CN115703799A

CN115703799A - Nitrogen heteroaryl compound, preparation method and application thereof

Info

Publication number: CN115703799A
Application number: CN202210962092.6A
Authority: CN
Inventors: 马世超
Original assignee: Shanghai Blueray Biopharma Co ltd
Current assignee: Shanghai Blueray Biopharma Co ltd
Priority date: 2021-08-11
Filing date: 2022-08-11
Publication date: 2023-02-17
Also published as: WO2023016511A1; TW202312995A

Abstract

The invention discloses a nitrogen heteroaryl compound, a preparation method and application thereof. The invention also provides a preparation method of the compound, a composition containing the compound and application of the compound in preparing a medicament for treating diseases or disorders related to the action mechanism of EED protein and/or PRC2 protein complex.

Description

Nitrogen heteroaryl compound, preparation method and application thereof

Technical Field

The invention relates to a nitrogen heteroaryl compound, a preparation method and application thereof.

Background

PcG (Polycomb Group) proteins are an important class of chromatin modifying enzymes. It can regulate the transcription of gene by modifying chromosome, so playing an important role in the growth and differentiation of stem cells and long-term cell memory. In mammalian cells, pcG proteins are largely divided into two classes of transcription repressing complexes, PRC1 (Polycomb regenerative Complex 1) and PRC2 (Polycomb regenerative Complex 2), respectively. Wherein PRC2 inhibits the expression of related genes by methylation modification of 27 th lysine (H3K 27) of histone 3 in the chromosome. The PRC2 protein complex is mainly composed of core proteins such as EZH2 (Enhancer of Zeste homo log 2) (or its very similar homologous protein EZH 1), EED (electrophoretic Development), and SUZ12 (super of Zeste 12). Wherein, EZH2 has enzyme catalytic activity, and can transfer methyl of substrate SAM (S-adenosyl-L-methionine) to H3K27 through a SET (Su (var), E (Z), and Trithorax) protein domain, thereby achieving one-to-three methylation modification of H3K 27. The enzymatic activity of EZH2 is also dependent on other components of PRC2, such as EED proteins belonging to the WD40 repeat structural protein family. The binding of EED and trimethylated H3K27Me3 has great allosteric promoting effect on the enzyme catalytic function of EZH2 and can position the PCR2 complex on chromatin needing modification. Dysfunction of PRC2, such as overexpression or gain-of-function mutation of EZH2, is clinically associated with a number of neoplastic diseases, including lung, breast, rectal, prostate, bladder, pancreatic, sarcoma, and lymph cancer, among others. PRC2 is also associated with a variety of cellular immune functions, such as EZH2 is involved in regulating lymphocyte activation and also can promote T cell responses to tumor cells in conjunction with glycolysis. Therefore, the PRC2 small molecule inhibitor has important and wide drug development value.

The development surrounding PRC2 inhibitors is mainly the development of two strategies, EZH2 inhibitors as well as EED inhibitors. Currently, the EZH2 inhibitors that are clinically used include EPZ-6438 (episzyme, second stage clinical), GSK2816126 (GSK, first stage clinical), and CPI-1205 (Constellation, first stage clinical), among others. Although EZH2 inhibitors have been developed in several phases of clinical research, these inhibitors all contain a common pharmacophore of 2-pyridones. Also, secondary mutations have begun to appear in clinical treatments with existing EZH2 inhibitors. The EED inhibitor has allosteric inhibition effect on the function of EZH2 enzyme, and can achieve the same or similar biological function as the EZH 2. And on one hand, the EED inhibitor well overcomes the drug resistance problem of the EZH2, and on the other hand, the EED inhibitor can be combined with the EZH2 inhibitor to achieve a better synergistic effect, so that the development of a novel EED inhibitor has a very important significance.

Disclosure of Invention

The technical problem to be solved by the invention is that the EED inhibitor in the prior art has a single structure. Therefore, the invention provides a nitrogen heteroaryl compound, a preparation method and application thereof. The nitrogen heterocyclic aryl compound of the invention shows good inhibitory activity to tumor cells, and has wide drug development prospect.

The invention provides a compound shown as a formula (I), pharmaceutically acceptable salt thereof or a stereoisomer thereof:

wherein A is independently

X is N or CR ² ；

R ¹ 、R ² And R ³ Independently is H, halogen, cyano, R ^5b Substituted or unsubstituted amino, R ^5b -O-、R ⁵ Substituted or unsubstituted C _1-4 Alkyl radical, R ⁵ Substituted or unsubstituted C _3-6 Cycloalkyl of, R ⁵ Substituted or unsubstituted C ₁ -C ₄ Haloalkyl, -C (= O) R ⁶ 、-CO ₂ R ⁶ 、-C(＝O)NR ⁶ R ⁷ 、-SO ₂ R ⁶ or-SO ₂ NR ⁶ R ⁷ (ii) a When the substituents are plural, the same or different;

R ⁴ independently of one another H, halogen, cyano, C _1-4 Haloalkyl, -C (= O) NR ^4a1 R ^4a2 、-S(＝O) ₂ R ^4b or-C (= O) R ^4c ；

R ^4a1 、R ^4a2 And R ^4b Independently is H or C _1-4 An alkyl group;

R ^4c independently of each other H, C _1-4 Alkyl or-O-C _1-4 An alkyl group;

R ⁵ independently is C _1-4 alkyl-O-, C _1-4 Alkyl radical, C _1-4 Haloalkyl, -N (R) ^5b R ^5b ) Amino, fluorine and hydroxyl protected by N protecting group;

R ^5b 、R ⁶ and R ⁷ Independently of each other H, C _1-4 Alkyl radical, C _1-4 A haloalkyl group;

"indicates that the carbon atom with" "is a chiral carbon atom, which is in the R configuration, S configuration, or a mixture thereof.

In certain preferred embodiments of the present invention, certain groups of the compounds of formula (I), pharmaceutically acceptable salts thereof, or stereoisomers thereof are defined as follows, and the groups not mentioned are as described in any of the embodiments herein (shortly, "in certain preferred embodiments of the present invention"):

the compound shown in the formula (I) is selected from the following compounds:

in the above compounds, X and R ¹ 、R ³ And R ⁴ Is as defined above.

In certain preferred embodiments of the invention:

is composed of

Or a mixture thereof.

In certain preferred embodiments of the invention:

when R is ¹ 、R ² 、R ³ And R ⁴ When independently halogen, the halogen is fluorine, chlorine, bromine or iodine, preferably chlorine.

In certain preferred embodiments of the invention:

when R is ¹ 、R ² And R ³ Independently is R ⁵ Substituted or unsubstituted C _1-4 When alkyl, said C _1-4 Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, preferably methyl or ethyl.

In certain preferred embodiments of the invention:

when R is ¹ 、R ² And R ³ Independently is R ⁵ Substituted or unsubstituted C _3-6 In the case of a cycloalkyl group of (A), said C _3-6 The cycloalkyl group of (a) is cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl.

In certain preferred embodiments of the invention:

when R is ¹ 、R ² And R ³ Independently is R ⁵ Substituted or unsubstituted C ₁ -C ₄ In the case of haloalkyl, the halogen is fluorine, chlorine, bromine or iodine, preferably fluorine.

In certain preferred embodiments of the invention:

when R is ¹ 、R ² And R ³ Independently is R ⁵ Substituted or unsubstituted C ₁ -C ₄ When halogenated alkyl, said C ₁ -C ₄ Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, preferably methyl or ethyl.

In certain preferred embodiments of the invention:

when R is ¹ 、R ² And R ³ Independently is R ⁵ Substituted or unsubstituted C ₁ -C ₄ When the alkyl is halogenated, the number of the halides is 1,2,3,4 or 5; for example 3.

In certain preferred embodiments of the invention:

when R is ¹ 、R ² And R ³ Independently is R ⁵ Substituted or unsubstituted C ₁ -C ₄ When halogenated with an alkyl group, theC is ₁ -C ₄ Haloalkyl is trifluoromethyl.

In certain preferred embodiments of the invention:

when R is ⁴ 、R ⁵ 、R ^5b 、R ⁶ And R ⁷ Independently is C ₁ -C ₄ When halogenated alkyl, the halogen is fluorine, chlorine, bromine or iodine, preferably fluorine.

In certain preferred embodiments of the invention:

when R is ⁴ 、R ⁵ 、R ^5b 、R ⁶ And R ⁷ Independently is C ₁ -C ₄ When halogenated with alkyl, C therein ₁ -C ₄ Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, preferably methyl or ethyl.

In certain preferred embodiments of the invention:

when R is ⁵ Independently is C ₁ -C ₄ When halogenated, C therein ₁ -C ₄ Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, preferably methyl or ethyl.

In certain preferred embodiments of the invention:

when R is ⁴ 、R ^5b 、R ⁶ And R ⁷ Independently is C ₁ -C ₄ When halogenated with alkyl, C therein ₁ -C ₄ Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, preferably methyl or ethyl.

In certain preferred embodiments of the invention:

when R is ⁴ 、R ⁵ 、R ^5b 、R ⁶ And R ⁷ Independently is C ₁ -C ₄ When the alkyl is halogenated, the number of the halides is 1,2,3,4 or 5; for example 3.

In certain preferred embodiments of the invention:

when R is ⁴ 、R ⁵ 、R ^5b 、R ⁶ And R ⁷ Independently is C ₁ -C ₄ When halogenated with an alkyl group, theC mentioned above ₁ -C ₄ Haloalkyl is trifluoromethyl.

In certain preferred embodiments of the invention:

when R is ^4a1 、R ^4a2 、R ^4b 、R ^5b 、R ⁶ And R ⁷ Independently is C _1-4 When alkyl, said C ₁ -C ₄ Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, preferably methyl or ethyl.

In certain preferred embodiments of the invention:

when R is ^4c Independently is C _1-4 Alkyl or-O-C _1-4 When alkyl, said C ₁ -C ₄ Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, preferably methyl or ethyl.

In certain preferred embodiments of the invention:

when R is ⁵ Independently is C _1-4 Alkyl radical, C _1-4 alkyl-O-, C _1-4 When halogenated alkyl, said C _1-4 Alkyl radical, C _1-4 alkyl-O-and C _1-4 C in haloalkyl _1-4 Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, preferably methyl.

In certain preferred embodiments of the invention:

when R is ⁵ When independently an amino group protected with an N-protecting group, the protecting group may be

R ^5a Is C _1-4 Alkyl or C _1-4 alkyl-O-; c _1-4 Alkyl and C _1-4 C in alkyl-O-radicals _1-4 Alkyl groups may independently be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl;

for example, in

It may be t-butoxycarbonyl.

In certain preferred embodiments of the invention:

x is N.

In certain preferred embodiments of the invention:

R ¹ selected from methyl, cyano, trifluoromethyl, difluoromethyl,

In certain preferred embodiments of the invention:

R ² is H.

In certain preferred embodiments of the invention:

R ³ selected from H, methyl, halogen or cyano.

In certain preferred embodiments of the invention:

R ⁴ selected from cyano, -C (= O) NR ^4a1 R ^4a2 、-S(＝O) ₂ R ^4b or-C (= O) R ^4c 。

In certain preferred embodiments of the invention:

R ^4a1 、R ^4a2 、R ^4b 、R ^4c 、R ^5b 、R ⁶ and R ⁷ Independently a methyl group.

