WO2016154998A1 - Pyrazolopyrimidine derivative, manufacturing method, pharmaceutical composition, and use thereof - Google Patents

Abstract

Disclosed are a pyrazolopyrimidine derivative, manufacturing method, pharmaceutical composition, and use thereof. Provided are a pyrazolopyrimidine derivative as represented by formula I, a stereoisomer, a solvate, a pharmaceutically acceptable salt, an active metabolite or a prodrug thereof. The pyrazolopyrimidine derivative as represented by formula I in the present invention has a good inhibitory activity with respect to a Bruton's tyrosine kinase (Btk), and particularly has a favorable in-vitro and in-vivo inhibit activity with respect to the growth of tumor cells. Thereby, the present invention has a good prospect of commercial application.

Description

Pyrazolopyrimidine derivative, preparation method, pharmaceutical composition and use thereof Technical field

The invention particularly relates to pyrazolopyrimidine derivatives, methods of preparation, pharmaceutical compositions and uses.

Background technique

Bruton's tyrosine kinase (Btk), a member of the Tec family of non-receptor tyrosine kinases, is key for expression in all hematopoietic cell types except T lymphocytes and natural killer cells. Signal enzyme. Btk plays a crucial role in the B cell signaling pathway that links cell surface B-cell receptor (BCR) to downstream cell responses.

Btk is a key regulator of B cell development, activation, signaling and survival (Kurosaki, Curr Op Imm, 2000, 276-281; Schaeffer and Schwartzberg, Curr Op Imm 2000, 282-288). In addition, Btk plays a role in numerous other hematopoietic signaling pathways, such as Toll like receptor (TLR) and cytokine receptor-mediated TNF-α production in macrophages, in mast cells. Immunoglobulin E receptor (FcεRI) signaling, signaling that inhibits apoptosis of Fas/APO-1 cells in B-lineage lymphoid cells, and collagen-stimulated platelet aggregation. See, for example, CA Jeffries et al (2003), Journal of Biological Chemistry 278: 26258-26264; NJ Horwood et al (2003), The Journal of Experimental Medicine 197: 1603-1611; Iwaki et al (2005), Journal of Biological Chemistry 280 (48): 40261 -40270; Vassilev et al. (1999), Journal of Biological Chemistry 274(3): 1646-1656; Quek et al. (1998), Current Biology 8(20): 1137-1140.

At present, BTK targeting inhibitors have been used in tumor therapy, but their inhibitory activity and efficacy are not significant enough. Clinically, larger doses are needed. Common side effects include thrombocytopenia, diarrhea, neutropenia, anemia and Upper respiratory tract infections, etc., and there is still a need to further develop compounds with better activity or pharmacological properties as newer, improved or more efficient BTK receptor inhibitors, thus enabling a deeper understanding of these drugs and BTK targets. The relationship between proteins and their anti-tumor mechanism are of great significance for the clinical treatment of tumors.

Summary of the invention

The technical problem to be solved by the present invention is to provide a pyrazolopyrimidine derivative in order to overcome the defects in the prior art that Bruton's tyrosine kinase inhibitor has insufficient biological activity and efficacy, large dosage, and side effects. , preparation methods, pharmaceutical compositions and uses. The pyrazolopyrimidine derivative of the present invention as shown in Formula I against Bruton's tyrosine The kinase (BTK) has good inhibitory activity, especially has good in vitro and in vivo inhibitory activity on the growth of tumor cells, and has a good market prospect.

The inventors have found through intensive studies that the pyrazolopyrimidine derivatives of the structure of formula I have an effective irreversible inhibitory effect on Btk.

