CN112424203A

CN112424203A - Bruton's tyrosine kinase inhibitors

Info

Publication number: CN112424203A
Application number: CN201980047159.XA
Authority: CN
Inventors: 张寅生; 任景; 吴喆阳; 王庆璘; 金超; 徐胜; 陆鹏; 施伟; 贺香依; 田晓萌; 汪杰
Original assignee: Chia Tai Tianqing Pharmaceutical Group Co Ltd
Current assignee: Chia Tai Tianqing Pharmaceutical Group Co Ltd
Priority date: 2018-07-20
Filing date: 2019-07-19
Publication date: 2021-02-26
Anticipated expiration: 2039-07-19
Also published as: CN112424203B; WO2020015735A1

Abstract

Discloses a Bruton's tyrosine kinase inhibitor, in particular to a compound shown as a formula (I) or a pharmaceutically acceptable salt thereof, a preparation method thereof, a pharmaceutical composition containing the compound and application thereof in treating Bruton's tyrosine kinase related diseases.

Description

Bruton's tyrosine kinase inhibitors

Cross Reference to Related Applications

This application claims priority and benefit to the chinese patent application No. 201810802823.4 filed on 2018, month 7 and day 20 with the intellectual property office of china, the disclosure of which is incorporated herein by reference in its entirety.

Technical Field

The present application relates to compounds containing an imidazopyrazine ring as BTK inhibitors, processes for their preparation, pharmaceutical compositions containing them and their use in the treatment of BTK related diseases.

Background

Bruton's Tyrosine Kinase (BTK) is mainly expressed in B cells, distributed in the lymphatic, hematopoietic and blood systems, first discovered in 1993, is a member of the non-receptor tyrosine kinase Tec family, which also includes TEC, ITK/TSK/EMT and BMX, which have high structural homology. BTK plays a crucial role in B cell signaling pathways that link cell surface B cell receptor (B-cell receptor) stimulation to downstream intracellular responses, and is a key regulator of B cell development, activation, signaling, and survival. In recent years, researches on B cells, particularly B cell non-Hodgkin lymphoma and rheumatoid arthritis show that BTK is often abnormally expressed.

The BTK signal transduction pathway is used for developing small molecule targeted drugs, and a brand new way is provided for treating B cell tumors such as leukemia and multiple myeloma and B cell immune diseases. Ibrutinib is the first approved BTK inhibitor to be marketed for the treatment of cll (chloroonly leukemia) and mcl (mantle cell lymphoma); in addition, acaraburtinib was approved for the treatment of MCL in 2017.

Currently, the BTK binding site of the currently marketed irreversible inhibitor, such as ibrutinib, often undergoes C481S mutation, resulting in decreased pharmaceutical activity and thus drug resistance, so that more BTK inhibitors are clinically needed, the problem of drug resistance caused by the mutation is overcome, and the irreversible inhibitor has a better inhibiting effect on both wild-type and mutant BTKs; meanwhile, the BTK has high selectivity, so that toxic and side effects caused by off-target effects are avoided.

Summary of The Invention

In one aspect, the present application provides a compound of formula (I) or a pharmaceutically acceptable salt thereof,

wherein the content of the first and second substances,

ring B is selected from 5-10 membered heteroaryl or C_6-10An aryl group;

R ¹independently selected from halogen, -OH, -NH₂、-CN、C _1-6Alkyl radical, C_1-6Alkoxy or halogen substituted C_1-6An alkyl group;

p is selected from 0, 1,2,3 or 4;

l is selected from-C (O) NH-, -CH₂NHC(O)-、-NHC(O)-、-O-、-NH-、-S-、-C(O)O-、-OC(O)-、-S(O) ₂O-or-OS (O)₂-；

R ²Independently selected from halogen, -OH, -NH₂、-CN、C _1-6Alkyl or C_1-6Alkoxy radical, said C_1-6Alkyl or C_1-6Alkoxy is optionally substituted with halogen;

m is selected from 0, 1,2,3 or 4;

is selected from

Wherein the content of the first and second substances,

X ¹independently selected from-NH-, -O-, -S-or-CH₂-；

n1 is selected from 1,2,3,4, 5 or 6;

n2 and n3 are each independently selected from 1,2 or 3;

n4 is selected from 0, 1,2 or 3;

R ³and R⁵Each independently selected from ═ O, halogen, -OH, -NH₂、-CN、C _1-6Alkyl radical, C_1-6Alkoxy radical, C_1-6Alkylamino radical, C_1-6Alkyl S-, -C (O) OR, -C (O) NHR OR-S (O)₂R, wherein R is independently selected from-H, C_1-6Alkyl radical, C_2-6Alkenyl radical, C_2-6Alkynyl, C_3-6Cycloalkyl, 3-6 membered heterocycloalkyl, 5-6 membered heteroaryl or C_6-10An aryl group;

q1, q2, q3 and q4 are each independently selected from 0, 1,2 or 3;

X ²is selected from N or CH;

X ³is selected from-CH₂-, -O-, -S-or-NH-.

In another aspect, the present application provides a pharmaceutical composition comprising a compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof.

In another aspect, the present application provides a method of treating a BTK-related disease in a mammal, comprising administering to a mammal, preferably a human, in need of such treatment a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof, as described herein, or a pharmaceutical composition as described herein.

In a further aspect, the present application provides the use of a compound of formula (I) or a pharmaceutically acceptable salt thereof, as described herein, or a pharmaceutical composition as described herein, in the manufacture of a medicament for the prevention or treatment of a BTK-related disease.

In yet another aspect, the present application provides a compound of formula (I) or a pharmaceutically acceptable salt thereof, as described herein, or a pharmaceutical composition as described herein, for use in the prevention or treatment of a BTK-related disease.

In yet another aspect, the present application provides a compound of formula (I) or a pharmaceutically acceptable salt thereof, as described herein, or a pharmaceutical composition as described herein, for the prevention or treatment of a BTK-related disease.

Detailed Description

The present application relates to compounds of formula (I) or a pharmaceutically acceptable salt thereof,

wherein the content of the first and second substances,

ring B is selected from 5-10 membered heteroaryl or C_6-10An aryl group;

R ¹independently selected from halogen, -OH, -NH₂、-CN、C _1-6Alkyl radical, C_1-6Alkoxy radicalRadicals or halogen substituted C_1-6An alkyl group;

p is selected from 0, 1,2,3 or 4;

l is selected from-C (O) NH-, -CH₂NHC(O)-、-NHC(O)-、-O-、-NH-、-S-、-C(O)O-、-OC(O)-、-S(O) ₂O-or-OS (O)₂-; or L is selected from the group consisting of-C (O) NH-, -NHC (O) -, -O-, -NH-, -S-, -C (O) O-, -OC (O) -, -S (O)₂O-or-OS (O)₂-；

m is selected from 0, 1,2,3 or 4;

is selected from

Wherein the content of the first and second substances,

X ¹independently selected from-NH-, -O-, -S-or-CH₂-；

n1 is selected from 1,2,3,4, 5 or 6;

n2 and n3 are each independently selected from 1,2 or 3;

n4 is selected from 0, 1,2 or 3;

q1, q2, q3 and q4 are each independently selected from 0, 1,2 or 3;

X ²is selected from N or CH;

X ³is selected from-CH₂-, -O-, -S-or-NH-.

In some embodiments, ring B is selected from 5-10 membered heteroaryl or phenyl; in some embodiments, ring B is selected from 5-6 membered heteroaryl or phenyl; in some embodiments, ring B is selected from pyridyl (e.g., pyridin-2-yl) or phenyl.

In some embodiments, R¹Independently selected from halogen, -CN, C_1-6Alkoxy, or C optionally substituted by halogen_1-6An alkyl group; in some embodiments, R¹Independently selected from halogen, -CN, C_1-3Alkoxy, or C optionally substituted by halogen_1-3An alkyl group; in some embodiments, R¹Independently selected from chloro, -CN, methoxy, methyl or-CF₃。

In some embodiments, R¹Independently selected from-F, -Cl, -Br, -I, -CN, C_1-3Alkoxy, or C optionally substituted by halogen_1-3An alkyl group; in some embodiments, R¹Independently selected from-F, -Cl, -CN, methoxy, methyl or-CF₃。

In some embodiments, R¹Independently selected from C optionally substituted by halogen_1-6An alkyl group; in some embodiments, R¹Independently selected from halogen substituted C_1-3An alkyl group; in some embodiments, R¹Is selected from-CF₃。

In some embodiments, p is selected from 0, 1 or 2; in some embodiments, p is selected from 0or 1.

In some embodiments, R¹Independently selected from halogen, -CN, C_1-6Alkoxy, or C optionally substituted by halogen_1-6Alkyl, and p is selected from0.1 or 2; in some embodiments, R¹is-F, -Cl, -CN, methoxy, methyl or-CF₃And p is 1.

In some embodiments, L is selected from the group consisting of-C (O) NH-, -NHC (O) -, -O-, -NH-, -S-, -C (O) O-, -OC (O) -, -S (O)₂O-or-OS (O)₂-。

In some embodiments, L is selected from the group consisting of-C (O) NH-, -CH₂NHC (O) -, -NHC (O) -or-O-; in some embodiments, L is selected from the group consisting of-C (O) NH-, -CH₂NHC (O) -, or-O-.

In some embodiments, L is selected from-C (O) NH-, -NHC (O) -or-O-; in some embodiments, L is selected from-C (O) NH-or-O-.

In some embodiments, L is-C (O) NH-. In some embodiments, L is-CH₂NHC (O) -. In some embodiments, L is-O-.

In some embodiments, R is relative to the attachment position of L to ring B¹At the ortho, meta or para position. In some embodiments, ring B is pyridyl and R is pyridinyl, relative to the position of attachment of L to ring B¹In the meta or para position. In some embodiments, ring B is phenyl and R is R relative to the position of attachment of L to ring B¹In ortho or meta position.

In some embodiments, R²Independently selected from halogen, -OH, -NH₂、-CN、C _1-3Alkyl or C_1-3Alkoxy radical, said C_1-3Alkyl or C_1-3Alkoxy is optionally substituted with halogen; in some embodiments, R²Independently selected from-F, -Cl, -Br, -I, -OH, -NH₂、-CN、-CH ₃or-CF₃(ii) a In some embodiments, R²Independently selected from-F, -Cl, -Br or-I; in some embodiments, R²Independently selected from-F or-Cl.

In some embodiments, m is selected from 0, 1 or 2; in some embodiments, m is selected from 0or 1.

In some embodiments, R²is-F or-Cl, and m is 1. In some embodiments, m is 0.

In some embodiments, X¹Independently selected from-NH-, -O-or-CH₂-。

In some embodiments, X¹Independently selected from-NH-, -O-or-CH₂-, wherein-NH-and-CH₂H in (A) is optionally substituted by R³And (4) substitution.

In some embodiments, X¹Independently selected from-NH-, -O-or-CH₂-, wherein the H atom in-NH-is optionally substituted by C_1-3Alkyl substitution and/or-CH₂Two H atoms in (a) are optionally substituted by ═ O.

In some embodiments, X¹Independently selected from-NH-, -O-or-CH₂-, wherein the H atom in-NH-is optionally substituted by methyl and/or-CH₂Two H atoms in (a) are optionally substituted by ═ O.

In some embodiments, n1 is selected from 1,2,3, or 4; in some embodiments, n1 is selected from 2,3, or 4.

In some embodiments, when n1 is 2,3,4, 5 or 6, one of X is¹Selected from-NH-, -O-, -S-or-CH₂-, wherein the H atom in-NH-is optionally substituted by C_1-3Alkyl substituted, the remainder of X¹is-CH₂-。

In some embodiments, when n1 is 2,3, or 4, one of X is¹Selected from-NH-, -O-or-CH₂-, wherein the H atom in-NH-is optionally substituted by C_1-3Alkyl substituted, the remainder of X¹is-CH₂-。

In some embodiments, when n1 is 2,3, or 4, one of X is¹Selected from-NH-, -O-, -CH₂-, in which the H atom in-NH-is optionally substituted by methyl, the remainder of X¹is-CH₂-。

In some embodiments, n2 is selected from 1 or 2 and n3 is selected from 2 or 3. In some embodiments, both n2 and n3 are 2. In some embodiments, n2 is 1 and n3 is 2 or 3.

In some embodiments, n4 is selected from 0, 1 or 2; in some embodiments, n4 is selected from 0or 1; in some embodiments, n4 is selected from 1.

In some embodiments, R³Independently selected from ═ O, halogen, -OH, -NH₂、-CN、C _1-3Alkyl radical, C_1-3Alkoxy radical, C_1-3Alkylamino radical, C_1-3Alkyl S-, -C (O) OR, -C (O) NHR OR-S (O)₂R; in some embodiments, R³Selected from ═ O, halogen, -OH, -NH₂-CN or C_1-3An alkyl group; in some embodiments, R³Is selected from ═ O or C_1-3An alkyl group; in some embodiments, R³Selected from ═ O or methyl.

