Dihydropyrido[3,4-d]pyrimidine compound, preparation method therefor, and use thereof

Abstract

The present invention relates to a dihydropyrido[3,4-d]pyrimidine compound as a multi-target inhibitor of BRG1 / BRM bromodomain and proton-sensing receptor GPR65. The present invention relates to a compound represented by formula (I) or a pharmaceutically acceptable salt, stereoisomer, solvate, or isotopic derivative thereof, wherein R1, R2, R3, X, Y, Z, W, and n are as defined in the description. The present invention further relates to a pharmaceutical composition comprising the compound and a use of the compound and the pharmaceutical composition comprising the compound for preventing and / or treating diseases associated with abnormal activity of BRG1 / BRM and / or proton-sensing receptor GPR65, including tumors, in particular glioblastoma.

Description

Dihydropyrido[3,4-d]pyrimidine compounds, their preparation methods and uses Technical Field

[0001] This invention relates to novel dihydropyrido[3,4-d]pyrimidine compounds or their pharmaceutically acceptable salts, stereoisomers, solvates or isotopic derivatives that can be used as multi-target inhibitors of the BRG1 / BRM bromide domain and the proton-sensing receptor GPR65, pharmaceutical compositions comprising such compounds, and the use of such compounds and pharmaceutical compositions comprising such compounds for the prevention and / or treatment of diseases, including tumors, particularly glioblastoma, associated with aberrant activity of BRG1 / BRM and / or the proton-sensing receptor GPR65. Background Technology

[0002] Glioblastoma (GBM) is the most common malignant primary brain tumor in adults, belonging to the WHO grade IV glioma, the most malignant type, accounting for nearly 75% of all glioma cases. Currently, the annual incidence of glioma in China is high (approximately 4.03 / 100,000), and its 5-year mortality rate is second only to pancreatic and lung cancer among all cancers, with a survival rate of only 4.7%. Traditional GBM treatment involves maximal surgical resection followed by postoperative chemoradiotherapy. The standard regimen of temozolomide (TMZ) combined with radiotherapy has, to some extent, prolonged patient survival. However, almost all GBM cases inevitably face recurrence, with relapsed patients having an average overall survival of only 12-15 months. Poor prognosis, complex tumor subtypes, and a relatively limited range of clinical treatment drugs make GBM a key focus and challenge in the treatment of neurosurgical diseases.

[0003] Bromodomains (BRDs) are an evolutionarily conserved family of protein sequences. 61 BRDs are known to exist in human genes and have been identified in 46 proteins. They are primarily involved in regulating histone acetylation, and are associated with tumor development and drug resistance (M. Liu, et al., Recent Advances on Small-Molecule Bromodomain-Containing Histone Acetyltransferase Inhibitors. J Med Chem 2023, 66, 1678-1699). BRG1 (also known as SMARCA4) and BRM (also known as SMARCA2) are two functionally exclusive ATPase subunits in the SWI / SNF chromatin remodeling complex, playing crucial roles in gene regulation, organismal development, DNA replication, and repair. TCGA and other cancer databases show that poor prognosis and high invasiveness in a variety of cancers may be associated with BRG1 mutations and overexpression (JA Guerrero-Martinez, JCReyes. High expression of SMARCA4 or SMARCA2 is frequently associated with an opposite prognosis in cancer. Sci. Rep. 2018, 8, 2043). In GBM, researchers have found that BRG1 / BRM gene mutations are relatively rare and structurally conserved. However, compared with adjacent normal brain tissue, BRG1 expression in GBM cells is significantly increased, which may be involved in the proliferation, migration and invasion of GBM cells (J. Bai, et al., BRG1 expression is increased in human glioma and controls glioma cell proliferation, migration and invasion in vitro. J. Cancer. Res. Clin. Oncol. 2012, 138, 991-998; SMNavickas, et al., The role of chromatin remodeler SMARCA4 / BRG1 in brain cancers: a potential therapeutic target. Oncogene 2023, 42, 2363–2373).Meanwhile, as research on BRG1 / BRM deepened, it was discovered that the homologous proteins BRG1 and BRM have a synthetic lethality relationship. That is, cancer cells that lose BRG1 expression due to mutations are highly dependent on BRM for survival (V. Previtali, et al., New Horizons of Synthetic Lethality in Cancer: Current Development and Future Perspectives. J. Med. Chem. 2024, 67, 11488-11521).

[0004] In recent years, Miller's team at the University of Tennessee Health Sciences Center reported a series of novel inhibitors targeting the BRG1 / BRM bromodomain for combination therapy of glioblastoma. Compounds IV-255 and IV-275, when combined with the DNA alkylating agent TMZ, showed superior efficacy to TMZ alone in both in vivo and in vitro experiments. These compounds further enhanced TMZ's inhibitory effect on GBM cell proliferation and delayed the development of GBM resistance to TMZ (Y. He, et al., Novel structural-related analogs of PFI-3 (SRAPs) that target the BRG1 catalytic subunit of the SWI / SNF complex increase the activity of temozolomide in glioblastoma cells. Bioorg. Med. Chem. 2022, 53, 116533; C. Yang, et al., Targeting the Bromodomain of BRG-1 / BRM Subunit of the SWI / SNF Complex Increases the Anticancer Activity of Temozolomide in Glioblastoma. Pharmaceuticals (Basel) 2021, 14, 904; C. Yang, et al., Next-generation bromodomain inhibitors of the SWI / SNF complex enhance DNA damage and cell death in glioblastoma. J Cell Mol Med 2023, 27, 2770-2781). However, the above compounds can only be used as synergists in combination with TMZ; they do not have significant antitumor effects when used alone, thus limiting their clinical application.

[0005] The proton-sensing receptor GPR65 is mainly distributed in immune cells and is one of the main transporters that interfere with the normal functioning of immune cells in the acidic tumor microenvironment. Inhibitors targeting GPR65 can improve immune escape by cancer cells in the tumor microenvironment, reduce the production of M2 macrophages that promote cancer cell proliferation, and increase CD8+. +The number of T cells and NK cells, reversing the inhibitory effect of the tumor microenvironment on immune cells, and restoring / enhancing the killing effect of immune cells on tumors in vivo (H. Sekine, M. Yamamoto, H. Motohashi. Tumors sweeten macrophages with acids. Nat. Immunol. https: / / doi.org / 10.1038 / s41590-018-0258-0 (2018)).

[0006] In 2018, the team led by Chen Juxiang at Shanghai Changhai Hospital used immunohistochemistry to detect the expression of GPR65 protein in tissue microarrays of tumor samples from hundreds of patients with different grades of glioma. They found that the expression level of GPR65 in brain tumor tissue was positively correlated with the grade of glioma, and that high expression of GPR65 was positively correlated with poor patient prognosis (H.-X. Wang, et al., Overexpression of G-protein-coupled receptors 65 in glioblastoma predicts poor patient prognosis. Clin. Neurool. Neurosur. 2018, 164, 132–137). In 2024, Zhang Xiaobiao's team at Zhongshan Hospital affiliated with Fudan University further discovered that GPR65 in brain tumor tissue is highly expressed in tumor-associated macrophages (TAMs) and induces macrophage polarization towards the M2 subtype, thereby promoting the proliferation, migration, invasion, and mesenchymal transformation of glioma cells. By knocking out GPR65 or inhibiting related pathways, the progression of glioblastoma can be inhibited in vivo (CLYan, et al., GPR65 sensing tumor-derived lactate induces HMGB1 release from TAM via the cAMP / PKA / CREB pathway to promote glioma progression. J.Exp.Clin.Cancer Res.2024,43,105-123). In the same year, Yang Xuejun's team at Tianjin Medical University General Hospital further confirmed that GPR65 is highly expressed in high-grade glioblastoma and is positively correlated with the malignancy and poor prognosis of glioma. At the same time, single-cell data analysis and immunofluorescence verified that GPR65 is mainly expressed in macrophages in the glioma microenvironment. Furthermore, inhibiting GPR65 is sufficient to reduce the polarization response of macrophages to glioma cells and break the malignant synergy between macrophages and glioma cells (JKFan, et al., GPR65 contributes to constructing immunosuppressive microenvironment in glioma. Neurosurg. Rev. 2024, 47, 417-428).

[0007] WO2021 / 245426 discloses N-(phenylaminocarbonyl)tetrahydroisoquinoline compounds and related compounds as regulators of GPR65. These compounds can be used to prevent or treat proliferative diseases, immune diseases, asthma, chronic obstructive pulmonary disease and acute respiratory distress syndrome, but do not disclose compounds with BRG1 / BRM inhibitory activity.

[0008] Given the important biological roles of BRG1 / BRM and proton-sensing receptor GPR65 in tumor progression, the development of multi-target inhibitors of BRG1 / BRM and GPR65 is expected to provide new treatment options for tumors, especially glioblastoma. Summary of the Invention

[0009] This invention provides a class of multi-target inhibitors of the BRG1 / BRM bromine domain and the proton-sensing receptor GPR65.

[0010] The first aspect of the present invention provides compounds of formula I or pharmaceutically acceptable salts, stereoisomers, solvates or isotopic derivatives thereof:

[0011] in

[0012] X, Y, and Z are each independently CH, CR, or N.

[0013] W can be -NH-, -O-, -CH2-, or -C(O)-.

[0014] R and R1 are each independently selected from halogen, cyano, nitro, hydroxy, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, and -NR4R5, or

[0015] The two R and / or R1 atoms on adjacent carbon atoms, together with the carbon atoms they are attached to, form a 5-6 membered ring optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur.

[0016] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NR4R5, -S(O)2-C1-C6 alkyl, -C(O)2-C1-C6 alkyl, C3-C8 cycloalkyl, 3 to 8-membered heterocyclic groups, C6-C 10 Aryl, 5- to 12-membered heteroaryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heterocyclic and heteroaryl groups contain 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur, and the cycloalkyl, heterocyclic, aryl and heteroaryl groups are optionally substituted by 1 to 3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy, provided that R2 and R3 are not simultaneously hydrogen;

[0017] R4 and R5 are each independently selected from hydrogen, C1-C6 alkyl, C3-C8 cycloalkyl, and C2-C6 alkynyl groups, and

[0018] n is 0, 1, or 2.

[0019] In some embodiments, the compound of formula I has the following formula I-1:

[0020] R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I.

[0021] In some embodiments, the compound of formula I has the following formula I-2:

[0022] R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I.

[0023] In some embodiments, the compound of formula I has the following formulas I-3:

[0024] R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I.

[0025] In some embodiments, the compound of formula I has the following formulas I-4:

[0026] R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I.

[0027] A second aspect of the invention relates to pharmaceutical compositions comprising a compound of formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, and one or more pharmaceutically acceptable carriers.

[0028] The third aspect of the invention relates to a compound of formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, or a pharmaceutical composition comprising a compound of formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, and one or more pharmaceutically acceptable carriers, for the prevention and / or treatment of diseases associated with abnormal activity of the BRG1 / BRM bromine domain and / or proton-sensing receptor GPR65.

[0029] The fourth aspect of the invention relates to the use of a compound of Formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, or a pharmaceutical composition comprising a compound of Formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, and one or more pharmaceutically acceptable carriers for the prevention and / or treatment of diseases associated with abnormal activity of the BRG1 / BRM bromine domain and / or the proton-sensing receptor GPR65.

[0030] The fifth aspect of the invention relates to the use of a compound of Formula I or a pharmaceutically acceptable salt, stereoisomer, solvate, prodrug, metabolite or isotope derivative thereof, or a pharmaceutical composition comprising a compound of Formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotope derivative thereof, and one or more pharmaceutically acceptable carriers in the preparation of a medicament for the prevention and / or treatment of diseases associated with abnormal activity of the BRG1 / BRM bromine domain and / or proton-sensing receptor GPR65.

[0031] A sixth aspect of the invention relates to a method for preventing and / or treating diseases associated with abnormal activity of the BRG1 / BRM bromine domain and / or the proton-sensing receptor GPR65 in subjects with such need, the method comprising administering to the subject a compound of Formula I or a pharmaceutically acceptable salt, stereoisomer, solvate, or isotopic derivative thereof, or a pharmaceutical composition comprising a compound of Formula I or a pharmaceutically acceptable salt, stereoisomer, solvate, or isotopic derivative thereof, and one or more pharmaceutically acceptable carriers. Attached Figure Description

[0032] Figure 1 shows the affinity results of the compounds of the present invention with BRG1 / BRM protein, where the horizontal axis represents the compound number and the vertical axis represents the affinity value (μM).

[0033] Figure 2 shows the affinity results between the compounds of the present invention and the GPR65 protein, where the horizontal axis represents the compound number and the vertical axis represents the affinity value (μM).

[0034] Figure 3 shows the in vitro glioblastoma cell proliferation inhibition test results of the compounds of the present invention, where the horizontal axis represents the compound number and the vertical axis represents the IC50 value. 50 Value (μM);

[0035] Figure 4 shows the results of the in vitro glioblastoma cell proliferation inhibition experiment of compound 1, where the horizontal axis represents the compound concentration (μM) and the vertical axis represents the inhibition percentage (%).

[0036] Figure 5 shows the results of the in vitro glioblastoma cell proliferation inhibition experiment of compound 10, where the horizontal axis represents the compound concentration (μM) and the vertical axis represents the inhibition percentage (%).

[0037] Figure 6 shows the results of the in vitro glioblastoma cell proliferation inhibition experiment of compound 17, where the horizontal axis represents the compound concentration (μM) and the vertical axis represents the inhibition percentage (%).

[0038] Figure 7 shows the results of the in vitro glioblastoma cell proliferation inhibition assay of compound 30, where the horizontal axis represents the compound concentration (μM) and the vertical axis represents the inhibition percentage (%).

[0039] Figure 8 shows the results of the in vitro glioblastoma cell proliferation inhibition experiment of compound 31, where the horizontal axis represents the compound concentration (μM) and the vertical axis represents the inhibition percentage (%).

[0040] Figure 9 shows the results of the in vitro glioblastoma cell proliferation inhibition assay of compound 36, where the horizontal axis represents the compound concentration (μM) and the vertical axis represents the inhibition percentage (%).

[0041] Figure 10 shows the results of the in vitro glioblastoma cell proliferation inhibition assay of compound 37, where the horizontal axis represents the compound concentration (μM) and the vertical axis represents the inhibition percentage (%).

[0042] Figure 11 shows the results of the in vitro glioblastoma cell proliferation inhibition experiment of compound 42, where the horizontal axis represents the compound concentration (μM) and the vertical axis represents the inhibition percentage (%).

[0043] Figure 12 shows the changes in body weight of mice in each group during the in vivo efficacy evaluation experiment of the B104-1-1 mouse-derived glioblastoma subcutaneous animal model;

[0044] Figure 13 shows the percentage change in body weight of mice in each group during the in vivo efficacy evaluation experiment of the B104-1-1 mouse-derived glioblastoma subcutaneous animal model.

[0045] Figure 14 shows the tumor volume growth curve in the in vivo efficacy evaluation experiment of the B104-1-1 mouse glioblastoma subcutaneous animal model;

[0046] Figure 15 shows the changes in body weight of mice in the in vivo drug efficacy evaluation experiment of the MC38 mouse-derived subcutaneous colon cancer animal model;

[0047] Figure 16 shows the percentage change in mouse body weight in the in vivo efficacy evaluation experiment of the MC38 mouse-derived subcutaneous colon cancer animal model.

[0048] Figure 17 shows the tumor volume growth curve in the in vivo efficacy evaluation experiment of the MC38 mouse-derived colon cancer subcutaneous animal model. Detailed Implementation

[0049] definition

[0050] Unless otherwise stated, the following terms used in this specification and claims have the following meanings. It should be understood that, in the absence of a specific definition herein, a term should be given its meaning as is known in the art. Furthermore, it should be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the scope of the invention in any way. Unless otherwise stated, in the event of a discrepancy between the structural formula and chemical name of a compound described herein, the structural formula shall prevail.

[0051] The group prefix "C" used in this article x -C y The "" indicates the range of the number of carbon atoms in the group, where x and y are both integers. For example, C3-C8 cycloalkyl means a cycloalkyl group with 3 to 8 carbon atoms, i.e., cycloalkyl groups with 3, 4, 5, 6, 7, or 8 carbon atoms. It should also be understood that "C3-C8" includes any sub-ranges within this range, such as C3-C7, C3-C6, C4-C7, C4-C6, C5-C6, etc.

[0052] As used herein, the term "alkyl" refers to a straight-chain or branched saturated monovalent hydrocarbon group having a specified number of carbon atoms. Alkyl groups typically contain 1-6 carbon atoms ("C1-C6 alkyl"), preferably 1-5 carbon atoms ("C1-C5 alkyl"), more preferably 1-4 carbon atoms ("C1-C4 alkyl"), 1-3 carbon atoms ("C1-C3 alkyl"), or 1-2 carbon atoms ("C1-C2 alkyl"). Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl, isopentyl, neopentyl, and n-hexyl.

[0053] As used herein, the term "alkynyl" refers to a straight-chain or branched monovalent hydrocarbon group having a specified number of carbon atoms and at least one carbon-carbon triple bond. Alynyl groups typically contain 2-6 carbon atoms ("C2-C6 alkynyl"), preferably 2-5 carbon atoms ("C2-C5 alkynyl"), more preferably 2-4 carbon atoms ("C2-C4 alkynyl") or 2-3 carbon atoms ("C2-C3 alkynyl"). Examples of alkynyl groups include, but are not limited to, ethynyl, propynyl, n-butynyl, isobutynyl, pentyynyl, and hexynyl.

[0054] As used herein, the term "alkoxy" refers to an alkyl group (i.e., "-O-alkyl") attached to a parent molecule via an oxygen atom, wherein the alkyl group is as defined above. Alkoxy groups typically contain 1-6 carbon atoms ("C1-C6 alkoxy"), more preferably 1-4 carbon atoms ("C1-C4 alkoxy"). Examples of alkoxy groups include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy, pentoxy, and hexoxy.

[0055] The term "halogen" as used in this article refers to fluorine, chlorine, bromine, and iodine, with fluorine and chlorine being preferred.