The present invention provides a pyrazolopyrimidine derivative of the formula I, a stereoisomer, a solvate thereof, a pharmaceutically acceptable salt, an active metabolite or a prodrug,

或

当L为O、S、

或

时，Z是

或

当L为

或

时，Z为

或

其中，X是卤素原子(例如氟原子、氯原子或溴原子)，氰基，

或

R为C₁～C₄的烷基(所述的“C₁～C₄的烷基”例如甲基、乙基、丙基、异丙基、丁基、异丁基或叔丁基，优选甲基)；Where: L is O, S,

or

When L is O, S,

or

When Z is

or

When L is

or

When Z is

or

Wherein X is a halogen atom (for example, a fluorine atom, a chlorine atom or a bromine atom), a cyano group,

or

R is a C ₁ -C ₄ alkyl group (the "C ₁ -C ₄ alkyl group" such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl or tert-butyl, preferably methyl);

或

所述的“被一个或多个甲基取代的苯基”例如4-甲基苯基；所述的“被一个或多个氰基取代的苯基”例如2-氰基苯基；所述的“被一个或多个三氟甲基取代的苯基”例如4-三氟甲基苯基；所述的“被一个或多个甲氧基取代的苯基”例如3-甲氧基苯基；所述的“被一个或多个乙烯基取代的苯基”例如

所述的“被一个或多个乙炔基取代的苯基”例如

所述的“被一个或多个苯基取代的苯基”例如

或者取代或未取代的杂芳基(所述的“取代或未取代的杂芳基”优选杂原子为氮原子、杂原子数为1-2个的取代或未取代的C₃～C₉的杂芳基；所述的“杂原子为氮原子、杂原子数为1-2个的取代或未取代的C₃～C₉的杂芳基”优选杂原子为氮原子、杂原子数为1个的取代或未取代的C₃～C₅的杂芳基，所述的“杂原子为氮原子、杂原子数为1个的取代或未取代的C₃～C₅的杂芳基”优选取代或未取代的吡啶基；所述的“未取代的吡啶基”例如

所述的“取代或未取代的芳基或者取代或未取代的杂芳基”中所述的“取代”是指被一个或多个选自卤素原子(例如氟原子、氯原子或溴原子)、氰基、取代或未取代的烷基(优选取代或未取代的C₁～C₄的烷基，所述的“未取代的烷基C₁～C₄的烷基”优选甲基，所述的“取代的C₁～C₄的烷基”优选三氟甲基)、烷氧基(优选C₁～C₄的烷氧基，所述的“C₁～C₄的烷氧基”优选甲氧基)、烯基(优选C₂～C₄的烯基，所述的“C₂～C₄的烯基”优选乙烯基)、炔基(优选C₂～C₄的炔基，所述的“C₂～C₄的炔基”优选乙炔基)和芳基(优选C₅～C₁₀的芳基，所述的“C₅～C₁₀的芳基”优选苯基)的取代基所取代，所述的“取代或未取代的烷基”中所述的“取代”是指被一个或多个卤素原子(例如氟原子、氯原子或溴原子)所取代(所述的“取代的烷基”优选三氟甲基)，当存在多个取代基时，所述的取代基可以相同或不同； Ar is a substituted or unsubstituted aryl group (preferably "substituted or unsubstituted C ₅ -C ₁₀ aryl group"; the "substituted or unsubstituted C ₅ -C ₁₀ aryl group" is preferably substituted or unsubstituted Phenyl; said "substituted phenyl" may be phenyl substituted by one or more fluorine atoms, phenyl substituted by one or more methyl groups, phenyl substituted by one or more cyano groups a phenyl substituted by one or more trifluoromethyl groups, a phenyl group substituted by one or more methoxy groups, a phenyl group substituted by one or more vinyl groups, a benzene substituted by one or more ethynyl groups a phenyl group substituted by one or more phenyl groups; said "phenyl group substituted by one or more fluorine atoms", for example

or

The "phenyl substituted by one or more methyl groups" such as 4-methylphenyl; the "phenyl substituted by one or more cyano groups" such as 2-cyanophenyl; "Phenyl substituted by one or more trifluoromethyl groups" such as 4-trifluoromethylphenyl; said "phenyl substituted by one or more methoxy groups" such as 3-methoxybenzene The "phenyl group substituted by one or more vinyl groups", for example

Said "phenyl substituted by one or more ethynyl groups", for example

Said "phenyl substituted by one or more phenyl groups", for example

Or a substituted or unsubstituted heteroaryl group (the "substituted or unsubstituted heteroaryl group" preferably has a hetero atom of a nitrogen atom and a substituted or unsubstituted C ₃ -C ₉ hetero atomic number of 1-2 a heteroaryl group; the "hetero atom is a nitrogen atom, and the number of hetero atoms is 1-2, or the unsubstituted C ₃ - C ₉ heteroaryl group" preferably has a hetero atom of a nitrogen atom and a hetero atom number of 1 The substituted or unsubstituted C ₃ -C ₅ heteroaryl group, wherein the "hetero atom is a nitrogen atom and the number of hetero atoms is one or the unsubstituted C ₃ - C ₅ heteroaryl group" is preferably Substituted or unsubstituted pyridyl; said "unsubstituted pyridyl", for example