In some embodiments, R³Independently selected from ═ O, -F, -Cl, -Br, -I, -OH, -NH₂or-CN; in some embodiments, R³Selected from ═ O.

In some embodiments, R is independently selected from-H, C_1-3Alkyl radical, C_2-3Alkenyl radical, C_2-3Alkynyl, C_3-6Cycloalkyl, 3-6 membered heterocycloalkyl, 5-6 membered heteroaryl or phenyl; in some embodiments, R is independently selected from-H, C_1-3Alkyl radical, C_2-3Alkenyl or C_2-3Alkynyl.

In some embodiments, q1 is selected from 1 or 2, q3 is selected from 1; in some embodiments, q1 is selected from 2 and q3 is selected from 1.

In some embodiments, q2 is selected from 1 or 2, q4 is selected from 1; in some embodiments, q2 is selected from 2 and q4 is selected from 1.

In some embodiments, X²Is selected from N.

In some embodiments, X³Selected from-O-, -S-or-NH-; at one endIn some embodiments, X³Selected from-O-, -S-or-NH-, wherein the H atom in-NH-is optionally substituted by C_1-3Alkyl substitution; in some embodiments, X³Is selected from-NH-.

In some embodiments of the present invention, the substrate is,

selected from the following structures:

in some embodiments, one X of the five structures described above¹Selected from-NH-, -O-, -S-, -CH₂Or by C_1-3Alkyl substituted-NH-and the remainder X¹is-CH₂-。

In some embodiments, one X of the five structures described above¹Selected from-NH-, -O-, -CH₂-or-NH-substituted by methyl, the remainder X¹is-CH₂-。

In some embodiments, the five structures are X adjacent to or spaced from the point of spiro union¹is-CH substituted by ═ O₂- (i.e., -C (O)).

In some embodiments, the X in the five structures is adjacent to the screw joint¹is-CH substituted by ═ O₂- (i.e., -C (O)), X at a position spaced from the point of screwing¹is-NH-, -O-, -CH₂-or-NH-substituted by methyl, the remainder X¹is-CH₂-。

In some embodiments, the five structures have an X at a position spaced from the point of screwing¹is-NH-or substituted by methylof-NH-, with the remainder of X¹is-CH₂-。

In some embodiments, R⁵Selected from ═ O, halogen, -OH, -NH₂-CN or C_1-3An alkyl group; in some embodiments, R⁵Is selected from ═ O or C_1-3An alkyl group; in some embodiments, R⁵Selected from ═ O.

In some embodiments of the present invention, the substrate is,

is selected from

In some embodiments, X in the above structures²Is N, X³Is NH, and/or R⁵Is ═ O.

In some embodiments of the present invention, the substrate is,

is composed of

In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt thereof of the present application is selected from a compound of formula (II-1) or (II-2) or a pharmaceutically acceptable salt thereof,

wherein the content of the first and second substances,

R ¹、p、R ²and m is as defined above.

In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt thereof of the present application is a compound of formula (II-3) or a pharmaceutically acceptable salt thereof,

wherein the content of the first and second substances,

R ¹、p、R ²and m is as defined above.

In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt thereof of the present application is selected from the following compounds or pharmaceutically acceptable salts thereof:

in another aspect, the present application relates to a pharmaceutical composition comprising a compound of formula (I), formula (II-1), formula (II-2), formula (II-3), or a specific compound described above, or a pharmaceutically acceptable salt thereof, of the present application. In some embodiments, the pharmaceutical compositions of the present application further comprise a pharmaceutically acceptable excipient.

In another aspect, the present application relates to a method of treating a BTK-associated disease in a mammal, comprising administering to a mammal, preferably a human, in need of such treatment a therapeutically effective amount of a compound of formula (I), formula (II-1), formula (II-2), formula (II-3), or a specific compound thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof.

In another aspect, the application relates to the use of a compound of formula (I), formula (II-1), formula (II-2), formula (II-3), or a specific compound described above, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof, in the manufacture of a medicament for the prevention or treatment of a BTK-associated disease.

In another aspect, the present application relates to the use of a compound of formula (I), formula (II-1), formula (II-2), formula (II-3), or a specific compound described above, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof, in the prevention or treatment of a BTK-associated disease.

In another aspect, the present application relates to a compound of formula (I), formula (II-1), formula (II-2), formula (II-3), or a specific compound thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof, for preventing or treating a BTK-associated disease.

In some embodiments, the BTK-related disease is a BTK-mediated disease. In some embodiments, the BTK-related disease is selected from an autoimmune disease, an inflammatory disease, or a cancer, wherein the cancer includes, but is not limited to, diffuse large B-cell lymphoma.

Definition of

The following terms used in the present application have the following meanings, unless otherwise specified. A particular term should not be considered as ambiguous or unclear without special definition, but rather construed according to ordinary meaning in the art. When a trade name appears herein, it is intended to refer to its corresponding commodity or its active ingredient.

The term "substituted" means that any one or more hydrogen atoms on a specified group is replaced with a substituent, so long as the valence of the specified group is normal and the substituted compound is stable. When the substituent is oxo (i.e., ═ O), meaning that two hydrogen atoms are substituted, oxo does not occur on the aryl.

The terms "optionally" or "optionally" mean that the subsequently described event or circumstance may or may not occur, and that the description includes instances where said event or circumstance occurs and instances where it does not. For example, ethyl is "optionally" substituted with halo, meaning that ethyl may be unsubstituted (CH)₂CH ₃) Monosubstituted (e.g. CH)₂CH ₂F) Polysubstituted (e.g. CHFCH)₂F、CH ₂CHF ₂Etc.) or completely substituted (CF)₂CF ₃). It will be appreciated by those skilled in the art that any group containing one or more substituents will not incorporate any substitution or substitution pattern which is not sterically impossible and/or cannot be synthesized.

Herein C_m-nIt is the moiety that has an integer number of carbon atoms in the given range. E.g. "C_1-6By "is meant that the group can have 1 carbon atom, 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, or 6 carbon atoms.

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. For example, if a group contains 2R, then each R has a separate option.

When the number of one linking group is 0, e.g. - (CH)₂) ₀-, indicates that the linking group is a covalent bond.

When one of the variables is selected from a covalent bond, it means that the two groups to which it is attached are directly linked, for example, where L represents a covalent bond in A-L-Z, it means that the structure is actually A-Z.

For the

When L is selected from-CO (NH) -, it represents

Is composed of

When a bond of a substituent is cross-linked to a ring, such substituent may be bonded to any atom on the ring. For example,

when X is present¹NH, wherein n1, n2, n3 and n4 are 1

The term "halo" or "halogen" refers to fluorine, chlorine, bromine and iodine.

The term "hydroxy" refers to an-OH group.

The term "amino" refers to the group-NH₂A group.

The term "alkyl" refers to a group of formula C_nH _2n+1Of hydrocarbon radicals, e.g. C_1-6Alkyl radical, C_1-3An alkyl group. The alkyl group may be linear or branched. For example, the term "C_1-6Alkyl "means an alkyl group having 1 to 6 carbon atoms (e.g., methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl, neopentyl, hexyl, 2-methylpentyl, and the like). Similarly, the alkyl portion (i.e., alkyl) of alkoxy, alkylamino, dialkylamino, alkylsulfonyl and alkylthio groups have the same definitions as above.

The term "alkoxy" refers to-O-alkyl, for example-O-C _1-6Alkyl, -O-C_1-3An alkyl group.

The term "alkylamino" refers to-NH-alkyl, e.g., -NH-C_1-6Alkyl, -NH-C_1-3An alkyl group.

The term "alkenyl" refers to a straight or branched chain unsaturated aliphatic hydrocarbon group having at least one double bond consisting of carbon and hydrogen atoms, e.g., C_2-6Alkenyl radical, C_2-3An alkenyl group. Non-limiting examples of alkenyl groups include, but are not limited to, ethenyl, 1-propenyl, 2-propenyl, 1-butenyl, isobutenyl, 1, 3-butadienyl, and the like.

The term "alkynyl" refers to a straight or branched chain unsaturated aliphatic hydrocarbon group having at least one triple bond consisting of carbon and hydrogen atoms, e.g., C_2-6Alkynyl, C_2-3Alkynyl. Non-limiting examples of alkynyl groups include, but are not limited to, ethynyl (-C ≡ CH), 1-propynyl (-C ≡ C-CH)₃) 2-propynyl (-CH)₂-C.ident.CH), 1, 3-butadiynyl (-C.ident.C-C.ident.CH), and the like.

The term "cycloalkyl" refers to a carbon ring that is fully saturated and may exist as a single ring, a bridged ring, or a spiro ring. Unless otherwise indicated, the carbocycle is typically a 3 to 10 membered ring, and may also be a 3 to 6 membered ring. Non-limiting examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, norbornyl (bicyclo [2.2.1] heptyl), bicyclo [2.2.2] octyl, adamantyl, and the like.

The term "heterocycloalkyl" refers to a cyclic group that is fully saturated and may exist as a single ring, a bridged ring, or a spiro ring. Unless otherwise indicated, the heterocyclic ring is typically a 3 to 7 membered ring containing 1 to 3 heteroatoms (preferably 1 or 2 heteroatoms) independently selected from sulfur, oxygen and/or nitrogen. The heterocycloalkyl group can be a 3 to 6 membered ring containing 1 or 2 heteroatoms independently selected from oxygen and nitrogen. Examples of 3-membered heterocycloalkyl include, but are not limited to, oxiranyl, thietanyl, cycloazenyl, non-limiting examples of 4-membered heterocycloalkyl include, but are not limited to, azetidinyl, oxetanyl, thiabutinyl, examples of 5-membered heterocycloalkyl include, but are not limited to, tetrahydrofuranyl, tetrahydrothienyl, pyrrolidinyl, isoxazolidinyl, oxazolidinyl, isothiazolidinyl, thiazolidinyl, imidazolidinyl, examples of tetrahydropyrazolyl, 6-membered heterocycloalkyl include, but are not limited to, piperidinyl, tetrahydropyranyl, tetrahydrothiopyranyl, morpholinyl, piperazinyl, 1, 4-thialkyl, 1, 4-dioxanyl, thiomorpholinyl, 1, 3-dithianyl, 1, 4-dithianyl, and examples of 7-membered heterocycloalkyl include, but are not limited to, azepanyl, oxepanyl, thiepanyl. Monocyclic heterocycloalkyl groups having 5 or 6 ring atoms are preferred.

The term "aryl" refers to an all-carbon monocyclic or fused polycyclic aromatic ring group having a conjugated pi-electron system. For example, the aryl group can have 6 to 20 carbon atoms, 6 to 14 carbon atoms, or 6 to 12 carbon atoms. Non-limiting examples of aryl groups include, but are not limited to, phenyl, naphthyl, anthracenyl, and 1,2,3, 4-tetrahydronaphthalene, and the like.

The term "heteroaryl" refers to a monocyclic or fused polycyclic ring system containing at least one ring atom selected from N, O, S, the remaining ring atoms being C, and having at least one aromatic ring. For example, heteroaryl groups may contain 1,2 or 3 heteroatoms selected from N, O, S. Preferred heteroaryl groups have a single 4 to 8 membered ring, especially a 5 to 8 membered ring, or a fused polycyclic ring containing 6 to 14, especially 6 to 10 ring atoms. Non-limiting examples of heteroaryl groups include, but are not limited to, pyrrolyl, furanyl, thienyl, imidazolyl, oxazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyrazinyl, quinolinyl, isoquinolinyl, tetrazolyl, triazolyl, triazinyl, benzofuranyl, benzothienyl, indolyl, isoindolyl, and the like.

The term "treating" means administering a compound or formulation described herein to ameliorate or eliminate a disease or one or more symptoms associated with the disease, and includes:

(i) inhibiting the disease or disease state, i.e., arresting its development;

(ii) alleviating the disease or condition, i.e., causing regression of the disease or condition.

The term "preventing" means administering a compound or formulation described herein to prevent a disease or one or more symptoms associated with the disease, and includes: prevention of a disease or condition occurs in a mammal, particularly when such mammal is susceptible to the disease condition, but has not yet been diagnosed as having the disease condition.

The term "therapeutically effective amount" means an amount of a compound of the present application that (i) treats or prevents a particular disease, condition, or disorder, (ii) alleviates, ameliorates, or eliminates one or more symptoms of a particular disease, condition, or disorder, or (iii) prevents or delays the onset of one or more symptoms of a particular disease, condition, or disorder described herein. The amount of a compound of the present application that constitutes a "therapeutically effective amount" depends on the compound, the disease state and its severity, the mode of administration, and the age of the mammal to be treated, but can be routinely determined by those skilled in the art with their own knowledge and this disclosure.