[0056] As used herein, the term "haloalkyl" means an alkyl group in which one or more hydrogen atoms are replaced by one or more identical or different halogen atoms as defined above. For example, "C1-C6 haloalkyl" refers to a "C1-C6 alkyl" in which one or more hydrogen atoms are replaced by one or more identical or different halogen atoms. Examples of C1-C6 haloalkyl include, but are not limited to, trifluoromethyl, trichloromethyl, difluoromethyl, and dichloromethyl.

[0057] As used in this article, the term "cyano" refers to the -CN group.

[0058] The term "nitro" as used in this article refers to the -NO2 group.

[0059] The term "hydroxyl group" as used in this article refers to the -OH group.

[0060] The term "trifluoromethyl" as used in this article refers to the -CF3 group.

[0061] The term "difluoromethyl" as used in this article refers to the -CF2H group.

[0062] The term "trichloromethyl" as used in this article refers to the -CCl3 group.

[0063] As used in this article, the term "dichloromethyl" refers to the -CCl2H group.

[0064] As used herein, the term "aryl" refers to a monovalent hydrocarbon group derived from a monocyclic or fused bicyclic or polycyclic ring system (where at least one ring contains a fully conjugated π-electron system) having well-known aromatic characteristics. Fused aryl groups can include aryl rings fused with a saturated or partially unsaturated carbocyclic or heterocyclic ring, or fused with another aryl or heteroaryl ring, provided that the connection point to the parent molecule in such a fused ring system is an atom of the aromatic portion of the ring system. Aryl groups typically contain 6–14 ("C6-C7") atoms. 14 Aryl group), more preferably 6-10 carbon atoms ("C6-C") 10 Aryl groups include, but are not limited to, phenyl, naphthyl, anthracene, phenanthryl, indanyl, indenyl, and tetrahydronaphthyl.

[0065] As used herein, the term "cycloalkyl" refers to a monovalent hydrocarbon group derived from a non-aromatic saturated carbocyclic system containing a specified number of carbon atoms. Cycloalkyl groups typically contain 3-8 carbon atoms ("C3-C8 cycloalkyl"), preferably 3-7 carbon atoms ("C3-C7 cycloalkyl") or 3-6 carbon atoms ("C3-C6 cycloalkyl"). Examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.

[0066] The term "heteroatom" as used in this article refers to N, O, or S atoms.

[0067] As used herein, the term "heterocyclic group" refers to a monovalent group derived from a saturated or partially unsaturated non-aromatic ring structure containing a specified number of ring atoms and including at least one heteroatom, preferably one to four heteroatoms as ring members. Heterocyclic groups include spirocyclic, bridged, or fused rings formed with one or more other heterocycles or carbocyclic rings, provided that the connection point with the parent molecule is an atom of the heterocyclic portion of such a ring system. Heterocyclic groups typically contain 3 to 12 ring atoms (i.e., 3 to 12-membered heterocyclic groups), preferably 3 to 8 ring atoms (i.e., 3 to 8-membered heterocyclic groups), and most preferably 5 or 6 ring atoms (i.e., 5 or 6-membered heterocyclic groups). Examples of heterocyclic groups include, but are not limited to, azirropropyl, oxacyclopropyl, thiohexacyclopropyl, azirrobutyl, oxacyclobutyl, thiohexacyclobutyl, tetrahydrofuranyl, tetrahydropyranyl, tetrahydrothiophene, tetrahydrothiophene, pyrrolyl, imidazoalkyl, pyrazolyl, oxazolyl, isoxazolyl, thiazoalkyl, isothiazolyl, dihydrofuranyl, dihydrothiophene, dihydrooxazolyl, dihydrothiazolyl, isodihydrothiazolyl, dihydropyrroleyl, dihydroimidazolyl, dihydropyridyl, dihydropyridinyl, dihydropyrimidinyl, dihydropyrazinyl, dihydropyridazinyl, piperidinyl, piperazinyl, dioxacyclohexyl, oxothiohexyl, azirroheptanyl, diazirroheptanyl, morpholinyl, thiomorpholinyl, indololinyl, and isoindololinyl.

[0068] As used herein, the term "heteroaryl" refers to a monovalent group derived from an aromatic ring structure containing a specified number of ring atoms and including at least one heteroatom, preferably one to four heteroatoms as ring members. Heteroaryls typically contain 5-12 ring atoms ("5-12-membered heteroaryls"), preferably 5-10 ring atoms ("5-10-membered heteroaryls"), and more preferably 5 or 6 ring atoms ("5 or 6-membered heteroaryls"). Heteroaryls may also be fused with another aryl or heteroaryl ring, or with a saturated or partially unsaturated carbocyclic or heterocyclic ring, provided that the connection point to the parent molecule in such a fused ring system is an atom of the heteroaryl portion of the ring system. Examples of heteroaryl groups include, but are not limited to, pyrrole, furanyl, thiophene, pyrazolyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, triazolyl, tetrazolyl, pyridinyl, pyrimidinyl, pyrazinyl, triazinyl, indole, isoyindole, indazolyl, benzofuranyl, benzothiophene, indazole, benzimidazolyl, benzoxazolyl, benzoisoxazolyl, benzothiazolyl, and benzoisoxazolyl. Thiazolyl, benzoxadiazolyl, benzothiadiazolyl, benzotriazolyl, imidazopyridyl, imidazopyrimidinyl, imidazopyridazinyl, purine, furan-pyridyl, thienopyridyl, benzopyranyl, quinolinyl, isoquinolinyl, quinazinyl, quinazolinyl, quinoxolinyl, benzopyridazinyl, cinolinyl, naphridyl, pteridinyl, carbazoyl, carolinyl, phenanthridine, acridineyl, phenanthroxazinyl, and phenothiazinyl.

[0069] As used herein, the term "optional" means that the condition immediately following the term may or may not occur. For example, "optionally halogen-substituted phenyl" encompasses both "unsubstituted phenyl" and "halogen-substituted phenyl".

[0070] As used herein, the term "stereoisomer" refers to isomers resulting from differences in the spatial arrangement of atoms in a molecule. Enantiomers are produced when a compound contains asymmetric carbon atoms; cis-trans isomers are produced when a compound contains carbon-carbon double bonds or cyclic structures; and trans-isomers may be produced when rotation around a single bond in a compound is impeded. The scope of this invention includes all enantiomers, diastereomers, racemates, optical isomers, geometric isomers, trans-isomers, epimers, and mixtures thereof of compounds of Formula I. When the bonds to chiral carbons are depicted as straight lines in the formulas of this invention, it should be understood that the (R) and (S) configurations of the chiral carbons, and therefore the two enantiomers and mixtures thereof, are included in the formula.

[0071] As used herein, the term "solvent" refers to a molecular complex comprising a compound of formula I and one or more pharmaceutically acceptable solvent molecules (e.g., ethanol). When the solvent is water, the term "hydrate" is used.

[0072] As used herein, the term "prodrug" refers to a compound that is inactive or has very low activity in vitro but releases a pharmacologically active compound in vivo via enzymatic or non-enzymatic transformation. This transformation can occur through various mechanisms, such as hydrolysis, oxidation, and reduction. Examples of prodrugs include compounds in which the amino group is acylated, alkylated, or phosphorylated, or where the hydroxyl group is acylated, alkylated, or phosphorylated, or where the carboxyl group is esterified or amidated. This invention includes prodrugs of compounds of Formula I, their pharmaceutically acceptable salts, or their stereoisomers. Specific examples of prodrugs include:

[0073] Where R' is an optionally substituted C1-C6 alkyl, C3-C8 cycloalkyl, or C6-C 10 Aryl, 3 to 8-membered heterocyclic, 5 to 12-membered heteroaryl or dimethylamino.

[0074] As used herein, the term "metabolite" refers to the active substance obtained by the metabolic transformation of a compound of Formula I in vivo. This invention includes active metabolites of compounds of Formula I, their pharmaceutically acceptable salts, or their stereoisomers, such as primary and / or secondary metabolites in vivo. Examples are as follows:

[0075] Primary metabolites in the body:

[0076] Secondary metabolites in the body:

[0077] Primary and secondary metabolites in the body:

[0078] As used herein, the term "isotope derivative" refers to an isotopically labeled compound, that is, a compound of formula I in which one or more atoms are replaced by atoms having the same atomic number but with a different atomic mass or mass number than those commonly found in nature. Examples of isotopes suitable for incorporation into compounds include, but are not limited to, hydrogen (such as...). 2 H and 3 H), carbon (such as ... 11 C 13 C and 14 C), chlorine (such as...) 36 Cl), fluorine (such as Cl), 18 F), iodine (such as F), 123 I, 124 I and 125 I) Nitrogen (such as 13 N and 15 N), oxygen (such as N), 15 O、 17 O and 18 O), phosphorus (such as O), phosphorus (such as 32 P) and sulfur (such as P) and sulfur (such as P)35 S). Without excessive experimentation, isotopically labeled compounds of the present invention can be prepared using conventional techniques known to those skilled in the art or methods similar to those described in the schemes and examples herein, using appropriate isotopically labeled reagents and / or intermediates. The present invention particularly includes deuterated derivatives of compounds of formula I, such as the following:

[0079] As used in this article, "pharmaceutically acceptable" means that a substance or material is suitable for contact with the tissues of a subject, such as a human or other mammal, without causing excessive toxicity, irritation, allergic reactions or other problems, within the bounds of reasonable medical judgment, and has a proportionate benefit / risk ratio.

[0080] As used herein, the term "pharmaceutically acceptable salt" refers to a salt formed by the reaction of a pharmaceutically non-toxic acid with the basic portion of a compound of formula I of the present invention, including, for example, hydrochlorides, acetates, hydrobromates, sulfates, bisulfates, carbonates, bicarbonates, sulfites, phosphates, hydrogen phosphates, oxalates, malonates, valerates, borates, p-toluenesulfonates, methanesulfonates, tartrates, benzoates, lactates, citrates, maleates, fumarates, malates, salicylates, mandelates, succinates, gluconates, lactobionates, etc. Such salts can be prepared by methods well known to those skilled in the art.

[0081] The term “prevention” as used in this article refers to reducing or eliminating the possibility of disease.

[0082] The term “treatment” as used in this article refers to the complete or partial elimination of the disease and / or its accompanying symptoms.

[0083] As used herein, the term “object” means an animal, preferably a mammal, that is intended to be an individual for experimentation or treatment, including but not limited to primates (e.g., apes and humans), equines (e.g., horses), canines (e.g., dogs), felines, domesticated livestock (e.g., pigs, goats, sheep, etc.), as well as domestic pets and animals kept in zoos, preferably humans.

[0084] Detailed description of the technical solution of the present invention

[0085] The first aspect of this invention relates to compounds of formula I or pharmaceutically acceptable salts, stereoisomers, solvates, or isotopic derivatives thereof:

[0086] in

[0087] X, Y, and Z are each independently CH, CR, or N.

[0088] W can be -NH-, -O-, -CH2-, or -C(O)-.

[0089] R and R1 are each independently selected from halogen, cyano, nitro, hydroxy, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, and -NR4R5, or

[0090] The two R and / or R1 atoms on adjacent carbon atoms, together with the carbon atoms they are attached to, form a 5-6 membered ring optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur.

[0091] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NR4R5, -S(O)2-C1-C6 alkyl, -C(O)2-C1-C6 alkyl, C3-C8 cycloalkyl, 3 to 8-membered heterocyclic groups, C6-C 10 Aryl, 5- to 12-membered heteroaryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heterocyclic and heteroaryl groups contain 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur, and the cycloalkyl, heterocyclic, aryl and heteroaryl groups are optionally substituted by 1 to 3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy, provided that R2 and R3 are not simultaneously hydrogen;

[0092] R4 and R5 are each independently selected from H, C1-C6 alkyl, C3-C8 cycloalkyl, and C2-C6 alkynyl groups, and

[0093] n is 0, 1, or 2.

[0094] In some embodiments, the compound of formula I has the following formula I-1:

[0095] R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I.

[0096] In some embodiments, the compound of formula I has the following formula I-2:

[0097] R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I.

[0098] In some embodiments, the compound of formula I has the following formulas I-3:

[0099] R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I.

[0100] In some embodiments, the compound of formula I has the following formulas I-4:

[0101] R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I.

[0102] In some implementations, no more than two of X, Y, and Z are N.

[0103] In some implementations, no more than one of X, Y, and Z is N.

[0104] In some embodiments, X, Y, and Z are all CH. In some embodiments, X is CH, and Y and Z are each CR. In some embodiments, Y is CH, and X and Z are each CR. In some embodiments, Z is CH, and X and Y are each CR. In some embodiments, X and Y are CH, and Z is CR. In some embodiments, X and Z are CH, and Y is CR. In some embodiments, Y and Z are CH, and X is CR. In some embodiments, X, Y, and Z are all CR. In each of the above embodiments, R is independently selected from halogen, cyano, nitro, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, and -N(C1-C6 alkyl)2.

[0105] In some embodiments, only one of X, Y, and Z is N, and the rest are CH or CR. In some embodiments, X is N, and Y and Z are each CH or CR. In some embodiments, Y is N, and X and Z are each CH or CR. In some embodiments, Z is N, and X and Y are each CH or CR. In all the above embodiments, R is independently selected from halogen, cyano, nitro, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, and -N(C1-C6 alkyl)2.

[0106] In some embodiments, R and R1 are each independently selected from halogen, cyano, nitro, hydroxy, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, and -N(C1-C6 alkyl)2. In some embodiments, R and R1 are each independently selected from halogen, cyano, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy.

[0107] In some embodiments, R and R1 are each independently selected from fluorine, chlorine, bromine, iodine, cyano, nitro, hydroxyl, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, trifluoromethyl, trichloromethyl, difluoromethyl, dichloromethyl, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy, -NH2, -NH(CH3)2, and -N(CH3)2. In some embodiments, R and R1 are each independently selected from fluorine, chlorine, bromine, cyano, methyl, ethyl, isopropyl, trifluoromethyl, trichloromethyl, methoxy, ethoxy, and isopropoxy. In some embodiments, R and R1 are each independently selected from fluorine, chlorine, bromine, cyano, methyl, trifluoromethyl, and methoxy.

[0108] In some embodiments, the two R and / or R1 on adjacent carbon atoms, together with the carbon atom to which they are attached, form a 5-6 membered ring optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur. In some embodiments, the two R on adjacent carbon atoms, together with the carbon atom to which they are attached, form a 5-6 membered ring optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur. In some embodiments, the two R1 on adjacent carbon atoms, together with the carbon atom to which they are attached, form a 5-6 membered ring optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur. In some embodiments, the two R1 on adjacent carbon atoms, together with the carbon atom to which they are attached, form a 5-6 membered ring optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur. In some embodiments, the R and R1 on adjacent carbon atoms, together with the carbon atom to which they are attached, form a 5-membered ring containing 2 heteroatoms independently selected from oxygen.

[0109] In some embodiments, R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, -S(O)2-C1-C6 alkyl, -C(O)2-C1-C6 alkyl, C3-C8 cycloalkyl, 3- to 8-membered heterocyclic groups, C6-C 10 Aryl, 5- to 12-membered heteroaryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heterocyclic and heteroaryl groups contain 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur, and the cycloalkyl, heterocyclic, aryl and heteroaryl groups are optionally substituted with 1 to 3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy, provided that R2 and R3 are not simultaneously hydrogen.

[0110] In some embodiments, R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, C6-C 10 Aryl, 5- to 12-membered heteroaryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur, and the cycloalkyl, aryl and heteroaryl are optionally substituted by 1 to 3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy, provided that R2 and R3 are not simultaneously hydrogen.

[0111] In some embodiments, R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, or C6-C substituted with 1-3 halogens. 10 Aryl and -O-C1-C6 alkyl-5-12 heteroaryl, wherein the heteroaryl contains 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur, provided that R2 and R3 are not simultaneously hydrogen.

[0112] In some embodiments, R2 and R3 are each independently selected from hydrogen, fluorine, chlorine, bromine, cyano, hydroxyl, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, trifluoromethyl, trichloromethyl, difluoromethyl, dichloromethyl, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy, -NH2, -NH(CH3), -N(CH3)2, -NH-cyclopropyl, -NH-cyclobutyl, -NH-cyclopentyl, -NH-cyclohexyl, -NH-ethynyl, -NH-(1-propynyl), -NH-(2-propynyl), phenyl, 4-fluorophenyl, pyridyl, and -O-CH2-pyridyl, provided that R2 and R3 are not simultaneously hydrogen. In some embodiments, R2 and R3 are each independently selected from hydrogen, fluorine, chlorine, cyano, hydroxyl, trifluoromethyl, methoxy, isopropoxy, -NH-cyclopropyl, -NH-(2-propynyl), phenyl, 4-fluorophenyl and -O-CH2-pyridyl, provided that R2 and R3 are not simultaneously hydrogen.

[0113] In some embodiments, R2 is hydrogen, and R3 is selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, -S(O)2-C1-C6 alkyl, -C(O)2-C1-C6 alkyl, C3-C8 cycloalkyl, 3- to 8-membered heterocyclic groups, C6-C 10 Aryl, 5- to 12-membered heteroaryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heterocyclic and heteroaryl groups contain 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur, and the cycloalkyl, heterocyclic, aryl and heteroaryl groups are optionally substituted with 1 to 3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy.

[0114] In some embodiments, R2 is hydrogen, and R3 is selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, C6-C 10 Aryl, 5- to 12-membered heteroaryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur, and the cycloalkyl, aryl and heteroaryl are optionally substituted by 1 to 3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy.

[0115] In some embodiments, R2 is hydrogen, and R3 is selected from halogens, cyano groups, hydroxyl groups, C1-C6 haloalkyl groups, C1-C6 alkoxy groups, -NH-C3-C8 cycloalkyl groups, -NH-C2-C6 alkynyl groups, and C6-C groups optionally substituted with 1-3 halogens. 10 Aryl and -O-C1-C6 alkyl-5-12 heteroaryl, wherein the heteroaryl contains 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur.