In certain preferred embodiments of the invention:

x is N;

R ¹ selected from methyl, cyano, trifluoromethyl, difluoromethyl,

R ³ Selected from H, methyl, halogen or cyano;

R ⁴ selected from cyano, -C (= O) NR ^4a1 R ^4a2 、-S(＝O) ₂ R ^4b or-C (= O) R ^4c ；

R ^4a1 、R ^4a2 、R ^4b And R ^4c Is as defined above.

In certain preferred embodiments of the invention:

x is N;

R ¹ is cyano, R ^5b Substituted or unsubstituted amino, R ⁵ Substituted or unsubstituted C _1-4 An alkyl group; for example R ⁵ Substituted or unsubstituted amino, R ⁵ Substituted or unsubstituted C _1-4 An alkyl group;

R ³ is H, R ⁵ Substituted or unsubstituted C _1-4 Alkyl, halogen or cyano; such as H;

R ⁵ independently is C _1-4 Alkyl, -N (R) ^5b R ^5b ) A hydroxyl group;

R ⁴ selected from cyano, -C (= O) NR ^4a1 R ^4a2 、-S(＝O) ₂ R ^4b or-C (= O) R ^4c (ii) a For example R ⁴ Selected from cyano or-S (= O) ₂ R ^4b (ii) a For example R ^4b Independently is C _1-4 An alkyl group;

R ^5b independently is H or C _1-4 An alkyl group.

In certain preferred embodiments of the invention:

x is N;

R ¹ is R ⁵ Substituted or unsubstituted C _1-4 An alkyl group;

R ³ is H;

R ⁵ independently is C _1-4 Alkyl, -N (R) ^5b R ^5b ) A hydroxyl group;

R ⁴ selected from cyano or-S (= O) ₂ R ^4b ；

R ^4b Independently is C _1-4 An alkyl group;

R ^5b independently is H or C _1-4 An alkyl group.

In certain preferred embodiments of the present invention, the compound of formula (I) is selected from the group consisting of the compounds shown below:

in the above compounds, R ¹ Is determined byAs defined above.

the invention also provides a preparation method of the compound shown in the formula (I), which comprises the following steps:

halogenated intermediate B ₀ With intermediate E ₀ Carrying out coupling reaction to obtain a compound shown as a formula (I);

wherein W represents halogen (e.g. Br); r ^x is-B (OH) ₂ Or

A、R ¹ 、R ³ 、R ⁴ And X is as defined above.

The invention also provides an intermediate compound B ₀ The preparation method can be any scheme as follows:

the method comprises the following steps:

in the step (1), the compound SM-5 is subjected to ring closing under the action of a catalyst (such as phosphorus oxychloride) to obtain a compound SM-6,

the compound SM-6 in the step (2) reacts under the action of an oxidant (such as m-chloroperoxybenzoic acid) to obtain a compound SM-7,

step (3) Compounds SM-7 and H ₂ N-CH ₂ Reaction of-A in the presence of a base (e.g. sodium carbonate) to give compound B ₀ ；

Scheme two, which is shown as step (2) to step (3) in scheme one;

scheme III, which is shown as step (3) in scheme I;

the reaction equation is as follows:

wherein, W, A, R ⁴ Is as defined above.

The solvent to which the present invention relates may be selected, for example, from: methanol, ethanol, isopropanol, toluene, xylene, chlorobenzene, water, dichloromethane, chloroform, carbon tetrachloride, 1, 2-dichloroethane, dioxane, DMF, acetonitrile, DMSO, NMP, THF, or a combination thereof; for example selected from dichloromethane, chloroform, 1, 2-dichloroethane, dioxane, DMF, acetonitrile, DMSO, NMP, THF or combinations thereof.

The base to which the present invention relates may include organic bases and inorganic bases.

The organic base to which the present invention relates may be selected, for example, from: TEA, DIPEA, or a combination thereof.

The inorganic base to which the present invention relates may be selected from, for example: sodium hydride, sodium methoxide, potassium carbonate, sodium carbonate, cesium carbonate, potassium tert-butoxide, sodium tert-butoxide, lithium tert-butoxide, liHMDS, LDA, butyllithium, potassium hydroxide, potassium acetate, lithium aluminum hydride or a combination thereof; for example selected from sodium hydride, potassium carbonate, sodium carbonate, cesium carbonate, potassium tert-butoxide, sodium tert-butoxide, liHMDS, LDA, butyllithium or combinations thereof.

The invention also provides a pharmaceutical composition, which comprises a therapeutically effective amount of the compound shown as the formula (I), pharmaceutically acceptable salts or stereoisomers thereof and pharmaceutically acceptable auxiliary materials. The pharmaceutically acceptable adjuvant may be one or more of a diluent, an absorbent, a wetting agent, a binder, a disintegrant, and a lubricant.

The invention also provides the compound shown in the formula (I) or the pharmaceutically acceptable salt or the stereoisomer thereof, or the application of the pharmaceutical composition in preparing medicines. The medicament is preferably a medicament for treating cancer.

The invention also provides the application of the compound shown in the formula (I) or the pharmaceutically acceptable salt or the stereoisomer thereof, or the pharmaceutical composition in preparing the medicines for treating cancers mediated by the EED protein and/or the PRC2 protein complex.

Preferably, the cancer includes, but is not limited to, diffuse large B-cell lymphoma, follicular lymphoma, non-hodgkin's lymphoma and like lymphoma, leukemia, multiple myeloma, mesothelioma, gastric cancer, malignant rhabdoid tumor, liver cancer, prostate cancer, breast cancer, brain tumors including neuroblastoma, glioma, glioblastoma and astrocytoma, cervical cancer, colon cancer, melanoma, endometrial cancer, esophageal cancer, head and neck cancer, lung cancer, nasopharyngeal cancer, ovarian cancer, pancreatic cancer, renal cancer, rectal cancer, thyroid cancer, parathyroid tumor, uterine tumor, and soft tissue sarcoma, and the like.

Preferably, the compound, its pharmaceutically acceptable salt or its stereoisomer is used in combination with other drugs; more preferably, the other drug is selected from an anticancer drug, a tumor immunity drug, an antiallergic drug, an antiemetic, an analgesic, or a cytoprotective drug.

The invention also provides a pharmaceutical preparation which comprises the compound shown as the formula (I) as described above, pharmaceutically acceptable salts thereof or stereoisomers thereof, or a pharmaceutical composition as described above. The pharmaceutical preparations are preferably in the form of suspensions, syrups, emulsions, solutions and the like in the form of tablets, capsules (e.g., sustained-release or timed-release capsules), pills, powders, granules (e.g., small granules), elixirs, tinctures, suspensions (e.g., nanosuspensions, microsuspensions) and spray-dried dispersions and the like. The preferred modes of administration of the pharmaceutical preparation are oral, sublingual, injection including subcutaneous injection, intravenous injection, intramuscular injection, intrasternal injection, infusion, etc., nasal (e.g., nasal inhalation), topical (e.g., creams and ointments), rectal (e.g., suppositories), and the like. The compounds disclosed herein may be administered alone or in combination with a suitable pharmaceutical carrier.

The invention also provides the medicine preparation which can be formulated into proper dosage of medicine so as to be convenient and control the dosage of the medicine. The dosage regimen of the compounds disclosed herein will vary with such factors as the pharmacodynamics and mode of administration, the subject, sex, age, health, weight, condition, other concurrent conditions, frequency of administration, liver and kidney function, and the effect desired, etc. The compounds disclosed herein may be administered in a single dose per day, or may be administered in a total dose divided into multiple doses (e.g., two to four times per day).

The present invention also provides a method of treating cancer comprising administering to a patient in need of such treatment a therapeutically effective amount of a compound of formula (I), a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, as described above, or a pharmaceutical composition as described above. Such cancers include, but are not limited to, diffuse large B-cell lymphoma, follicular lymphoma, non-hodgkin's lymphoma and like lymphoid cancers, leukemia, multiple myeloma, mesothelioma, gastric cancer, malignant rhabdoid tumor, liver cancer, prostate cancer, breast cancer, brain tumors including neuroblastoma, glioma, glioblastoma and astrocytoma, cervical cancer, colon cancer, melanoma, endometrial cancer, esophageal cancer, head and neck cancer, lung cancer, nasopharyngeal cancer, ovarian cancer, pancreatic cancer, renal cancer, rectal cancer, thyroid cancer, parathyroid tumor, uterine tumor, and soft tissue sarcoma, and the like.

The invention also provides a compound shown in the formula (I), pharmaceutically acceptable salt thereof or stereoisomer thereof, and other medicaments which are selected from the following medicaments: anticancer drugs, tumor immunity drugs, antiallergic drugs, antiemetics, analgesics, cytoprotective drugs, and the like, which are used in combination, have a better effect.

In the present invention, the cancer is preferably a cancer mediated by the complex of the EED protein and/or the PRC2 protein.

The present invention also provides a method of inhibiting the activity of an EED protein and/or a PRC2 protein complex, comprising administering to a subject a therapeutically effective amount of a compound according to formula (I), a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, as described above, or a pharmaceutical composition as described above.

The present invention also provides a method of blocking the binding of EED to H3K27 (e.g., H3K27me 3), comprising administering to a subject a therapeutically effective amount of a compound according to formula (I), a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, as described above, or a pharmaceutical composition as described above.

It is to be understood that within the scope of the present invention, the above-described features of the present invention and those specifically described below (e.g., in the examples) may be combined with each other to form new or preferred embodiments. Not to be reiterated herein, but to the extent of space.

Description of the terms

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

As used herein, the term "about" when used in reference to a specifically recited value means that the value may vary by no more than 1% from the recited value. For example, as used herein, the expression "about 100" includes 99 and 101 and all values in between (e.g., 99.1, 99.2, 99.3, 99.4, etc.).

As used herein, the term "comprising" or "includes" can be open, semi-closed, and closed. In other words, the term also includes "consisting essentially of or" consisting of 823030A ".

Radical definition

Definitions for standardized chemical terms can be found in the literature references, including Carey and Sundberg "ADVANCED ORGANIC CHEMISTRY 4TH ED." vols. A (2000) and B (2001), plenum Press, new York. Unless otherwise indicated, conventional methods within the skill of the art are employed, such as mass spectrometry, NMR, IR and UV/VIS spectroscopy, and pharmacological methods. Unless a specific definition is set forth, the terms used herein in the pertinent description of analytical chemistry, organic synthetic chemistry, and pharmaceutical chemistry are known in the art. Standard techniques can be used in chemical synthesis, chemical analysis, pharmaceutical preparation, formulation and delivery, and treatment of patients. For example, the reaction and purification can be carried out using the instructions of the kit from the manufacturer, or according to the methods known in the art or the instructions of the present invention. The techniques and methods described above can generally be practiced according to conventional methods well known in the art, as described in various general and more specific documents referred to and discussed in this specification. In the present specification, groups and substituents thereof may be selected by one skilled in the art to provide stable moieties and compounds.

When a substituent is described by a general formula written from left to right, the substituent also includes chemically equivalent substituents obtained when the formula is written from right to left. For example, -CH ₂ O-is equivalent to-OCH ₂ -。

The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described. All documents, or portions of documents, cited in this application, including but not limited to patents, patent applications, articles, books, operating manuals, and treatises, are hereby incorporated by reference in their entirety.