The term "pharmaceutically acceptable" is intended to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.

As the pharmaceutically acceptable salt, for example, a metal salt, an ammonium salt, a salt with an organic base, a salt with an inorganic acid, a salt with an organic acid, a salt with a basic or acidic amino acid, or the like can be included.

The term "pharmaceutical composition" refers to a mixture of one or more compounds of the present application or salts thereof and pharmaceutically acceptable excipients. The purpose of the pharmaceutical composition is to facilitate administration of the compounds of the present application to an organism.

The term "pharmaceutically acceptable adjuvants" refers to those adjuvants which do not have a significant irritating effect on the organism and do not impair the biological activity and properties of the active compound. Suitable adjuvants are well known to those skilled in the art, such as carbohydrates, waxes, water-soluble and/or water-swellable polymers, hydrophilic or hydrophobic materials, gelatin, oils, solvents, water, and the like.

The words "comprise" or "comprise" and variations thereof such as "comprises" or "comprising," are to be understood in an open, non-exclusive sense, i.e., "including but not limited to.

The compounds and intermediates of the present application may also exist in different tautomeric forms, and all such forms are included within the scope of the present application. The term "tautomer" or "tautomeric form" refers to structural isomers of different energies that can interconvert via a low energy barrier. For example, proton tautomers (also referred to as proton transfer tautomers) include interconversion via proton migration, such as keto-enol and imine-enamine isomerizations. A specific example of a proton tautomer is an imidazole moiety, wherein the proton can migrate between two ring nitrogens. Valence tautomers include interconversion by recombination of some of the bonding electrons.

The present application also includes isotopically-labeled compounds of the present application, which are identical to those recited herein, but for the fact that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature. Examples of isotopes that can be incorporated into compounds of the present application include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, iodine and chlorine, such as respectively²H、 ³H、 ¹¹C、 ¹³C、 ¹⁴C、 ¹³N、 ¹⁵N、 ¹⁵O、 ¹⁷O、 ¹⁸O、 ³¹P、 ³²P、 ³⁵S、 ¹⁸F、 ¹²³I、 ¹²⁵I and³⁶cl, and the like.

Certain isotopically-labelled compounds of the present application (e.g. with³H and¹⁴c-labeled ones) can be used in compound and/or substrate tissue distribution assays. Tritiated (i.e. by tritiation)³H) And carbon-14 (i.e.¹⁴C) Isotopes are particularly preferred for their ease of preparation and detectability. Positron emitting isotopes, such as¹⁵O、 ¹³N、 ¹¹C and ¹⁸f can be used in Positron Emission Tomography (PET) studies to determine substrate occupancy. Isotopically labeled compounds of the present application can generally be prepared by following procedures analogous to those disclosed in the schemes and/or in the examples below, by substituting an isotopically labeled reagent for a non-isotopically labeled reagent.

In addition, heavier isotopes are used (such as deuterium (i.e., deuterium)²H) Substitution may provide certain therapeutic advantages resulting from greater metabolic stability (e.g., increased in vivo half-life or reduced dosage requirements), and thus may be preferred in certain circumstances, wherein deuterium substitution may be partial or complete, with partial deuterium substitution meaning that at least one hydrogen is substituted with at least one deuterium.

The compounds of the present application may be asymmetric, e.g., having one or more stereoisomers. Unless otherwise indicated, all stereoisomers include, for example, enantiomers and diastereomers. The compounds of the present application containing asymmetric carbon atoms can be isolated in optically active pure form or in racemic form. The optically active pure form can be resolved from a racemic mixture or synthesized by using chiral starting materials or chiral reagents.

The pharmaceutical compositions of the present application can be prepared by combining the compounds of the present application with suitable pharmaceutically acceptable excipients, for example, can be formulated into solid, semi-solid, liquid or gaseous formulations, such as tablets, pills, capsules, powders, granules, ointments, emulsions, suspensions, suppositories, injections, inhalants, gels, microspheres, aerosols, and the like.

Typical routes of administration of a compound of the present application or a pharmaceutically acceptable salt thereof or a pharmaceutical composition thereof include, but are not limited to, oral, rectal, topical, inhalation, parenteral, sublingual, intravaginal, intranasal, intraocular, intraperitoneal, intramuscular, subcutaneous, intravenous administration.

The pharmaceutical compositions of the present application can be manufactured by methods well known in the art, such as conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, lyophilizing, and the like.

In some embodiments, the pharmaceutical composition is in an oral form. For oral administration, the pharmaceutical compositions may be formulated by mixing the active compounds with pharmaceutically acceptable excipients well known in the art. These adjuvants enable the compounds of the present application to be formulated as tablets, pills, lozenges, dragees, capsules, liquids, gels, slurries, suspensions and the like, for oral administration to a patient.

Solid oral compositions may be prepared by conventional mixing, filling or tableting methods. For example, it can be obtained by the following method: the active compounds are mixed with solid adjuvants, optionally the mixture obtained is milled, if desired with further suitable adjuvants, and the mixture is then processed to granules, to give tablets or dragee cores. Suitable excipients include, but are not limited to: binders, diluents, disintegrants, lubricants, glidants, sweeteners or flavoring agents, and the like.

The pharmaceutical compositions may also be adapted for parenteral administration, as sterile solutions, suspensions or lyophilized products in suitable unit dosage forms.

In all methods of administration of the compounds of the general formula I described herein, the daily dose is from 0.01 to 200mg/kg body weight, in single or divided doses.

The compounds of the present application may be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combinations thereof with other chemical synthetic methods, and equivalents thereof known to those skilled in the art, with preferred embodiments including, but not limited to, the examples of the present application.

The chemical reactions of the embodiments herein are carried out in a suitable solvent that is compatible with the chemical changes of the present application and the reagents and materials required therefor. In order to obtain the compounds of the present application, it is sometimes necessary for a person skilled in the art to modify or select the synthesis steps or reaction schemes based on the existing embodiments.

An important consideration in the art of synthetic route planning is the selection of suitable protecting Groups for reactive functional Groups (such as amino Groups in the present application), for example, reference may be made to Greene's Protective Groups in Organic Synthesis (4th Ed.) Hoboken, New Jersey: John Wiley & Sons, Inc. all references cited herein are incorporated herein in their entirety.

In some embodiments, the compounds of formula (I) herein may be prepared by one skilled in the art of organic synthesis using standard methods in the art by the following routes:

the following abbreviations are used in this application:

PE represents petroleum ether; EA represents ethyl acetate; DMSO represents dimethyl sulfoxide; DMF represents N, N-dimethylformamide; DCM represents dichloromethane; NBS represents N-bromosuccinimide; DIPEA stands for diisopropylethylamine; MeOH represents methanol; NMP stands for N-methylpyrrolidone; EDTA represents ethylene diamine tetraacetic acid; DTT represents dithiothreitol; EGTA stands for ethylene glycol bis (2-aminoethyl ether) tetraacetic acid; ATP stands for adenosine triphosphate.

For clarity, the invention is further illustrated by examples, which do not limit the scope of the application. All reagents used herein were commercially available and used without further purification.

Example 1

Step 1: n- ((3-chloropyrazin-2-yl) methyl) carboxamide

Compound 1-1(45g) and trimethyl orthoformate (318g,328ml) were added to the reaction flask in this order, and the mixture was heated at 110 ℃ and stirred overnight. The reaction mixture was concentrated, and the residue was purified by chromatography (PE: EA 67:33 to 20:80) to give intermediate 1-2(31.81 g).

¹H NMR(500MHz,DMSO-d ₆):δ8.67-8.64(d,J＝14.5Hz,1H),8.58(br,1H),8.45(s,1H),8.18(s,1H),4.58(s,2H).MS(ESI):m/z 194.0[M+Na] ⁺.

Step 2: 8-chloroimidazo [1,5-a ] pyrazines

To a reaction flask were added intermediates 1-2(11.8g), acetonitrile (80ml) and DMF (2ml), N in that order₂Protection, adding POCl after 10min under ice-water bath condition₃(42.1g), after the addition was completed, the temperature was returned to room temperature and the mixture was stirred for 1 hour. In an ice bath, ammonia water is slowly dropped to quench the reaction, DCM is used for extraction, saturated salt water is used for washing, anhydrous sodium sulfate is used for drying, filtration is carried out, and the filtrate is concentrated to obtain an intermediate 1-3(10.25 g).

¹H NMR(500MHz,DMSO-d ₆):δ8.70(s,1H),8.40-8.39(d,J＝4.5hz,1H),7.87(s,1H),7.43-7.42(d,J＝5hz,1H). ¹³C NMR(125MHz,DMSO-d ₆):δ144.10,132.74,126.95,124.87,124.01,117.04.MS(ESI):m/z 153.9[M+H] ⁺.

And step 3: 1, 3-dibromo-8-chloroimidazo [1,5-a ] pyrazine

Intermediate 1-3(10g) and DMF (60ml) were added to the reaction flask. NBS (25.3g) was added portionwise in ice bath, supplemented with DMF (20ml) in N₂The mixture was allowed to return to room temperature under protection and stirred overnight. The reaction mixture was successively charged with 500mL of water, 500mL of EA and about 200mL of a saturated sodium thiosulfate solution in an ice-water bath, the organic layer was separated, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to give intermediate 1 to 4(20.19 g).

¹H NMR(500MHz,DMSO-d ₆):δ8.18-8.16(m,1H),7.52-7.51(d,J＝5hz,1H). ¹³C NMR(125MHz,DMSO-d ₆):δ143.48,128.51,123.27,116.50.113.35,110.12.MS(ESI):m/z 310.1[M+H] ⁺.

And 4, step 4: 1, 3-dibromoimidazo [1,5-a ] pyrazin-8-amine

To a reaction flask, intermediates 1 to 4(20g), sec-butanol (150ml) and ammonia water (169g,187ml) were added in this order, and the reaction apparatus was closed and heated to 90 ℃ for 4 hours. After the reaction solution was cooled to room temperature, the reaction solution was concentrated to remove most of the sec-butanol. To the residue were added water and EA, the organic phase was separated, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to give intermediate 1 to 5(17.35 g).

¹H NMR(500MHz,DMSO-d ₆):δ7.43-7.42(d,J＝5Hz,1H),7.14-7.13(d,J＝5Hz,1H),6.81(br,2H). ¹³C NMR(125MHz,DMSO-d ₆):δ150.99,130.68,118.34,111.18,107.09,106.90.MS(ESI):m/z 291.1[M+H] ⁺.

And 5: 8- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -2, 8-diazaspiro [4.5] decan-1-one

Intermediate 1-5(0.459g) was added to a microwave tube and dissolved in NMP (8mL), and 2, 8-diazaspiro [4.5] -1-decanone hydrochloride (0.2g) and DIPEA (0.407g) were added thereto with stirring at room temperature for 1 minute, and then placed in a microwave reactor 100W at 150 ℃ for 5 hours. The reaction solution was concentrated and purified by silica gel column chromatography, DCM: MeOH (95:5) elution, combined eluates and concentrated to give intermediates 1-6(0.15 g).

¹H NMR(500MHz,DMSO)δ7.60(s,1H),7.29-7.28(d,J＝5Hz,1H),6.91-6.90(d,J＝4.5Hz,1H),3.22-3.19(m,2H),2.97-2.92(m,2H),2.03-2.00(m,2H),1.92-1.88(m,2H),1.48-1.45(m,2H),1.24-1.20(m,2H).MS(ESI):m/z 365.3[M+H] ⁺.

Step 6: 4- (8-amino-3- (1-carbonyl-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

Adding the intermediate 1-6(0.1g) and 3-fluoro-4- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -N- (4- (trifluoromethyl) pyridin-2-yl) benzamide (i.e. the compound 1-7, 0.09g) into a microwave tube, dissolving in dioxane (5mL), adding potassium carbonate (0.15g) and water (1mL), stirring for 1 minute, adding 1, 1' -bis (diphenylphosphino) ferrocene palladium (II) dichlorodichloromethane complex (0.05g), blowing nitrogen by a bubbling method, and heating to 80 ℃ under a microwave reactor of 100W for 20 minutes. After the reaction is finished, the reaction solution is dissolved in 50mL of water, DCM is used for extraction, the organic phase is dried by anhydrous sodium sulfate, filtration is carried out, the filtrate is concentrated, and the concentrate is purified by a silica gel column, wherein DCM: MeOH eluted (97:3) to give Compound 1(0.07 g).

¹H NMR(500MHz,DMSO)δ11.42(s,1H),8.72-8.71(d,J＝4Hz,1H),8.56(s,1H),8.03-8.00(m,2H),7.68-7.65(m,1H),7.59-7.58(d,J＝7Hz,2H),7.32-7.31(m,1H),7.02-7.02(d,J＝2Hz,1H),6.01(s,2H),3.41-3.38(m,2H),3.23-3.21(m,2H),3.01-2.96(m,2H),2.06-2.03(m,2H),1.99-1.94(m,2H),1.52-1.50(d,J＝12.5Hz,2H).MS(ESI):m/z 569.5[M+H] ⁺.