[0116] In some embodiments, R2 is hydrogen, and R3 is selected from fluorine, chlorine, bromine, cyano, hydroxyl, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, trifluoromethyl, trichloromethyl, difluoromethyl, dichloromethyl, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy, -NH2, -NH(CH3), -N(CH3)2, -NH-cyclopropyl, -NH-cyclobutyl, -NH-cyclopentyl, -NH-cyclohexyl, -NH-ethynyl, -NH-(1-propynyl), -NH-(2-propynyl), phenyl, 4-fluorophenyl, pyridyl, and -O-CH2-pyridyl. In some embodiments, R2 is hydrogen, and R3 is selected from fluorine, chlorine, cyano, hydroxyl, trifluoromethyl, methoxy, isopropoxy, -NH-cyclopropyl, -NH-(2-propynyl), phenyl, 4-fluorophenyl and -O-CH2-pyridyl.

[0117] In some embodiments, R3 is hydrogen, and R2 is selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, -S(O)2-C1-C6 alkyl, -C(O)2-C1-C6 alkyl, C3-C8 cycloalkyl, 3- to 8-membered heterocyclic groups, C6-C 10 Aryl, 5- to 12-membered heteroaryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heterocyclic and heteroaryl groups contain 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur, and the cycloalkyl, heterocyclic, aryl and heteroaryl groups are optionally substituted with 1 to 3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy.

[0118] In some embodiments, R3 is hydrogen, and R2 is selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, C6-C 10 Aryl, 5- to 12-membered heteroaryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur, and the cycloalkyl, aryl and heteroaryl are optionally substituted by 1 to 3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy.

[0119] In some embodiments, R3 is hydrogen, and R2 is selected from halogens, cyano groups, hydroxyl groups, C1-C6 haloalkyl groups, C1-C6 alkoxy groups, -NH-C3-C8 cycloalkyl groups, -NH-C2-C6 alkynyl groups, and C6-C groups optionally substituted with 1-3 halogens. 10 Aryl and -O-C1-C6 alkyl-5-12 heteroaryl, wherein the heteroaryl contains 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur.

[0120] In some embodiments, R3 is hydrogen, and R2 is selected from fluorine, chlorine, bromine, cyano, hydroxyl, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, trifluoromethyl, trichloromethyl, difluoromethyl, dichloromethyl, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy, -NH2, -NH(CH3), -N(CH3)2, -NH-cyclopropyl, -NH-cyclobutyl, -NH-cyclopentyl, -NH-cyclohexyl, -NH-ethynyl, -NH-(1-propynyl), -NH-(2-propynyl), phenyl, 4-fluorophenyl, pyridyl, and -O-CH2-pyridyl. In some embodiments, R3 is hydrogen, and R2 is selected from fluorine, chlorine, cyano, hydroxyl, trifluoromethyl, methoxy, isopropoxy, -NH-cyclopropyl, -NH-(2-propynyl), phenyl, 4-fluorophenyl and -O-CH2-pyridyl.

[0121] In some embodiments, R2 and R3 are each independently selected from halogens, cyano groups, hydroxyl groups, C1-C6 alkyl groups, C1-C6 haloalkyl groups, C1-C6 alkoxy groups, -NH2, -NH-C1-C6 alkyl groups, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl groups, -NH-C2-C6 alkynyl groups, -S(O)2-C1-C6 alkyl groups, -C(O)2-C1-C6 alkyl groups, C3-C8 cycloalkyl groups, 3- to 8-membered heterocyclic groups, C6-C 10 Aryl, 5- to 12-membered heteroaryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heterocyclic and heteroaryl groups contain 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur, and the cycloalkyl, heterocyclic, aryl and heteroaryl groups are optionally substituted with 1 to 3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy.

[0122] In some embodiments, R2 and R3 are each independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, C6-C 10Aryl, 5- to 12-membered heteroaryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur, and the cycloalkyl, aryl and heteroaryl are optionally substituted by 1 to 3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy.

[0123] In some embodiments, R2 and R3 are each independently selected from halogens, cyano groups, hydroxyl groups, C1-C6 haloalkyl groups, C1-C6 alkoxy groups, -NH-C3-C8 cycloalkyl groups, -NH-C2-C6 alkynyl groups, and C6-C groups optionally substituted with 1-3 halogens. 10 Aryl and -O-C1-C6 alkyl-5-12 heteroaryl, wherein the heteroaryl contains 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur.

[0124] In some embodiments, R2 and R3 are each independently selected from fluorine, chlorine, bromine, cyano, hydroxyl, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, trifluoromethyl, trichloromethyl, difluoromethyl, dichloromethyl, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy, -NH2, -NH(CH3), -N(CH3)2, -NH-cyclopropyl, -NH-cyclobutyl, -NH-cyclopentyl, -NH-cyclohexyl, -NH-ethynyl, -NH-(1-propynyl), -NH-(2-propynyl), phenyl, 4-fluorophenyl, pyridyl, and -O-CH2-pyridyl. In some embodiments, R2 and R3 are each independently selected from fluorine, chlorine, cyano, hydroxyl, trifluoromethyl, methoxy, isopropoxy, -NH-cyclopropyl, -NH-(2-propynyl), phenyl, 4-fluorophenyl and -O-CH2-pyridyl.

[0125] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0126] X, Y, and Z are each independently CH, CR, or N;

[0127] R and R1 are each independently selected from halogen, cyano, nitro, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, and -N(C1-C6 alkyl)2, or

[0128] The two R and / or R1 on adjacent carbon atoms together with the carbon atoms to which they are attached form a 5-6 membered ring, optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur;

[0129] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, -S(O)2-C1-C6 alkyl, -C(O)2-C1-C6 alkyl, C3-C8 cycloalkyl, 3- to 8-membered heterocyclic groups, C6-C 10 Aryl, 5- to 12-membered heteroaryl, and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heterocyclic and heteroaryl groups contain 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, and the cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally substituted with 1-3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy groups, provided that R2 and R3 are not simultaneously hydrogen; and

[0130] n is 0, 1, or 2.

[0131] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0132] X, Y, and Z are each independently CH, CR, or N;

[0133] R and R1 are each independently selected from halogen, cyano, nitro, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, and -N(C1-C6 alkyl)2, or

[0134] The two R and / or R1 on adjacent carbon atoms together with the carbon atoms to which they are attached form a 5-6 membered ring, optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur;

[0135] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, C6-C 10 Aryl, 5- to 12-membered heteroaryl, and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, and the cycloalkyl, aryl, and heteroaryl groups are optionally substituted with 1-3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy, provided that R2 and R3 are not simultaneously hydrogen; and

[0136] n is 0, 1, or 2.

[0137] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0138] X, Y, and Z are each independently CH, CR, or N;

[0139] R and R1 are each independently selected from halogen, cyano, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy, or

[0140] The two R and / or R1 on adjacent carbon atoms together with the carbon atoms to which they are attached form a 5-6 membered ring, optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur;

[0141] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, or C6-C alkyl optionally substituted with 1-3 halogens. 10 Aryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, provided that R2 and R3 are not simultaneously hydrogen; and

[0142] n is 0, 1, or 2.

[0143] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0144] X, Y, and Z are each independently CH, CR, or N;

[0145] R and R1 are each independently selected from fluorine, chlorine, bromine, iodine, cyano, nitro, hydroxyl, methyl, ethyl, n-propyl, isopropyl, trifluoromethyl, trichloromethyl, difluoromethyl, dichloromethyl, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy, -NH2, -NH(CH3) and -N(CH3)2, or

[0146] The two R and / or R1 on adjacent carbon atoms together with the carbon atoms to which they are attached form a 5-6 membered ring, optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur;

[0147] R2 and R3 are each independently selected from hydrogen, fluorine, chlorine, bromine, cyano, hydroxyl, methyl, ethyl, n-propyl, isopropyl, trifluoromethyl, trichloromethyl, difluoromethyl, dichloromethyl, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy, -NH2, -NH(CH3), -N(CH3)2, -NH-cyclopropyl, -NH-cyclobutyl, -NH-cyclopentyl, -NH-cyclohexyl, -NH-ethynyl, -NH-(1-propynyl), -NH-(2-propynyl), phenyl, 4-fluorophenyl, pyridyl, and -O-CH2-pyridyl, provided that R2 and R3 are not simultaneously hydrogen; and

[0148] n is 0, 1, or 2.

[0149] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0150] X, Y, and Z are each independently CH, CR, or N;

[0151] R and R1 are each independently selected from fluorine, chlorine, bromine, cyano, methyl, trifluoromethyl, and methoxy, or

[0152] The two R and / or R1 on adjacent carbon atoms together with the carbon atoms to which they are attached form a 5-6 membered ring, optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur;

[0153] R2 and R3 are each independently selected from hydrogen, fluorine, chlorine, cyano, hydroxyl, trifluoromethyl, methoxy, isopropoxy, -NH-cyclopropyl, -NH-(2-propynyl), phenyl, 4-fluorophenyl, and -O-CH2-pyridyl, provided that R2 and R3 are not simultaneously hydrogen; and

[0154] n is 0, 1, or 2.

[0155] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0156] No more than one of X, Y and Z is N and the rest are CH or CR;

[0157] R and R1 are each independently selected from halogen, cyano, nitro, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl and -N(C1-C6 alkyl)2;

[0158] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, -S(O)2-C1-C6 alkyl, -C(O)2-C1-C6 alkyl, C3-C8 cycloalkyl, 3- to 8-membered heterocyclic groups, C6-C 10 Aryl, 5- to 12-membered heteroaryl, and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heterocyclic and heteroaryl groups contain 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, and the cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally substituted with 1-3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy groups, provided that R2 and R3 are not simultaneously hydrogen; and

[0159] n is 0, 1, or 2.

[0160] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0161] No more than one of X, Y and Z is N and the rest are CH or CR;

[0162] R and R1 are each independently selected from halogen, cyano, nitro, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl and -N(C1-C6 alkyl)2;

[0163] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, C6-C 10 Aryl, 5- to 12-membered heteroaryl, and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, and the cycloalkyl, aryl, and heteroaryl groups are optionally substituted with 1-3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy, provided that R2 and R3 are not simultaneously hydrogen; and

[0164] n is 0, 1, or 2.

[0165] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0166] No more than one of X, Y and Z is N and the rest are CH or CR;

[0167] R and R1 are each independently selected from halogen, cyano, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy;

[0168] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, or C6-C alkyl optionally substituted with 1-3 halogens. 10 Aryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, provided that R2 and R3 are not simultaneously hydrogen; and

[0169] n is 0, 1, or 2.

[0170] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0171] X, Y, and Z are each independently CH or CR;

[0172] R and R1 are each independently selected from halogen, cyano, nitro, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, and -N(C1-C6 alkyl)2, or

[0173] The two R and / or R1 on adjacent carbon atoms together with the carbon atoms to which they are attached form a 5-6 membered ring, optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur;

[0174] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, -S(O)2-C1-C6 alkyl, -C(O)2-C1-C6 alkyl, C3-C8 cycloalkyl, 3- to 8-membered heterocyclic groups, C6-C 10 Aryl, 5- to 12-membered heteroaryl, and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heterocyclic and heteroaryl groups contain 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, and the cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally substituted with 1-3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy groups, provided that R2 and R3 are not simultaneously hydrogen; and

[0175] n is 0, 1, or 2.

[0176] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0177] X, Y, and Z are each independently CH or CR;

[0178] R and R1 are each independently selected from halogen, cyano, nitro, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, and -N(C1-C6 alkyl)2, or

[0179] The two R and / or R1 on adjacent carbon atoms together with the carbon atoms to which they are attached form a 5-6 membered ring, optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur;

[0180] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, C6-C 10 Aryl, 5- to 12-membered heteroaryl, and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, and the cycloalkyl, aryl, and heteroaryl groups are optionally substituted with 1-3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy, provided that R2 and R3 are not simultaneously hydrogen; and

[0181] n is 0, 1, or 2.

[0182] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0183] X, Y, and Z are each independently CH or CR;

[0184] R and R1 are each independently selected from halogen, cyano, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy, or

[0185] The two R and / or R1 on adjacent carbon atoms together with the carbon atoms to which they are attached form a 5-6 membered ring, optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur;

[0186] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, or C6-C alkyl optionally substituted with 1-3 halogens. 10Aryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, provided that R2 and R3 are not simultaneously hydrogen; and

[0187] n is 0, 1, or 2.

[0188] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0189] Only one of X, Y, and Z is N and the rest are CH or CR;

[0190] R and R1 are each independently selected from halogen, cyano, nitro, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl and -N(C1-C6 alkyl)2;

[0191] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, -S(O)2-C1-C6 alkyl, -C(O)2-C1-C6 alkyl, C3-C8 cycloalkyl, 3- to 8-membered heterocyclic groups, C6-C 10 Aryl, 5- to 12-membered heteroaryl, and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heterocyclic and heteroaryl groups contain 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, and the cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally substituted with 1-3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy groups, provided that R2 and R3 are not simultaneously hydrogen; and

[0192] n is 0, 1, or 2.

[0193] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0194] Only one of X, Y, and Z is N and the rest are CH or CR;

[0195] R and R1 are each independently selected from halogen, cyano, nitro, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl and -N(C1-C6 alkyl)2;

[0196] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, C6-C 10 Aryl, 5- to 12-membered heteroaryl, and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, and the cycloalkyl, aryl, and heteroaryl groups are optionally substituted with 1-3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy, provided that R2 and R3 are not simultaneously hydrogen; and

[0197] n is 0, 1, or 2.

[0198] In some embodiments of compounds of formula I, I-1, I-2, I-3, or I-4:

[0199] Only one of X, Y, and Z is N and the rest are CH or CR;

[0200] R and R1 are each independently selected from halogen, cyano, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy;

[0201] R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, or C6-C alkyl optionally substituted with 1-3 halogens. 10 Aryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, provided that R2 and R3 are not simultaneously hydrogen; and

[0202] n is 0, 1, or 2.

[0203] In some embodiments, compounds selected from the following or pharmaceutically acceptable salts, stereoisomers, solvates, or isotopic derivatives thereof are provided:

[0204] A second aspect of the invention provides a pharmaceutical composition comprising a compound of formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, and one or more pharmaceutically acceptable carriers.

[0205] In some embodiments, the pharmaceutical composition comprises 0.1 wt% to 99.5 wt% of a compound of formula I or a pharmaceutically acceptable salt, stereoisomer, solvate, prodrug, metabolite, or isotope derivative thereof as an active ingredient, preferably 0.5 wt% to 99.5 wt%, more preferably 1 wt% to 50 wt%, for example 1 wt%, 1.5 wt%, 2 wt%, 5 wt%, 10 wt%, 15 wt%, 20 wt%, 25 wt%, 30 wt%, or 50 wt% of the active ingredient. The remainder of the pharmaceutical composition is a pharmaceutically acceptable carrier.

[0206] In some embodiments, the pharmaceutical composition comprises two, three, or more pharmaceutically acceptable carriers. These pharmaceutically acceptable carriers include those conventional in the pharmaceutical field, such as diluents, fillers, binders, disintegrants, lubricants, wetting agents, solubilizers, solvents, colorants, fragrances, absorption enhancers, surfactants, and adsorbents. Examples include, but are not limited to, starch, pregelatinized starch, sodium carboxymethyl starch, powdered sugar, lactose, calcium phosphate, magnesium stearate, talc, micronized silica gel, dextrin, cellulose and its derivatives (e.g., hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose (HPMC), sodium carboxymethyl cellulose, etc.), microcrystalline cellulose, mannitol, sorbitol, polysorbate 80, polyethylene glycol, water, water for injection, physiological saline, glucose solution, etc. The pharmaceutical composition may also contain various other commonly used additives, such as preservatives, emulsifiers, suspending agents, and flavoring agents.

[0207] The pharmaceutical composition can be prepared into any pharmaceutically acceptable dosage form using any conventional technique in the art, including but not limited to tablets, capsules, pills, granules, syrups, injections, solutions, suspensions, powders (including sterile powders for injection), etc. The pharmaceutical composition of the present invention can be administered to a subject (e.g., human or non-human mammal) via any route of administration, including, for example, oral, intravenous, intraperitoneal, intramuscular, local, transdermal, ocular, nasal, inhalation, subcutaneous, oral, sublingual, rectal, etc. The effective amount of the compound of formula I of the present invention or a pharmaceutically acceptable salt thereof depends on a variety of factors, including but not limited to: the specific compound to be administered; the species, size, age, and general health condition of the mammal; the severity of the disease; the individual patient's response; the route of administration; the bioavailability characteristics of the administered formulation; the selected dosage regimen; the use of other concomitant medications, etc., which can generally be determined by the attending physician according to routine practice. Generally, the effective amount is typically in the range of about 0.001 to about 100 mg / kg body weight / day, preferably about 0.01 to about 50 mg / kg body weight / day. In some cases, dose levels below the lower limit of the above range may be more than sufficient, while in other cases, a larger dose may be required without causing any harmful side effects. Such a larger dose is usually divided into several smaller doses to be administered throughout the day.

[0208] A third aspect of the invention provides a pharmaceutical composition comprising a compound of Formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, or comprising a compound of Formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, and one or more pharmaceutically acceptable carriers, for the prevention and / or treatment of diseases associated with abnormal activity of the BRG1 / BRM bromine domain and / or proton-sensing receptor GPR65.

[0209] A fourth aspect of the invention provides the use of a compound of Formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, or a pharmaceutical composition comprising a compound of Formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, and one or more pharmaceutically acceptable carriers for the prevention and / or treatment of diseases associated with abnormal activity of the BRG1 / BRM bromine domain and / or the proton-sensing receptor GPR65.

[0210] The fifth aspect of the invention provides the use of a compound of Formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, or a pharmaceutical composition comprising a compound of Formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, and one or more pharmaceutically acceptable carriers, in the preparation of a medicament for the prevention and / or treatment of diseases associated with abnormal activity of the BRG1 / BRM bromine domain and / or the proton-sensing receptor GPR65.

[0211] A sixth aspect of the invention provides a method for preventing and / or treating diseases associated with abnormal activity of the BRG1 / BRM bromine domain and / or the proton-sensing receptor GPR65 in subjects with such need, the method comprising administering to the subject an effective amount of a compound of formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, or a pharmaceutical composition comprising a compound of formula I or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, and one or more pharmaceutically acceptable carriers.