Certain chemical groups defined herein are preceded by a shorthand notation to indicate the total number of carbon atoms present in the group. E.g. C _1-6 Alkyl refers to an alkyl group as defined below having a total of 1 to 6 carbon atoms (e.g., 1,2,3,4,5,6 carbon atoms). The total number of carbon atoms in the shorthand notation excludes carbons that may be present in a substituent of the group.

When none of the listed substituents are indicated by which atom it is attached to a general chemical structure (including but not specifically mentioned compounds), such substituent may be bonded through any atom thereof. Combinations of substituents and/or variants thereof are permissible only if such combinations result in stable compounds.

In addition to the foregoing, the following terms, when used in the specification and claims of this application, have the meanings indicated below, unless otherwise specifically indicated.

In the various parts of this specification, substituents of the disclosed compounds are disclosed in terms of group type or range. Particularly point outThe invention includes each and every independent subcombination of the various members of these groups classes and ranges. The term "C _x -C _y Alkyl "or" C _x-y Alkyl "refers to straight or branched chain saturated hydrocarbons containing from x to y carbon atoms. For example, the term "C ₁ -C ₆ Alkyl "or" C _1-6 Alkyl "means in particular independently disclosed methyl, ethyl, C ₃ Alkyl radical, C ₄ Alkyl radical, C ₅ Alkyl and C ₆ An alkyl group; ' C _1-4 Alkyl "or" C ₁ -C ₄ Alkyl refers specifically to independently disclosed methyl, ethyl, C ₃ Alkyl (i.e. propyl, including n-propyl and isopropyl), C ₄ Alkyl (i.e., butyl, including n-butyl, isobutyl, sec-butyl, and tert-butyl).

When no substituent is explicitly indicated in the listed group, such group is simply referred to as unsubstituted. For example when "C ₁ -C ₄ When an alkyl group is "without the limitation of" substituted or unsubstituted ", it means only" C ₁ -C ₄ Alkyl "by itself or unsubstituted C ₁ -C ₄ Alkyl groups ".

In each of the sections of the present invention, linking substituents are described. Where the structure clearly requires a linking group, the markush variables listed for that group are understood to be linking groups. For example, if the structure requires a linking group and the markush group definition for this variable recites "alkyl," it is to be understood that this "alkyl" represents an attached alkylene group.

In some specific structures, when an alkyl group is explicitly indicated as a linking group, then the alkyl group represents a linked alkylene group, e.g., the group "halo-C ₁ -C ₆ C in alkyl ₁ -C ₆ Alkyl is understood to mean C ₁ -C ₆ An alkylene group.

In addition, unless otherwise expressly indicated, the description of "\8230" used herein, independently, "is to be understood in a broad sense and means that each individual entity described is independent of the other and may be independently the same or different specific groups. In more detail, the description "\8230, independently" means that the specific options expressed between the same symbols in different groups do not affect each other; it may also be indicated that the specific options expressed between the same symbols do not affect each other in the same group.

It should be understood that as used herein, singular forms such as "a", "an", include plural references unless specified otherwise.

The term "one or more" or "one or more" means 1,2,3,4,5,6, 7,8,9 or more.

In this application, the term "halogen" refers to fluorine, chlorine, bromine or iodine.

When represents a single bond, it means

And

and (3) mixing.

It will be understood by those skilled in the art that, in accordance with the convention used in the art, the structural formulae used in the radicals described herein

And

means that the corresponding group R is linked to other fragments, groups in the compound via this site.

"hydroxy" refers to an-OH group.

"carbonyl" means a-C (= O) -group. When in use

When R in (A) is a carbonyl group,

is composed of

"cyano" means-CN.

"amino" means-NH ₂ 。

“R ^5b Substituted amino group ", R ^5b May be one or two, when R is ^5b When two groups are used, they may be the same group or different groups.

"carboxyl" means-COOH.

In this application, the term "alkyl" as a group or as part of another group (e.g., as used in halo-substituted alkyl and the like groups) refers to a fully saturated straight or branched hydrocarbon chain radical consisting only of carbon and hydrogen atoms, having, for example, 1 to 12 (preferably 1 to 8, more preferably 1 to 6, more preferably 1 to 4) carbon atoms, and attached to the remainder of the molecule by a single bond, including, for example, but not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, 2-methylbutyl, 2-dimethylpropyl, n-hexyl, heptyl, 2-methylhexyl, 3-methylhexyl, octyl, nonyl, and decyl, and the like.

In this application, the term "haloalkyl" as a group or part of another group means that one or more hydrogen atoms in an alkyl group (as defined herein) are replaced by a halogen (as defined herein), which may be one or more in number; when the number of the halogen is plural, the halogens are the same or different. For example, fluoroalkyl refers to an alkyl group substituted with one or more fluorines. Examples of haloalkyl include, but are not limited to, trifluoromethyl, difluoromethyl, and monofluoromethyl.

In the present application, preferably the number of carbon atoms in the "heterocycloalkyl" is 3,4,5 or 6, the number of heteroatoms is selected from N, O and S, the number of heteroatoms is 1,2,3 or 4, further preferably the number of carbon atoms is 4 or 5, the number of heteroatoms is selected from N and O, the number of heteroatoms is 1 or 2; for example

In the present application, preferably, the number of carbon atoms in the "heterospirocycloalkyl" is 4,5 or 6, the number of heteroatoms is selected from N, O and S, the number of heteroatoms is 1,2,3 or 4, further preferably, the number of heteroatoms is selected from N and O, the number of heteroatoms is 1 or 2; for example

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

In this application, the term "cycloalkyl" as a group or part of another group means a saturated cyclic hydrocarbon group.

As used herein, the term "cycloalkenyl" as a group or part of another group means a cyclic hydrocarbon group having at least one double bond (e.g., a carbon-carbon double bond). Cycloalkenyl groups can be attached to the rest of the molecule through the atoms of the double bond therein.

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

In this application, the term "aryl" as a group or as part of another group means a conjugated hydrocarbon ring system group having 6 to 18 carbon atoms, preferably having 6 to 10 carbon atoms. For the purposes of the present invention, an aryl group may be a monocyclic, bicyclic, tricyclic or higher polycyclic ring system and may also be fused to a cycloalkyl or heterocyclic group as defined above, provided that the aryl group is attached to the remainder of the molecule by a single bond via an atom on the aromatic ring. Examples of aryl groups include, but are not limited to, phenyl, naphthyl, anthryl, phenanthryl, fluorenyl, 2, 3-dihydro-1H-isoindolyl, 2-benzoxazolinone, 2H-1, 4-benzoxazin-3 (4H) -on-7-yl, and the like.

In the present application, the term "arylalkyl" refers to an alkyl group as defined above substituted with an aryl group as defined above.

In this application, the term "heteroaryl" as a group or part of another group means a 5-to 16-membered conjugated ring system group having 1 to 15 carbon atoms (preferably having 1 to 10 carbon atoms) and 1 to 6 heteroatoms selected from nitrogen, oxygen and sulfur within the ring. Unless otherwise specifically indicated in the specification, a heteroaryl group may be a monocyclic, bicyclic, tricyclic or higher ring system, and may also be fused to a cycloalkyl or heterocyclic group as defined above, provided that the heteroaryl group is attached to the rest of the molecule by a single bond via an atom on the aromatic ring. The nitrogen, carbon or sulfur atoms in the heteroaryl group may be optionally oxidized; the nitrogen atoms may optionally be quaternized. For the purposes of the present invention, heteroaryl groups are preferably stable 5-to 12-membered aromatic groups containing 1 to 5 heteroatoms selected from nitrogen, oxygen and sulfur, more preferably stable 5-to 10-membered aromatic groups containing 1 to 4 heteroatoms selected from nitrogen, oxygen and sulfur or 5-to 6-membered aromatic groups containing 1 to 3 heteroatoms selected from nitrogen, oxygen and sulfur (e.g. heteroaryl is C) ₁ -C ₅ Wherein the heteroatoms are selected from N, O and S, the number of heteroatoms being 1,2,3 or 4). Examples of heteroaryl groups include, but are not limited to, thienyl, imidazolyl, pyrazolyl, thiazolyl, oxazolyl, oxadiazolyl, isoxazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, benzimidazolyl, benzopyrazolyl, indolyl, furyl, pyrrolyl, triazolyl, tetrazolyl, triazinyl, indolizinyl, isoindolyl, indazolyl, isoindolyl, purinyl, quinolyl, isoquinolyl, naphthyridinyl, quinoxalinyl, pteridinyl, carbazolyl, carbolinyl, phenanthridinyl, phenanthrolinyl, acridinyl, phenazinylIsothiazolyl, benzothiazolyl, benzothienyl, oxatriazolyl, cinnolinyl, quinazolinyl, thiophenyl, indolizinyl, phenanthroline, isoxazolyl, phenoxazinyl, phenothiazinyl, 4,5,6,7-tetrahydrobenzo [ b ] b]Thienyl, naphthopyridyl, [1,2,4]Triazolo [4,3-b ]]Pyridazine, [1,2,4]]Triazolo [4,3-a]Pyrazine, [1,2,4]]Triazolo [4,3-c ]]Pyrimidines, [1,2,4]]Triazolo [4,3-a ]]Pyridine, imidazo [1,2-a ]]Pyridine, imidazo [1,2-b ]]Pyridazine, imidazo [1,2-a ]]Pyrazine, and the like.

In this application, "optionally" means that the subsequently described event or circumstance may or may not occur, and that the description includes instances where the event or circumstance occurs and instances where it does not. For example, "optionally substituted aryl" means that the aryl group is substituted or unsubstituted, and the description includes both substituted and unsubstituted aryl groups. The "optionally" substituents described in the claims and the description section of the present invention are selected from alkyl, alkenyl, alkynyl, halogen, haloalkyl, haloalkenyl, haloalkynyl, cyano, nitro, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl.

In the present invention, the term "substituted" or "substituent" means that one or more hydrogen atoms are replaced by the specified group. When the number of the substituents is not specified, the substituents may be one or more; when the position of substitution is not indicated, the substitution may be at any position, but formation of a stable or chemically feasible chemical is permissible.

When any variable (e.g., R) occurs more than one time in the composition or structure of a compound, its definition in each case is independent. Thus, for example, if a group is substituted with 0-2R, the group may optionally be substituted with up to two R, and there are separate options for R in each case. For example, in

Wherein when n is 2, it means that the phenyl ring is substituted with 2R and each R has independent options, i.e.The 2R's may be the same or different. Furthermore, combinations of substituents and/or variants thereof are permissible only if such combinations result in stable compounds.

The terms "moiety," "structural moiety," "chemical moiety," "group," "chemical group" as used herein refer to a specific fragment or functional group in a molecule. Chemical moieties are generally considered to be chemical entities that are embedded in or attached to a molecule.

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

"tautomer" refers to an isomer formed by the transfer of a proton from one atom of a molecule to another atom of the same molecule. All tautomeric forms of the compounds of the invention are also intended to be included within the scope of the invention.

The compounds of the present invention or pharmaceutically acceptable salts thereof may contain one or more chiral carbon atoms and may therefore give rise to enantiomers, diastereomers and other stereoisomeric forms. Each chiral carbon atom may be defined as (R) -or (S) -, based on stereochemistry. The present invention is intended to include all possible isomers, as well as racemates and optically pure forms thereof. The compounds of the invention may be prepared by selecting as starting materials or intermediates racemates, diastereomers or enantiomers. Optically active isomers may be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques, e.g., crystallization and chiral chromatography. Stereoisomers of the compounds of the present invention may be present as (R) -or (S) -isomers.