Example 2

Step 1: 8- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) hexahydro-2H-pyrazino [1,2-a ] pyrazin-1 (6H) -one

Reference example 1 reaction step 5, substituting hexahydro-2H-pyrazino [1,2-a ] pyrazin-1 (6H) -one dihydrochloride for 2, 8-diazaspiro [4.5] -1-decanone hydrochloride as a reaction starting material gave intermediate 2-6(121 mg).

¹H NMR(500MHz,DMSO-d ₆)δ7.79(s,1H),7.28(d,J＝5.0Hz,1H),6.93(d,J＝5.0Hz,1H),6.60(brs,2H),3.56(d,J＝12.0Hz,1H),3.29–3.23(m,1H),3.10(dt,J＝11.9,3.9Hz,1H),3.01(td,J＝11.5,2.9Hz,1H),2.95–2.88(m,4H),2.80(t,J＝11.0Hz,1H),2.56–2.52(m,1H),2.47(dd,J＝11.1,3.0Hz,1H).MS(ESI):m/z366.4[M+H] ⁺.

Step 2: 4- (8-amino-3- (9-carbonyloctahydro-2H-pyrazino [1,2-a ] pyrazin-2-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

Referring to reaction step 6 of example 1, intermediate 2-6 was used instead of intermediate 1-6 as a reaction starting material to give compound 2(26 mg).

¹H NMR(500MHz,DMSO-d ₆)δ11.43(s,1H),8.72(d,J＝5.1Hz,1H),8.56(s,1H),8.06–7.98(m,2H),7.79(d,J＝3.4Hz,1H),7.67(t,J＝7.8Hz,1H),7.58(d,J＝5.1Hz,1H),7.36(d,J＝5.0Hz,1H),7.05(d,J＝5.0Hz,1H),6.13(brs,2H),3.67(d,J＝11.6Hz,1H),3.42-3.36(m,2H),3.16–3.04(m,2H),3.01–2.85(m,4H),2.58-2.52(m,2H).MS(ESI):m/z 570.4[M+H] ⁺.

Example 3

Step 1: 4- (8-amino-3- (1-carbonyl-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -N- (pyridin-2-yl) benzamide

Referring to reaction step 6 of example 1, intermediates 1 to 6 and compounds 3 to 7 were used as starting materials to give compound 3(50 mg).

¹H NMR(500MHz,DMSO-d ₆):δ10.80(s,1H),8.41-8.40(m,1H),8.23-8.21(m,1H),8.16-8.14(m,2H),7.88-7.85(m,1H),7.77-7.75(m,2H),7.61(s,1H),7.31-7.30(m,1H),7.19-7.17(m,1H),7.03-7.02(m,1H),6.11(br,2H),3.41-3.38(m,2H),3.23-3.20(m,2H),3.02-2.98(m,2H),2.06-2.04(m,2H),1.99-1.93(m,2H),1.52-1.50(m,2H).HR-MS(ESI):m/z 483.2233[M+H] ⁺.

Example 4

Step 1: 8- (8-amino-1- (4-phenoxyphenyl) imidazo [1,5-a ] pyrazin-3-yl) -2, 8-diazaspiro [4.5] decan-1-one

Referring to reaction step 6 of example 1, (4-phenoxyphenyl) boronic acid (i.e., compound 4-7) was used as a reaction material in place of compound 1-7 to give compound 4(37 mg).

¹H NMR(500MHz,DMSO-d ₆):δ7.61-7.60(m,3H),7.44-7.41(m,2H),7.25-7.24(m,1H),7.19-7.16(m,1H),.7.12-7.10(m,4H),6.96-6.95(m,1H),6.00(br,2H),3.37-3.36(m,2H),3.22-3.20(m,2H),2.99-2.95(m,2H),2.08-2.01(m,2H),2.0-1.94(m,2H),1.51-1.48(m,2H).HR-MS(ESI):m/z 455.2201[M+H] ⁺.

Example 5

Step 1: 8- (8-amino-1- (2-chloro-4-phenoxyphenyl) imidazo [1,5-a ] pyrazin-3-yl) -2, 8-diazaspiro [4.5] decan-1-one

Reference example 1 reaction step 6, substitution of 2- (2-chloro-4-phenoxyphenyl) -4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolane (i.e., compound 5-7, whose preparation reference is US20090286768) for compound 1-7 was used as a reaction raw material to obtain compound 5(69 mg).

¹H NMR(500MHz,DMSO-d ₆):δ7.59(s,1H),7.500-7.45(m,3H),7.25-7.22(m,2H),7.20-7.18(m,3H),7.05-7.02(m,1H),6.95-6.94(m,1H),5.79(br,2H),3.37-3.36(m,2H),3.22-3.19(m,2H),2.97-2.92(m,2H),2.04-1.96(m,2H),1.94-1.91(m,2H),1.50-1.47(m,2H).HR-MS(ESI):m/z 489.1816[M+H] ⁺.

Example 6

Step 1: 8- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -2-oxa-8-azaspiro [4.5] decan-1-one

Reference example 1 reaction step 5 with 2-oxa-8-azaspiro [4.5]Replacement of 2, 8-diazaspiro [4.5] by-1-decanone hydrochloride]-1-decanone hydrochloride was used as the starting material for the reaction to give intermediate 6-6(196 mg). MS (ESI) M/z 366.3[ M + H ]] ⁺.

Step 2: 4- (8-amino-3- (1-carbonyl-2-oxa-8-azaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

Referring to reaction step 6 of reference example 1,8- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -2-oxa-8-azaspiro [4.5] decan-1-one (intermediate 6-6) was used as a reaction starting material in place of intermediate 1-6 to give compound 6(31 mg).

¹H NMR(500MHz,DMSO-d ₆):δ11.43(s,1H),8.72-8.71(m,1H),8.56(s,1H),8.03-8.00(m,2H),7.67-7.64(m,1H),7.58-7.57(m,1H),7.35-7.34(m,1H),7.02-7.01(m,1H),6.01(br,2H),4.34-4.32(m,2H),3.41-3.38(m,2H),3.05-3.00(m,2H),2.30-2.27(m,2H),2.02-1.97(m,2H),1.76-1.73(m,2H).HR-MS(ESI):m/z 570.1891[M+H] ⁺.

Example 7

Step 1: 8-azaspiro [4.5] decan-1-ones

To the sealed tube were added tert-butyl 1-oxo-8-azaspiro [4.5] decane-8-carboxylate (i.e., compound 7-3, 0.5g) and a 4M solution of hydrogen chloride in dioxane (0.144g,0.987ml,3.95mmol) and the reaction was stirred at room temperature for 1 h. The reaction solution was subjected to rotary evaporation under reduced pressure to remove the solvent. Dissolving with 5ml dioxane solution, concentrating, repeating for 3 times to obtain 8-azaspiro [4.5] decan-1-one hydrochloride. To the resulting hydrochloride salt was added 10mL of saturated sodium bicarbonate solution, stirred vigorously, extracted 3 times with DCM, and the combined organic layers were separated. Dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated to give intermediate 7-4(340 mg).

¹H NMR(500MHz,DMSO-d6):δ2.84-2.81(m,2H),2.58-2.54(m,2H),2.22-2.19(m,2H),1.88-1.85(m,2H),1.83-1.79(m,2H),1.45-1.39(m,2H),1.30-1.20(m,2H).MS(EI):m/z 153[M] ⁺.

Step 2: 8- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -8-azaspiro [4.5] decan-1-one

Reference example 1 reaction step 5, replacement of 2, 8-diazaspiro [4.5] with intermediate 7-4]-1-decanone hydrochloride was used as the starting material for the reaction to give intermediate 7-6(175 mg). MS (ESI) M/z 364.4[ M + H] ⁺.

And step 3: 4- (8-amino-3- (1-carbonyl-8-azaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

Referring to reaction step 6 of example 1, intermediate 7-6 was used instead of intermediate 1-6 as a reaction starting material to give compound 7(39 mg).

¹H NMR(500MHz,DMSO-d ₆):δ11.43(s,1H),8.72-8.71(m,1H),8.56(s,1H),8.03-8.00(m,2H),7.67-7.64(m,1H),7.58-7.57(m,1H),7.31-7.30(m,1H),7.01-7.00(m,1H),5.99(br,2H),3.02-2.98(m,2H),2.31-2.28(m,2H),2.03-1.95(m,4H),1.89-1.81(m,2H),1.79-1.78(m,2H),1.53-1.51(m,2H).HR-MS(ESI):m/z 568.2096[M+H] ⁺.

Example 8

Step 1: 8- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -1, 8-diazaspiro [4.5] decan-2-one

Reference example 1 reaction step 5 with 1, 8-diazaspiro [4.5]Replacement of 2, 8-diazaspiro [4.5] by Decan-2-one hydrochloride]-1-decanone hydrochloride was used as the starting material for the reaction to give intermediate 8-6(119 mg). MS (ESI) M/z 365.4[ M + H ]] ⁺。

Step 2: 4- (8-amino-3- (2-carbonyl-1, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

The intermediate 8 to 6(0.11g), the compound 1 to 7(0.25g) and potassium phosphate (128mg) were sequentially added to a microwave tube, and dissolved in DMF (6mL), and after bubbling nitrogen for 1 minute, tetrakis (triphenylphosphine) palladium (87mg) was added, and after continuing bubbling nitrogen for 1 minute, the mixture was placed in a microwave reactor 100W and heated to 80 ℃ for 60 minutes. The reaction was concentrated, passed through silica gel column, DCM: MeOH elution (97:3) finally afforded compound 8(40 mg).

1H NMR(500MHz,DMSO-d6)δ11.42(s,1H),8.72-8.71(d,J＝5Hz,1H),8.56(s,1H),8.06-8.00(m,2H),7.67-7.64(m,1H),7.58-7.57(d,J＝5Hz,1H),7.48-7.47(m,1H),7.28-7.27(d,J＝5Hz,1H),7.02-7.01(d,J＝4.5Hz,1H),6.02(s,2H),3.15-3.11(m,2H),2.25-2.22(m,2H),1.95-1.92(m,2H),1.83-1.78(m,4H).HR-MS(ESI):m/z 569.2044[M+H]+。

Example 9

Step 1: 8- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -2, 8-diazaspiro [4.5] decan-3-one

Reference example 1 reaction step 5, substituting 2, 8-diazaspiro [4.5] decan-3-one hydrochloride for 2, 8-diazaspiro [4.5] -1-decanone hydrochloride as a reaction starting material gave intermediate 9-6(253 mg).

1H NMR(500MHz,DMSO-d6)δ7.54(s,1H),7.20-7.20(d,J＝5Hz,1H),6.91-6.90(d,J＝5Hz,1H),6.56(s, 2H),3.12-3.00(m,6H),2.13-2.11(m,2H),1.74-1.713(m,4H).

MS(ESI):m/z 365.3[M+H]+。

Step 2: 4- (8-amino-3- (3-carbonyl-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

Reference example 8 reaction step 2, intermediate 9-6 was substituted for intermediate 8-6 as a reaction starting material to give compound 9(16.8 mg).

1H NMR(500MHz,DMSO-d6)δ11.42(s,1H),8.72-8.71(d,J＝5Hz,1H),8.56(s,1H),8.03-8.00(m,2H),7.67-7.64(m,1H),7.58-7.55(m,2H),7.28-7.27(d,J＝5Hz,1H),7.02-7.01(d,J＝5Hz,1H),5.98(s,2H),3.19-3.10(m,6H),2.15(s,2H),1.79-1.76(m,4H).HR-MS(ESI):m/z 569.2040[M+H]+。

Example 10

Step 1: 7- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -2, 7-diazaspiro [3.5] nonan-1-one

Reference example 1 reaction step 5, substituting 2, 7-diazaspiro [3.5] nonan-1-one hydrochloride for 2, 8-diazaspiro [4.5] -1-decanone hydrochloride as a reaction starting material gave intermediate 10-6(124 mg). MS (ESI) M/z 351.3[ M + H ] +.

Step 2: 4- (8-amino-3- (1-carbonyl-2, 7-diazaspiro [3.5] nonan-7-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

Referring to reaction step 2 of example 8, intermediate 10-6 was used instead of intermediate 8-6 as a starting material to give compound 10(13 mg).

¹H NMR(500MHz,DMSO-d ₆)δ11.43(s,1H),8.72(d,J＝5.1Hz,1H),8.56(s,1H),8.07–7.93(m,2H),7.81(s,1H),7.65(t,J＝7.8Hz,1H),7.58(d,J＝5.1Hz,1H),7.32(d,J＝5.0Hz,1H),7.02(d,J＝5.0Hz,1H),6.00(brs,2H),3.42–3.34(m,2H),3.13(s,2H),3.09-3.00(m,2H),2.06–1.97(m,2H),1.93-1.85(m,2H).HR-MS(ESI):m/z 555.1876[M+H] ⁺.