The term "substituted" as used in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means one or more selected from a halogen atom (for example, a fluorine atom, a chlorine atom or a bromine atom). a cyano group, a substituted or unsubstituted alkyl group (preferably a substituted or unsubstituted C ₁ -C ₄ alkyl group, and the "unsubstituted alkyl C ₁ -C ₄ alkyl group" is preferably a methyl group. The "substituted C ₁ -C ₄ alkyl group" is preferably a trifluoromethyl group, an alkoxy group (preferably a C ₁ -C ₄ alkoxy group, and the "C ₁ -C ₄ alkoxy group"" Preferably, methoxy), alkenyl (preferably C ₂ -C ₄ alkenyl, said "C ₂ -C ₄ alkenyl" is preferably vinyl), alkynyl (preferably C ₂ -C ₄ alkynyl, The "C ₂ -C ₄ alkynyl group" is preferably an ethynyl group and an aryl group (preferably a C ₅ - C ₁₀ aryl group, and the "C ₅ - C ₁₀ aryl group" is preferably a phenyl group). The term "substituted" as used in the "substituted or unsubstituted alkyl group" means substituted by one or more halogen atoms (for example, a fluorine atom, a chlorine atom or a bromine atom). Substituted alkyl group "preferably trifluoromethyl", when stored When a plurality of substituents, the substituents may be the same or different;

V is hydrogen or a fluorine atom;

或

)、环烷基(优选C₄～C₇的环烷基，所述的“C₄～C₇的环烷基”优选C₅～C₆的环烷基；所述的“C₆的环烷基”例如环己基)或杂环烷基(优选杂原子为氧、硫或氮原子、杂原子数为1-3个的C₄～C₇的杂环烷基，所述的“杂原子为氧、硫或氮原子、杂原子数为1-3个的C₄～C₇的杂环烷基”优选杂原子为氮原子、杂原子数为1-2个的C₄～C₅的杂环烷基；所述的“杂原子为氮原子、杂原子数为1-2个的C₄～C₅的杂环烷基”优选吡咯烷基或者哌啶基；所述的“吡咯烷基”优选

或

所述的“哌啶基”优选

或

)；所述的“取代或未取代的亚烷基”中所述的取代是指被“杂原子为氧、硫或氮原子、杂原子数为1-3个的C₄～C₇的杂环烷基”(优选杂原子为氮原子、杂原子数为1-2个的C₄～C₅的杂环烷基，所述的“杂原子为氮原子、杂原子数为1-2个的C₄～C₅的杂环烷基”为吡咯烷基)、C₁～C₆的烷氧基、C₁～C₆的烷硫基和C₁～C₆的烷氨基中的一个或多个所取代。Y is a substituted or unsubstituted alkylene group (preferably a substituted or unsubstituted C ₁ -C ₆ alkylene group, and the "substituted or unsubstituted C ₁ -C ₆ alkylene group" is preferably substituted or not The substituted methylene group, the "substituted methylene group" is preferably a pyrrolidinyl-substituted methylene group; the "pyrrolidinyl-substituted methylene group" is, for example,

or

a cycloalkyl group (preferably a C ₄ -C ₇ cycloalkyl group, said "C ₄ -C ₇ cycloalkyl group" preferably a C ₅ -C ₆ cycloalkyl group; said "C ₆ ring" An alkyl group such as a cyclohexyl group or a heterocycloalkyl group (preferably a hetero atom having a hetero atom of an oxygen, sulfur or nitrogen atom, a C _1-4 C ₇ hetero atom having a number of 1-3), said "hetero atom" a C ₄ -C ₇ heterocycloalkyl group which is an oxygen, sulfur or nitrogen atom and has 1 to 3 hetero atoms" preferably has a hetero atom of a nitrogen atom and a C ₄ - C ₅ hetero atomic number of 1-2 a heterocycloalkyl group; the "heteroatom is a nitrogen atom, a C ₄ -C ₅ heterocycloalkyl group having 1-2 hetero atoms" is preferably a pyrrolidinyl group or a piperidinyl group; Base