Conventional techniques for the preparation/separation of individual isomers include Chiral synthesis from suitable optically pure precursors, or resolution of the racemates (or racemates of salts or derivatives) using, for example, chiral high performance liquid chromatography, as described, for example, in Gerald Gubitz and Martin G.Schmid (eds.), chiral Separations, methods and Protocols, methods in Molecular Biology, vol.243,2004; m. stalup, chiral Separations, annu. Rev. Anal. Chem.3:341-63,2010; fumiss et al (EDs.), VOGEL' S ENCYCOPEDIA OF PRACTICAL ORGANIC CHEMISTRY 5.Sup.TH ED., longman Scientific and Technical Ltd., essex,1991,809-816; heller, acc, chem, res, 1990,23,128.

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

"pharmaceutically acceptable acid addition salts" refers to salts with inorganic or organic acids which retain the biological effectiveness of the free base without other side effects. Inorganic acid salts include, but are not limited to, hydrochloride, hydrobromide, sulfate, nitrate, phosphate, and the like; organic acid salts include, but are not limited to, formates, acetates, 2-dichloroacetates, trifluoroacetates, propionates, caproates, caprylates, caprates, undecenates, glycolates, gluconates, lactates, sebacates, adipates, glutarates, malonates, oxalates, maleates, succinates, fumarates, tartrates, citrates, palmitates, stearates, oleates, cinnamates, laurates, malates, glutamates, pyroglutamates, aspartates, benzoates, methanesulfonates, benzenesulfonates, p-toluenesulfonates, alginates, ascorbates, salicylates, 4-aminosalicylates, napadisylates, and the like. These salts can be prepared by methods known in the art.

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

In the present application, "pharmaceutical composition" refers to a formulation of a compound of the present invention with a vehicle generally accepted in the art for delivering biologically active compounds to a mammal (e.g., a human). The medium includes a pharmaceutically acceptable carrier. The purpose of the pharmaceutical composition is to facilitate administration to an organism, facilitate absorption of active ingredients and exert biological activity.

The term "pharmaceutically acceptable" as used herein refers to a substance (e.g., carrier or diluent) that does not affect the biological activity or properties of the compounds of the present invention and is relatively non-toxic, i.e., the substance can be administered to an individual without causing an adverse biological response or interacting in an adverse manner with any of the components contained in the composition.

As used herein, "pharmaceutically acceptable adjuvant" includes, but is not limited to, any adjuvant, carrier, excipient, glidant, sweetener, diluent, preservative, dye/colorant, flavoring agent, surfactant, wetting agent, dispersing agent, suspending agent, stabilizer, isotonizing agent, solvent, or emulsifier that is approved by the relevant governmental regulatory agency for human or livestock use.

The terms "preventing," "prevention," and "prevention" as used herein include reducing the likelihood of occurrence or worsening of a disease or disorder in a patient.

The term "treatment" and other similar synonyms as used herein include the following meanings:

(i) Preventing the occurrence of a disease or condition in a mammal, particularly when such mammal is susceptible to the disease or condition, but has not been diagnosed as having the disease or condition;

(ii) Inhibiting the disease or disorder, i.e., arresting its development;

(iii) Alleviating the disease or condition, i.e., causing the state of the disease or condition to resolve; or

(iv) Alleviating the symptoms caused by the disease or disorder.

The terms "effective amount," "therapeutically effective amount," or "pharmaceutically effective amount" as used herein, refer to an amount of at least one agent or compound sufficient to alleviate one or more symptoms of the disease or condition being treated to some extent upon administration. The result may be a reduction and/or alleviation of signs, symptoms, or causes, or any other desired change in a biological system. For example, an "effective amount" for treatment is the amount of a composition comprising a compound disclosed herein that is clinically necessary to provide a significant remission effect of the condition. An effective amount suitable in any individual case can be determined using techniques such as a dose escalation assay.

The terms "administering," "administration," "administering," and the like, as used herein, refer to a method of delivering a compound or composition to a desired site for a biological effect. These methods include, but are not limited to, oral routes, via the duodenal route, parenteral injection (including intravenous, subcutaneous, intraperitoneal, intramuscular, intraarterial injection or infusion), topical administration, and rectal administration. Administration techniques useful for The compounds and methods described herein are well known to those skilled in The art, for example, in Goodman and Gilman, the pharmaceutical Basis of Therapeutics, current ed.; pergamon; and Remington's, pharmaceutical Sciences (current edition), mack Publishing co. In preferred embodiments, the compounds and compositions discussed herein are administered orally.

The terms "drug combination", "administering other treatment", "administering other therapeutic agent" and the like as used herein refer to a drug treatment obtained by mixing or combining more than one active ingredient, including fixed and unfixed combinations of active ingredients. The term "fixed combination" refers to the simultaneous administration of at least one compound described herein and at least one synergistic agent to a patient in the form of a single entity or a single dosage form. The term "non-fixed combination" means that at least one compound described herein and at least one synergistic formulation are administered to a patient as separate entities either simultaneously, in combination, or sequentially at variable intervals. These also apply to cocktail therapy, for example the administration of three or more active ingredients.

It will also be appreciated by those skilled in the art that in the processes described below, the functional groups of the intermediate compounds may need to be protected by suitable protecting groups. Such functional groups include hydroxyl, amino, mercapto, and carboxylic acid. Suitable hydroxy protecting groups include trialkylsilyl or diarylalkylsilyl groups (e.g.tert-butyldimethylsilyl, tert-butyldiphenylsilyl or trimethylsilyl), tetrahydropyranyl, benzyl, and the like. Suitable protecting groups for amino, amidino and guanidino include t-butyloxycarbonyl, benzyloxycarbonyl and the like. Suitable thiol protecting groups include-C (O) -R "(where R" is alkyl, aryl or aralkyl), p-methoxybenzyl, trityl and the like. Suitable carboxyl protecting groups include alkyl, aryl or aralkyl esters.

Protecting groups may be introduced and removed according to standard techniques known to those skilled in the art and as described herein. The use of protecting Groups is described in detail in Greene, T.W. and P.G.M.Wuts, protective Groups in organic Synthesis, (1999), 4th Ed., wiley. The protecting group may also be a polymeric resin.

The above preferred conditions may be combined arbitrarily to obtain preferred embodiments of the present invention without departing from the general knowledge in the art.

The reagents and starting materials used in the present invention are commercially available.

The positive progress effects of the invention are as follows: the anti-cell proliferation activity of the compounds of the invention is increased by about 10-fold compared to the compound N- ((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) -8- (2-methylpyridin-3-yl) - [1,2,4] triazolo [4,3-c ] pyrimidin-5-amine in US20160176882A1, WO2017219948 A1. When the compound disclosed by the invention is combined with the EED protein, a bicyclic structure outside a combined 'pocket' enables the compound to have better metabolic stability.

Detailed Description

The invention is further illustrated by the following examples, which are not intended to limit the scope of the invention. The experimental methods without specifying specific conditions in the following examples were selected according to the conventional methods and conditions, or according to the commercial instructions.

The starting materials used in the following examples are commercially available from chemical vendors such as Aldrich, TCI, alfa Aesar, bidey, annelgie, etc., or can be synthesized by known methods.

In the following examples, the ice bath is-5 ℃ to 0 ℃, the room temperature is 10 ℃ to 30 ℃, and the reflux temperature is generally the reflux temperature of the solvent under normal pressure. The reaction is carried out overnight in a period of 8 to 15 hours. In the following examples, the operation temperature is not limited and is carried out at room temperature.

In the following examples, the separation and purification of intermediates and final products are by normal phase or reverse phase chromatographic column separation or other suitable methods. The normal phase flash chromatography column uses ethyl acetate and n-hexane or methanol and dichloromethane as mobile phase. Reverse phase preparative High Pressure Liquid Chromatography (HPLC) was carried out using a C18 column with UV 214nm and 254nm detection, with mobile phases A (water and 0.1% formic acid), B (acetonitrile) or mobile phases A (water and 0.1% ammonium bicarbonate), B (acetonitrile).

In each example:

LCMS apparatus: pump Agilent 1260 UV detector: agilent 1260 DAD

Mass Spectrometer API 3000

A chromatographic column: waters sunfire C18, 4.6X 50mm,5 μm

Mobile phase: A-H ₂ O (0.1% HCOOH); b-acetonitrile

NMR instrument: bruker Ascend 400M ( ¹ H NMR:400MHz； ¹³ C NMR:100MHz)。

Example 1 intermediate 4- (aminomethyl) -5-fluoro-2, 3-dihydrobenzofuran-3-ol (a):

the method comprises the following steps: intermediate 2-bromo-3, 6-difluorobenzaldehyde (a-2):

a-1 (22g, 114mmol) and dry THF (200 mL) were added to a 500mL dry three-necked flask and cooled to-70 ℃. Lithium diisopropylamide (2M, 68.4 mL) was added slowly dropwise to the reaction. After the reaction mixture was stirred at the same temperature for 45 minutes, DMF (17.8mL, 228mmol) was added thereto. The reaction solution was stirred at the same temperature for two hours and then warmed to 0 ℃. Saturated ammonium chloride (200 mL) was added to the reaction. The reaction was extracted with EtOAc (200mL X2). The combined organic phases were washed once with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified on silica gel (petroleum ether: ethyl acetate = 100).

¹ H NMR(400MHz,CDCl ₃ )δ10.34(s,1H),7.34(ddd,J＝9.2,7.4,4.5Hz,1H),7.16(td,J＝9.3,4.0Hz,1H)ppm.

Step two: intermediate 2-bromo-3-fluoro-6-methoxybenzaldehyde (a-3):

a-2 (20g, 90.5mmol) was charged in a 2L three-necked flask and dissolved with stirring in anhydrous THF (1000 ml) and MeOH (200 ml). Sodium methoxide (5.87g, 108.6 mmol) was added thereto, and the reaction mixture was stirred at 60 ℃ for 18 hours. Most of the solvent was removed by concentration under reduced pressure, and 500mL of water was added. The suspension was stirred for 30 minutes and then filtered to collect the solid. The solid was slurried with a mixture of petroleum ether and ethyl acetate (5).

LC-MS:m/z 233.1[M+H] ⁺ .

Step three: intermediate 2-bromo-3-fluoro-6-hydroxybenzaldehyde (a-4):

A1L one-necked flask was charged with A-3 (16.8g, 72.1mmol) and methylene chloride (300 mL). Boron tribromide (21.7 g,86.5 mmol) was slowly added dropwise at-78 deg.C, the reaction was warmed to room temperature, and stirred for 18 hours. The reaction was diluted with dichloromethane (300 mL) and saturated sodium bicarbonate (300 mL) was added slowly. The organic phase was washed twice with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified on silica gel (petroleum ether: ethyl acetate =50: 1) to give a-4 (10g, 63.3% yield) as a pale yellow solid.

¹ H NMR(400MHz,CDCl ₃ )δ11.77(s,1H),10.34(s,1H),7.29(dt,J＝12.7,6.3Hz,1H),6.94(dd,J＝9.3,4.1Hz,1H)ppm.