Example 11

Step 1: 9- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -2, 9-diazaspiro [5.5] undecan-1-one

Reference example 1 reaction step 5, substituting 2, 9-diazaspiro [5.5] undecane-1-one hydrochloride for 2, 8-diazaspiro [4.5] -1-decanone hydrochloride as a reaction starting material gave intermediate 11-6(90 mg).

¹H NMR(500MHz,DMSO-d ₆)δ7.40-7.37(m,2H),6.91-6.90(d,J＝5.5Hz,1H),3.14-3.10(m,4H),2.17-2.11(m,2H),1.79-1.71(m,2H),1.54-1.51(m,2H),1.24-1.23(m,2H).MS(ESI):m/z 379.4[M+H] ⁺。

Step 2: 4- (8-amino-3- (1-carbonyl-2, 9-diazaspiro [5.5] undecan-9-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

Reference example 8 reaction step 2, intermediate 11-6 was substituted for intermediate 8-6 as a reaction starting material to give compound 11(9 mg).

¹H NMR(500MHz,DMSO-d ₆)δ11.46(s,1H),8.72-8.71(d,J＝5Hz,1H),8.56(s,1H),8.05-8.02(m,2H),7.70-7.69(d,J＝7.5Hz,1H),7.59-7.58(d,J＝4.5Hz,1H),7.01-7.00(d,J＝5Hz,1H),3.15-3.08(m,4H),2.23-2.17(m,2H),2.02(m,2H),1.81-1.72(m,2H),1.58-1.56(m,2H).HR-MS(ESI):m/z 583.2225[M+H] ⁺。

Example 12

Step 1: 7- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -2, 7-diazaspiro [4.4] nonan-1-one

Reference example 1 reaction step 5, substituting 2, 7-diazaspiro [4.4] nonan-1-one hydrochloride for 2, 8-diazaspiro [4.5] -1-decanone hydrochloride as a reaction starting material gave intermediates 12-6(0.31 g).

¹H NMR(500MHz,DMSO-d6)δ7.802(s,1H),7.393-7.382(d,J＝5.5Hz,1H),6.745-6.734(d,J＝5.5Hz,1H),3.658-3.611(m,3H),3.477-3.457(m,1H),3.238-3.211(m,2H),2.116-2.019(m,3H),1.920-1.885(m,1H).MS(ESI):m/z 351.3[M+H] ⁺.

Step 2: 4- (8-amino-3- (6-carbonyl-2, 7-diazaspiro [4.4] nonan-2-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

Referring to reaction step 6 of example 1, intermediate 12-6 was used instead of intermediate 1-6 as a starting material to give compound 12(102 mg).

¹H NMR(500MHz,DMSO)δ11.415(s,1H),8.717-8.707(d,J＝5Hz,1H),8.561(s,1H),8.025-7.992(m,2H),7.775(s,1H),7.663-7.631(m,1H),7.580-7.570(d,J＝5Hz,1H),7.404-7.394(d,J＝5Hz,1H),6.895-6.885(d,J＝5Hz,1H),5.868(s,2H),3.719-3.611(m,3H),3.478-3.458(d,J＝10Hz,1H),3.252-3.225(m,2H),2.156-2.069(m,3H),1.938-1.929(m,1H).MS(ESI):m/z 555.4[M+H] ⁺.

Example 13

Step 1: 7- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -2, 7-diazaspiro [4.5] decan-1-one

Reference example 1 reaction step 5, substituting 2, 7-diazaspiro [4.5] -1-decanone hydrochloride for 2, 8-diazaspiro [4.5] -1-decanone hydrochloride as a reaction starting material gave intermediate 13-6(0.079 g).

¹H NMR(500MHz,DMSO)δ7.663(s,1H),7.278-7.267(d,J＝5.5Hz,1H),6.938-6.927(d,J＝5.5Hz,1H),3.197-3.171(m,2H),3.044-3.021(m,1H),2.948-2.924(m,1H),2.803-2.758(m,1H),2.191-2.140(m,1H),2.012-1.958(m,2H),1.831-1.781(m,1H),1.745-1.719(m,1H),1.672-1.556(m,1H),1.530-1.508(m,1H).MS(ESI):m/z 365.3[M+H] ⁺.

Step 2: 4- (8-amino-3- (1-carbonyl-2, 7-diazaspiro [4.5] decan-7-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

Referring to reaction step 6 of example 1, intermediate 13-6 was used instead of intermediate 1-6 as a starting material to give compound 13(156 mg).

¹H NMR(500MHz,DMSO)δ11.423(s,1H),8.719-8.709(d,J＝5Hz,1H),8.561(s,1H),8.032-7.999(m,2H),7.673-7.647(m,2H),7.580-7.571(d,J＝4.5Hz,1H),7.286-7.277(d,J＝4.5Hz,1H),7.044-7.035(d,J＝4.5Hz,1H),6.017(s,2H),3.376-3.351(m,1H),3.223-3.197(m,2H),3.074-3.051(d,J＝11.5Hz,1H),2.995-2.971(d,J＝12Hz,1H),2.799-2.753(m,1H),2.275-2.250(m,1H),2.057-1.994(m,1H),1.888-1.863(m,1H),1.776-1.751(m,1H),1.700-1.647(m,1H),1.579-1.553(m,1H).MS(ESI):m/z 569.4[M+H] ⁺.

Example 14

The method comprises the following steps: (4- ((2-methoxybenzamido) methyl) phenyl) boronic acid

4-Aminomethylbenzeneboronic acid (1.36g), 2-methoxybenzoic acid (1.00g), tetrahydrofuran (30mL) and N, N-diisopropylethylamine (4.25g) were sequentially charged into a round-bottomed flask, nitrogen was replaced three times, and propylphosphoric anhydride (3.14g, 6.16mL) was added. The mixture was heated to 70 ℃ and stirred overnight. After cooling to room temperature, 300ml of water was added to the reaction solution, and a solid precipitated, which was filtered to obtain a filter cake. The filter cake was scraped off and washed again with 300ml of water to give compound 14-7(1.19 g).

¹H NMR(500MHz,DMSO-d6)δ8.68(t,J＝5.7Hz,1H),7.76(ddd,J＝11.9,6.9,5.3Hz,3H),7.51–7.44(m,1H),7.31(d,J＝7.7Hz,2H),7.15(d,J＝8.3Hz,1H),7.05(t,J＝7.4Hz,1H),4.52(d,J＝6.0Hz,2H),3.90(s,3H).MS(ESI):m/z 284.3[M-H] ^-

Step two: n- (4- (8-amino-3- (1-carbonyl-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) benzyl) -2-methoxybenzamide

Intermediate 1 to 6(200mg) and compound 14 to 7(148mg) were weighed in a microwave reaction tube, dissolved in dioxane (5mL), and potassium carbonate (0.18g) and water (1mL) were added, followed by stirring for 1 minute, followed by addition of 1, 1' -bis (diphenylphosphino) ferrocene palladium (II) dichlorodichloromethane complex (0.10g), followed by reaction step 6 of example 1 to obtain compound 14(157 mg).

¹H NMR(500MHz,DMSO-d6)δ8.75(t,J＝5.7Hz,1H),7.78(d,J＝7.5Hz,1H),7.66–7.54(m,3H),7.48(dd,J＝17.9,7.8Hz,3H),7.25(d,J＝4.7Hz,1H),7.16(d,J＝8.3Hz,1H),7.05(t,J＝7.4Hz,1H),6.97(d,J＝4.7Hz,1H),5.96(s,2H),4.58(d,J＝5.8Hz,2H),3.91(s,3H),3.38(s,2H),3.21(t,J＝6.6Hz,2H),2.97(t,J＝11.5Hz,2H),2.03(t,J＝6.6Hz,2H),1.95(t,J＝10.7Hz,2H),1.50(d,J＝12.8Hz,2H).HR-MS(ESI):m/z526.2475[M+H] ⁺。

Example 15

The method comprises the following steps: (4- ((5-fluoro-2-methoxybenzamido) methyl) phenyl) boronic acid

Referring to reaction step 1 of example 14, 4-aminomethylphenylboronic acid hydrochloride (242mg) and 5-fluoro 2-methoxybenzoic acid (200mg) were used as starting materials to give compounds 15 to 7(196 mg).

¹H NMR(500MHz,DMSO-d6)δ8.78(t,J＝5.7Hz,1H),7.76(d,J＝7.8Hz,2H),7.51(dd,J＝9.2,3.2Hz,1H),7.36–7.26(m,3H),7.18(dd,J＝9.1,4.2Hz,1H),4.51(d,J＝6.0Hz,2H),3.89(s,3H).MS(ESI):m/z 302.4[M-H] ^-.

Step two: n- (4- (8-amino-3- (1-carbonyl-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) benzyl) -5-fluoro-2-methoxybenzamide

Reference example 1 reaction step 6, starting from intermediates 1-6 and compounds 15-7, gave compound 15(166mg)

¹H NMR(500MHz,DMSO-d6)δ8.85(t,J＝5.8Hz,1H),7.67–7.56(m,3H),7.54(dd,J＝9.1,3.0Hz,1H),7.46(d,J＝7.9Hz,2H),7.34(td,J＝8.7,3.1Hz,1H),7.26(d,J＝4.7Hz,1H),7.19(dd,J＝9.0,4.1Hz,1H),6.97(d,J＝3.5Hz,1H),5.98(s,2H),4.58(d,J＝5.9Hz,2H),3.91(s,3H),3.36(s,2H),3.21(t,J＝6.7Hz,2H),2.97(t,J＝11.3Hz,2H),2.03(t,J＝6.7Hz,2H),1.99–1.88(m,2H),1.50(d,J＝13.0Hz,2H).HR-MS(ESI):m/z 544.2374[M+H] ⁺.

Example 16

Step 1: 8- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -2, 8-diazaspiro [4.5] decane-2-carboxylic acid tert-butyl ester

Reference example 1 reaction step 5 with 2, 8-diazaspiro [4.5]Tert-butyl decane-2-carboxylate was used as the starting material for the reaction to give intermediate 16-6(2.48 g). MS (ESI) M/z 451.4[ M + H ]] ⁺.

Step 2: 8- (8-amino-1- (2-fluoro-4- ((4- (trifluoromethyl) pyridin-2-yl) carbamoyl) phenyl) imidazo [1,5-a ] pyrazin-3-yl) tert-butyl-2, 8-diazaspiro [4.5] decane-2-carboxylic acid methyl ester

Referring to reaction step 6 of example 1, intermediate 16-6 was used instead of intermediate 1-6 as a starting material to give compound 16-8(822 mg). MS (ESI) M/z 655.5[ M + H ]] ⁺.

And step 3: 4- (8-amino-3- (2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

16-8(700mg) and dichloromethane (10mL) were sequentially added to the reaction flask, the mixture was stirred and dissolved in an ice bath, and trifluoroacetic acid (3.3mL) was added to the reaction system to react at room temperature for 16 hours. After the reaction is finished, adding a saturated sodium bicarbonate solution to adjust the pH to be alkalescent, extracting with ethyl acetate, concentrating the extract, purifying by silica gel column chromatography, and adding DCM: MeOH (93:7) elution, combined eluates and concentrated to give compound 16(327 mg).

¹H NMR(500MHz,DMSO-d6):δ8.71-8.70(m,1H),8.55(s,1H),8.02-7.99(m,2H),7.66-7.63(m,1H),7.57-7.56(m,1H),7.26-7.25(m,1H),7.01-7.00(m,1H),5.99(br,2H),3.11-3.09(m,4H),2.88-2.85(m,2H),2.68(s,2H),1.69-1.68(m,4H),1.60-1.57(m,2H)

HR-MS(ESI):m/z 555.2259[M+H] ⁺.

Example 17

Step 1: 7- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -2, 7-diazaspiro [3.5] nonane-2-carboxylic acid tert-butyl ester

Reference example 1 reaction step 5, starting from tert-butyl 2, 7-diazaspiro [3.5] nonane-2-carboxylate, gave intermediate 17-6(1.46 g).

¹H NMR(500MHz,DMSO-d6):δ7.19-7.18(m,1H),6.90-6.89(m,1H),6.57(br,2H),3.61(br,4H),3.34-3.3.28(m,4H),1.93-1.87(m,2H),1.84-1.82(m,2H),1.38(s,9H).

¹³C NMR(125MHz,DMSO-d6):174.2,156.1,151.4,144.5,128.4,113.4,106.2,103.9,78.8,48.94,47.2,34.8,33.3,30.6,29.5,28.5.MS(ESI):m/z 437.5[M+H] ⁺.