[0212] In the above three to six aspects, the diseases associated with the abnormal activity of the BRG1 / BRM bromine domain and / or the proton-sensing receptor GPR65 are selected from proliferative diseases, immune diseases, asthma, chronic obstructive pulmonary disease (COPD), and acute respiratory distress syndrome (ARDS), wherein the proliferative diseases are tumors selected from melanoma, renal cell carcinoma, gastrointestinal cancer, acute myeloid leukemia, pancreatic cancer, triple-negative breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, and glioma. Glioblastoma is preferred; the immune disease is selected from psoriasis, psoriatic arthritis, rheumatoid arthritis, multiple sclerosis, systemic lupus erythematosus, Hashimoto's thyroiditis, Graves' disease, uveitis, ulcerative colitis, Crohn's disease, autoimmune uveoretinitis, systemic vasculitis, polymyositis and dermatomyositis, scleroderma, Sjögren's syndrome, ankylosing spondylitis, sarcoidosis, autoimmune hemolytic anemia, autoimmune myocarditis, type I diabetes, and atopic dermatitis.

[0213] The compounds or pharmaceutical compositions of the present invention can be administered to a subject (e.g., a human or a non-human mammal) via any route of administration, including, for example, oral, intravenous, intraperitoneal, intramuscular, local, transdermal, ocular, nasal, inhalation, subcutaneous, oral, sublingual, rectal, etc.

[0214] The attending physician may adjust the dosage and frequency of administration of the compounds or pharmaceutical compositions of the present invention, taking into account factors such as the patient's age, general health condition, weight, and severity of the symptoms to be treated. Generally, the total daily dose of the compounds or pharmaceutical compositions of the present invention is typically from about 0.1 to about 1000 mg of active ingredient per day, for example, from about 1 to about 800 mg per day, from about 10 to about 600 mg per day, or from about 50 to about 500 mg per day, administered as a single dose or in 2, 3, or 4 divided doses.

[0215] The main beneficial effects of the compounds of this invention include the following aspects:

[0216] (1) The compounds of the present invention have good affinity for both the BRG1 / BRM bromine domain and GPR65, and are a class of multi-target inhibitors, which can combine different anti-tumor mechanisms to achieve greater therapeutic efficacy and reduce the development of drug resistance.

[0217] (2) The compounds of the present invention exhibit broad-spectrum anti-proliferative activity against a variety of patient-derived glioblastoma cell lines in vitro and have the potential to be developed into a universal glioblastoma therapeutic.

[0218] (3) The compounds of the present invention showed good in vivo anti-brain tumor activity in a mouse glioblastoma in vivo model and have the potential to be developed into an effective glioblastoma therapeutic.

[0219] (4) The compounds of the present invention also showed a good anti-tumor trend in a mouse-derived colon cancer in vivo model, and are expected to show therapeutic effects in a variety of solid tumors;

[0220] (5) As a novel small molecule tumor inhibitor targeting BRG1 / BRM, the compound of the present invention has a direct anti-GBM cell proliferation effect, thus having a wider range of clinical application value.

[0221] (6) As a new type of small molecule immunosuppressant, the compounds of the present invention have more universal targets and are more universally applicable to the treatment of tumors of different subtypes. Compared with traditional biological immunotherapies such as immune checkpoint inhibitors, CAR-T, oncolytic viruses, etc., small molecule therapeutic agents have better blood-brain barrier permeability and pharmacokinetic properties, simpler administration methods, easier control of treatment doses, and help reduce immune-related adverse reactions.

[0222] General method for preparing the compounds of the present invention

[0223] Compounds of Formula I can be prepared from commercially available or readily prepared starting materials according to synthetic and purification methods known to those skilled in the art of organic synthesis. Exemplary methods for preparing compounds of Formula I are described in the following schemes and examples. It should be understood that these exemplary methods do not constitute any limitation on the invention, and those skilled in the art of organic synthesis will understand alternative synthetic routes.

[0224] Option 1

[0225] Scheme 1 illustrates a general method for preparing compounds of formula I-1, wherein the variables R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I. The method is as follows:

[0226] Commercially available mono- or poly-substituted aryl isocyanates or heteroaryl isocyanates react with intermediate 1 (Int 1) in an organic solvent (e.g., any one or more of dichloromethane, chloroform, tetrahydrofuran, N,N-dimethylformamide) in the presence of a base (e.g., pyridine, triethylamine, or 4-dimethylaminopyridine) to form a compound of formula I-1.

[0227] Option 2

[0228] Scheme 2 illustrates another method for preparing compounds of formula I-1, wherein the variables R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I. The method includes the following steps:

[0229] Step 1: React mono- or poly-substituted aryl amines or heteroaryl amines with triphosgene in an organic solvent (e.g., any one or more of dichloromethane, chloroform, tetrahydrofuran, N,N-dimethylformamide) in the presence of a base (e.g., pyridine, triethylamine, or 4-dimethylaminopyridine) to obtain the corresponding mono- or poly-substituted aryl isocyanate or heteroaryl isocyanate intermediate, which can be used directly for the preparation in Step 2 without separation and purification;

[0230] Step 2: Add intermediate 1 (Int 1) and a base (e.g., pyridine, triethylamine or 4-dimethylaminopyridine) to the reaction solution obtained in step 1, and generate a compound of formula I-1 by condensation reaction.

[0231] Option 3

[0232] Scheme 3 illustrates a general method for preparing compounds of formula I-2, wherein the variables R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I. The method includes the following steps:

[0233] Step 1: React mono- or poly-substituted aromatic or heterocyclic phenols with triphosgene in an organic solvent (e.g., any one or more of dichloromethane, chloroform, tetrahydrofuran, N,N-dimethylformamide) in the presence of a base (e.g., pyridine, triethylamine, or 4-dimethylaminopyridine) to obtain the corresponding mono- or poly-substituted aryl carbamate or heteroaryl carbamate intermediate, which can be used directly in the preparation of Step 2 without separation and purification;

[0234] Step 2: Add intermediate 1 (Int 1) and a base (e.g., pyridine, triethylamine or 4-dimethylaminopyridine) to the reaction solution obtained in step 1, and generate a compound of formula I-2 by condensation reaction.

[0235] Option 4

[0236] Scheme 4 illustrates a general method for preparing compounds of formula I-3, wherein the variables R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I. The method includes the following steps:

[0237] Step 1: React mono- or poly-substituted aryl or heteroaryl acetic acid with an acylation agent (e.g., thionyl chloride or oxalyl chloride) in an organic solvent (e.g., dichloromethane, chloroform, tetrahydrofuran, N,N-dimethylformamide) to obtain the corresponding mono- or poly-substituted arylacetyl chloride or heteroarylacetyl chloride intermediate, which can be used directly in the preparation of Step 2 without separation and purification;

[0238] Step 2: Add intermediate 1 (Int 1) and a base (e.g., pyridine, triethylamine or 4-dimethylaminopyridine) to the reaction solution obtained in step 1, and generate a compound of formula I-3 by condensation reaction.

[0239] Option 5

[0240] Example 5 illustrates a general method for preparing compounds of formula I-4, wherein the variables R1, R2, R3, X, Y, Z, and n are as defined above for compounds of formula I. The method includes the following steps:

[0241] Step 1: React the mono- or poly-substituted aryl or heteroaryl acetoacetic acid with an acylation reagent (e.g., sulfoxide or oxalyl chloride) in an organic solvent (e.g., dichloromethane, chloroform, tetrahydrofuran, N,N-dimethylformamide) to obtain the corresponding 2-oxo-2-mono- or poly-substituted arylacetyl chloride or 2-oxo-2-mono- or poly-substituted heteroarylacetyl chloride intermediate (Int 2), which can be used directly in the preparation of Step 2 without separation and purification;

[0242] Step 2: Add intermediate 1 (Int 1) and a base (e.g., pyridine, triethylamine or 4-dimethylaminopyridine) to the reaction solution obtained in step 1, and generate a compound of formula I-4 through condensation reaction.

[0243] Example

[0244] The present invention will be further described in detail below with reference to embodiments. Obviously, the described embodiments are only a part of the present invention, and not all of it. These embodiments are only used to illustrate the present invention and should not be construed as limiting the scope of protection of the present invention. Based on the embodiments of the present invention, all other technical solutions obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0245] List of abbreviations:

[0246] Unless otherwise specified, abbreviations not listed below have the meanings commonly understood by those skilled in the art.

[0247] In the examples, the abbreviations PE represent petroleum ether, EA represent ethyl acetate, DCM represent dichloromethane, MeOH represent methanol, EtOH represent ethanol, THF represent tetrahydrofuran, DCE represent 1,2-dichloroethane, DMSO-d6 represents deuterated dimethyl sulfoxide, DMF represents N,N-dimethylformamide, DCE represents ethylene glycol dimethyl ether, LHMDS represents lithium bis(trimethylsilyl)amino, Et3N represents triethylamine, iPrOH represents isopropanol, TLC represents thin-layer chromatography, and mp represents melting point.

[0248] General experimental conditions:

[0249] The starting materials and solvents used in the examples were commercially available or prepared according to literature methods. Unless otherwise specified, the reaction was carried out at room temperature (20°C); silica gel column chromatography was performed using an automated medium-pressure rapid column chromatography system, employing a Teledyne ISCO / CombiFlash NextGen 300+; thin-layer chromatography was performed using silica gel 60F254 plates (Merck); analytical HPLC-MS was performed using an Agilent / 1260 Infinity II liquid chromatography system and an Agilent / G6125B mass spectrometry system; NMR was performed using a Bruker AVANCE II 400MHz nuclear magnetic resonance spectrometer, where s represents a singlet, bs or brs represents a broad singlet, d represents a doublet, t represents a triplet, and m represents a multiplet. Melting points, except for solid products, were determined using a Buchi / Melting Point M-565 melting point apparatus.

[0250] HPLC-MS analytical method:

[0251] Reversed-phase column: Agilent Poroshell 120 EC-C18, 2.7μm, 4.6 x 100mm, column temperature 40℃

[0252] Detector: Ultraviolet detection wavelength 254 nm, mass spectrometry ionization source electrospray ionization (ESI)

[0253] Mobile phase: A: 0.1% formic acid aqueous solution, B: 0.1% formic acid acetonitrile solution

[0254] Gradient elution method:

[0255] I. Preparation of intermediates

[0256] Preparation of intermediate Int1-1 (4-chloro-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine-7-onium chloride)

[0257] Step 1: Synthesis of Int1-9

[0258] Compound 70 (18.4 mmol, 5.0 g) was completely dispersed in 90 mL of ethanol in a 200 mL round-bottom flask with stirring at room temperature. Sodium ethoxide (58.5 mmol, 19.9 mL, 20% wt in EtOH) and formamidin hydrochloride (43.8 mmol, 3.5 g) were then added sequentially. After the additions were complete, the reaction mixture was transferred to 70 °C and reacted for 5 hours. The reaction was monitored by TLC (evolving solvent: PE / EA = 3:1, R...). f =0.2). After the reaction was complete, the temperature was lowered to room temperature, and excess ethanol was removed under reduced pressure. The residue was diluted with EA (50 mL) and saturated ammonium chloride aqueous solution (50 mL) and extracted, and the organic and aqueous phases were collected separately. The pH of the aqueous phase was adjusted to 5 with acetic acid, and then the aqueous phase was extracted with EA (4 × 50 mL). The organic phases were combined and washed once with water and once with brine. The organic phase was dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to obtain a yellow solid crude product. After purification by slurrying at room temperature (n-hexane / EA = 8 / 1) for 12 h, filtration yielded 3.9 g of pale yellow solid compound Int1-9, yield 84%, mp 132–135 °C.

[0259] 1 ¹H NMR (400MHz, deuterated chloroform) δ 8.05 (s, 1H), 4.42 (s, 2H), 3.64 (t, J = 5.6Hz, 2H), 2.63 (s, 2H), 1.49 (s, 9H).

[0260] Step 2: Synthesis of Int1-10

[0261] At room temperature under nitrogen protection, Int1-9 (19.9 mmol, 5.0 g) and triphenylphosphine (39.8 mmol, 10.4 g) were dispersed and dissolved evenly in 120 mL of DCE. Carbon tetrachloride (59.7 mmol, 9.18 g, 5.76 mL) was added, and the reaction was carried out at 70 °C for 4 hours. The reaction was monitored by TLC (developing solvent: EA / MeOH = 10:1, R f =0.9). After the reaction was complete, the temperature was lowered to room temperature, diluted with DCM and water, and extracted. The aqueous phase was extracted with DCM (3 × 50 mL), washed once with brine, and the organic phase was collected and dried over anhydrous sodium sulfate. The solvent was removed under reduced pressure to give a brown solid crude product. The crude product was purified by rapid silica gel column chromatography (PE / EA = 20 / 3) to give 4.27 g of white solid Int1-10, yield 80%, mp 95–97 °C.

[0262] 1 ¹H NMR (400MHz, deuterated chloroform) δ 8.79 (s, 1H), 4.64 (s, 2H), 3.74 (t, J = 6.0Hz, 2H), 2.87 (t, J = 6.0Hz, 2H), 1.49 (s, 9H).

[0263] Step 3: Synthesis of Int1-1

[0264] Compound Int1-10 (15.8 mmol, 4.27 g) was dispersed in 40.0 mL of ethyl acetate under ice bath conditions. The solution was then gradually added dropwise to a 40.0 mL solution of ethyl hydrochloric acid. After the addition was complete, the mixture was stirred at room temperature for 48 hours. The reaction solution became white and turbid. The reaction was monitored by TLC (developing solvent: DCM / MeOH = 20:1, R...). f =0.4). After the reaction was complete, the reaction solution was directly filtered, the filter cake was washed with a small amount of EA, and dried under vacuum at room temperature to obtain an off-white solid Int1-13.1 g, yield 95%, mp>200℃.

[0265] 1 H NMR (400MHz, DMSO-d6) δ10.13(s,2H),8.92(s,1H),4.32(s,2H),3.45(t,J=6.0Hz,2H),3.02(t,J=6.0Hz,2H).

[0266] Preparation of intermediate Int1-4 (4-cyano-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine-7-onium chloride)

[0267] Step 1: Synthesis of Int1-11

[0268] Compound Int1-10 (5.57 mmol, 1.5 g) was dispersed in 20.0 mL of DMF under nitrogen protection at room temperature. Tetra(triphenylphosphine)palladium (0.557 mmol, 643 mg) and zinc cyanide (6.96 mmol, 817 mg) were added sequentially. After addition, the mixture was stirred at 90 °C for 2 hours. The reaction solution turned red. The reaction was monitored by TLC (evolving solvent: DCM / MeOH = 50:1, R0). f =0.6). After the reaction was complete, the sample was extracted with saturated sodium bicarbonate and EA. The aqueous phase was extracted with EA (4 × 20 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to give 2.4 g of crude product. The crude product was purified by rapid silica gel column chromatography (DCM / MeOH = 100 / 1) to give 111.2 g of pale yellow solid Int1-1, yield 80%.

[0269] 1 ¹H NMR (400MHz, deuterated chloroform) δ 9.12 (s, 1H), 4.72 (s, 2H), 3.78 (t, J = 6.0Hz, 2H), 3.05 (t, J = 6.0Hz, 2H), 1.50 (s, 9H).

[0270] Step 2: Synthesis of Int1-4

[0271] Under electromagnetic stirring, compound Int1-11 (0.77 mmol, 200 mg) and dioxane hydrochloride solution (2 mL, 4 M) were added sequentially to a 10 mL round-bottom flask. The mixture was reacted at room temperature for 3 hours (TLC monitoring, developing solvent: DCM / MeOH = 50:1). After the reaction was complete, the solvent was removed under reduced pressure to obtain the crude product Int1-4, which was directly used in the next step.

[0272] Preparation of intermediate Int1-5 (4-(trifluoromethyl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine-7-onium chloride)

[0273] Step 1: Synthesis of Int1-12

[0274] Compound 71 (15.1 mmol, 3.0 g) was dispersed and pre-cooled in 60.0 mL of DME at -78 °C under nitrogen protection. LHMDS (18.1 mmol, 18 mL, 1.0 M in THF) was slowly added dropwise, and the reaction was continued at -78 °C for 1 hour after the addition was complete. Ethyl trifluoroacetate (19.9 mmol, 2.4 mL) was then slowly added dropwise to the reaction solution, and the reaction was continued at -78 °C for another 1 hour after the addition was complete. The reaction was then continued at room temperature for 2.5 hours. The reaction solution was a clear, deep yellow solution. The reaction was monitored by TLC (developing solvent: DCM / MeOH = 20:1, R...). f=0.4). After the reaction was complete, the reaction was quenched with saturated ammonium chloride, and the aqueous phase was extracted with EA (4 × 50 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was then purified by rapid silica gel column chromatography (DCM / MeOH = 100 / 1) to give 1.5 g of a brown oily substance of Int1-12, with a yield of 34%.

[0275] 1 ¹H NMR (400MHz, deuterated chloroform) δ 14.61 (s, 1H), 4.23 (s, 2H), 3.56 (t, J = 5.6Hz, 2H), 2.58 (t, J = 5.6Hz, 2H), 1.48 (s, 9H).

[0276] Step 2: Synthesis of Int1-13

[0277] Under stirring at room temperature, Int1-12 (3.4 mmol, 1.0 g) was completely dispersed in 20 mL of ethanol in a 50 mL round-bottom flask. Sodium ethoxide (10.8 mmol, 3.6 mL, 20% wt in EtOH) and formamidin hydrochloride (8.2 mmol, 0.65 g) were then added sequentially. After the additions were complete, the reaction mixture was transferred to 70 °C and reacted for 5 hours. The reaction was monitored by TLC (evolving solvent: PE / EA = 3:1, R...). f =0.2). After the reaction was complete, the temperature was lowered to room temperature, and excess ethanol was removed under reduced pressure. The residue was diluted with EA (10 mL) and saturated ammonium chloride aqueous solution (10 mL) and extracted, and the organic and aqueous phases were collected separately. The aqueous phase was then extracted with EA (4 × 10 mL), the organic phases were combined, washed once with water and once with brine, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to obtain a yellow solid crude product. The crude product was purified by rapid silica gel column chromatography to obtain 141.7 mg of a brown oily substance of Int1-13, with a yield of 17%.