Step four: intermediate 4-bromo-5-fluoro-2, 3-dihydrobenzofuran-3-ol (A-5):

trimethyl sulfoxide iodide (9.73g, 44.2mmol) and DMSO (50 ml) were charged in a dry three-necked flask. Sodium tert-butoxide (4.25g, 44.2mmol) was added under an ice-water bath. The reaction mixture was stirred at room temperature for 2 hours, and A-4 (8.8g, 40.2mmol) was added thereto. After the reaction mixture was stirred at room temperature for 18 hours, ethyl acetate (250 ml) and water (250 ml) were added to the mixture, followed by extraction with ethyl acetate (250ml X2). The organic phase was washed once with water and once with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified on silica gel (petroleum ether: ethyl acetate =10 = 1) to give a-5 (6.2g, 66.2% yield) as a white solid.

¹ H NMR(400MHz,CDCl ₃ )δ7.03(t,J＝8.7Hz,1H),6.76(dd,J＝8.8,3.5Hz,1H),5.51-5.40(m,1H),4.58(ddd,J＝13.4,10.8,4.6Hz,2H),2.33(d,J＝4.9Hz,1H)ppm.

Step five: intermediate 5-fluoro-3-hydroxy-2, 3-dihydrobenzofuran-4-carbonitrile (A-6):

a-5 (2.7 g,11.6 mmol), zinc cyanide (2.04g, 17.4 mmol), DMF (50 ml) and tetrakis triphenylphosphine palladium (1.34g, 1.16mmol) were added sequentially to a dry one-neck flask. The reaction solution was heated to 120 ℃ under nitrogen protection and stirred for 18 hours. The reaction solution was cooled to room temperature, and extracted with ethyl acetate (200mL X3) and water (200 ml). The organic phase was washed once with water and brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified on silica gel (petroleum ether: ethyl acetate = 5) to give a-6 (1.6 g,77% yield) as a white solid.

¹ H NMR(400MHz,CDCl ₃ )δ7.20-6.99(m,2H),5.63(dd,J＝6.9,2.9Hz,1H),4.69(dd,J＝10.8,7.1Hz,1H),4.55(dd,J＝10.8,3.1Hz,1H),2.74(s,1H)ppm.

Step six: intermediate (5-fluoro-2, 3-dihydrobenzofuran-4-yl) methylamine (A2):

a-6 (1.55g, 8.65mmol), trifluoroacetic acid (1.09g, 8.65mmol), methanol (20 mL) and 10% palladium on carbon (2 g, 50% water) were added sequentially in a 100mL single-neck flask. The reaction mixture was bubbled with hydrogen for 5 minutes, purged three times with a hydrogen balloon and stirred at 60 ℃ for 48 hours under a hydrogen balloon. The mixture was filtered through celite, washed with methanol (50mL X2), the filtrate was concentrated under reduced pressure to which dioxane (10 mL) and 10M aqueous sodium hydroxide (1 mL) were added, extracted with dioxane (10 mL X2), and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. Compound A2 (1.4 g,90% pure, 87% yield) was obtained and used directly in the next step.

¹ H NMR(400MHz,CD ₃ OD)δ3.27(t,2H),3.77(s,2H),4.56(t,2H),6.59(dd 1H),6.81(dd,1H)ppm；LC-MS:m/z 168.1[M+H] ⁺ .

Example 2- (5- ((dimethylamino) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) imidazo [1,5-c ] pyrimidine-1-carbonitrile (BR-1)

The method comprises the following steps: 2- (5-bromo-2- (methylthio) pyrimidin-4-yl) -2- ((diphenylmethylene) amino) acetonitrile (2)

To a solution of sodium hydride (3.63g, 90.8mmol,60% purity) in N, N-dimethylformamide (60.0 mL) was added a solution of N- (diphenylmethylene) aminoacetonitrile 1 (10.0 g, 45.4mmol) in N, N-dimethylformamide (50.0 mL) at zero degrees, and after 2 minutes, 5-bromo-4-chloro-2- (methylthio) pyrimidine (10.8g, 45.4mmol) in N, N-dimethylformamide (30.0 mL) was added at zero degrees, and the reaction was carried out at 40 ℃ for 2 hours. The reaction was monitored by LCMS and quenched with aqueous ammonium chloride, extracted with ethyl acetate and dried to give crude 2- (5-bromo-2- (methylthio) pyrimidin-4-yl) -2- ((diphenylmethylene) amino) acetonitrile 2 (15.0 g, crude) which was used directly in the next step.

¹ H NMR(400MHz,DMSO-d ₆ )δ7.58-7.52(m,7H),7.44-7.40(m,3H),7.26-7.23(m,2H),4.37(s,3H)ppm；LCMS m/z 424.7[M+H] ⁺ .

Step two: 2-amino-2- (5-bromo-2- (methylthio) pyrimidin-4-yl) acetonitrile (3)

2- (5-bromo-2- (methylthio) pyrimidin-4-yl) -2- ((diphenylmethylene) amino) acetonitrile 2 (15.0 g, crude), tetrahydrofuran (200 mL) and aqueous solution (150 mL) were added to concentrated hydrochloric acid (12M, 23.6 mL) at zero degrees and reacted at 20 ℃ for 2 hours. The reaction was monitored by TLC for completion, after the tetrahydrofuran was decanted from the reaction solution, aqueous sodium hydroxide (1.00M) was added to adjust the pH to about 8.0, and the mixture was extracted with ethyl acetate, dried and dried to give crude 2-amino-2- (5-bromo-2- (methylthio) pyrimidin-4-yl) acetonitrile 3 (9.00 g, crude) which was used directly in the next step.

¹ H NMR(400MHz,DMSO-d ₆ )δ8.86(s,1H),5.28(s,1H),2.89(s,2H),2.57(s,3H)ppm；LCMS m/z260.9[M+H] ⁺ .

Step three: n- ((5-bromo-2- (methylthio) pyrimidin-4-yl) (cyano) methyl) carboxamide (4)

Formic acid (39.0 g, 848mmol) and acetic anhydride (34.8g, 341mmol) were mixed, reacted at 50 ℃ for 1 hour, cooled to 0 ℃ and added to a solution of 2-amino-2- (5-bromo-2- (methylthio) pyrimidin-4-yl) acetonitrile 3 (9.00 g, crude) in methylene chloride (100 mL). The reaction was carried out at 20 ℃ for 2h. The reaction was monitored by TLC, adjusted to 8 with aqueous sodium hydroxide (1.00M) at zero degrees, extracted with ethyl acetate, dried, and column chromatographed to give crude N- ((5-bromo-2- (methylthio) pyrimidin-4-yl) (cyano) methyl) carboxamide 4 (7.00g, 22.6mmol,73.2% yield).

Step four: 8-bromo-5- (methylthio) imidazo [1,5-c ] pyrimidine-1-carbonitrile (5)

Phosphorus oxychloride (28.8g, 188mmol) was added to N- ((5-bromo-2- (methylthio) pyrimidin-4-yl) (cyano) methyl) formamide 4 (7.00g, 24.3mmol) and the reaction was carried out at 110 ℃ for 1 hour. TLC is used for monitoring the reaction completion, part of phosphorus oxychloride is removed from the reaction liquid, the reaction liquid is quenched by water at the temperature of less than 30 ℃, finally the temperature is reduced to 8 by NaOH aqueous solution (6.00M), ethyl acetate is used for extraction, drying is carried out, the reaction liquid is dried to obtain crude products, the crude products are pulped by petroleum ether/ethyl acetate (5.

¹ H NMR(400MHz,DMSO-d ₆ )δ8.81(s,1H),8.06(s,1H),2.76(s,3H)ppm；LCMS m/z 270.9[M+H] ⁺ .

Step five: 8-bromo-5- (methylsulfonyl) imidazo [1,5-c ] pyrimidine-1-carbonitrile (6)

8-bromo-5- (methylthio) imidazo [1,5-c ] pyrimidine-1-carbonitrile 5 (0.60g, 2.23mmol) was dissolved in dichloromethane (5 mL), 85% m-chloroperoxybenzoic acid (1.40g, 6.91mmol) was added at 0 deg.C and stirred at 20 deg.C for 12 hours. The reaction was monitored by LCMS for completion and the reaction was used directly in the next step.

LCMS m/z 302.9[M+H] ⁺ .

Step six: 8-bromo-5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) imidazo [1,5-c ] pyrimidine-1-carbonitrile (7)

8-bromo-5- (methylsulfonyl) imidazo [1,5-c ] pyrimidine-1-carbonitrile 6 (671mg, 2.23mmol) and (5-fluoro-2, 3-dihydrobenzofuran-4-yl) methylamine A (453mg, 2.23mmol) were dissolved in dichloromethane (5 mL), sodium carbonate (1.18g, 11.1mmol) was added, and the mixture was stirred at 20 ℃ for 4 hours. The reaction was monitored by LCMS for completion, and the reaction was diluted with dichloromethane, washed with brine, dried on the organic phase and dried by spin-drying and column chromatography to give 8-bromo-5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) imidazo [1,5-c ] pyrimidine-1-carbonitrile 7 as a brown solid (400mg, 46.2% yield).

¹ H NMR(400MHz,DMSO-d ₆ )δ8.90-8.77(m,2H),7.75(s,1H),6.93(t,J＝9.4Hz,1H),6.69(dd,J＝4.0,8.8Hz,1H),4.67(s,2H),4.53(t,J＝8.8Hz,2H),3.29-3.24(m,3H)ppm；LCMS m/z 388.0[M+H] ⁺ .

Step seven: 8-bromo-5- (bromomethyl) - [1,2,4] triazolo [1,5-a ] pyridine (9)

A100 mL round bottom flask was charged with 8-bromo-5-methyl- [1,2,4] triazolo [1,5-a ] pyridine 8 (2g, 9.43mmol) and carbon tetrachloride (20 mL), and NBS (2g, 11.3 mmol) and AIBN (310mg, 0.2 mmol) were added under an ice-water bath. After the addition was complete, the reaction was allowed to warm to 80 deg.C and held overnight for 16h. The reaction solution was washed with an aqueous sodium sulfite solution and then spin-dried to give a reddish-brown solid, which was purified by a flash column (eluent petroleum ether: ethyl acetate = 3. After vacuum drying, pure 8-bromo-5- (bromomethyl) - [1,2,4] triazolo [1,5-a ] pyridine 9 was obtained (1.7g, 62% yield).

¹ H NMR(400MHz,CDCl ₃ )δ8.48(s,1H),7.77(d,J＝7.7Hz,1H),7.07(d,J＝7.7Hz,1H),4.90(s,2H)ppm；LCMS:m/z 291.9[M+H] ⁺ .

Step eight: 1- (8-bromo- [1,2,4] triazolo [1,5-a ] pyridin-5-yl) -N, N-dimethylmethylamine (10)

A100 mL round bottom flask was charged with 8-bromo-5- (bromomethyl) - [1,2,4] triazolo [1,5-a ] pyridine 9 (1g, 3.44mmol) and tetrahydrofuran (15 mL), and a 1M solution of dimethylamine in tetrahydrofuran (17mL, 17.2mmol) and potassium carbonate (0.59g, 1.43mmol) were added under an ice-water bath. After the addition was complete, the reaction was maintained at 80 ℃ for 3h. After completion of the reaction, a tan solid was obtained after liquid spin-drying, and 1- (8-bromo- [1,2,4] triazolo [1,5-a ] pyridin-5-yl) -N, N-dimethylmethylamine 10 (0.6 g,68.3% yield) was obtained after purification by elution with a rapid separation column (petroleum ether: ethyl acetate = 5).