Step 2: 7- (8-amino-1- (2-fluoro-4- ((4- (trifluoromethyl) pyridin-2-yl) carbamoyl) phenyl) imidazo [1,5-a ] pyrazin-3-yl) tert-butyl-2, 7-diazaspiro [3.5] nonane-2-carboxylic acid tert-butyl ester

Referring to reaction step 6 of example 1, intermediate 17-6 was used instead of intermediate 1-6 as a starting material to give compound 17-8(715 mg). MS (ESI) M/z 641.6[ M + H ]] ⁺.

And step 3: 4- (8-amino-3- (2, 7-diazaspiro [3.5] non-7-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

17-8(607mg) and dichloromethane (10mL) were sequentially added to the reaction flask, the mixture was stirred and dissolved in an ice bath, and then trifluoroacetic acid (2.0mL) was added to the reaction system to react at room temperature for 3 hours. After the reaction is finished, adding a saturated sodium bicarbonate solution to adjust the pH to be alkalescent, extracting with ethyl acetate, concentrating the extract, purifying by silica gel column chromatography, and adding DCM: MeOH (93:7) elution, combined eluates and concentrated to give compound 17(333 mg).

¹H NMR(500MHz,DMSO-d6):δ8.71-8.70(m,1H),8.56(s,1H),8.02-7.99(m,2H),7.65-7.62(m,1H),7.58-7.57(m,1H),7.27-7.26(m,1H),7.01-7.00(m,1H),5.99(br,2H),3.05-3.03(m,8H),1.90-1.88(m,4H).HR-MS(ESI):m/z 541.2119[M+H] ⁺.

Example 18

Step 1: 4- (8-amino-3- (2-methyl-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

After compound 16(100mg) and methylene chloride (2.5mL) were sequentially added to a reaction flask and washed, 37% aqueous formaldehyde (0.054mL), acetic acid (10.83mg) and sodium cyanoborohydride (45.32mg) were added thereto and the reaction was terminated at room temperature for 3 hours. Adding sodium bicarbonate saturated solution to quench the reaction and adjusting the pH to be alkalescent, and adding ethyl acetate for extraction. The extract was concentrated and purified by silica gel column chromatography, DCM: MeOH (96:4) elution, combined eluates and concentrated to give compound 18(42 mg).

¹H NMR(500MHz,DMSO-d6):δ11.43(br,1H),8.71-8.70(m,1H),8.56(s,1H),8.03-7.99(m,2H),7.66-7.63(m,1H),7.58-7.57(m,1H),7.26-7.25(m,1H),7.01-7.00(m,1H),5.98(br,2H),3.10-3.07(m,4H),2.48-2.47(m,2H),2.36(s,2H),2.23(m,3H),1.73-1.63(m,6H).

HR-MS(ESI):m/z 569.2444[M+H] ⁺.

Example 19

Step 1: 4- (8-amino-3- (2-methyl-2, 7-diazaspiro [3.5] non-7-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

Reference example 18 reaction step 1, starting with 17, gave compound 19(50 mg).

¹H NMR(500MHz,DMSO-d6):δ11.43(br,1H),8.71-8.70(m,1H),8.56(s,1H),8.02-7.99(m,2H),7.65-7.62(m,1H),7.58-7.57(m,1H),7.26-7.25(m,1H),7.01-7.00(m,1H),5.99(br,2H),3.06-3.03(m,4H),2.99(s,4H),2.27-2.23(m,3H),1.87-1.85(m,4H)

¹³C NMR(125MHz,DMSO-d6):δ165.6,160.5,158.5,153.6,151.9,150.4,145.6,138.7,135.3,133.0,127.9,126.8,126.6,124.6,124.5,116.4,116.2,115.8,113.8,110.4,106.0,66.0,47.6,46.2,35.7,34.1.HR-MS(ESI):m/z 555.2279[M+H] ⁺.

Example 20

Step 1: 2-methyl-1-oxo-2, 8-diazaspiro [4.5] decane-8-carboxylic acid tert-butyl ester

1-oxo-2, 8-diazaspiro [1,4] decane-8-carboxylic acid tert-butyl ester (2.5g) and DMF (100mL) were added to a reaction flask, sodium hydrogen (1.664g) was added under ice bath, methyl iodide (2.6mL) was added thereto and reacted at room temperature for 16 hours, and after completion of the reaction, a saturated sodium bicarbonate solution was added thereto and quenched, and extracted three times with dichloromethane. The extract was concentrated to give compound 20-1(3.90 g).

¹H NMR(500MHz,DMSO-d6):δ3.624-3.577(m,1H),3.18-3.17(m,2H),3.10(s,2H),2.72(s,3H),1.66-1.63(m,4H),1.40(s,9H).

¹³C NMR(125MHz,DMSO-d6):154.5,150.4,121.8,79.2,54.4,53.4,28.5。

Step 2: 2-methyl-2, 8-diazaspiro [4.5] decan-1-one

Reference example 16 reaction step 3, starting from compound 20-1, gave intermediate 20-2(2.461 g).

¹H NMR(500MHz,DMSO-d6):δ3.25-3.22(m,2H),2.86-2.83(m,2H),2.70(s,3H),2.56-2.53(m,2H),1.90-1.87(m,2H),1.59-1.53(m,2H),1.21-1.18(m,2H).

And step 3: 8- (8-amino-1-bromoimidazo [1,5-a ] pyrazin-3-yl) -2-methyl-2, 8-diazaspiro [4.5] decan-1-one

Referring to reaction step 5 of example 1, intermediate 20-2 was used as a reaction starting material to give intermediate 20-6(98 mg).

¹H NMR(500MHz,DMSO-d6):δ7.25-7.24(m,1H),6.90-6.89(m,1H),6.56(br,2H),3.30-3.25(m,6H),2.75(s,3H),2.03-1.99(m,2H),1.97-1.86(m,2H),1.50-1.41(m,2H).

¹³C NMR(125MHz,DMSO-d6):177.3,151.3,144.7,130.1,128.3,113.3,106.3,103.9,101.9,46.7,45.7,42.5,32.0,29.8,29.0.MS(ESI):m/z 379.4[M+H] ⁺.

And 4, step 4: 4- (8-amino-3- (2-methyl-1-oxo-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

Referring to reaction step 6 of example 1, intermediate 20-6 was used instead of intermediate 1-6 as a starting material to give compound 20(47 mg).

¹H NMR(500MHz,DMSO-d6):δ11.43(s,1H),8.72-8.71(m,1H),8.56(s,1H),8.03-8.00(m,2H),7.67-7.64(m,1H),7.58-7.57(m,1H),7.33-7.32(m,1H),7.01-7.00(m,1H),6.13(br,2H),3.40-3.37(m,4H),3.01-2.96(m,2H),2.76(s,3H),2.01-1.96(m,4H),1.50-1.48(m,2H).

¹³C NMR(125MHz,DMSO-d6):177.4,165.5,158.5,153.5,151.7,150.4,145.8,138.9,135.5,133.0,126.8,124.7,116.5,116.2,115.8,113.7,110.4,106.3,46.9,45.7,42.6,32.2,29.8,29.1.HR-MS(ESI):m/z 583.2222[M+H] ⁺.

Example 21

Step 1: (2-fluoro-4- (pyridin-2-ylcarbamoyl) phenyl) boronic acid

4-carboxy-2-fluorophenylboronic acid (2g), 2-aminopyridine (1.228g), DMF (18mL), DIPEA (3.79mL) were added to the reaction flask in this order, HATU (4.96g) was added thereto at room temperature, and the reaction was carried out at 80 ℃ for 2 hours. After the reaction is finished, pouring the system into ice water, separating out solid, and performing suction filtration and drying to obtain an intermediate 21-7(1.055 g).

¹H NMR(500MHz,DMSO-d6)δ10.87(s,1H),8.41(s,3H),8.19-8.18(d,J＝8.5Hz,1H),7.88-7.83(m,2H),7.75-7.73(d,J＝5Hz,1H),7.68-7.65(m,1H),7.20-7.18(m,1H).

¹³C NMR(125MHz,DMSO-d6)166.4,165.2,164.4,152.4,148.4,138.6,137.6,135.6,123.6,120.4,115.3,114.8.MS(ESI):m/z 259.2[M-H] ^-.

Step 2: 4- (8-amino-3- (1-oxo-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (pyridin-2-yl) benzamide

Referring to reaction step 6 of example 1, intermediates 21-7 were used instead of intermediates 1-7 as starting materials to give compound 21(154 mg).

¹H NMR(500MHz,DMSO-d6)δ10.92(s,1H),8.43-8.42(d,J＝4.5Hz,1H),8.22-8.20(d,J＝8.5Hz,1H),8.01-7.99(m,2H),7.89-7.86(m,1H),7.65-7.60(m,2H),7.31-7.30(d,J＝4.5Hz,1H),7.22-7.19(m,1H),7.02-7.01(d,J＝4.5Hz,1H),5.97(s,2H),3.40-3.38(m,2H),3.20(m,2H),3.01-2.96(m,2H),2.06-2.03(m,2H),1.98-1.94(m,2H),1.55-1.50(m,2H).

¹³C NMR(125MHz,DMSO-d6)180.5,164.9,160.6,158.6,152.5,151.9,148.5,145.6,138.6,136.1,132.9,127.9,126.4,124.6,120.5,116.3,116.1,115.4,113.9,106.1,55.4,47.0,41.9,38.4,32.0,31.5.HR-MS(ESI):m/z501.2132[M+H] ⁺.

Example 22

Step 1: (2-fluoro-4- (pyridin-2-ylcarbamoyl) phenyl) boronic acid

Referring to synthetic step 1 of example 21, starting from 4-carboxyphenylboronic acid and 2-amino-4-trifluoromethylpyridine, compound 22-7(772mg) was obtained. MS (ESI) M/z 309.3[ M-H ]] ^-.

Step 2: 4- (8-amino-3- (1-oxo-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -N- (4- (trifluoromethyl) pyridin-2-yl) benzamide

Referring to reaction step 6 of example 1, intermediate 22-7 was used instead of intermediate 1-7 as a starting material to give compound 22(105 mg).

¹H NMR(500MHz,DMSO-d6)δ11.32(s,1H),8.70-8.69(d,J＝5Hz,1H),8.58(s,1H),8.18-8.16(d,J＝8Hz,2H),7.79-7.78(d,J＝8Hz,2H),7.62(s,1H),7.56-7.55(d,J＝5Hz,1H),7.32-7.31(d,J＝5Hz,1H),7.04-7.03(d,J＝5Hz,1H),6.12(s,2H),3.42-3.39(m,2H),3.24-3.24(m,2H),3.02-2.98(m,2H),2.06-2.04(m,2H),2.00-1.94(m,2H),1.53-1.50(m,2H).HR-MS(ESI):m/z 551.2129[M+H] ⁺.

Example 23

Step 1: (2-fluoro-4- ((4-methylpyridin-2-yl) carbamoyl) phenyl) boronic acid

Referring to Synthesis step 1 of example 21, 2-amino-4-methylpyridine was used as a reaction starting material in place of 2-aminopyridine to obtain compounds 23-7(2.063 g).

¹H NMR(500MHz,DMSO-d6)δ10.78(s,1H),8.42-8.41(d,J＝7.5Hz,2H),8.26-8.25(d,J＝5Hz,1H),8.04(s,1H),7.84-7.82(m,1H),7.74-7.72(d,J＝10Hz,1H),7.68-7.65(m,1H),7.04-7.03(d,J＝5Hz,1H),2.37(s,3H).

¹³C NMR(125MHz,DMSO-d6)166.3,165.1,164.4,152.5,149.3,148.1,137.7,135.7,123.6,121.5,115.7,114.7,21.4.MS(ESI):m/z 273.3[M-H] ^-.

Step 2: 4- (8-amino-3- (1-oxo-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4-methylpyridin-2-yl) benzamide

Reference example 1 reaction step 6, intermediate 23-7 was substituted for intermediate 1-7 as a reaction starting material to give compound 23(193 mg).

¹H NMR(500MHz,DMSO-d6)δ10.85(s,1H),8.28-8.27(d,J＝5Hz,1H),8.07(s,1H),8.00-7.98(m,2H),7.65-7.61(m,2H),7.31-7.30(d,J＝5Hz,1H),7.05-7.04(d,J＝5Hz,1H),7.01-7.00(d,J＝5Hz,1H),5.99(s,2H),3.40-3.37(m,2H),3.23-3.20(m,2H),3.00-2.95(m,2H),2.38(s,3H),2.05-2.03(m,2H),1.99-1.93(m,2H),1.52-1.50(m,2H).HR-MS(ESI):m/z 515.2222[M+H] ⁺.