or

Said "piperidinyl" is preferred

or

The substitution described in the "substituted or unsubstituted alkylene group" means a C ₄ to C ₇ hetero atom which is a hetero atom which is an oxygen, sulfur or nitrogen atom and has 1-3 hetero atoms. a cycloalkyl group (preferably, the hetero atom is a nitrogen atom, a C ₄ -C ₅ heterocycloalkyl group having 1-2 hetero atoms, and the "hetero atom is a nitrogen atom, and the number of hetero atoms is 1-2" One or more of a C ₄ -C ₅ heterocycloalkyl group "pyrrolidinyl group", a C ₁ -C ₆ alkoxy group, a C ₁ -C ₆ alkylthio group or a C ₁ -C ₆ alkylamino group or Multiple replaced.

或

进一步优选O、S和

In the present invention, the L is preferably O, S,

or

Further preferred O, S and

乙炔基

和苯基的取代基所取代；“被一个或多个氟原子取代的苯基”例如

或

“被一个或多个甲基取代的苯基”例如4-甲基苯基；“被一个或多个氰基取代的苯基”例如2-氰基苯基；“被一个或多个三氟甲基取代的苯基”例如4-三氟甲基苯基；“被一个或多个甲氧基取代的苯基”例如3-甲氧基苯基；“被一个或多个乙烯基取代的苯基”例如

“被一个或多个乙炔基取代的苯基”例如

“被一个或多个苯基取代的苯基”例如

In the present invention, the Ar is preferably a substituted or unsubstituted aryl group, and the "substituted or unsubstituted aryl group" is preferably a "substituted or unsubstituted C ₅ - C ₁₀ aryl group"; The substituted or unsubstituted C ₅ -C ₁₀ aryl group" is preferably a substituted or unsubstituted phenyl group; the "substituted phenyl group" is preferably one or more selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, and three Fluoromethyl, cyano, methyl, methoxy, vinyl

Ethynyl

Substituted with a substituent of a phenyl group; "phenyl group substituted by one or more fluorine atoms", for example

or

"Phenyl substituted by one or more methyl groups" such as 4-methylphenyl; "phenyl substituted by one or more cyano groups" such as 2-cyanophenyl; "by one or more trifluoro Methyl substituted phenyl" such as 4-trifluoromethylphenyl; "phenyl substituted by one or more methoxy" such as 3-methoxyphenyl; "substituted by one or more vinyl groups"Phenyl" for example

"Phenyl substituted by one or more ethynyl groups", for example

"Phenyl substituted by one or more phenyl groups", for example

In the present invention, the V is preferably hydrogen.

或

)或杂环烷基(优选杂原子为氧、硫或氮原子、杂原子数为1-3个的C₄～C₇的杂环烷基，所述的“杂原子为氧、硫或氮原子、杂原子数为1-3个的C₄～C₇的杂环烷基”优选杂原子为氮原子、杂原子数为1-2个的C₄～C₅的杂环烷基；所述的杂原子为氮原子、杂原子数为1-2个的C₄～C₅的杂环烷基优选吡咯烷基或者哌啶基；所述的“吡咯烷基”优选

或

所述的“哌啶基”优选

或

)。In the present invention, Y is preferably a substituted or unsubstituted alkylene group (preferably a substituted or unsubstituted C ₁ -C ₆ alkylene group, said "substituted or unsubstituted C ₁ -C ₆ sub- The alkyl group is preferably a substituted or unsubstituted methylene group, and the "substituted methylene group" is preferably a pyrrolidinyl-substituted methylene group; the "pyrrolidinyl-substituted methylene group" is, for example,

or

Or a heterocycloalkyl group (preferably a C ₄ -C ₇ heterocycloalkyl group having a hetero atom of oxygen, sulfur or nitrogen, 1-3 hetero atoms, said "hetero atom" being oxygen, sulfur or nitrogen a C ₄ -C ₇ heterocycloalkyl group having 1 to 3 atoms and a hetero atom is preferably a C ₄ -C ₅ heterocycloalkyl group having a nitrogen atom and a number of 1-2 hetero atoms; The C ₄ -C ₅ heterocycloalkyl group in which the hetero atom is a nitrogen atom and the number of hetero atoms is 1-2 is preferably a pyrrolidinyl group or a piperidinyl group; and the "pyrrolidinyl group" is preferably used.

or

Said "piperidinyl" is preferred

or

).