[0278] 1 ¹H NMR (400MHz, deuterated chloroform) δ 9.12 (s, 1H), 4.75 (s, 2H), 3.74 (t, J = 6.0Hz, 2H), 3.04 (t, J = 6.0Hz, 2H), 1.50 (s, 9H).

[0279] Step 3: Synthesis of Int1-5

[0280] Under electromagnetic stirring, compound Int1-13 (0.27 mmol, 83 mg) and hydrochloric acid-ethyl acetate solution (1.5 mL, 4 M) were added sequentially to a 10 mL round-bottom flask. The mixture was reacted at room temperature for 3 h (TLC monitoring, developing solvent: DCM / MeOH = 50:1). After the reaction was complete, the solvent was removed under reduced pressure to obtain the crude product Int1-5, which was directly used in the next step.

[0281] Preparation of intermediate Int1-6 (4-fluoro-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine-7-onium chloride)

[0282] Step 1: Synthesis of Int1-14

[0283] Compound Int1-10 (5.57 mmol, 1.5 g) was dispersed in 20.0 mL DMSO under nitrogen protection at room temperature. Cesium fluoride (8.36 mmol, 1.27 g) was added, and the mixture was stirred at 70 °C for 3 hours after the addition was complete. The reaction solution turned brown. The reaction was monitored by TLC (evolving solvent: DCM / MeOH = 50:1, R0). f =0.6, the same point as the starting material (low-resolution LC-MS is needed for auxiliary judgment). After the reaction was complete, the sample was extracted with saturated sodium bicarbonate and EA. The aqueous phase was extracted with EA (4 × 20 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to obtain 1.4 g of crude product. The crude product was purified by rapid silica gel column chromatography (DCM / MeOH = 100 / 1) to obtain 182 mg of a colorless oily substance of Int1-14, with a yield of 13%.

[0284] 1 ¹H NMR (400MHz, deuterated chloroform) δ 8.70 (d, J = 1.6 Hz, 1H), 4.67 (s, 2H), 3.73 (t, J = 6.0 Hz, 2H), 2.82 (t, J = 6.0 Hz, 2H), 1.49 (s, 9H).

[0285] Step 2: Synthesis of Int1-6

[0286] Under electromagnetic stirring, compound Int1-14 (0.72 mmol, 182 mg) and dioxane hydrochloride solution (1.5 mL, 4 M) were added sequentially to a 10 mL round-bottom flask. The mixture was reacted at room temperature for 3 h (TLC monitoring, developing solvent: DCM / MeOH = 50:1). After the reaction was complete, the solvent was removed under reduced pressure to obtain crude Int1-6, which was directly used in the next step.

[0287] Preparation of intermediate Int1-7 (5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine-7-onium chloride)

[0288] Step 1: Synthesis of Int1-15

[0289] Compound Int1-10 (1.92 mmol, 500 mg) was dispersed in 4 mL of methanol under nitrogen protection at room temperature. Zinc powder (7.68 mmol, 500 mg) and acetic acid (19.2 mmol, 1.1 mL) were added. After the addition was complete, the mixture was stirred and refluxed at 70 °C for 4 hours. The reaction solution turned yellow. The reaction was monitored by TLC (evolving solvent: PE / EA = 1:1, R...). f =0.2,). After the reaction was complete, the insoluble matter was removed by filtration, the filter cake was washed with a small amount of EA, the filtrate was collected, and extracted with saturated NaHCO3 and EA. The aqueous phase was extracted with EA (4 × 20 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to give 613 mg of crude product. The crude product was purified by rapid silica gel column chromatography to give 124 mg of a pale yellow oily substance of Int1-15, with a yield of 28%.

[0290] 1 ¹H NMR (400MHz, deuterated chloroform) δ 9.00 (s, 1H), 8.50 (s, 1H), 4.64 (s, 2H), 3.71 (t, J = 6.0Hz, 2H), 2.84 (t, J = 6.0Hz, 2H), 1.49 (s, 9H).

[0291] Step 2: Synthesis of Int1-7

[0292] Under electromagnetic stirring, compound Int1-15 (0.30 mmol, 71 mg) and hydrochloric acid-ethyl acetate solution (1.5 mL, 4 M) were added sequentially to a 10 mL round-bottom flask. The mixture was reacted at room temperature for 3 h (TLC monitoring, developing solvent: DCM / MeOH = 50:1). After the reaction was complete, the solvent was removed under reduced pressure to obtain the crude product Int1-7, which was directly used in the next step.

[0293] Preparation of intermediate Int1-8 (4-hydroxy-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine-7-onium chloride)

[0294] Under electromagnetic stirring, compound Int1-14 (0.72 mmol, 182 mg) and hydrochloric acid-methanol solution (1.5 mL, 4 M) were added sequentially to a 10 mL round-bottom flask. The mixture was reacted at room temperature for 3 h (TLC monitoring, developing solvent: DCM / MeOH = 50:1). After the reaction was complete, the solvent was removed under reduced pressure to obtain the crude product Int1-8, which was directly used in the next step. LC-MS (ESI) m / z C7H 10 N3O + [M+H] + Calculated value: 152.1, Measured value: 152.1.

[0295] Preparation of intermediate Int1-16 (2,4-diphenyl-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine)

[0296] Under nitrogen protection at room temperature, compounds Int1-2 (0.42 mmol, 100 mg), phenylboronic acid 72 (0.99 mmol, 121.7 mg), and tetrakis(triphenylphosphine)palladium (0.04 mmol, 48 mg) were dispersed in ethylene glycol dimethyl ether. Then, 0.6 mL of a 2M sodium carbonate aqueous solution was added and dispersed again. The mixture was then transferred to a microwave reactor and microwaved at 150 °C for 1 hour. The reaction was monitored by TLC (evolving solvent: DCM / MeOH = 20:1). After the reaction was complete, the mixture was extracted with water and DCM. The aqueous phase was extracted with DCM (4 × 10 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to give 187 mg of a greenish-yellow oily crude product. The crude product was then purified by rapid silica gel column chromatography (DCM / MeOH = 20 / 1) to give 42 mg of a white solid, Int1-16, in 35% yield.

[0297] 1 ¹H NMR (400MHz, deuterated chloroform) δ 8.47 (dd, J = 7.2, 2.8Hz, 2H), 7.76–7.65 (m, 2H), 7.56–7.41 (m, 6H), 4.23 (s, 2H), 3.13 (t, J = 5.6Hz, 2H), 2.89 (t, J = 5.6Hz, 2H). LC-MS (ESI) m / z C 19 H 18 N3 + [M+H] + Calculated value: 288.1, Measured value: 288.1.

[0298] Preparation of intermediate Int1-17 (2,4-bis(4-fluorophenyl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine)

[0299] Under nitrogen protection at room temperature, compounds Int1-2 (0.42 mmol, 100 mg), p-fluorophenylboronic acid 73 (0.99 mmol, 140 mg), and tetrakis(triphenylphosphine)palladium (0.04 mmol, 48 mg) were dispersed in ethylene glycol dimethyl ether. Then, 0.6 mL of a 2M sodium carbonate aqueous solution was added and dispersed again. The mixture was then transferred to a microwave reactor and microwaved at 150 °C for 1 hour. The reaction was monitored by TLC (evolving solvent: DCM / MeOH = 20:1). After the reaction was complete, the mixture was extracted with water and DCM. The aqueous phase was extracted with DCM (4 × 10 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to give 195 mg of a brown oily crude product. The crude product was then purified by rapid silica gel column chromatography (DCM / MeOH = 20 / 1) to give 57 mg of a white solid, Int1-17, in 43% yield.

[0300] 1 ¹H NMR (400MHz, deuterated chloroform) δ 8.47 (dd, J = 8.7, 5.8Hz, 2H), 7.70 (dd, J = 8.6, 5.5Hz, 2H), 7.16 (dt, J = 23.2, 8.6Hz, 4H), 4.20 (s, 2H), 3.13 (t, J = 5.7Hz, 2H), 2.87 (t, J = 5.7Hz, 2H). LC-MS (ESI) m / z C 19 H 16 F2N3 + [M+H] + Calculated value: 324.1, Measured value: 324.1.

[0301] Preparation of intermediate Int1-18 (4-phenyl-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine)

[0302] Under nitrogen protection at room temperature, compound Int1-1 (0.42 mmol, 85.7 mg), phenylboronic acid 72 (0.99 mmol, 121.7 mg), and tetrakis(triphenylphosphine)palladium (0.04 mmol, 48 mg) were dispersed in ethylene glycol dimethyl ether. Then, 0.6 mL of sodium carbonate aqueous solution (2 M) was added and dispersed again. The mixture was then transferred to a microwave reactor and microwaved at 150 °C for 1 hour. The reaction was monitored by TLC (evolving solvent: DCM / MeOH = 20:1). After the reaction was complete, the mixture was extracted with water and DCM. The aqueous phase was extracted with DCM (4 × 10 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to give 125 mg of a yellow oily crude product. The crude product was then purified by rapid silica gel column chromatography (DCM / MeOH = 20 / 1) to give 25 mg of a yellow-green oily substance, Int1-18, in 28% yield.

[0303] 1¹H NMR (400MHz, deuterated chloroform) δ 9.03 (s, 1H), 7.62–7.55 (m, 2H), 7.47 (dd, J = 5.2, 2.4Hz, 3H), 4.16 (s, 2H), 3.09 (t, J = 5.6Hz, 2H), 2.85 (t, J = 5.6Hz, 2H). LC-MS (ESI) m / z C 13 H 14 N3 + [M+H] + Calculated value: 212.1, Measured value: 212.1.

[0304] Preparation of intermediate Int1-19 (4-(4-fluorophenyl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine)

[0305] Under nitrogen protection at room temperature, compound Int1-1 (0.42 mmol, 85.7 mg), p-fluorophenylboronic acid 73 (0.99 mmol, 140 mg), and tetrakis(triphenylphosphine)palladium (0.04 mmol, 48 mg) were dispersed in ethylene glycol dimethyl ether. Then, 0.6 mL of sodium carbonate aqueous solution (2 M) was added and dispersed again. The mixture was then transferred to a microwave reactor and microwaved at 150 °C for 1 hour. The reaction was monitored by TLC (evolving solvent: DCM / MeOH = 20:1). After the reaction was complete, the mixture was extracted with water and DCM. The aqueous phase was extracted with DCM (4 × 10 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to give 146 mg of a yellow oily crude product. The crude product was then purified by rapid silica gel column chromatography (DCM / MeOH = 20 / 1) to give 40.5 mg of a yellow-green oily substance, Int1-19, in 42% yield.

[0306] 1 ¹H NMR (400MHz, deuterated chloroform) δ 9.01 (s, 1H), 7.60 (dd, J = 8.6, 5.4Hz, 2H), 7.17 (t, J = 8.6Hz, 2H), 4.15 (s, 2H), 3.10 (t, J = 5.7Hz, 2H), 2.84 (t, J = 5.7Hz, 2H). LC-MS (ESI) m / z C 13 H 13 FN3 + [M+H] + Calculated value: 230.1, Measured value: 230.1.

[0307] Preparation of intermediate Int1-20 (2-phenyl-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine)

[0308] Under nitrogen protection at room temperature, compounds Int1-3 (0.42 mmol, 85.7 mg), phenylboronic acid 72 (0.99 mmol, 121.7 mg), and tetrakis(triphenylphosphine)palladium (0.04 mmol, 48 mg) were dispersed uniformly in ethylene glycol dimethyl ether. Then, 0.6 mL of sodium carbonate aqueous solution (2 M) was added and dispersed again. The mixture was then transferred to a microwave reactor and microwaved at 150 °C for 1 hour. The reaction was monitored by TLC (evolving solvent: DCM / MeOH = 20:1). After the reaction was complete, the mixture was extracted with water and DCM. The aqueous phase was extracted with DCM (4 × 10 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to give 150 mg of a yellow oily crude product. The crude product was then purified by rapid silica gel column chromatography (DCM / MeOH = 20 / 1) to give 64 mg of a white solid Int1-20, with a yield of 73%.

[0309] 1 H NMR (400MHz, DMSO-d6) δ9.05 (s, 1H), 8.74 (s, 1H), 8.37 (dd, J=6.8, 2.8Hz, 2H), 7.90 (d, J= 2.0Hz,1H),7.60–7.45(m,5H),4.79(s,2H),3.81(t,J=5.6Hz,2H),2.90(t,J=5.6Hz,2H). LC-MS(ESI)m / z C 13 H 14 N3 + [M+H] + Calculated value: 212.1, Measured value: 212.1.

[0310] Preparation of intermediate Int1-21 (2-(4-fluorophenyl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine)

[0311] Under nitrogen protection at room temperature, compounds Int1-3 (0.42 mmol, 85.7 mg), p-fluorophenylboronic acid 73 (0.99 mmol, 140 mg), and tetrakis(triphenylphosphine)palladium (0.04 mmol, 48 mg) were dispersed in ethylene glycol dimethyl ether. Then, 0.6 mL of a 2M sodium carbonate aqueous solution was added and dispersed again. The mixture was then transferred to a microwave reactor and microwaved at 150 °C for 1 hour. The reaction was monitored by TLC (evolving solvent: DCM / MeOH = 20:1). After the reaction was complete, the mixture was extracted with water and DCM. The aqueous phase was extracted with DCM (4 × 10 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to give 200 mg of a yellow oily crude product. The crude product was then purified by rapid silica gel column chromatography (DCM / MeOH = 20 / 1) to give 78 mg of a white solid, Int1-21, in 81% yield.

[0312] 1 H NMR (400MHz, DMSO-d6) δ9.09(d,J=3.2Hz,1H),8.73(s,1H),8.41(dd,J=8.8,5.6Hz,2H),7.90(d,J=2.4Hz ,1H),7.59–7.43(m,2H),7.35(t,J=8.8Hz,2H),4.78(s,2H),3.81(t,J=5.6Hz,2H),2.90(t,J=5.6Hz,2H). LC-MS(ESI)m / z C 13 H 13 FN3 + [M+H] + Calculated value: 230.1, Measured value: 230.1.

[0313] Preparation of intermediate Int1-22 (4-(prop-2-yn-1-ylamino)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine-7-onium chloride)

[0314] Step 1: Synthesis of Int1-23

[0315] At room temperature, compound Int1-10 (7.42 mmol, 2.0 g) was dispersed in 15 mL of DMF, followed by the sequential addition of compound 74 (22.26 mmol, 1.2 g) and potassium carbonate (23.74 mmol, 3.28 g). After the addition was complete, the mixture was stirred at 105 °C for 3 hours. The reaction solution turned brown. The reaction was monitored by TLC (evolving solvent: PE / EA = 1:2, R...). f =0.5). After the reaction was complete, the mixture was extracted with water and EA. The aqueous phase was extracted with EA (4 × 20 mL), and the organic phases were combined. The mixture was dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to give 2.5 g of crude oil. The crude product was purified by rapid silica gel column chromatography (PE / EA = 1 / 1) to give 1.4 g of yellow oil of Int1-23, with a yield of 65%.

[0316] 1 ¹H NMR (400MHz, deuterated chloroform) δ 8.51 (s, 1H), 4.75 (br s, 1H), 4.48 (s, 2H), 4.32 (dd, J = 5.6, 2.4Hz, 2H), 3.72 (t, J = 6.0Hz, 2H), 2.45 (t, J = 6.0Hz, 2H), 2.26 (t, J = 2.4Hz, 1H), 1.47 (s, 9H).

[0317] Step 2: Synthesis of Int1-22

[0318] Compound Int1-23 (3.47 mmol, 1 g) was dispersed in 5 mL of ethyl acetate under electromagnetic stirring, followed by the addition of a hydrochloric acid-ethyl acetate solution (17.33 mmol, 4.3 mL, 4 M). The mixture was reacted at room temperature for 12 h (TLC monitoring, developing solvent: DCM / MeOH = 50:1). After the reaction was complete, the solvent was removed under reduced pressure to obtain 835 mg of crude Int1-22, a light brownish-white solid powder (hygroscopic), which was directly used in the next step.

[0319] II. Compound Preparation Examples

[0320] Example 1. Preparation of compound 1 (4-chloro-N-(3,4-difluorophenyl)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide) (General Synthetic Method A)

[0321] In an ice bath under nitrogen protection, Int1-1 (1.46 mmol, 300 mg) was dispersed and dissolved in 30 mL of dichloromethane. Triethylamine (7.3 mmol, 1.0 mL) and 3,4-difluorophenyl isocyanate 75 (2.9 mmol, 452 mg) were then slowly added sequentially to the system. The mixture was stirred at room temperature for 3 hours. The reaction solution was a pale yellow and clear. The reaction was monitored by TLC (evolving solvent: DCM / MeOH = 30:1, R...). f =0.5). After the reaction was complete, the mixture was extracted with water and EA. The aqueous phase was extracted with EA (4 × 10 mL). The organic phases were combined and dried over anhydrous sodium sulfate. The solvent was removed under reduced pressure to give 680 mg of crude pale yellow solid. The crude product was purified by rapid silica gel column chromatography (DCM / MeOH = 100 / 1) to give 345 mg of compound 1 as a white solid, mp 193–195 °C, yield 73%.

[0322] 1 H NMR(400MHz, DMSO-d6)δ8.98(s,1H),8.86(s,1H),7.63(ddd,J=14.0,7.6,2.4Hz,1H),7.36–7.28(m ,1H),7.27–7.22(m,1H),4.72(s,2H),3.81(t,J=6.0Hz,2H),2.85(t,J=6.0Hz,2H).LC-MS(ESI)m / z C 14 H 12 ClF2N4O + [M+H] + Calculated value: 325.1, Measured value: 325.1.

[0323] Example 2. Preparation of compound 20 (4-chloro-N-(pyridin-3-yl)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide) (General Synthetic Method B)

[0324] Under nitrogen protection in an ice bath, 3-aminopyridine 76 (2.18 mmol) was dispersed and dissolved in 5 mL of dichloromethane. Triphosgene (1.09 mmol) in 3 mL of dichloromethane solution and triethylamine (2.18 mmol) were then slowly added sequentially to the system. The mixture was stirred at room temperature for 6 hours. The reaction solution was used directly in the next reaction without purification.