Example 24

Step 1: (2-fluoro-4- ((4-methoxypyridin-2-yl) carbamoyl) phenyl) boronic acid

Reference example 21 Synthesis of step 1 with 2-amino-4-methoxypyrazinePyridine is used as a reaction raw material to replace 2-aminopyridine, and then compound 24-7(2.050g) is obtained. MS (ESI) M/z 289.3[ M-H ]] ^-.

Step 2: 4- (8-amino-3- (1-oxo-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -3-fluoro-N- (4-methoxypyridin-2-yl) benzamide

Referring to reaction step 6 of example 1, intermediate 24-7 was used instead of intermediate 1-7 as a starting material to give compound 24(310 mg).

¹H NMR(500MHz,DMSO-d6)δ10.90(s,1H),8.24-8.23(d,J＝5.5Hz,1H),8.00-7.98(m,2H),7.86(s,1H),7.65-7.61(m,2H),7.31-7.30(d,J＝5Hz,1H),7.01-7.00(d,J＝4.5Hz,1H),6.82-6.81(m,1H),5.98(s,2H),3.88(s,3H),3.40-3.37(m,2H),3.23-3.17(m,2H),3.00-2.95(m,2H),2.05-2.03(m,2H),1.99-1.93(m,2H),1.52-1.49(m,2H).HR-MS(ESI):m/z 531.2267[M+H] ⁺.

Example 25

Step 1: (4- ((5-Chloropyridin-2-yl) carbamoyl) -2-fluorophenyl) boronic acid

Referring to synthetic step 1 of example 21, 2-amino-5-chloropyridine was used as a reaction material in place of 2-aminopyridine to obtain compounds 25-7(965 mg). MS (ESI) M/z 293.2[ M-H ]] ^-.

Step 2: 4- (8-amino-3- (1-oxo-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -N- (5-chloropyridin-2-yl) -3-fluorobenzamide

Referring to reaction step 6 of example 1, intermediates 25-7 were used instead of intermediates 1-7 as starting materials to give compound 25(230 mg).

¹H NMR(500MHz,DMSO-d6)δ11.14(s,1H),8.48(d,J＝2.4Hz,1H),8.26(d,J＝8.9Hz,1H),8.06–7.91(m,3H),7.71–7.57(m,2H),7.31(d,J＝4.9Hz,1H),7.01(d,J＝4.9Hz,1H),5.99(s,2H),3.46–3.38(m,2H),3.22(t,J＝6.8Hz,2H),2.98(t,J＝11.8Hz,2H),2.04(t,J＝6.9Hz,2H),2.02–1.89(m,2H),1.51(d,J＝13.0Hz,2H).

HR-MS(ESI):m/z 535.1768[M+H] ⁺.

Example 26

Step 1: (4- ((4-cyanopyridin-2-yl) carbamoyl) -2-fluorophenyl) boronic acid

Referring to synthetic step 1 of example 21, compound 26-7(1.13g) was obtained by substituting 2-aminopyridine with 2-amino-4-cyanopyridine as a reaction starting material. MS (ESI) M/z 284.2[ M-H ]] ^-.

Step 2: 4- (8-amino-3- (1-oxo-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -N- (4-cyanopyridin-2-yl) -3-fluorobenzamide

Referring to reaction step 6 of example 1, intermediate 26-7 was used instead of intermediate 1-7 as a starting material to give compound 26(28 mg).

¹H NMR(500MHz,DMSO-d6)δ11.41(s,1H),8.68(dd,J＝5.0,0.9Hz,1H),8.53(t,J＝1.1Hz,1H),8.09–7.88(m,2H),7.82–7.52(m,3H),7.31(d,J＝5.0Hz,1H),7.01(d,J＝4.9Hz,1H),6.00(s,2H),3.40(d,J＝3.9Hz,2H),3.22(t,J＝6.8Hz,2H),2.98(td,J＝12.1,2.6Hz,2H),2.04(t,J＝6.8Hz,2H),1.96(m,2H),1.51(d,J＝13.2Hz,2H).

¹³C NMR(126MHz,DMSO-d6)δ180.5,165.5,160.5,158.6,153.2,151.9,150.2,145.7,133.0,128.0,124.7,124.5,121.9,121.4,117.4,116.9,116.4,116.2,113.9,106.1,47.0,41.9,38.4,32.0,31.5.HR-MS(ESI):m/z 526.2125[M+H] ⁺.

Example 27

Step 1: (4- ((5-chloro-4- (trifluoromethyl) pyridin-2-yl) carbamoyl) -2-fluorophenyl) boronic acid

Referring to synthetic step 1 of example 21, 2-amino-4-trifluoromethyl-5-chloropyridine was used as a reaction material in place of 2-aminopyridine to obtain compound 27-7(320 mg).

¹H NMR(500MHz,DMSO-d6)δ11.52(s,1H),8.78(s,1H),8.67(s,1H),8.44(s,2H),7.85-7.83(d,J＝8Hz,1H),7.76-7.74(d,J＝10Hz,1H),7.70-7.67(m,1H).MS(ESI):m/z 363.4[M+H] ^-.

Step 2: 4- (8-amino-3- (1-oxo-2, 8-diazaspiro [4.5] decan-8-yl) imidazo [1,5-a ] pyrazin-1-yl) -N- (5-chloro-4- (trifluoromethyl) pyridin-2-yl) -3-fluorobenzamide

Referring to reaction step 6 of example 1, intermediates 27-7 were used instead of intermediates 1-7 as starting materials to give compound 27(11 mg).

¹H NMR(500MHz,DMSO)δ11.57(s,1H),8.79(s,1H),8.69(s,1H),8.02-7.99(m,2H),7.68-7.65(m,1H),7.61(s,1H),7.31-7.30(d,J＝5Hz,1H),7.02-7.01(d,J＝4.5Hz,1H),6.00(s,2H),3.23-3.20(m,2H),3.00-2.95(m,2H),2.05-2.03(m,2H),1.98-1.93(m,2H),1.52-1.49(m,2H),1.23(m,2H).HR-MS(ESI):m/z603.1655[M+H] ⁺.

Test example 1: in vitro Activity

1.1BTK inhibitory Activity screening

With kinase buffer (50mM HEPES, 10mM MgCl)₂2mM DTT, 1mM EGTA, 0.01% Tween 20) 350 ng/. mu.L of BTK stock solution was diluted, 6. mu.l of 1.67 X0.134 ng/. mu.l working solution (final concentration of 0.08 ng/. mu.l) was added to each well, and DMSO-dissolved different compounds were added to the wells using a nanoliter loading apparatus to give a final concentration of 1000nM-0.244nM, 4-fold gradient, 7 concentrations, blank control empty (no enzyme) and negative control wells (enzyme-containing, vehicle DMSO-loaded) were set, and 2 duplicate wells were set. After 30min of reaction of the enzyme with the compound or vehicle, 5 × 250 μ M ATP (final concentration 50 μ M) and 5 × 0.5 μ M substrate (final concentration 0.1 μ M, ULight-poly GT) in kinase buffer were mixed in a ratio of 1:1 mixing, adding 4 mu L of the mixture into each hole; after the plate was covered with a membrane, the reaction was carried out at room temperature for 2 hours, and 5. mu.L of 4X 40mM EDTA (final concentration: 10mM) was added to each well for 5 minutes at room temperature, followed byAdd 5. mu.L of 4X 8nM detection reagent (final concentration 2nM, Ab) to each well and incubate for 1 hour at room temperature; and (3) reading the plate (excitation is 620nm, emission is 665nm) by using a PE Envision multifunctional microplate reader, and calculating IC50 by adopting four-parameter fitting.

1.2BTK (C481S) inhibition Activity Screen

With kinase buffer (50mM HEPES, 10mM MgCl)₂2mM DTT, 1mM EGTA, 0.01% Tween 20) 50 ng/. mu.L of BTK (C481S) stock was diluted, 6. mu.l of 1.67 X0.25 ng/. mu.l working solution (final concentration of 0.15 ng/. mu.l) was added to each well, DMSO-dissolved different compounds were added to the wells using a nanoliter loader to give a final concentration of 1000nM to 0.244nM, 4-fold gradient, 7 concentrations, blank control empty (no enzyme) and negative control wells (enzyme-containing, vehicle DMSO) were set, and 2 duplicate wells were set. After 30min of reaction of the enzyme with the compound or vehicle, 5 × 250 μ M ATP (final concentration 50 μ M) and 5 × 0.5 μ M substrate (final concentration 0.1 μ M, ULight-poly GT) in kinase buffer were mixed in a ratio of 1:1 mixing, adding 4 mu L of the mixture into each hole; after the plate was sealed with a membrane, after reacting at room temperature for 2 hours, 5. mu.L of 4X 40mM EDTA (final concentration: 10mM) was added to each well for 5 minutes at room temperature, and 5. mu.L of 4X 8nM detection reagent (final concentration: 2nM, Ab) was added to each well and incubated at room temperature for 1 hour; and (3) reading the plate (excitation is 620nm, emission is 665nm) by using a PE Envision multifunctional microplate reader, and calculating IC50 by adopting four-parameter fitting.

1.3EGFR (epidermal growth factor receptor) inhibitory Activity Screen

With kinase buffer (50mM HEPES, 10mM MgCl)₂2mM DTT, 1mM EGTA, 0.01% Tween 20) 50 ng/. mu.L of EGFR (WT) stock solution was diluted, 6. mu.l of 1.67 X0.8 ng/. mu.l working solution (final concentration of 0.5 ng/. mu.l) was added to each well, DMSO-dissolved different compounds were added to the wells using a nanoliter loading apparatus to give a final concentration of 1000nM-0.48nM, 4-fold gradient for 7 concentrations, blank control empty (no enzyme) and negative control wells (enzyme-containing, vehicle DMSO) were set up in 2 duplicate wells. After the enzyme has reacted with the compound or vehicle for 10min, 5X 25. mu.M ATP (final concentration of 5. mu.M) and 5X 0.5. mu.M substrate (final concentration of 0.1. mu.M, ULight-poly G) prepared in kinase buffer are mixed withT), mixing according to a ratio of 1:1, and adding 4 mu L of the mixture into each hole; after the plate is sealed and the plate is covered with a membrane, after the reaction is carried out for 1h at room temperature, 5 muL of 4 multiplied 40mM EDTA (10mM final concentration) is added into each hole, the reaction is carried out for 5min at room temperature, 5 muL of 4 multiplied 8nM detection reagent (2 nM final concentration, Eu-anti-phosphorus-tyrosinane antibody) is added into each hole, the incubation is carried out for 1h at room temperature, a PE Envision multifunctional enzyme-linked immunosorbent assay is used for reading the plate (excitation 320nM, emission 665nM), and the IC50 is calculated by adopting four-parameter fitting.

1.4ITK (Interleukin-2-inhibitor T-cell kinase) inhibition activity screening

With kinase buffer (50mM HEPES, 10mM MgCl)₂2mM DTT, 1mM EGTA, 0.01% Tween 20) 50 ng/. mu.L of ITK stock solution was diluted, 6. mu.L of 1.67X 0.0835 g/. mu.L working solution (final concentration of 0.05 ng/. mu.L) was added to each well, and DMSO-dissolved different compounds were added to the wells using a nanoliter applicator to give a final concentration of 1000nM to 0.24nM, 4-fold gradient, 7 concentrations, while blank control empty (no enzyme) and negative control wells (enzyme-containing, vehicle DMSO) were set. After the enzyme reacts with the compound or the solvent for 30min, 5 × 50 μ M ATP (final concentration of 10 μ M) prepared by using a kinase buffer solution and 5 × 0.5 μ M substrate (final concentration of 0.1 μ M, ULight-poly GT) are mixed according to a ratio of 1:1 and added into the wells according to 4 μ L per well; after the plate is sealed and the plate is covered with a membrane, after reacting for 2h at room temperature, 5. mu.L of 4X 40mM EDTA (final concentration 10mM) is added to each well for 5min at room temperature, and 5. mu.L of 4X 8nM detection reagent (final concentration 2nM, Eu-anti-phosphorus-tyrosine antibody) is added to each well and incubated for 1h at room temperature; and (3) reading the plate (excitation is 320nm, emission is 665nm) by using a PE Envision multifunctional microplate reader, and calculating IC50 by adopting four-parameter fitting.