或

时，Z优选为

其中，X优选Cl、Br或CN，Y优选吡咯烷基或者哌啶基；所述的“吡咯烷基”优选

或

所述的“哌啶基”优选

或

)。In the present invention, when L is O, S,

or

When Z is preferred

Wherein X is preferably Cl, Br or CN, and Y is preferably pyrrolidinyl or piperidinyl; said "pyrrolidinyl" is preferred

or

Said "piperidinyl" is preferred

or

).

或

时，Z优选为

或

其中，X优选Cl、Br或CN，Y优选吡咯烷基或者哌啶基；所述的“吡咯烷基”优选

或

所述的“哌啶基”优选

或

)。In the present invention, when L is

or

When Z is preferred

or

Wherein X is preferably Cl, Br or CN, and Y is preferably pyrrolidinyl or piperidinyl; said "pyrrolidinyl" is preferred

or

Said "piperidinyl" is preferred

or

).

In the present invention, the pyrazolopyrimidine derivative of the formula I is preferably a compound of the formula I-A:

Among them, the definitions of Ar, L, Y, Z and V are the same as described above.

In the present invention, the pyrazolopyrimidine derivative represented by Formula I is further preferably one of the following compounds:

(R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl ]-2-bromoprop-2-en-1-one (compound 1);

(R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl ]-2-chloroprop-2-en-1-one (compound 2);

(R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl ]-2-fluoroprop-2-en-1-one (compound 3);

(R,E)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine-1 -yl]-4,4,4-trifluoro-but-2-en-1-one (compound 4);

(R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl ]-2-trifluoromethylprop-2-en-1-one (compound 5);

(R,Z)-4-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine-1 -yl]-4-oxobut-2-enyl cyanide (compound 6);

(R,E)-4-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine-1 -yl]methyl-4-oxobut-2-enoate (compound 7);

(R,E)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine-1 -yl]-3-chloroprop-2-en-1-one (compound 8);

(R,Z)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine-1 -yl]-3-bromoprop-2-en-1-one (compound 9);

(R,E)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine-1 -yl]-3-bromoprop-2-en-1-one (compound 10);

(R,E)-4-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine-1 -yl]-4-oxobut-2-enamide (Compound 11);

(R)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl]tetrahydrol Pyrrol-1-yl]-2-fluoroprop-2-en-1-one (compound 12);

(R)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl]tetrahydrol Pyrrol-1-yl]-2-bromoprop-2-en-1-one (compound 13);

(R)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl]tetrahydrol Pyrrol-1-yl]-2-chloroprop-2-en-1-one (compound 14);

(R,E)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl] Tetrahydropyrrol-1-yl]-4,4,4-trifluorobut-2-en-1-one (Compound 15);

(R,E)-4-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl] Methyl tetrahydropyrrol-1-yl]-4-oxobut-2-enoate (Compound 16);

(R,Z)-4-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl] Tetrahydropyrrol-1-yl]-4-oxobut-2-enyl cyanide (Compound 17);

(S)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl]tetrahydrol Pyrrol-1-yl]-2-fluoroprop-2-en-1-one (compound 18);

(S)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl]tetrahydrol Pyrrol-1-yl]-2-chloroprop-2-en-1-one (Compound 19);

(S)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl]tetrahydrol Pyrrol-1-yl]-2-bromoprop-2-en-1-one (compound 20);

(S,E)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl] Tetrahydropyrrol-1-yl]-4,4,4-trifluorobut-2-en-1-one (Compound 21);

(S,E)-4-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl] Methyl tetrahydropyrrol-1-yl]-4-oxobut-2-enoate (Compound 22);