[0325] The reaction solution was pre-cooled again in an ice bath, and 4 mL of dichloromethane solution containing triethylamine (7.28 mmol) and Int1-1 (1.46 mmol) was added sequentially. The reaction was stirred at room temperature for 12 hours, and the reaction was monitored by TLC (evolving solvent: DCM / MeOH = 20:1). After the reaction was complete, the mixture was extracted with water and DCM. The aqueous phase was extracted with DCM (4 × 10 mL), and the organic phases were combined and dried over anhydrous sodium sulfate. The solvent was removed under reduced pressure to obtain the crude product. The crude product was purified by rapid silica gel column chromatography (DCM / MeOH = 50 / 1) to give 144 mg of a pale yellow solid of compound 20, with a yield of 34%.

[0326] 1 H NMR (400MHz, DMSO-d6) δ8.98(s,1H),8.86(s,1H),8.66(d,J=2.8Hz,1H),8.17(dd,J=4.8,1.6Hz,1H),7.89(ddd ,J=8.4,2.8,1.6Hz,1H),7.28(dd,J=8.4,4.8Hz,1H),4.75(s,2H),3.83(t,J=6.0Hz,2H),2.86(t,J=6.0Hz,2H). LC-MS(ESI)m / zC 13 H 13 ClN5O + [M+H] + Calculated value: 290.1, measured value: 290.1.

[0327] Example 3. Preparation of compound 35 (2,4-dichloro-N-(3,4-difluorophenyl)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide).

[0328] Using Int1-2 instead of Int1-1, 20 mg of compound 35 was prepared as a white solid according to the general synthesis method A described in Example 1, with a yield of 13%.

[0329] 1 H NMR (400MHz, DMSO-d6) δ8.98 (s, 1H), 7.63 (ddd, J=13.6, 7.5, 2.8Hz, 1H), 7.33 (q, J=9.2Hz, 1H), 7.25 (q, J=6.0, 4.8Hz, 1H), 4.73 (s, 2H), 3.80 (t, J=6.0Hz, 2H), 2.82 (t, J=6.0Hz, 2H). LC-MS(ESI)m / z C 14 H 11 C l2 F2N4O + [M+H] + Calculated value: 359.0, Measured value: 359.0.

[0330] Example 4. Preparation of compound 38 (2-chloro-N-(3,4-difluorophenyl)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide).

[0331] Using Int1-3 instead of Int1-1, 60 mg of compound 38 was prepared as a yellow solid by general synthesis method A described in Example 1, with a yield of 55%.

[0332] 1 H NMR (400MHz, DMSO-d6) δ8.93 (s, 1H), 8.61 (s, 1H), 7.63 (ddd, J = 13.6, 7.6, 2.4Hz, 1H),7.36–7.21(m,2H),4.70(s,2H),3.75(t,J=5.6Hz,2H),2.85(t,J=5.6Hz,2H). LC-MS(ESI)m / z C 14 H 12 ClF2N4O + [M+H] + Calculated value: 325.1, Measured value: 325.1.

[0333] Example 5. Preparation of compound 25 (4-cyano-N-(3,4-difluorophenyl)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide)

[0334] Using Int1-4 instead of Int1-1, 215 mg of compound 25 was prepared as a light brown solid by general synthesis method A described in Example 1, with a yield of 89%.

[0335] 1H NMR(400MHz,DMSO-d6)δ9.19(s,1H),9.05(s,1H),7.64(dd,J=14.0,7.6Hz,1H) ,7.42–7.14(m,2H),4.79(s,2H),3.84(t,J=5.6Hz,2H),3.03(t,J=5.6Hz,2H). LC-MS(ESI)m / z C 15 H 12 F2N5O + [M+H] + Calculated value: 316.1, Measured value: 316.1.

[0336] Example 6. Preparation of compound 26 (N-(3,4-difluorophenyl)-4-(trifluoromethyl)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide)

[0337] Using Int1-5 instead of Int1-1, 53 mg of compound 26 was prepared as a white solid according to the general synthesis method A described in Example 1, with a yield of 36%.

[0338] 1 H NMR (400MHz, DMSO-d6) δ9.22 (s, 1H), 8.98 (s, 1H), 7.64 (ddd, J = 13.6, 7.6, 2.4Hz, 1H), 7.38–7.18 (m, 2H), 4.83 (s, 2H), 3.81 (t, J = 6.0Hz, 2H), 3.01 (d, J = 6.0Hz, 2H). LC-MS(ESI)m / z C 15 H 12 F5N4O + [M+H] + Calculated value: 359.1, Measured value: 359.1.

[0339] Example 7. Preparation of compound 27 (N-(3,4-difluorophenyl)-4-fluoro-5,8-dihydropyro[3,4-d]pyrimidine-7(6H)-formamide)

[0340] Compound 27 was prepared as a 38 mg white solid, yielding 17%, by replacing Int1-1 with Int1-6 and following the general synthesis method A described in Example 1.

[0341] 1H NMR (400MHz, DMSO-d6) δ8.97 (s, 1H), 8.78 (d, J = 1.6 Hz, 1H), 7.63 (ddd, J = 13.6, 7.6, 2. 4Hz,1H),7.38–7.21(m,2H),4.74(s,2H),3.79(t,J=5.6Hz,2H),2.81(t,J=5.6Hz,2H). LC-MS(ESI)m / zC 14 H 12 F3N4O + [M+H] + Calculated value: 309.1, Measured value: 309.1.

[0342] Example 8. Preparation of compound 28 (N-(3,4-difluorophenyl)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide)

[0343] Using Int1-7 instead of Int1-1, 83 mg of compound 28 was prepared as a white solid by general synthesis method A described in Example 1, with a yield of 55%.

[0344] 1 H NMR (400MHz, DMSO-d6) δ8.98(s,1H),8.93(s,1H),8.63(s,1H),7.64(ddd,J=13.6,7.6,2 .4Hz,1H),7.37–7.22(m,2H),4.69(s,2H),3.76(t,J=5.6Hz,2H),2.87(t,J=5.6Hz,2H). LC-MS(ESI)m / zC 14 H 13 F2N4O + [M+H] + Calculated value: 291.1, Measured value: 291.1.

[0345] Example 9. Synthesis of compound 41 (N-(3,4-difluorophenyl)-4-hydroxy-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide)

[0346] Using Int1-8 instead of Int1-1, 15 mg of compound 41 was prepared as a white solid according to the general synthesis method A described in Example 1, with a yield of 15%.

[0347] 1H NMR (400MHz, DMSO-d6) δ12.44(s,1H),8.87(s,1H),8.07(s,1H),7.63(ddd,J=13.6, 7.6, 2.4Hz, 1H), 7.37–7.21 (m, 2H), 4.39 (s, 2H), 3.65 (t, J = 5.6Hz, 2H), 2.46 (s, 3H). LC-MS(ESI)m / z C 14 H 13 F2N4O2 + [M+H] + Calculated value: 307.1, Measured value: 307.1.

[0348] Example 10. Synthesis of Compound 54 (N-(3,4-dichlorophenyl)-2,4-diphenyl-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide)

[0349] 22 mg of compound 54 was prepared as a white solid, yielding 44%, by replacing 3,4-difluorophenyl isocyanate 52 and Int1-1 with 3,4-dichlorophenyl isocyanate 77 and Int1-16 respectively, according to the general synthesis method A described in Example 1.

[0350] 1 H NMR (400MHz, DMSO-d6) δ9.35(s,1H),8.42(dd,J=6.8,2.8Hz,2H),7.98(d,J=2.4Hz,1H),7.79(dd ,J=6.8,2.8Hz,2H),7.65–7.46(m,8H),4.88(s,2H),3.78(t,J=5.6Hz,2H),2.95(t,J=5.6Hz,2H). LC-MS(ESI)m / z C 26 H 21 Cl2N4O + [M+H] + Calculated value: 475.1, Measured value: 475.1.

[0351] Example 11. Synthesis of Compound 55 (N-(3,4-dichlorophenyl)-2,4-bis(4-fluorophenyl)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide)

[0352] 27 mg of compound 55 was prepared as a white solid by replacing 3,4-dichlorophenyl isocyanate 77 and Int1-17 with 3,4-difluorophenyl isocyanate 52 and Int1-1 respectively, according to the general synthesis method A described in Example 1. The yield was 57%.

[0353] 1 H NMR (400MHz, DMSO-d6) δ9.14(s,1H),8.47(dd,J=8.8,5.8Hz,2H),7.94(d,J=2.4Hz,1H),7.88(dd,J=8.8,5.6Hz, 2H),7.59–7.48(m,2H),7.38(dt,J=12.8,8.8Hz,4H),4.87(s,2H),3.75(t,J=5.6Hz,2H),2.95(t,J=5.6Hz,2H). LC-MS(ESI)m / z C 26 H 19 Cl2F2N4O + [M+H] + Calculated value: 511.1, Measured value: 511.1.

[0354] Example 12. Synthesis of Compound 56 (N-(3,4-dichlorophenyl)-4-phenyl-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide)

[0355] 30 mg of compound 56 was prepared as a white solid by replacing 3,4-dichlorophenyl isocyanate 77 and Int1-18 with 3,4-difluorophenyl isocyanate 52 and Int1-1 respectively, according to the general synthesis method A described in Example 1. The yield was 80%.

[0356] 1 H NMR(400MHz,DMSO-d6)δ9.07(s,1H),9.02(s,1H),7.93–7.88(m,1H),7.70(dd,J=6.8,2.8Hz ,2H),7.53(dd,J=7.2,3.6Hz,5H),4.78(s,2H),3.70(t,J=5.6Hz,2H),2.92(t,J=5.6Hz,2H). LC-MS(ESI)m / z C 20 H 17 Cl2N4O + [M+H] + Calculated value: 399.1, Measured value: 399.1.

[0357] Example 13. Synthesis of Compound 57 (N-(3,4-dichlorophenyl)-4-(4-fluorophenyl)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide)

[0358] 71 mg of compound 57 was prepared as a white solid, with a yield of 98%, by replacing 3,4-dichlorophenyl isocyanate 52 and Int1-19 with 3,4-difluorophenyl isocyanate 77 and Int1-19 respectively, according to the general synthesis method A described in Example 1.

[0359] 1 H NMR (400MHz, DMSO-d6) δ9.13(s,1H),9.06(s,1H),7.92(d,J=2.4Hz,1H),7.78(dd,J=8.8,5.6Hz,2H),7.55(dd,J=8.8 ,2.4Hz,1H),7.50(d,J=8.8Hz,1H),7.36(t,J=8.8Hz,2H),4.78(s,2H),3.71(t,J=5.6Hz,2H),2.92(t,J=5.6Hz,2H). LC-MS(ESI)m / z C 20 H 16 Cl2FN4O + [M+H] + Calculated value: 417.1, Measured value: 417.1.

[0360] Example 14. Synthesis of Compound 58 (N-(3,4-dichlorophenyl)-2-phenyl-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide)

[0361] 29 mg of compound 58 was prepared as a white solid by replacing 3,4-dichlorophenyl isocyanate 77 and Int1-20 with 3,4-difluorophenyl isocyanate 52 and Int1-1 respectively, according to the general synthesis method A described in Example 1. The yield was 51%.

[0362] 1 H NMR (400MHz, DMSO-d6) δ9.05 (s, 1H), 8.74 (s, 1H), 8.37 (dd, J=6.8, 3.2Hz, 2H), 7.90 (d, J= 2.0Hz,1H),7.60–7.45(m,5H),4.79(s,2H),3.81(t,J=5.6Hz,2H),2.90(t,J=5.6Hz,2H). LC-MS(ESI)m / z C 20 H 17 Cl2N4O + [M+H] + Calculated value: 399.1, Measured value: 399.1.

[0363] Example 15. Synthesis of Compound 59 (N-(3,4-dichlorophenyl)-2-(4-fluorophenyl)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide)

[0364] 47 mg of compound 59 was prepared as a white solid by replacing 3,4-dichlorophenyl isocyanate 77 and Int1-21 with 3,4-difluorophenyl isocyanate 52 and Int1-1 respectively, according to the general synthesis method A described in Example 1. The yield was 87%.

[0365] 1 H NMR (400MHz, DMSO-d6) δ9.09(d,J=3.2Hz,1H),8.73(s,1H),8.41(dd,J=8.8,5.6Hz,2H),7.90(d,J=2.0Hz ,1H),7.59–7.43(m,2H),7.35(t,J=8.8Hz,2H),4.78(s,2H),3.81(t,J=5.6Hz,2H),2.90(t,J=5.6Hz,2H). LC-MS(ESI)m / z C 20 H 16 Cl2FN4O + [M+H] + Calculated value: 417.1, Measured value: 417.1.

[0366] The following compounds were prepared by using the isocyanates or aromatic amines and other intermediate compounds listed in the table below as starting materials, according to general synthetic methods A or B described in Examples 1 and 2.

[0367] Example 53. Preparation of compound 2 (N-(3,4-difluorophenyl)-4-methoxy-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide) (General Synthetic Method C)

[0368] In an ice bath under nitrogen protection, methanol (1.8 mL) was dispersed and dissolved in dry tetrahydrofuran (1.0 mL). Sodium hydride (5.6 mmol, 224 mg, 60 wt% in oil) was slowly added in portions to the system. After the addition was complete, the reaction solution was a grayish-yellow paste, producing a large amount of gas. The reaction was continued at 0 °C for 30 minutes. A tetrahydrofuran solution of compound 1 (0.25 mmol, 80 mg) (1 mL) was slowly added dropwise to the reaction system. After the addition was complete, the reaction solution became a pale yellow, slightly turbid liquid. The mixture was stirred for another 10 minutes and then stirred at room temperature for 2 hours. The reaction was monitored by TLC (developing solvent: DCM / MeOH = 30:1, R f =0.3). After the reaction was complete, the mixture was extracted with water and EA. The aqueous phase was extracted with EA (4 × 5 mL). The organic phases were combined and dried over anhydrous sodium sulfate. The solvent was removed under reduced pressure to give 118 mg of a pale yellow oily crude product. The crude product was purified by rapid silica gel column chromatography (DCM / MeOH = 50 / 1) to give 70 mg of compound 2 as a white solid, yield 89%, mp 189–191 °C.

[0369] 1 ¹H NMR (400MHz, deuterated chloroform) δ 8.58 (s, 1H), 7.41 (ddd, J = 12.0, 7.2, 2.0Hz, 1H), 7.04 (q, J = 9.2Hz, 1H), 6.98–6.91 (m, 1H), 6.73 (s, 1H), 4.58 (s, 2H), 4.01 (s, 3H), 3.77 (t, J = 5.6Hz, 2H), 2.74 (t, J = 5.6Hz, 2H). LC-MS (ESI) m / z C 15 H 15 F2N4O2 + [M+H] + Calculated value: 321.1, Measured value: 321.1.

[0370] Example 54. Preparation of compound 3 (N-(3,4-difluorophenyl)-4-isopropoxy-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide).

[0371] Compound 3 was prepared in 57 mg as a white solid, yield 69%, using i-PrOH instead of CH3OH, according to the general synthesis method C described in Example 53, with an mp of 123–125 °C.

[0372] 1¹H NMR (400MHz, deuterated chloroform) δ 8.56 (s, 1H), 7.45 (ddd, J = 12.4, 7.2, 2.4Hz, 1H), 7.11–7.01 (m, 1H), 6.98–6.92 (m, 1H), 6.55 (s, 1H), 5.42 (p, J = 6.0Hz, 1H), 4.57 (s, 2H), 3.77 (t, J = 6.0Hz, 2H), 2.74 (t, J = 6.0Hz, 2H), 1.38 (s, 3H), 1.36 (s, 3H). LC-MS (ESI) m / z C 17 H 19 F2N4O2 + [M+H] + Calculated value: 349.1, measured value: 349.2.

[0373] Example 55. Preparation of compound 4 (N-(3,4-difluorophenyl)-4-(pyridin-4-ylmethoxy)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide)

[0374] 31 mg of compound 4 was prepared as a white solid, yield 32%, mp 175–177 °C, by replacing CH3OH with compound 78, according to the general synthetic method C described in Example 53.

[0375] 1 ¹H NMR (400MHz, deuterated chloroform) δ 8.66–8.56 (m, 3H), 7.45 (ddd, J = 12.4, 7.2, 2.4Hz, 1H), 7.33 (d, J = 5.4Hz, 2H), 7.07 (q, J = 9.2Hz, 1H), 6.98–6.92 (m, 1H), 6.53 (s, 1H), 5.50 (s, 2H), 4.63 (s, 2H), 3.82 (t, J = 5.6Hz, 2H), 2.87 (t, J = 5.6Hz, 2H).

[0376] Example 56. Preparation of compound 5 (4-(cyclopropylamino)-N-(3,4-difluorophenyl)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide) (General Synthetic Method D)

[0377] Compound 1 (0.25 mmol, 80 mg) was dissolved and dispersed in 2 mL of anhydrous ethanol at room temperature under nitrogen protection. Cyclopropylamine 79 (1.00 mmol, 68.6 μL) was added dropwise, and the reaction mixture was refluxed at 80 °C for 5 hours. The reaction was monitored by TLC (developing solvent: DCM / MeOH = 20:1, R...). f=0.4). After the reaction was complete, the mixture was extracted with water and EA. The aqueous phase was extracted with EA (4 × 5 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to give 87 mg of crude brownish-yellow oil. The crude product was purified by rapid silica gel column chromatography (DCM / MeOH = 30 / 1) to give 33 mg of compound 5 as a white oil, with a yield of 39%.

[0378] 1 ¹H NMR (400MHz, deuterated chloroform) δ 8.55 (s, 1H), 7.46 (ddd, J = 12.4, 7.2, 2.4Hz, 1H), 7.06 (q, J = 9.2Hz, 1H), 6.97–6.91 (m, 1H), 6.65 (s, 1H), 4.89 (s, 1H), 4.50 (s, 2H), 3.83 (t, J = 5.6Hz, 2H), 2.91–2.81 (m, 1H), 2.45 (t, J = 6.0Hz, 2H), 0.92 (q, J = 6.8Hz, 2H), 0.62–0.55 (m, 2H).