1.5FAK (focal addition Kinase) inhibition Activity Screen

mu.L of FAK stock was diluted with kinase buffer (50mM HEPES, 10mM MgCl2, 2mM DTT, 1mM EGTA, 0.01% Tween 20), 6. mu.L of 1.67X 0.0334 ng/. mu.L working solution (final concentration of 0.02ng/ml) was added to each well, DMSO-solubilized different compounds were added to the wells using a nanoliter loader, 7 concentration gradients of compounds from 1000nM to 0.244nM were applied, 4-fold dilutions were made, and blank control empty (no enzyme) and negative control wells (enzyme-containing, vehicle DMSO) were run in parallel. After the enzyme reacts with the compound or the solvent for 120min, 5 × 50 μ M ATP (final concentration of 10 μ M) prepared by using a kinase buffer solution and 5 × 0.5 μ M substrate (final concentration of 0.1 μ M, ULight-poly GT) are mixed according to a ratio of 1:1 and added into the wells according to 4 μ L per well; after sealing the plate and membrane, after reacting for 2h at room temperature, 5. mu.L of 4X 40mM EDTA (final concentration 10mM) was added to each well for 5min at room temperature, and 5. mu.L of 4X 8nM detection reagent (final concentration 2nM, E. mu. -anti-phospho-tyrosine antibody) was added to each well and incubated for 1h at room temperature; and (3) reading the plate by using a PE Envision multifunctional microplate reader (excitation 320or 340nm and emission 665nm), and calculating IC50 by adopting four-parameter fitting.

1.6 proliferation inhibition of TMD-8 cells by Compounds

Taking TMD-8 cells in an exponential growth phase, collecting the cells to a centrifuge tube, centrifuging for 3min at 1500 rpm by using a low-speed desktop centrifuge, discarding the supernatant, and adding 2mL of plating medium (RPMI basic medium + 5% FBS +0.05mM 2-mercaptoethanol) for cell resuspension. The cells were counted using a cell counter, and the required amount of cells was adjusted to a density of 5X 104 cells/mL, seeded on a 96-well plate using a line gun, 100. mu.L/well, and cultured in a cell culture chamber containing 5% CO2 saturated humidity at 37 ℃. After 24h of incubation, compound loading was performed using a nanoliter loading apparatus, 2 duplicate wells were set for each concentration, cells without compound were used as negative controls, CCK-8 was added at 10 μ L/well after 72 hours, absorbance was measured at 450nm with an Envision microplate reader after 4 hours, inhibition was calculated, inhibition (%) (negative control mean-experimental mean)/(negative control mean-blank mean) × 100%), compound logarithm was used as abscissa, inhibition was used as ordinate, four parameter analysis, dose-effect curve was fitted, and IC50 was calculated.

The results of the above tests are shown in Table 1.

TABLE 1

Test example 2: in vitro hepatic microsomal metabolic stability

2.1 human liver microsome experiments

300 μ L of final incubation system: the solution contained 30. mu.L of human liver microsomes (protein concentration: 5mg/mL, XENOTECH, USA), 30. mu.L of NADPH (10mM) + MgCl2(5mM), 3. mu.L of the substrate, i.e., the compound of example (dissolved in 50% acetonitrile aqueous solution, 100. mu.M), and 237. mu.L of PBS buffer, in which the proportion of the organic solvent (acetonitrile) was 0.5%. Each tube is prepared with a substrate and enzyme mixing solution with a total volume of 270 mu L, and after pre-incubation for 5min at 37 ℃, 30 mu L NADPH + MgCl2 is added, and 50 mu L diazepam glacial acetonitrile (20ng/mL) containing an internal standard is taken out for 300 mu L to terminate the reaction at 0min, 15 min, 30min and 60 min. After vortexing for 5min, the cells were centrifuged (13000rpm, 4 ℃) for 10 min. Draw 100. mu.L of supernatant into the injection bottle, inject 1. mu.L of supernatant, perform LC-MS/MS analysis, calculate the remaining percentage, see Table 2.

2.2 mouse liver microsome experiments

300 μ L of final incubation system: the sample contained 30. mu.L of mouse liver microsomes (protein concentration: 5mg/mL, XENOTECH, USA), 30. mu.L of NADPH (10mM) + MgCl2(5mM), 3. mu.L of the substrate, i.e., the compound of example (dissolved in 50% acetonitrile aqueous solution, 100. mu.M), and 237. mu.L of PBS buffer, in which the proportion of the organic solvent (acetonitrile) was 0.5%. Each tube is prepared with a substrate and enzyme mixing solution with a total volume of 270 mu L, and after pre-incubation for 5min at 37 ℃, 30 mu L NADPH + MgCl2 is added, and 50 mu L diazepam glacial acetonitrile (20ng/mL) containing an internal standard is taken out for 300 mu L to terminate the reaction at 0min, 15 min, 30min and 60 min. After vortexing for 5min, the cells were centrifuged (13000rpm, 4 ℃) for 10 min. Draw 100. mu.L of supernatant into the injection bottle, inject 1. mu.L of supernatant, perform LC-MS/MS analysis, calculate the remaining percentage, see Table 2.

2.3 rat liver microsome experiments

300 μ L of final incubation system: the sample contained 30. mu.L of rat liver microsomes (protein concentration: 5mg/mL, XENOTECH, USA), 30. mu.L of NADPH (10mM) + MgCl2(5mM), 3. mu.L of the substrate, i.e., the compound of example (dissolved in 50% acetonitrile aqueous solution, 100. mu.M), and 237. mu.L of PBS buffer, in which the proportion of the organic solvent (acetonitrile) was 0.5%. Each tube is prepared with a substrate and enzyme mixing solution with a total volume of 270 mu L, and after pre-incubation for 5min at 37 ℃, 30 mu L NADPH + MgCl2 is added, and 50 mu L diazepam glacial acetonitrile (20ng/mL) containing an internal standard is taken out for 300 mu L to terminate the reaction at 0min, 15 min, 30min and 60 min. After vortexing for 5min, the cells were centrifuged (13000rpm, 4 ℃) for 10 min. Draw 100. mu.L of supernatant into the injection vial, 1. mu.L of injection was analyzed by LC-MS/MS, and the remaining percentage was calculated as shown in Table 2.

TABLE 2

Claims

A compound of formula (I) or a pharmaceutically acceptable salt thereof,

wherein the content of the first and second substances,

ring B is selected from 5-10 membered heteroaryl or C_6-10An aryl group;

R ¹independently selected from halogen, -OH, -NH₂、-CN、C _1-6Alkyl radical, C_1-6Alkoxy or halogen substituted C_1-6An alkyl group;

p is selected from 0, 1,2,3 or 4;

l is selected from-C (O) NH-, -CH₂NHC(O)-、-NHC(O)-、-O-、-NH-、-S-、-C(O)O-、-OC(O)-、-S(O) ₂O-or-OS (O)₂-；

R ²Independently selected from halogen, -OH, -NH₂、-CN、C _1-6Alkyl or C_1-6Alkoxy radical, said C_1-6Alkyl or C_1-6Alkoxy is optionally substituted with halogen;

m is selected from 0, 1,2,3 or 4;

is selected from
Wherein the content of the first and second substances,

X ¹independently selected from-NH-, -O-, -S-or-CH₂-；

n1 is selected from 1,2,3,4, 5 or 6;

n2 and n3 are each independently selected from 1,2 or 3;

n4 is selected from 0, 1,2 or 3;

R ³and R⁵Each independently selected from ═ O, halogen, -OH, -NH₂、-CN、C _1-6Alkyl radical, C_1-6Alkoxy radical, C_1-6Alkylamino radical, C_1-6Alkyl S-, -C (O) OR, -C (O) NHR OR-S (O)₂R, wherein R is independently selected from-H, C_1-6Alkyl radical, C_2-6Alkenyl radical, C_2-6Alkynyl, C_3-6Cycloalkyl, 3-6 membered heterocycloalkyl, 5-6 membered heteroaryl or C_6-10An aryl group;

q1, q2, q3 and q4 are each independently selected from 0, 1,2 or 3;

X ²is selected from N or CH;

X ³is selected from-CH₂-, -O-, -S-or-NH-.
A compound of formula (I) or a pharmaceutically acceptable salt thereof as claimed in claim 1 wherein ring B is selected from 5-10 membered heteroaryl or phenyl; or ring B is selected from 5-6 membered heteroaryl or phenyl; or ring B is selected from pyridyl or phenyl.
A compound of formula (I) or (la) as claimed in claims 1-2A pharmaceutically acceptable salt thereof, wherein R¹Independently selected from halogen, -CN, C_1-6Alkoxy, or C optionally substituted by halogen_1-6An alkyl group; or R¹Independently selected from-F, -Cl, -Br, -I, -CN, C_1-3Alkoxy, or C optionally substituted by halogen_1-3An alkyl group; or R¹Independently selected from-F, -Cl, -CN, methoxy, methyl or-CF₃。
A compound of formula (I), or a pharmaceutically acceptable salt thereof, as claimed in any one of claims 1-3 wherein p is selected from 0, 1 or 2; or p is selected from 0or 1.
A compound of formula (I) or a pharmaceutically acceptable salt thereof as claimed in any one of claims 1 to 4 wherein L is selected from-C (O) NH-, -CH₂NHC (O) -, -NHC (O) -or-O-; or L is selected from-C (O) NH-, -CH₂NHC (O) -or-O-.
A compound of formula (I), or a pharmaceutically acceptable salt thereof, as claimed in any one of claims 1-5 wherein R²Independently selected from halogen, -OH, -NH₂、-CN、C _1-3Alkyl or C_1-3Alkoxy radical, said C_1-3Alkyl or C_1-3Alkoxy is optionally substituted with halogen; or R²Independently selected from-F, -Cl, -Br, -I, -OH, -NH₂、-CN、-CH ₃or-CF₃(ii) a Or R²Independently selected from-F, -Cl, -Br or-I; or R²Independently selected from-F or-Cl.
A compound of formula (I), or a pharmaceutically acceptable salt thereof, as claimed in any one of claims 1-6 wherein m is selected from 0, 1 or 2; or m is selected from 0or 1.
A compound of formula (I) as claimed in any one of claims 1 to 7 or a pharmaceutical thereofThe above acceptable salt, wherein X¹Independently selected from-NH-, -O-or-CH₂-; or when n1 is 2,3,4, 5 or 6, one X therein¹Selected from-NH-, -O-, -S-or-CH₂-, wherein the H atom in-NH-is optionally substituted by C_1-3Alkyl substituted, the remainder of X¹is-CH₂-。
A compound of formula (I), or a pharmaceutically acceptable salt thereof, as claimed in any one of claims 1-8 wherein n1 is selected from 1,2,3 or 4; or n1 is selected from 2,3 or 4.
A compound of formula (I), or a pharmaceutically acceptable salt thereof, as claimed in any one of claims 1-9 wherein n2 is selected from 1 or 2, n3 is selected from 2 or 3; or both n2 and n3 are 2; or n2 is 1 and n3 is 2 or 3.
A compound of formula (I), or a pharmaceutically acceptable salt thereof, as claimed in any one of claims 1-10 wherein n4 is selected from 0, 1 or 2; or n4 is selected from 0or 1; or n4 is selected from 1.
A compound of formula (I), or a pharmaceutically acceptable salt thereof, as claimed in any one of claims 1-11 wherein R³Independently selected from ═ O, halogen, -OH, -NH₂、-CN、C _1-3Alkyl radical, C_1-3Alkoxy radical, C_1-3Alkylamino radical, C_1-3Alkyl S-, -C (O) OR, -C (O) NHR OR-S (O)₂R; or R³Selected from ═ O, halogen, -OH, -NH₂-CN or C_1-3An alkyl group; or R³Independently selected from ═ O or C_1-3An alkyl group; or R³Selected from ═ O or methyl.
A compound of formula (I), or a pharmaceutically acceptable salt thereof, as claimed in any one of claims 1-12 wherein R is independently selected from H, C_1-3Alkyl radical, C_2-3Alkenyl radical, C_2-3Alkynyl, C_3-6Cycloalkyl, 3-6 membered heterocycloalkyl, 5-6 membered heteroaryl or phenyl; or R is independently selected from-H, C_1-3Alkyl radical, C_2-3Alkenyl or C_2-3Alkynyl.
A compound of formula (I), or a pharmaceutically acceptable salt thereof, as claimed in any one of claims 1-13 wherein
Selected from the following structures:
a compound of formula (I), or a pharmaceutically acceptable salt thereof, as claimed in any one of claims 1-13 wherein
Is selected from
A compound of formula (I), or a pharmaceutically acceptable salt thereof, as claimed in any one of claims 1-13 wherein
Is composed of
A compound of formula (I) or a pharmaceutically acceptable salt thereof as claimed in any one of claims 1 to 16 selected from a compound of formula (II-1), formula (II-2) or formula (II-3) or a pharmaceutically acceptable salt thereof,
a compound of formula (I) as claimed in claim 1, or a pharmaceutically acceptable salt thereof, selected from the following compounds, or pharmaceutically acceptable salts thereof:
a pharmaceutical composition comprising a compound of any one of claims 1-18, or a pharmaceutically acceptable salt thereof.
A method of treating a BTK-related disorder in a mammal, comprising administering to a mammal in need thereof a therapeutically effective amount of a compound of any one of claims 1-18, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition of claim 19; preferably, the BTK-related disease is selected from an autoimmune disease, an inflammatory disease or cancer.