteroalkyl, 5-7 membered aryl or 5-7 membered heteroaryl, wherein Ra is C1-C5 alkyl or Ra is C2-C4 alkenyl.

9. A compound, a pharmaceutically acceptable salt, hydrate, isomer, prodrug or mixture thereof according to any one of claims 1-8, characterized in that, R6 is selected from the following substituted or unsubstituted groups:

10. The compound, pharmaceutically acceptable salt, hydrate, isomer, prodrug or mixture thereof according to claim 9, characterized in that, R6 is optionally substituted at any possible position by one or more groups selected from oxygen, hydroxyl, amino, carboxyl, cyano, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, methoxy, ethoxy, chloromethyl, trifluoromethyl, 2,2,2-trifluoroethyl, perfluoroethyl, formyl, acetyl, acryloyl, methylphosphonyl, dimethylphosphonyl, methanesulfonyl, ethanesulfonyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, furyl, thienyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, triazole, tetrazole, phenyl, pyridyl.

11. The following compounds, their pharmaceutically acceptable salts, hydrates, isomers, prodrugs or mixtures thereof, 12. A pharmaceutical composition, characterized in that, including the compounds, their pharmaceutically acceptable salts, hydrates, isomers, prodrugs or mixtures thereof as described in any one of claims 1-11, and pharmaceutically acceptable excipients.

13. Use of the compounds, their pharmaceutically acceptable salts, hydrates, isomers, prodrugs or mixtures thereof as described in any one of claims 1-11, or the pharmaceutical composition as described in claim 12, for the preparation of a drug for preventing, alleviating or treating diseases associated with the interaction of Menin-MLL protein.

14. The use according to claim 13, characterized in that, Diseases associated with the interaction of Menin-MLL protein include malignancies, diabetes or complications associated with said diseases, wherein malignancies include hematological tumors, lymphomas, solid tumors.

15. The use according to claim 14, wherein Hematological tumors include leukemia and myeloma, including but not limited to acute lymphoblastic leukemia, chronic lymphocytic leukemia, acute myeloid leukemia, chronic myeloid leukemia, acute monocytic leukemia, chronic monocytic leukemia, childhood leukemia, acute granulocytic leukemia, chronic granulocytic leukemia, mixed lineage leukemia, hairy cell leukemia, precursor T-cell lymphoblastic leukemia, large granular lymphocytic leukemia, meningeal leukemia, myelodysplastic syndrome, myeloproliferative diseases, myeloproliferative neoplasia, plasmacytoma, multiple myeloma; Lymphomas include but not limited to cutaneous T-cell lymphoma, lymphoid neoplasm, AIDS-related lymphoma, Hodgkin lymphoma, non-Hodgkin lymphoma or malignant lymphoma; Solid tumors include but not limited to pancreatic cancer, colon cancer, rectal cancer, liver cancer, gastric cancer, glioblastoma, lung cancer, breast cancer, prostate cancer; Related complications include but not limited to leukemic meningitis.

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