¹ H NMR(400MHz,DMSO-d ₆ )δ8.74-8.38(m,1H),8.14-7.85(m,1H),7.31-6.90(m,1H),4.03-3.78(m,2H),2.38-2.20(m,6H)ppm；LCMS:m/z 255.0,257.0[M+H] ⁺ .

Step nine: (5- ((dimethylamino) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) boronic acid (11)

1- (8-bromo- [1,2,4] triazolo [1,5-a ] pyridin-5-yl) -N, N-dimethylmethylamine 10 (100mg, 391. Mu. Mol), pinacol diboron ester (159mg, 627. Mu. Mol), potassium acetate (115mg, 1.18mmol), [1, 1-bis (diphenylphosphino) ferrocene ] dichloropalladium dichloromethane (64.0 mg, 78.4. Mu. Mol) were dissolved in dioxane (6 mL) in a glove box and reacted at 125 ℃ for 5 hours. The reaction was monitored by LCMS for completion and the reaction was used directly in the next reaction.

LCMS m/z 221.1[M+H] ⁺ .

Step ten: 8- (5- ((dimethylamino) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) imidazo [1,5-c ] pyrimidine-1-carbonitrile (BR-1)

To the boric acid reaction solution in the previous step, 8-bromo-5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) imidazo [1,5-c ] pyrimidine-1-carbonitrile 7 (151mg, 390. Mu. Mol), potassium carbonate (108mg, 781. Mu. Mol), [1, 1-bis (diphenylphosphino) ferrocene ] dichloropalladium dichloromethane (31.9mg, 39.0. Mu. Mol), water (1 mL), dioxane (3 mL) was added and reacted at 100 ℃ for 2 hours. The reaction was monitored by LCMS for completion, the reaction was concentrated and the solution obtained by column chromatography purification and perp-HPLC was adjusted to pH 8 with saturated aqueous sodium bicarbonate solution, concentrated to remove the organic solvent and filtered to give 8- (5- ((dimethylamino) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) imidazo [1,5-c ] pyrimidine-1-carbonitrile BR-1 as a white solid (70.0 mg,36.8% yield).

¹ H NMR(400MHz,DMSO-d ₆ )δ＝8.93(s,1H),8.83(s,1H),8.50(s,1H),7.85-7.76(m,2H),7.32(d,J＝7.6Hz,1H),6.96(t,J＝9.6Hz,1H),6.72(dd,J＝4.0,8.8Hz,1H),4.77(s,2H),4.56(t,J＝8.8Hz,2H),4.03(s,2H),3.35-3.35(m,2H),2.34(s,6H)ppm；LCMS m/z 484.4[M+H] ⁺ .

EXAMPLE 3- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) -8- (5- (2-hydroxypropan-2-yl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) imidazo [1,5-c ] pyrimidine-1-carbonitrile (BR-2)

The method comprises the following steps: (E) -5-bromo-6- (((dimethylamino) methylene) amino) -pyridine-2-carboxylic acid methyl ester E-2

Dissolving (E) -5-bromo-6- (((dimethylamino) methylene) amino) -pyridine-2-carboxylic acid methyl ester (5.0g, 21.6 mmol) and 1, 1-dimethoxy-N, N-dimethylmethylamine (5.16g, 43.3mmol) in toluene (100 mL), heating the reaction solution to 110 ℃ for reaction for 4 hours, detecting by TLC that no raw material remains, and concentrating the reaction solution to obtain crude (E) -5-bromo-6- (((dimethylamino) methylene) amino) -pyridine-2-carboxylic acid methyl ester E-2 (6.00 g).

LCMS:m/z 585.7[M+H] ⁺ .

Step two: (E) -5-bromo-6- (((hydroxylamino) methylene) amino) -pyridine-2-carboxylic acid methyl ester E-3

To a reaction flask were added (E) -5-bromo-6- (((dimethylamino) methylene) amino) -pyridine-2-carboxylic acid methyl ester E-2 (6.0g, 20.97mmol), hydroxylamine hydrochloride (2.91g, 41.94mmol), sodium acetate (3.44g, 41.9mmol), and ethanol (60 mL) in this order. The reaction solution was heated to 50 ℃ under nitrogen atmosphere for 4 hours, and after completion of the reaction, the reaction solution was monitored by LCMS, cooled to room temperature, and filtered to obtain (E) -5-bromo-6- (((hydroxylamino) methylene) amino) -pyridine-2-carboxylic acid methyl ester E-3 (5.9g, 85.0% yield).

¹ H NMR(400MHz,CDCl ₃ )δ8.27(br d,J＝8.5Hz,1H),8.19-8.13(m,1H),7.85(d,J＝7.9Hz,1H),7.50(d,J＝7.9Hz,1H),3.90(s,3H)ppm；LCMS:m/z 273.7[M+H] ⁺ .

Step three: 8-bromo- [1,2,4] triazolo [1,5-a ] pyridine-5-carboxylic acid methyl ester E-4

Trifluoroacetic anhydride (23.0 g, 109mmol) was added dropwise to a solution of (E) -5-bromo-6- (((hydroxylamino) methylene) amino) -pyridine-2-carboxylic acid methyl ester E-3 (10 g, 36.49mmol) in tetrahydrofuran (100 mL) at room temperature. Then slowly raised to 75 ℃ and stirred for 3 hours. After the reaction was complete, the mixture was quenched by addition of saturated aqueous sodium bicarbonate (100 mL) at 0 ℃, extracted with ethyl acetate (100ml × 3), and the combined organic phases were dried and concentrated. The concentrate was purified by a silica gel column (petroleum ether: ethyl acetate =5:1 to 4: 1) to give the product methyl 8-bromo- [1,2,4] triazolo [1,5-a ] pyridine-5-carboxylate E-4 as a white solid (5.00 g, yield 53.5%).

Step four: 2- (8-bromo- [1,2,4] triazolo [1,5-a ] pyridin-5-yl) propan-2-ol E-5

To a solution of methyl 8-bromo- [1,2,4] triazolo [1,5-a ] pyridine-5-carboxylate E-4 (500mg, 1.95mmol) in tetrahydrofuran (10 mL) at-30 deg.C was added dropwise methylmagnesium bromide (3M, 2.60mL) and stirred at this temperature for three hours. After completion of the reaction, the reaction mixture was quenched by addition of water (10 mL), extracted with ethyl acetate (10 mL. Times.3), and the combined organic phases were dried, concentrated and purified by flash silica gel column to give 2- (8-bromo- [1,2,4] triazolo [1,5-a ] pyridin-5-yl) propan-2-ol E-5 (400mg, 80.0% yield).

¹ H NMR(400MHz,DMSO-d ₆ )δ8.61(s,1H),8.05(d,J＝8.0Hz,1H),7.28(d,J＝8.0Hz,1H),5.91(s,1H),1.72(s,6H)ppm；LCMS:m/z 255.9[M+H] ⁺ .

Step five: (5- (2-hydroxypropan-2-yl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) boronic acid (12)

2- (8-bromo- [1,2,4] triazolo [1,5-a ] pyridin-5-yl) tertiary alcohol E-5 (100mg, 390. Mu. Mol), pinacol diborate (158mg, 624. Mu. Mol), potassium acetate (114mg, 1.17mmol), [1, 1-bis (diphenylphosphine) ferrocene ] dichloropalladium dichloromethane (63.7 mg, 78.0. Mu. Mol) were dissolved in dioxane (6 mL) in a glove box and reacted at 125 ℃ for 6 hours. The reaction was monitored by LCMS for completion and the reaction was used directly for the next reaction.

LCMS m/z 222.2[M+H] ⁺ .

Step six: 5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) -8- (5- (2-hydroxypropan-2-yl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) imidazo [1,5-c ] pyrimidine-1-carbonitrile (BR-2)

To the reaction solution of boric acid 12 in the previous step were added 8-bromo-5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) imidazo [1,5-c ] pyrimidine-1-carbonitrile 7 (151mg, 389. Mu. Mol), potassium carbonate (107mg, 778. Mu. Mol), [1, 1-bis (diphenylphosphino) ferrocene ] dichloropalladium dichloromethane (31.7 mg, 38.9. Mu. Mol), water (1 mL), dioxane (3 mL) and reacted at 100 ℃ for 2 hours. The reaction was monitored by LCMS for completion, the reaction was concentrated and the solution obtained by column chromatography purification and perp-HPLC was adjusted to pH 8 with saturated aqueous sodium bicarbonate solution, concentrated to remove the organic solvent and filtered to give 5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) -8- (5- (2-hydroxypropan-2-yl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) imidazo [1,5-c ] pyrimidine-1-carbonitrile as a white solid (50.0 mg,26.0% yield).

¹ H NMR(400MHz,DMSO-d ₆ )δ＝8.90(s,1H),8.84(s,1H),8.51(s,1H),7.83-7.78(m,2H),7.44(d,J＝7.6Hz,1H),6.95(t,J＝9.4Hz,1H),6.75-6.67(m,1H),5.86(s,1H),4.77(s,2H),4.56(t,J＝8.6Hz,2H),3.37-3.32(m,2H),1.77(s,6H)ppm；LCMS m/z 485.4[M+H] ⁺ .

Example 4- (5- (dimethylamino) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) imidazo [1,5-c ] pyrimidine-1-carbonitrile (BR-3)

The method comprises the following steps: 8-iodo-N, N-dimethyl- [1,2,4] triazolo [1,5-a ] pyridin-5-amine 14

To a tetrahydrofuran solution of dimethylamine (4 mL, 2M/L) was added 5-chloro-8-iodo- [1,2,4] triazolo [1,5-a ] pyridine 13 (200mg, 0.72mmol). The mixture was stirred at 90 ℃ for 16h. LCMS showed the reaction was complete, concentrated and purified (petroleum ether: ethyl acetate = 1) to give 8-iodo-N, N-dimethyl- [1,2,4] triazolo [1,5-a ] pyridin-5-amine 14 (190mg, 90%).

LCMS:m/z 288.9[M+H] ⁺ .

Step two: (5- (dimethylamino) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) boronic acid (15)

8-iodo-N, N-dimethyl- [1,2,4] triazolo [1,5-a ] pyridin-5-amine 14 (100mg, 414. Mu. Mol), pinacol diborate (168mg, 663. Mu. Mol), potassium acetate (122mg, 1.24mmol), [1, 1-bis (diphenylphosphine) ferrocene ] dichloropalladium dichloromethane (67.7 mg, 82.9. Mu. Mol) were dissolved in dioxane (5 mL) in a glove box and reacted at 125 ℃ for 6 hours. The reaction was monitored by LCMS for completion and the reaction was used directly for the next reaction.

LCMS m/z 207.2[M+H] ⁺ .

Step three: 8- (5- (dimethylamino) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) imidazo [1,5-c ] pyrimidine-1-carbonitrile

To the boric acid 15 reaction solution in the previous step were added 8-bromo-5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) imidazo [1,5-c ] pyrimidine-1-carbonitrile (160mg, 412. Mu. Mol), potassium carbonate (114mg, 825. Mu. Mol), [1, 1-bis (diphenylphosphino) ferrocene ] palladium dichloride dichloromethane (33.6 mg, 41.2. Mu. Mol), water (1 mL), dioxane (3 mL) and reacted at 100 ℃ for 2 hours. The reaction was monitored by LCMS for completion, the reaction was concentrated and the solution obtained by column chromatography purification and perp-HPLC was adjusted to pH 8 with saturated aqueous sodium bicarbonate, concentrated to remove the organic solvent and filtered to give 8- (5- (dimethylamino) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) imidazo [1,5-c ] pyrimidine-1-carbonitrile BR-3 as a white solid (13.0 mg,6.61% yield).