[0379] Example 57. Preparation of compound 6 (N-(3,4-difluorophenyl)-4-(prop-2-yn-1-ylamino)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formamide)

[0380] 13 mg of compound 6 was prepared as a pale yellow oil by replacing compound 79 with compound 74, according to the general synthetic method D described in Example 56, with a yield of 16%.

[0381] 1 ¹H NMR (400MHz, deuterated chloroform) δ 8.52 (s, 1H), 7.50–7.43 (m, 1H), 7.11–7.03 (m, 1H), 6.96–6.91 (m, 1H), 6.48 (s, 1H), 4.82–4.77 (m, 1H), 4.51 (s, 2H), 4.36–4.32 (m, 1H), 3.86 (t, J = 5.6 Hz, 2H), 2.54 (t, J = 5.6 Hz, 2H), 2.29 (t, J = 2.4 Hz, 1H).

[0382] Example 58. Preparation of compound 42 (2,4-dichloro-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formic acid 3,4-dichlorophenyl ester) (General Synthetic Method E)

[0383] Compound 80 (2.18 mmol) was dispersed and dissolved in 5 mL of dichloromethane in an ice bath under nitrogen protection. Then, 3 mL of a dichloromethane solution containing triphosgene (1.09 mmol) and 2.18 mmol of triethylamine were slowly added sequentially to the system. After the addition was complete, the mixture was stirred at room temperature for 6 hours. The reaction solution was used directly in the next reaction without purification.

[0384] The reaction solution was pre-cooled again in an ice bath, and 4 mL of dichloromethane solution containing triethylamine (7.28 mmol) and Int1-2 (1.46 mmol) was added sequentially. After the addition was complete, the reaction was stirred at room temperature for 12 hours. The reaction was monitored by TLC (developing solvent: DCM / MeOH = 20:1). After the reaction was complete, the mixture was extracted with water and DCM. The aqueous phase was extracted with DCM (4 × 10 mL), and the organic phases were combined and dried over anhydrous sodium sulfate. The solvent was removed under reduced pressure to obtain the crude product. The crude product was purified by rapid silica gel column chromatography (DCM / MeOH = 50 / 1) to give compound 42 as a white solid, 97 mg, in a yield of 60%.

[0385] 1 H NMR(400MHz, DMSO-d6)δ7.68(d,J=8.8Hz,1H),7.64–7.56(m,1H),7.26(dd,J=8.8, 2.8Hz, 1H), 4.75 (d, J = 69.0Hz, 2H), 3.83 (d, J = 57.2Hz, 2H), 2.88 (d, J = 29.6Hz, 2H). LC-MS(ESI)m / z C 14 H 10 Cl4N3O2 + [M+H] + Calculated value: 393.9, measured value: 393.9.

[0386] Example 59. Preparation of compound 43 (2,4-dichloro-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formic acid 3,4-difluorophenyl ester).

[0387] 137 mg of compound 43 was prepared as a white solid, with a yield of 74%, by replacing compound 80 with compound 81, according to the general synthetic method E described in Example 58.

[0388] 1 H NMR (400MHz, DMSO-d6) δ7.47 (dt, J = 19.6, 9.8 Hz, 2H), 7.09 (d, J = 9.2 Hz, 1H), 4.75 (d, J = 67.6 Hz, 2H), 3.83 (d, J = 57.2 Hz, 2H), 2.88 (d, J = 28.4 Hz, 2H). LC-MS(ESI)m / z C14 H 10 Cl2F2N3O2 + [M+H] + Calculated value: 360.0, Measured value: 360.0.

[0389] Example 60. Synthesis of compound 44 (2,4-dichloro-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formic acid 4-(trifluoromethyl)phenyl ester).

[0390] 121 mg of compound 44 was prepared as a white solid, with a yield of 75%, by replacing compound 80 with compound 82, according to the general synthetic method E described in Example 58.

[0391] 1 H NMR (400MHz, DMSO-d6) δ7.80 (d, J = 8.6 Hz, 2H), 7.44 (d, J = 8.0 Hz, 2H), 4.78 (d, J = 76.4 Hz, 2H), 3.86 (d, J = 62.4 Hz, 2H), 2.90 (d, J = 29.6 Hz, 2H). LC-MS(ESI)m / z C 15 H9Cl2F3N3O2 - [MH] - Calculated value: 390.0, Measured value: 390.0.

[0392] Example 61. Preparation of compound 47 (3,4-dichlorophenyl 4-chloro-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formic acid).

[0393] Using Int1-1 instead of Int1-2, 121 mg of compound 47 was prepared as a white solid according to the general synthetic method E described in Example 58, with a yield of 83%.

[0394] 1 H NMR (400MHz, DMSO-d6) δ8.88(s,1H),7.67(d,J=8.8Hz,1H),7.62(d,J=9.6Hz,1H),7.26(d, J=8.8Hz, 1H), 4.75 (d, J=65.2Hz, 2H), 3.85 (dt, J=58.4, 6.0Hz, 2H), 2.92 (d, J= 29.6Hz, 2H). LC-MS(ESI)m / z C 14 H9Cl3N3O2 - [MH] - Calculated value: 356.0, Measured value: 356.0.

[0395] Example 62. Preparation of compound 48 (3,4-difluorophenyl 4-chloro-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formic acid).

[0396] 113 mg of compound 48 was prepared as a white solid, in 68% yield, by replacing compound 80 with compound 81 and Int1-1 with Int1-2, according to the general synthetic method E described in Example 58.

[0397] 1 H NMR (400MHz, DMSO-d6) δ8.88(s,1H),7.48(q,J=9.6Hz,2H),7.15–7.03(m,1H),4.75(d,J=64.4Hz,2H),3.99–3.71(m,2H),2.92(d,J=27.2Hz,2H). LC-MS(ESI)m / z C 14 H9ClF2N3O2 - [MH] - Calculated value: 324.0, Measured value: 324.0.

[0398] Example 63. Preparation of compound 49 (4-chloro-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formic acid 4-(trifluoromethyl)phenyl ester).

[0399] 94 mg of compound 49 was prepared as a white solid, in 64% yield, by replacing compound 80 with compound 82 and Int1-1 with Int1-2, according to the general synthetic method E described in Example 58.

[0400] 1 H NMR (400MHz, DMSO-d6) δ8.88(s,1H),7.79(d,J=8.4Hz,2H),7.44(d,J=8.4Hz,2H), 4.77(d,J=72.8Hz,2H), 3.88(dt,J=63.6,6.0Hz,2H), 2.93(dt,J=29.2,6.0Hz,2H). LC-MS(ESI)m / z C 15 H 10 ClF3N3O2 - [MH] - Calculated value: 356.0, Measured value: 356.0.

[0401] Example 64. Preparation of compound 68 (4-chlorophenyl 4-chloro-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formate).

[0402] 60 mg of compound 68 was prepared as a white solid, in 41% yield, by substituting compound 83 for compounds 80 and Int1-2, according to the general synthetic method E described in Example 58.

[0403] 1 H NMR (400MHz, DMSO-d6) δ7.46 (d, J = 8.6 Hz, 2H), 7.23 (d, J = 8.4 Hz, 2H), 4.75 (d, J = 71.6 Hz, 2H), 3.84 (d, J = 59.2 Hz, 2H), 2.88 (d, J = 25.6 Hz, 2H). LC-MS(ESI)m / z C 14 H 11 Cl3N3O2 + [M+H] + Calculated value: -358.0, Actual value: 358.0.

[0404] Example 65. Preparation of compound 69 (3,4-difluorophenyl 4-(prop-2-yn-1-ylamino)-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-formic acid).

[0405] 296 mg of compound 69 was prepared as a white solid, with a yield of 64%, by replacing compound 80 with compound 81 and Int1-22 with Int1-22, according to the general synthetic method E described in Example 58.

[0406] 1 H NMR (400MHz, DMSO-d6) δ8.37(s,1H),7.46(dd,J=19.6,9.6Hz,3H),7.08(d,J=9.2Hz,1H),4.49(d,J= 64.0Hz, 2H), 4.16 (dd, J = 5.6, 2.4Hz, 2H), 3.78 (d, J = 56.0Hz, 2H), 3.04 (d, J = 2.4Hz, 1H), 2.56 (s, 2H).

[0407] LC-MS(ESI)m / z C 18 H 16 F3N4O2 + [M+H] + Calculated value: 345.1, Measured value: 345.1.

[0408] Example 66 Preparation of compound 45 (1-(2,4-dichloro-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-2-(3,4-dichlorophenyl)ethane-1,2-dione) (General Synthetic Method F)

[0409] Step 1: Synthesis of Int2-1

[0410] 3,4-Dichloroacetophenone 84 (5.29 mmol) and selenium dioxide (7.93 mmol) were uniformly dispersed in anhydrous pyridine under nitrogen protection at room temperature. The mixture was then reacted in an oil bath at 110 °C for 1 hour, followed by cooling to 90 °C for 4 hours. The reaction was monitored by TLC (developing solvent: PE / EA = 10:1). After the reaction was complete, the reaction solution was cooled to room temperature and filtered to remove the black insoluble matter. The yellow filtrate was collected and concentrated. The concentrated residue was dispersed and dissolved in 20 mL of sodium hydroxide aqueous solution (2N) under ice bath conditions. 30 mL of hydrochloric acid aqueous solution (2N) was slowly added, resulting in the precipitation of a large amount of pinkish-purple solid. The filter cake was collected by suction filtration, washed with a small amount of water, dried under vacuum, reconstituted with EA, dried over anhydrous sodium sulfate, and then evaporated to dryness to obtain 873 mg of pinkish-purple solid Int2-1. LC-MS (ESI) m / z C8H3Cl2O3 - [MH] - Calculated value: 216.9, Measured value: 216.9.

[0411] Step 2: Synthesis of Compound 45

[0412] In an ice bath under nitrogen protection, Int2-1 (0.44 mmol) was dispersed and dissolved in 5 mL of tetrahydrofuran. Thionyl chloride (0.51 mmol) was slowly added to the system sequentially, and the reaction was stirred at 0 °C for 1.5 h. Then, 2 mL of tetrahydrofuran solution containing triethylamine (0.55 mmol) and Int1-2 (0.42 mmol) was added sequentially, and the reaction was stirred at 50 °C for 2 h. The reaction was monitored by TLC (evolving solvent: DCM / MeOH = 40:1). After the reaction was complete, the mixture was extracted with water and EA. The aqueous phase was extracted with EA (4 × 10 mL), and the organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was purified by rapid silica gel column chromatography (DCM / MeOH = 50 / 1) to give compound 45 as a white solid, 48 mg, in a yield of 28.6%.

[0413] Because compound 45 exhibits amide spin-blocked isomerism, two isomers are present in its 1H NMR spectrum: Isomer A: 1¹H NMR (400MHz, DMSO-d⁶) δ 8.18 (d, J = 2.0Hz, 1H), 7.95 (dd, J = 8.4, 2.0Hz, 1H), 7.90 (d, J = 8.4Hz, 1H), 4.85 (s, 2H), 3.70 (t, J = 6.0Hz, 2H), 2.75 (t, J = 6.0Hz, 2H). Isomer B: 1 H NMR (400MHz, DMSO-d6) δ8.10 (d, J = 2.0Hz, 1H), 7.88–7.84 (m, 2H), 4.66 (s, 2H), 3.98 (t, J = 6.0Hz, 2H), 2.92 (t, J = 6.0Hz, 2H). LC-MS(ESI)m / z C 15 H 10 Cl4N3O2 + [M+H] + Calculated value: 404.0, measured value: 403.9.

[0414] Example 67. Preparation of compound 50 (1-(4-chloro-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-2-(3,4-dichlorophenyl)ethane-1,2-dione)

[0415] Using Int1-1 instead of Int1-2, 26 mg of compound 50 was prepared as a white solid according to the general synthetic method F described in Example 66, with a yield of 17%.

[0416] Because compound 50 exhibits amide spin-blocked isomerism, two isomers are present in its 1H NMR spectrum: Isomer A: 1 ¹H NMR (400MHz, DMSO-d⁶) δ 8.90 (s, ¹H), 8.19 (d, J = 2.0Hz, ¹H), 7.95 (dd, J = 8.4, 2.0Hz, ¹H), 7.90 (d, J = 8.4Hz, ¹H), 4.85 (s, 2H), 3.71 (t, J = 6.0Hz, 2H), 2.79 (t, J = 6.0Hz, 2H). Isomer B: 1 H NMR (400MHz, DMSO-d6) δ8.84(s,1H),8.11(d,J=2.0Hz,1H),7.89–7.83(m,2H),4.64(s,2H),4.00(t,J=6.0Hz,2H),2.95(t,J=6.0Hz,2H). LC-MS(ESI)m / z C 15 H 11 Cl3N3O2 + [M+H] +Calculated value: 370.0, measured value: 370.0.

[0417] Example 68. Preparation of compound 46 (1-(2,4-dichloro-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-2-(3,4-dichlorophenyl)ethane-1-one) (General Synthetic Method G)

[0418] Compound 85 (0.44 mmol) was dispersed and dissolved in 5 mL of tetrahydrofuran under nitrogen protection in an ice bath. Thionyl chloride (0.51 mmol) was slowly added to the system sequentially, and the reaction was stirred at 0 °C for 1.5 h. Then, 2 mL of tetrahydrofuran solution containing triethylamine (0.55 mmol) and Int1-2 (0.42 mmol) was added sequentially, and the reaction was stirred at room temperature for 2 h. The reaction was monitored by TLC (evolving solvent: DCM / MeOH = 40:1). After the reaction was complete, the mixture was extracted with water and EA. The aqueous phase was extracted with EA (4 × 10 mL), and the organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was purified by rapid silica gel column chromatography (DCM / MeOH = 50 / 1) to give compound 46 as a pale yellow solid. LC-MS (ESI) m / z C 15 H 12 Cl4N3O + [M+H] + Calculated value: 390.0, Measured value: 390.0.

[0419] Example 69. Preparation of compound 51 (1-(4-chloro-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-2-(3,4-dichlorophenyl)ethane-1-one)

[0420] Compound 51, a pale yellow oil, was prepared by using Int1-1 instead of Int1-2 according to the general synthetic method G described in Example 68. 15 H 13 Cl3N3O + [M+H] + Calculated value: 356.0, Measured value: 356.0.

[0421] III. Formulation Preparation Examples

[0422] Formulation Example A: Preparation of Injectable Formulation

[0423] (I) Formula Composition:

[0424] (II) Preparation method:

[0425] According to the formulation, compound 23, polysorbate 80 and mannitol from Example 2 were added to 4000 ml of water for injection. After stirring and dissolving, water for injection was added to a total volume of 5000 ml. Stirring was continued, and the mixture was sterilized by filtering through a 0.22 μm microporous membrane. The filtrate was aseptically filled into 5 ml ampoules (25 mg / ampoule) at 5 ml each, sealed, and sterilized.

[0426] Formulation Example B: Preparation of Tablets

[0427] (I) Formulation

[0428] (II) Preparation process

[0429] Compound 23 from Example, lactose, and a portion of microcrystalline cellulose were micronized in a ratio of 200:100:40. The remaining microcrystalline cellulose, pregelatinized starch, micronized silica gel, and sodium carboxymethyl starch (which had passed through an 80-mesh sieve) were added according to the formula ratio. After mixing evenly, an appropriate amount of 0.3% HPMC solution was added to form a soft mass. The mass was granulated through an 18-mesh sieve and dried at 60°C (the moisture content of the granules was controlled to be around 3%). Magnesium stearate (which had passed through an 80-mesh sieve) was added and mixed evenly with the granules. The granules were then sized through a 16-mesh sieve, compressed into tablets, and packaged.

[0430] Formulation Example C: Preparation of Tablets

[0431] (I) Formulation

[0432] (II) Preparation process

[0433] The tablets of Formulation Example C were prepared in a similar manner to those of Formulation Example B above.

[0434] Formulation Example D: Preparation of Tablets

[0435] (I) Formulation

[0436] (II) Preparation process

[0437] The tablets of Formulation Example C were prepared in a similar manner to those of Formulation Example B above.

[0438] IV. Bioactivity Detection

[0439] 1. Evaluation of the affinity between small molecules and proteins

[0440] (1) Experimental materials

[0441] Biacore TM T200, from CM5 series chips (purchased from) Catalog number: 29149604); Human BRM recombinant protein (1350-1520, human, recombinant, expressed in HEK293, MCE), Human BRG1 recombinant protein (1480-1603, His tag human, recombinant, expressed in E. coli, Sigma-Aldrich) and Human GPR65 recombinant protein (full-length protein, His tag human, expressed in E. coli, MCE).

[0442] (2) Experimental Procedure

[0443] Step 1) pH screening: Select suitable ligand buffer pH from 10mM sodium acetate buffer solutions with pH values ​​of 5.5, 5.0, and 4.5. The ligand (protein) concentration is 50 μg / ml. The contact time with the chip is set to 180 s, and the flow rate is set to 5 μl / min. Then, the chip surface is regenerated with 50mM NaOH solution.

[0444] Step 2) Ligand (protein) immobilization: The ligand protein was immobilized on the chip surface using a direct amino-coupling method. First, the chip surface was activated with EDC / NHS reagent and the protein was coupled, then the chip surface was blocked with ethanolamine reagent. The process was performed in aim-for-immobilized-level mode, with the target level set to 1000 RU, for automated coupling. After coupling, the coupling amount was detected using a response bound.

[0445] Step 3) Development of Small Molecule-Protein Affinity Assay Method: Ten PFI-3 solutions (BRG1 / BRM positive control) and BTB09089 solution (GPR65 positive control) with concentrations of 125.0 μM, 62.5 μM, 31.25 μM, 16.125 μM, and 8.0625 μM, respectively, and 50% DMSO HBS-EP buffer (regeneration solution) were prepared and placed on the sample holder. The contact time between the small molecule solution and the chip was set to 180 s, and the duration was set to 60 s. The contact time between the regeneration solution and the chip was set to 30 s. Data results were analyzed by comparing the sensor map results of channel 2 with those of channel 1 (background reference channel) to examine the binding ability of small molecules and ligand proteins, and the affinity (K) was calculated by kinetic or affinity fitting methods. D The surface testing-regeneration process parameters were optimized based on preliminary experimental results to obtain a stable testing method.