¹ H NMR(400MHz,DMSO-d ₆ )δ＝8.90-8.68(m,2H),8.40(s,1H),7.70-7.61(m,2H),6.94(t,J＝9.2Hz,1H),6.74-6.66(m,1H),6.56(d,J＝8.0Hz,1H),4.74(s,2H),4.55(t,J＝8.6Hz,2H),3.37-3.33(m,2H),3.17(s,6H)ppm；LCMS m/z 470.4[M+H] ⁺ .

Example 5- ((dimethylamino) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -N- ((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) -1- (methylsulfonyl) imidazo [1,5-c ] pyrimidin-5-amine (BR-4)

Step one 8-bromo-N- ((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) imidazo [1,5-c ] pyrimidin-5-amine (17)

8-bromo-5- (methylthio) imidazo [1,5-c ] pyrimidine-16 (4.0g, 16.4 mmol) was dissolved in dichloromethane (60 ml), m-chloroperoxybenzoic acid (4.99g, 24.6mmol,85% purity) was added at 0 ℃ and reacted at 0 ℃ for 55 minutes, then triethylamine (6.63g, 65.5 mmol) was added dropwise at 0 ℃ and stirred for 5 minutes, and finally 5-fluoro-2, 3-dihydrobenzofuran-4-yl) methylamine A (3.34g, 16.4mmol, HCl) was added to the reaction solution, heated to 20 ℃ and stirred for 3 hours, TLC showed no remaining of raw material, water was added to the reaction solution and quenched, the organic phase was washed, dried, concentrated, and the crude product was purified with a flash silica gel column to give 8-bromo-N- ((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) imidazo [1,5-c ] pyrimidine-5-17 amine (4.0g, 3.40% yield).

¹ H NMR(400MHz,CDCl ₃ )δ8.23(s,1H),7.41-7.30(m,2H),6.87-6.77(m,1H),6.71-6.58(m,1H),6.10(br s,1H),4.79-4.71(m,2H),4.60(t,J＝8.8Hz,2H),3.37(t,J＝8.8Hz,2H)ppm；LCMS:m/z 363.0[M+H] ⁺

Step two: starting from 8-bromo-N- ((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) imidazo [1,5-c ] pyrimidin-5-amine (17) and (5- ((dimethylamino) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) boronic acid (11), the procedure of example two, step ten gives: 8- (5- ((dimethylamine) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -N- ((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) imidazo [1,5-c ] pyrimidin-5-amine (18):

¹ H NMR(400MHz,CDCl ₃ )δ8.22(s,1H),8.03(s,1H),7.78(s,1H),7.71(d,J＝7.4Hz,1H),7.30-7.27(m,1H),7.07(s,1H),6.87-6.72(m,1H),6.65(dd,J＝3.9,8.6Hz,1H),4.82(s,2H),4.60(t,J＝8.8Hz,2H),4.16(s,2H),3.67(t,J＝6.0Hz,1H),3.43(br t,J＝8.6Hz,2H),2.52(s,6H))ppm；LCMS:m/z 459.2[M+H] ⁺

step three: 8- (5- ((dimethylamine) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -N- ((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) -1-iodoimidazo [1,5-c ] pyrimidin-5-amine (19)

8- (5- ((dimethylamine) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -N- ((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) imidazo [1,5-c ] pyrimidin-5-amine (400mg, 0.782mmol) was dissolved in acetic acid (2 ml), N-iodosuccinimide (137mg, 0.62mmol) was added at 0 ℃, the reaction was warmed to 20 ℃ for 0.25 hours, TLC and LCMS were monitored for the end of the reaction, the reaction was adjusted to pH 7, extracted with ethyl acetate, the organic phase was washed, dried and concentrated, and the crude product was purified on a silica gel flash column to give 8- (5- ((dimethylamine) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -N- ((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) -1-iodoimidazo [1,5-c ] pyrimidine-5-amine (33.9% yield).

¹ H NMR(400MHz,DMSO-d ₆ )δ8.73(s,1H),8.53-8.47(m,1H),8.47-8.43(m,1H),7.64(d,J＝7.3Hz,1H),7.33-7.26(m,2H),6.95(t,J＝9.4Hz,1H),6.74-6.66(m,1H),4.74(d,J＝4.9Hz,2H),4.55(t,J＝8.8Hz,2H),4.03(s,2H),3.30(br s,2H),2.35(s,6H)ppm；LCMS:m/z 585.5[M+H] ⁺

Step four: 8- (5- ((dimethylamine) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -N- ((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) -1- (methylsulfonyl) imidazo [1,5-c ] pyrimidin-5-amine (BR-4)

8- (5- ((dimethylamine) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -N- ((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) -1-iodoimidazo [1,5-c ] pyrimidin-5-amine (100mg, 0.172mmol), cuprous iodide (98mg, 0.513mmol), sodium methanesulfinate (52.4mg, 0.513mmol) were dissolved in dimethyl sulfoxide (1 ml), nitrogen was bubbled for 5 minutes, microwave was applied at 120 ℃ for 20 minutes, then reaction was cooled to 100 ℃ for 3 hours, LCMS was monitored for completion of the reaction, the reaction solution was concentrated, and the crude product was purified by Pre-HPLC to give 8- (5- ((dimethylamine) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) -N- ((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) -1- (methylsulfonyl) imidazo [1,5-c ] pyrimidin-5-amine (3032 mg, yield).

¹ H NMR(400MHz,DMSO-d ₆ )δ8.94(s,1H),8.89-8.80(m,1H),8.36(br s,1H),7.66-7.56(m,2H),7.26-7.15(m,1H),6.96(t,J＝9.4Hz,1H),6.75-6.67(m,1H),4.82-4.72(m,2H),4.56(t,J＝8.7Hz,2H),3.99(br s,2H),3.31-3.31(m,2H),2.87(s,3H),2.33(br s,6H)ppm；LCMS:m/z 537.2[M+H] ⁺

Example 6 following the procedure of example 5, the replacement of (5- ((dimethylamino) methyl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) boronic acid (11) with (5- (2-hydroxypropan-2-yl) - [1,2,4] triazolo [1,5-a ] pyridin-8-yl) boronic acid (12) gave the following compound:

2- (8- (5- (((5-fluoro-2, 3-dihydrobenzofuran-4-yl) methyl) amino) -1- (methylsulfonyl) imidazo [1, 5-c)]Pyrimidin-8-yl) - [1,2,4]Triazolo [1,5-a ]]Pyridin-5-yl) propan-2-ol (BR-5) ¹ H NMR(400MHz,DMSO-d ₆ )δ8.94(s,1H),8.88-8.76(m,1H),8.38(s,1H),7.65-7.57(m,2H),7.34(d,J＝7.4Hz,1H),6.96(t,J＝9.4Hz,1H),6.80-6.62(m,1H),5.82(s,1H),4.85-4.68(m,2H),4.56(t,J＝8.7Hz,2H),3.32-3.31(m,2H),2.88(s,3H),1.76(s,6H)ppm；LCMS:m/z 538.2[M+H] ⁺

Pharmacology and uses

EED, one of the major components of the PRC2 protein complex, although not enzymatically active, plays an important role in the overall function of PRC 2. The effect of EED on PRC2 is embodied in two aspects: 1) EED directly with three methylated H3K27Me3 binding, so that can be PCR2 complexes on the need of modified chromatin; 2) EED has a great allosteric promoting effect on the enzymatic function of EZH 2. Therefore, the development of compounds as targets of allosteric protein EEDs provides a new strategy for inhibiting the activity of EZH2 enzymes. Furthermore, such inhibitors have the advantage of being better or complementary to inhibitors of the catalytic site of EZH2 enzyme, for example, EED inhibitors may also act to inhibit EZH2 enzyme activity when a patient is resistant to EZH2 enzyme inhibitors. The invention discloses that the compound can be used as an EED target inhibitor and has a therapeutic effect on diseases related to the EED and/or PRC2 action mechanism.

The biological function of the disclosed compounds is demonstrated in biochemical and cellular level assays. For example, in biochemical assays, the disclosed compounds are capable of strong competitive binding (IC) to H3K27Me3 polypeptides that bind EED proteins ₅₀ Can reach<1 nM). At the cellular level, the compounds disclosed herein can inhibit not only the methylation level of histone H3K27 but also the proliferation of cancer cells (IC) by this effect ₅₀ Can reach<1nM)。

Example 7: ELISA (H3K 27 Trimethylation) assay

Representative compounds of the disclosure were diluted in DMSO in 3-fold gradients, with 10 concentration gradients detected per compound, with the highest assay concentration being 10 μ M. Compounds were diluted 200-fold into G401 cells cultured in 96-well plates (final DMSO concentration 0.5%). After 72 hours of culture of the cells, the level of trimethylation of histone H3K27 was measured by ELISA.

Histone extraction cells treated with Compound in 96-well plates were treated with 1 XPBS (10 XPBS buffer (80 g NaCl (Sigma, product No. S3014), 2g KCl (Sigma, product No. 60128), 14.4g Na ₂ HP04 (Sigma, product number S5136), 2.4g KH ₂ P04 (Sigma, product No. P9791) to 1L water, pH to 7.4) three times, 100. Mu.L of 0.4N HCl per well, incubated at 4 ℃ for 2 hours with gentle shaking to lyse the cells. The cell lysate was then neutralized with 80 μ L of neutralization buffer (0.5M disodium hydrogen phosphate, pH 12.5,2.5mm dtt 1% cocktail (Sigma, product No. P8340) (cell lysate was mixed well with neutralization buffer).

ELISA assay method cell lysates were transferred in parallel to 2 384-well assay plates (Perkinelmer, optiPlate-384HB, product No. 6007290), one plate for detection of H3K27 trimethylation and the other plate for determination of H3 level, PBS adjusted to a final volume of 50. Mu.L/well and coated overnight at 4 ℃. The next day, the well-inside solution was discarded, washed 5 times with TBST buffer (L xTBS (10xTBS. Add 70. Mu.L of blocking buffer (TBST, 5% BSA) to the coated reaction wells and incubate at room temperature for 1 hour. The blocking buffer was discarded and primary antibody (30. Mu.L/well) was added. The required primary antibodies were diluted with blocking buffer in the following dilution ratios: anti-H3K 27me3 antibody (Cell Signaling Technology, product number 9733), 1; anti-H3 antibody (Cell Signaling Technology, product No. 4499), 1: and (4) 10000 dilution. After the addition of the primary antibody, the cells were incubated at room temperature for 1 hour. After washing 5 times with TBST, the water was drained off, and secondary antibody (30. Mu.L/well) was added to each well and incubated at room temperature for 1 hour. The secondary antibody (anti-rabbit antibody (Jackson ImmunoResearch, product No. 111-035-003)) was used after being diluted 2000-fold with blocking buffer. After 1 hour, the mixture was washed with TBST and drained. mu.L of ECL substrate (Pierce, product No. 34080) was added to each well and centrifuged at 2000rpm for 30 seconds. Various assays were performed using Molecular Devices, spectraMaxAnd (4) a product signal. Data processing: H3K27 methylation readings were first normalized with H3 signal, and 0.5% DMSO treated samples were used as controls to calculate the percent inhibition of the compound. Data were fit to dose response curves using GraphPad prisim5 program to obtain IC of test compound ₅₀ The value is obtained.