[0446] Step 4) Small molecule-ligand protein affinity test: Based on the experimental parameters and procedures developed in Step 3 above, the protein affinity of the small molecule to be tested is determined. The affinity is then calculated using either kinetic or affinity fitting methods based on the test results.

[0447] (3) Experimental Results

[0448] The affinity test results of the compounds of this invention as small molecules targeting the BRG1 / BRM bromine domain and the proton-sensing receptor GPR65 with three receptor proteins are shown in Table 1 below. Figure 1 shows the affinity (K) between the compounds and BRG1 / BRM proteins. D As a result, Figure 2 shows the affinity (K) between the compound and the GPR65 protein. D Results. In the figure, PFI-3 is the positive affinity control compound for BRG1 / BRM protein, and BTB09089 is the positive affinity control compound for GPR65 protein.

[0449] Table 1. Results of compound-protein affinity evaluation

[0450] Note: "++" refers to K. D Value less than 10 μM; "++" indicates K D Value greater than 10 μM but less than 200 μM; "+" indicates K D Values ​​greater than 200 μM; "-" indicates no affinity detected; "NT" indicates not tested.

[0451] The above affinity experiment results show that the compound of the present invention has good affinity for the BRG1 / BRM bromine domain and the proton-sensing acceptor GPR65.

[0452] 2. In vitro glioblastoma cell proliferation inhibition experiment

[0453] (1) Experimental Procedure

[0454] 1) Compound preparation

[0455] This experiment investigated the inhibitory activity of the compound of the present invention on the proliferation of brain tumor cells in vitro. The preparation method of the test compound was as follows: the volume of DMSO required for 10 mM solution was calculated according to the mass and molecular weight of the compound, DMSO was added in the specified amount, and the mixture was vortexed until completely dissolved to prepare a mother solution.

[0456] 2) Gradual dilution of the test compound

[0457] Each compound was diluted from 10 mM down to 9 concentration points, each point was diluted 3 times, and each concentration was vortexed and briefly centrifuged to the bottom of the tube. The mixtures were then added sequentially to a 96-well plate, with one DMSO control well added to each compound.

[0458] 3) Cell dilution

[0459] The cell lines used in this experiment were patient-derived glioma cell lines: G709, G118, GBM17, G755-5, G98, Mes28, MGG8, TT423, and TT274. Once the cells reached a sufficient number, they were plated. From 10cm diameter cell culture dishes, the cells were washed once with 3mL PBS, digested with 1mL Accutase at 37℃ for 1min, and then digested with 3mL PBS to stop the digestion. The cells were then centrifuged (250g) for 3min to resuspend in the culture medium and counted. Based on a cell density of 1500 cells / well, the corresponding amount of cells was resuspended in the corresponding volume of culture medium, plated, and 25μL of cell suspension was added to the bottom of each 384-well plate using a pipette. 50μL PBS was added to each of the outer wells to complete the filling. The plates were incubated at 37℃ for approximately 3 hours before the compound was added.

[0460] 4) Transfer of the test compound

[0461] Thaw the serially diluted 96-well plate of the compound, centrifuge briefly, dilute with culture medium, and add to the wells. The final concentration of the compound in the wells is 100 μM after a 100-fold dilution (the same applies to other concentration points).

[0462] Sample volume per compound: 0.5 μL * 9 cell types * 3 replicates * 1.2x

[0463] Each compound requires the following culture medium volume: 24.5 μL * 9 cell types * 3 replicates * 1.2 times.

[0464] Measure the required volume of compound using a pipette and add it to the required volume of culture medium. Mix thoroughly by pipetting and aspirating. Add 25 μL of the compound suspension to a 384-well plate with cells already coated. Briefly centrifuge the entire plate, place it on a horizontal shaker at 120 rpm for 10 min, centrifuge again briefly, and incubate at 37°C for 3 days.

[0465] 5) Testing

[0466] Add 10 μL of CellTiter-Glo reagent to each well, shake on a shaker for 2 min, briefly centrifuge, and detect the luminescence signal value using a microplate reader. Then calculate and fit the results to obtain the half-maximal inhibitory concentration (IC50) of each analyte against different glioblastoma cells. 50 value.

[0467] (2) Experimental Results

[0468] The inhibitory effects of the compounds of this invention on the proliferation of glioblastoma cells from various patient sources are shown in Tables 2-3 and Figure 3.

[0469] Table 2. Results of in vitro glioblastoma cell proliferation inhibition experiment

[0470] Note: "NT" means untested.

[0471] Table 3. Results of in vitro glioblastoma cell proliferation inhibition experiment

[0472] As shown in Figure 4-11, representative compounds 1, 10, 17, 30, 31, 36, 37, and 42 exhibited dose-dependent proliferative inhibitory activity against various glioblastoma cell lines, and all showed stronger tumor-suppressive activity, with half-maximal inhibitory concentrations (IC50) against most glioblastoma cells. 50 (less than 2μM)

[0473] The above results indicate that the compounds of the present invention have strong inhibitory activity against glioblastoma cell proliferation and have broad-spectrum anti-glioblastoma cell proliferation characteristics.

[0474] 3. Pharmacodynamic evaluation of the B104-1-1 murine subcutaneous glioblastoma animal model

[0475] (1) Experimental scheme

[0476] The mice used in the in vivo pharmacodynamic evaluation experiments were all female BALB / c mice, 6-8 weeks old. The mice were purchased from Spiford (Beijing) Biotechnology Co., Ltd., and housed at the Animal Center of Kanglong Pharmaceutical (Beijing) New Drug Technology Co., Ltd. (AAALAC certified unit). The ambient temperature of the mice was 23±3℃, the relative humidity was maintained at 40%-70%, and the normal 12-hour diurnal rhythm was maintained.

[0477] In the subcutaneous xenograft tumor model, the well-growing murine glioblastoma cell line B104-1-1 was used. Cells were resuspended in Matrigel at a rate of 5 million cells per 0.1 mL per mouse and injected subcutaneously into the right rib area of ​​60 mice. The tumors were implanted until they reached approximately 85 mm². 3Thirty animals with relatively uniform tumor volume were selected and divided into two groups: a solvent control group and multiple treatment groups (compounds 17 and 36). The drug solution was prepared according to the ratio of 5% compound / DMSO + 40% polyethylene glycol 300 + 5% polyoxyethylene ether castor oil + 50% physiological saline, and thoroughly vortexed and sonicated to form a homogeneous solution. Treatment compound 17 was administered intraperitoneally at 50 mg / kg daily. Treatment compound 36 was administered intraperitoneally at 50 mg / kg for the first two days, followed by 30 mg / kg daily. The solvent control group received the corresponding solvent. During the experiment, the length and width of the subcutaneous tumor were measured three times a week before administration, and the mice were weighed. When the weight of mice in a group dropped to below 90% (inclusive), they were given a nutritional gel. The tumor volume was calculated using the formula: Volume = 0.5 × Major Axis × Minor Axis. 2 The tumor growth inhibition rate (TGI) (an antitumor activity evaluation index) was calculated using tumor volume: TGI = (1 - T / C) × 100%, where T and C are the mean tumor volumes of the treatment group and control group, respectively. A tumor volume of 2000 mm² was considered normal. 3 The tumor was considered the endpoint of treatment; it was removed and fixed with formalin solution.

[0478] (2) Experimental Results

[0479] 1) Animal weight

[0480] The changes in animal body weight during the experiment are shown in Table 4, Figure 12 and Figure 13.

[0481] Table 4. Changes in body weight of mice in each group during the in vivo efficacy evaluation experiment of B104-1-1 mouse glioblastoma.

[0482] Note: a. Mean ± Standard Error

[0483] 2) Tumor growth inhibition rate

[0484] Table 5 shows a comparison of mean tumor volume, T / C%, and TGI%. Figure 14 shows the tumor growth curve.

[0485] Table 5. Changes in tumor volume in mice of different groups during the in vivo efficacy evaluation experiment of B104-1-1 mouse glioblastoma.

[0486] Note: a. Mean ± standard error; b. Compared with the solvent control group, LSD test was used.

[0487] The above results indicate that compounds 17 and 36 both exhibited good anti-tumor effects in the B104-1-1 murine glioblastoma cell subcutaneous animal model.

[0488] 4. Pharmacodynamic evaluation of the MC38 mouse subcutaneous colon cancer animal model

[0489] (1) Experimental scheme

[0490] The mice used in the in vivo pharmacodynamic evaluation experiments were all female C57BL / 6J mice, 6-8 weeks old. The mice were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd., and housed at the Animal Center of Kanglong Pharmaceutical (Beijing) Co., Ltd. (AAALAC certified unit). The ambient temperature of the mice was 23±3℃, the relative humidity was maintained at 40%-70%, and the normal 12-hour diurnal rhythm was maintained.

[0491] In a subcutaneous xenograft tumor model, the well-grown murine colon cancer cell line MC38 was used. Cells were resuspended in Matrigel at a rate of 1 million cells / 0.1 mL / mouse and injected subcutaneously into the right rib area of ​​experimental mice. A total of 40 animals were inoculated. The tumors were allowed to grow until they reached approximately 50 mm². 3 Twenty animals with relatively uniform tumor volume were selected and divided into a solvent control group and a treatment group (compound 52). The drug solution was prepared according to the ratio of 10% compound / methylpyrrolidone + 40% polyethylene glycol 400 + 5% polyoxyethylene ether castor oil + 45% (10% Soluplus, 0.2% sodium dodecyl sulfate aqueous solution), and thoroughly vortexed and sonicated to form a homogeneous solution. The treatment group received compound 52 intraperitoneally at 50 mg / kg daily. The solvent control group received the corresponding solvent. During the experiment, the length and width of the subcutaneous tumor were measured three times a week before administration using calipers, and the mice were weighed. When the weight of mice in the group dropped to below 90% (inclusive), they were given nutritional gels. The tumor volume was calculated using the formula: Volume = 0.5 × major axis × minor axis 2 The tumor growth inhibition rate (TGI) (an antitumor activity evaluation index) was calculated using tumor volume: TGI = (1 - T / C) × 100%, where T and C are the mean tumor volumes of the treatment group and control group, respectively. A tumor volume of 2000 mm² was considered normal. 3 The tumor was considered the endpoint of treatment; it was removed and fixed with formalin solution.

[0492] (2) Experimental Results

[0493] 1) Animal weight

[0494] The changes in animal body weight during the experiment are shown in Table 6, Figure 15, and Figure 16.

[0495] Table 6. Changes in body weight of mice in each group during the in vivo efficacy evaluation experiment of MC38 in murine colon cancer.

[0496] Note: a. Mean ± Standard Error

[0497] 2) Tumor growth inhibition rate

[0498] Table 7 shows a comparison of mean tumor volume, T / C%, and TGI%. Figure 17 shows the tumor growth curve.

[0499] Table 7. Tumor volume changes in mice of different groups during the in vivo efficacy evaluation experiment of MC38 in murine colon cancer.

[0500] Note: a. Mean ± standard error; b. Compared with the solvent control group, LSD test was used.

[0501] The above results indicate that compound 52 exhibits good anti-colon cancer effects in the MC38 mouse subcutaneous colon cancer animal model.

[0502] The present invention has been described through the above embodiments. However, it should be understood that the above embodiments are for illustrative purposes only and are not intended to limit the invention to the scope of the described embodiments. Those skilled in the art will also understand that the present invention is not limited to the above embodiments, and many more variations and modifications can be made based on the teachings of the present invention, all of which fall within the scope of protection claimed by the present invention. The scope of protection of the present invention is defined by the appended claims and their equivalents.

Claims

1. A compound of formula I or a pharmaceutically acceptable salt, stereoisomer, solvate, or isotopic derivative thereof: in X, Y, and Z are each independently CH, CR, or N. W can be -NH-, -O-, -CH2-, or -C(O)-. R and R1 are each independently selected from halogen, cyano, nitro, hydroxy, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, and -NR4R5; or The two R and / or R1 atoms on adjacent carbon atoms, together with the carbon atoms they are attached to, form a 5-6 membered ring optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur. R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NR4R5, -S(O)2-C1-C6 alkyl, -C(O)2-C1-C6 alkyl, C3-C8 cycloalkyl, 3 to 8-membered heterocyclic groups, C6-C 10 Aryl, 5- to 12-membered heteroaryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heterocyclic and heteroaryl groups contain 1 to 3 heteroatoms independently selected from nitrogen, oxygen and sulfur, and the cycloalkyl, heterocyclic, aryl and heteroaryl groups are optionally substituted by 1 to 3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy, provided that R2 and R3 are not simultaneously hydrogen; R4 and R5 are each independently selected from hydrogen, C1-C6 alkyl, C3-C8 cycloalkyl, and C2-C6 alkynyl groups, and n is 0, 1, or 2.

2. The compound according to claim 1, or a pharmaceutically acceptable salt, stereoisomer, solvate, or isotopic derivative thereof, wherein the compound has formula I-1: R1, R2, R3, X, Y, Z and n are defined as in claim 1.

3. The compound according to claim 1, or a pharmaceutically acceptable salt, stereoisomer, solvate, or isotopic derivative thereof, wherein the compound has formula I-2: R1, R2, R3, X, Y, Z and n are defined as in claim 1.

4. The compound according to claim 1, or a pharmaceutically acceptable salt, stereoisomer, solvate, or isotopic derivative thereof, wherein the compound has formula I-3: R1, R2, R3, X, Y, Z and n are defined as in claim 1.

5. The compound according to claim 1, or a pharmaceutically acceptable salt, stereoisomer, solvate, or isotopic derivative thereof, wherein the compound has formula I-4: R1, R2, R3, X, Y, Z and n are defined as in claim 1.

6. The compound according to any one of claims 1-5 or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, wherein no more than one of X, Y and Z is N.

7. The compound according to any one of claims 1-6 or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, wherein R and R1 are each independently selected from halogen, cyano, nitro, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl and -N(C1-C6 alkyl)2, preferably halogen, cyano, C1-C6 alkyl, C1-C6 haloalkyl and C1-C6 alkoxy.

8. The compound according to any one of claims 1-7, or a pharmaceutically acceptable salt, stereoisomer, solvate, or isotopic derivative thereof, wherein R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, C6-C 10 The cycloalkyl, 5- to 12-membered heteroaryl, and -O-C1-C6 alkyl-5-12-membered heteroaryl groups, wherein the heteroaryl group contains 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, and the cycloalkyl, aryl, and heteroaryl groups are optionally substituted with 1-3 substituents independently selected from halogen, cyano, hydroxy, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy groups. Preferably, R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxy, C1-C6 haloalkyl, C1-C6 alkoxy, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, and C6-C6 groups optionally substituted with 1-3 halogens. 10 Aryl and -O-C1-C6 alkyl-5-12 heteroaryl groups.

9. The compound according to any one of claims 1-5, or a pharmaceutically acceptable salt, stereoisomer, solvate, or isotopic derivative thereof, wherein... X, Y, and Z are each independently CH, CR, or N; R and R1 are each independently selected from halogen, cyano, nitro, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, and -N(C1-C6 alkyl)2, or The two R and / or R1 on adjacent carbon atoms together with the carbon atoms to which they are attached form a 5-6 membered ring, optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur; R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH2, -NH-C1-C6 alkyl, -N(C1-C6 alkyl)2, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, C6-C 10 Aryl, 5- to 12-membered heteroaryl, and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, and the cycloalkyl, aryl, and heteroaryl groups are optionally substituted with 1-3 substituents independently selected from halogen, cyano, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy, provided that R2 and R3 are not simultaneously hydrogen; and n is 0, 1, or 2.

10. The compound according to any one of claims 1-5, or a pharmaceutically acceptable salt, stereoisomer, solvate, or isotopic derivative thereof, wherein... X, Y, and Z are each independently CH, CR, or N; R and R1 are each independently selected from halogen, cyano, C1-C6 alkyl, C1-C6 haloalkyl, and C1-C6 alkoxy, or The two R and / or R1 on adjacent carbon atoms together with the carbon atoms to which they are attached form a 5-6 membered ring, optionally containing 1-3 heteroatoms independently selected from nitrogen, oxygen and sulfur; R2 and R3 are each independently selected from hydrogen, halogen, cyano, hydroxyl, C1-C6 haloalkyl, C1-C6 alkoxy, -NH-C3-C8 cycloalkyl, -NH-C2-C6 alkynyl, or C6-C alkyl optionally substituted with 1-3 halogens. 10 Aryl and -O-C1-C6 alkyl-5-12-membered heteroaryl, wherein the heteroaryl contains 1-3 heteroatoms independently selected from nitrogen, oxygen, and sulfur, provided that R2 and R3 are not simultaneously hydrogen; and n is 0, 1, or 2.

11. A compound or a pharmaceutically acceptable salt, stereoisomer, solvate, or isotopic derivative thereof, wherein the compound is selected from the following:

12. A pharmaceutical composition comprising the compound of any one of claims 1-11 or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, and one or more pharmaceutically acceptable carriers.

13. Use of the compound of any one of claims 1-11 or a pharmaceutically acceptable salt, stereoisomer, solvate or isotopic derivative thereof, or the pharmaceutical composition of claim 12, in the preparation of a medicament for the prevention and / or treatment of diseases associated with abnormal activity of the BRG1 / BRM bromine domain and / or proton-sensing receptor GPR65.

14. The use according to claim 13, wherein the disease is selected from proliferative diseases, immune diseases, asthma, chronic obstructive pulmonary disease (COPD) and acute respiratory distress syndrome (ARDS), preferably, the proliferative disease is a tumor selected from melanoma, renal cell carcinoma, gastrointestinal cancer, acute myeloid leukemia, pancreatic cancer, triple-negative breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer and glioma, more preferably glioblastoma; the immune disease is selected from psoriasis, psoriatic arthritis, rheumatoid arthritis, multiple sclerosis, systemic lupus erythematosus, Hashimoto's thyroiditis, Graves' disease, uveitis, ulcerative colitis, Crohn's disease, autoimmune uveoretinitis, systemic vasculitis, polymyositis and dermatomyositis, scleroderma, Sjögren's syndrome, ankylosing spondylitis, sarcoidosis, autoimmune hemolytic anemia, autoimmune myocarditis, type I diabetes and atopic dermatitis.

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