Table 1 below shows the IC of some of the compounds ₅₀ The value is obtained.

TABLE 1

Example 8: cell proliferation assay

Human B-cell non-Hodgkin lymphoma cells, KARPAS-422S, were cultured in culture flasks using standard cell culture conditions. The medium was 15% fetal bovine serum (FBS, invitrogen, product No. 10099-141), 1% penicillin/streptomycin solution (P/S) RPMI-1640 (Invitrogen, product No. 11875), and the flask was placed at a temperature of 37 ℃ and a relative humidity of 95% and 5% CO ₂ Cultured in a sterile incubator. To examine the effect of PRC2 inhibitors on cell proliferation, cells in exponential growth phase were taken at 1X10 ⁴ The density of cells/well was seeded into 96-well plates (Corning, product No. 3904) and 100 μ L of medium was added per well. Subsequently, different concentrations of compounds of the disclosure were added to the wells of seeded cells (9 concentration gradients were set for each compound, with a maximum assay concentration of 10 μ M, 3-fold gradient dilution), 2 replicates were set for each treatment concentration, and a final DMSO concentration of 0.5%. The number of viable CELLs was then determined every 3 to 4 days using Vi-CELL (Beckman Coulter). The cells counted each time are in equal density (1X 10) ⁴ Individual cells/well) were seeded into new 96-well plates, and fresh medium was replenished to 100 μ Ι _, while compounds at different concentrations were added. After culturing to day 13, 100. Mu.L of CellTiter-Glo (CellTiter-GloCellTiter-GloCellTiter-GloCTG) (Promega, product No. 3) was added to each wellG7573 And placed in the dark at room temperature for 10-20 minutes, and luminescence signals were read using Molecular Devices, spectraMax i 3X. The IC of the test compound was obtained by fitting the data to a dose response curve using GraphPad prism 5 ₅₀ The value is obtained.

Table 5 below shows the IC's of some of the compounds ₅₀ The value is obtained.

TABLE 2

The compounds disclosed herein above are useful for treating cancers associated with the mechanism of action of the EED protein and/or PRC2 protein complex, including but not limited to diffuse large B-cell lymphoma, follicular lymphoma, non-Hodgkins lymphoma and like lymphoma, leukemia, multiple myeloma, mesothelioma, gastric cancer, malignant rhabdoid tumor, liver cancer, prostate cancer, breast cancer, brain tumors including neuroblastoma, glioma, glioblastoma and astrocytoma, cervical cancer, colon cancer, melanoma, endometrial cancer, esophageal cancer, head and neck cancer, lung cancer, nasopharyngeal cancer, ovarian cancer, pancreatic cancer, renal cancer, rectal cancer, thyroid cancer, parathyroid tumor, uterine tumor, soft tissue sarcoma and the like.

Claims

1. A compound of formula (I), a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, wherein:

wherein A is independently

X is N or CR ² ；

R ¹ 、R ² And R ³ Independently is H, halogen, cyano, R ^5b Substituted or unsubstituted amino, R ^5b -O-、R ⁵ Substituted or unsubstituted C ₁ - ₄ Alkyl radical, R ⁵ Substituted or unsubstituted C _3-6 Cycloalkyl of, R ⁵ Substituted or unsubstituted C _1-4 Haloalkyl, -C (= O) R ⁶ 、-CO ₂ R ⁶ 、-C(＝O)NR ⁶ R ⁷ 、-SO ₂ R ⁶ or-SO ₂ NR ⁶ R ⁷ (ii) a When the substituents are plural, the same or different;

R ^4a1 、R ^4a2 And R ^4b Independently is H or C _1-4 An alkyl group;

R ^4c independently H, C _1-4 Alkyl or-O-C _1-4 An alkyl group;

R ^5b 、R ⁶ and R ⁷ Independently of each other H, C _1-4 Alkyl or C _1-4 A haloalkyl group;

"" indicates that when the carbon atom with "-" is a chiral carbon atom, it is in R configuration, S configuration, or a mixture thereof.

2. A compound of formula (I), a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, as claimed in claim 1, wherein:

the compound shown in the formula (I) is selected from the following compounds:

and/or when R ¹ 、R ² 、R ³ And R ⁴ When independently halogen, said halogen is fluorine, chlorine, bromine or iodine;

and/or when R ¹ 、R ² And R ³ Independently is R ⁵ Substituted or unsubstituted C _1-4 When alkyl, said C _1-4 Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl;

and/or when R ¹ 、R ² And R ³ Independently is R ⁵ Substituted or unsubstituted C _3-6 In the case of a cycloalkyl group of (A), said C _3-6 The cycloalkyl group of (a) is cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl;

and/or when R ¹ 、R ² And R ³ Independently is R ⁵ Substituted or unsubstituted C _1-4 When halogenated alkyl, the halogen is fluorine, chlorine, bromine or iodine;

and/or when R ¹ 、R ² And R ³ Independently is R ⁵ Substituted or unsubstituted C _1-4 When halogenated alkyl, said C _1-4 Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl;

and/or when R ¹ 、R ² And R ³ Independently is R ⁵ Substituted or unsubstituted C _1-4 When halogenated alkyl is adopted, the number of the halogenated groups is 1,2,3,4 or 5;

and/or when R ⁴ 、R ⁵ 、R ^5b 、R ⁶ And R ⁷ Independently is C _1-4 When halogenated alkyl, the halogen is fluorine, chlorine, bromine or iodine;

and/or when R ⁴ 、R ⁵ 、R ^5b 、R ⁶ And R ⁷ Independently is C _1-4 When halogenated with alkyl, C therein _1-4 Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl;

and/or when R ⁴ 、R ⁵ 、R ^5b 、R ⁶ And R ⁷ Independently is C _1-4 When halogenated alkyl is adopted, the number of the halogenated groups is 1,2,3,4 or 5;

and/or when R ^4a1 、R ^4a2 、R ^4b 、R ^5b 、R ⁶ And R ⁷ Independently is C _1-4 When alkyl, said C _1-4 Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl;

and/or when R ^4c Independently is C _1-4 Alkyl or-O-C _1-4 When alkyl, said C _1-4 Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl;

and/or when R ⁵ Independently is C _1-4 Alkyl radical, C _1-4 alkyl-O-, C _1-4 When halogenated alkyl, said C _1-4 Alkyl radical, C _1-4 alkyl-O-and C _1-4 C in haloalkyl _1-4 Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl;

and/or when R ⁵ When independently an amino group protected by an N-protecting group, said protecting group is

R ^5a Is C _1-4 Alkyl or C _1-4 An alkyl group-O-.

3. A compound of formula (I), a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, as claimed in claim 1 or 2, wherein:

when R is ⁵ Independently an amino group protected by an N protecting group, said protecting group being

R ^5a Is C _1-4 Alkyl or C _1-4 alkyl-O-or C _1-4 Alkyl and C _1-4 C in alkyl-O-radicals _1-4 Alkyl is independently methyl, ethyl, n-propyl, isopropyl, n-propylButyl, isobutyl or tert-butyl;

and/or, X is N;

and/or, R ¹ Selected from methyl, cyano, trifluoromethyl, difluoromethyl,

And/or, R ² Is H;

and/or, R ³ Selected from H, methyl, halogen or cyano;

and/or, R ⁴ Selected from cyano, -C (= O) NR ^4a1 R ^4a2 、-S(＝O) ₂ R ^4b or-C (= O) R ^4c ；

And/or, R ^4a1 、R ^4a2 、R ^4b 、R ^4c 、R ^5b 、R ⁶ And R ⁷ Independently a methyl group.

4. A compound of formula (I), a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, as claimed in claim 1, wherein: wherein the compound shown in the formula (I) is selected from the following compounds:

in the above compounds, R ¹ Is defined as in claim 1;

and/or the presence of a gas in the gas,

is composed of

Or a mixture thereof.

5. A compound of formula (I), a pharmaceutically acceptable salt thereof, or a stereoisomer thereof according to any one of claims 1 to 4, wherein: the compound shown in the formula (I) is selected from the following compounds:

6. a process for the preparation of a compound of formula (I) as claimed in any one of claims 1 to 5, characterized in that: which comprises the following steps:

wherein W represents halogen, such as Br; r ^x is-B (OH) ₂ Or

A、R ¹ 、R ³ 、R ⁴ X is as defined in any one of claims 1 to 5.

7. Intermediate compound B ₀ The preparation method is characterized by comprising the following steps: it is any one of the following schemes:

the method comprises the following steps:

in the step (1), a compound SM-5 is subjected to ring closing under the action of a catalyst to obtain a compound SM-6,

reacting the compound SM-6 in the step (2) under the action of an oxidant to obtain a compound SM-7,

step (3) Compounds SM-7 and H ₂ N-CH ₂ Reaction of-A in the presence of a base to give compound B ₀ ；

Scheme two, which is shown as step (2) to step (3) in scheme one;

scheme III, which is shown as step (3) in scheme I;

the reaction equation is as follows:

wherein, W, A, R ⁴ Is as defined in any one of claims 1 to 5.

8. A pharmaceutical composition characterized by: which comprises a therapeutically effective amount of a compound as shown in formula (I) as claimed in any one of claims 1 to 5, a pharmaceutically acceptable salt or a stereoisomer thereof and pharmaceutically acceptable auxiliary materials.

9. The application of a substance A in preparing a medicament is characterized in that: the substance A is a compound shown in the formula (I) as described in any one of claims 1 to 5, a pharmaceutically acceptable salt thereof or a stereoisomer thereof, or a pharmaceutical composition as described in claim 8; the medicament is preferably a medicament for treating cancer; more preferably, the drug is a drug for treating cancer mediated by the EED protein and/or PRC2 protein complex.

10. The use of claim 9, wherein:

the cancer is selected from the group consisting of diffuse large B-cell lymphoma, lymphoid cancer such as follicular lymphoma, leukemia, multiple myeloma, mesothelioma, gastric cancer, malignant rhabdoid tumor, liver cancer, prostate cancer, breast cancer, brain tumors including neuroblastoma, glioma, glioblastoma and astrocytoma, cervical cancer, colon cancer, melanoma, endometrial cancer, esophageal cancer, head and neck cancer, lung cancer, nasopharyngeal cancer, ovarian cancer, pancreatic cancer, renal cancer, rectal cancer, thyroid cancer, parathyroid tumor, uterine tumor and soft tissue sarcoma; preferably, lymphoma such as diffuse large B cell lymphoma and follicular lymphoma;

and/or the substance A is used in combination with other medicines; preferably, the other drug is selected from the group consisting of anticancer drugs, tumor immunity drugs, antiallergic drugs, antiemetics, analgesics, and cytoprotective drugs.

11. A pharmaceutical formulation characterized by: comprising a compound of formula (I), a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to any one of claims 1 to 5 or a pharmaceutical composition according to claim 8, preferably in the form of tablets, capsules, pills, powders, granules, elixirs, tinctures, suspensions, syrups, emulsions and solutions; more preferably, the pharmaceutical formulation is administered by a mode selected from the group consisting of oral, sublingual, subcutaneous, intravenous, intramuscular, intrasternal, nasal, topical and rectal;

and/or, the pharmaceutical formulation is administered once or more times per day.