(S,Z)-4-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl] Tetrahydropyrrole-1- 4-oxobut-2-enyl cyanide (compound 23);

(S,E)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl] Tetrahydropyrrol-1-yl]-4-(dimethylamino)but-2-en-1-one (Compound 24);

(R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl ]prop-2-en-1-one (compound 25);

(R)-[4-[4-Amino-1-(1-(vinylsulfone)piperidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl]phenyl] (phenyl)methanone (compound 26);

(R,E)-4-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine-1 -yl]-4-oxobut-2-enyl cyanide (Compound 27);

(R)-1-[3-[4-Amino-3-(4-(difluoro(phenyl)methyl)phenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl Piperidin-1-yl]prop-2-en-1-one (compound 28);

(R)-3-[4-(difluoro(phenyl)methyl)phenyl]-1-(1-(vinylsulfone)piperidin-3-yl)-1H-pyrazolo[3,4- d] pyrimidine-4-ammonia (compound 29);

[(R)-3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl]( (R)-oxiran-2-yl)methanone (compound 30);

[(R)-3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl]( (S)-oxiran-2-yl)methanone (Compound 31);

(R)-1-[3-[4-Amino-3-(4-phenylthiophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl ]-2-bromoprop-2-en-1-one (compound 32);

(R)-1-[3-[4-Amino-3-(4-benzylphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl] -2-bromoprop-2-en-1-one (compound 33);

(R)-1-[3-[4-Amino-3-(4-(4-fluorophenoxy)phenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidin Pyridin-1-yl]-2-bromoprop-2-en-1-one (compound 34);

[(R)-3-(4-Amino-3-(3-fluoro-4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)tetrahydropyrrole- 1-yl]((S)-oxiran-2-yl)methanone (compound 35);

(R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]tetrahydropyrrole-1- 2-bromoprop-2-en-1-one (compound 36);

(R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]tetrahydropyrrole-1- 2-chloroprop-2-en-1-one (compound 37);

(R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]tetrahydropyrrole-1- 2-fluoroprop-2-en-1-one (compound 38);

(R,Z)-4-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]tetrahydropyrrole- 1-yl]-4-oxobut-2-enyl cyanide (compound 39);

[(R)-3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)tetrahydropyrrole-1-yl] ((R)-oxiran-2-yl)methanone (Compound 40);

(S,E)-1-[3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)tetrahydropyrrole- 1-yl]-3-chloroprop-2-en-1-one (Compound 41);

(S,Z)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]tetrahydropyrrole- 1-yl]-3-bromoprop-2-en-1-one (compound 42);

(R)-1-[3-[4-Amino-3-(4-(pyridin-4-yloxy)phenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidin Pyridin-1-yl]-2-bromoprop-2-en-1-one (compound 43);

(S)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl ]-2-bromoprop-2-en-1-one (compound 44);

(R)-1-[3-[4-Amino-3-(4-p-tolyloxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]-piperidine-1 -yl]-2-bromoprop-2-en-1-one (compound 45);

(R)-2-[4-[4-Amino-1-[1-(2-bromoacryloyl)piperidin-3-yl]-1H-pyrazolo[3,4-d]pyrimidine-3- Phenyloxy]benzonitrile (Compound 46);

(R)-1-[-3-[4-amino-3-(4-(4-trifluoromethylphenoxy)phenyl)-1H-pyrazolo[3,4-d]pyrimidine-1 -yl]piperidin-1-yl]-2-bromoprop-2-en-1-one (compound 47);

(R)-1-[-3-[4-amino-3-(4-(4-vinylphenoxy)phenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl Piperidin-1-yl]-2-bromoprop-2-en-1-one (Compound 48);

(R)-1-[-3-[4-amino-3-(4-(4-ethynylphenoxy)phenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl Piperidin-1-yl]-2-bromoprop-2-en-1-one (Compound 49);

(R)-1-[3-[4-Amino-3-(4-(3-methoxyphenoxy)phenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl Piperidin-1-yl]-2-bromoprop-2-en-1-one (Compound 50);

(R)-1-[3-[4-Amino-3-(4-(biphenyl-4-oxy)phenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl] Piperidin-1-yl]-2-bromoprop-2-en-1-one (Compound 51);

(R)-1-[3-[4-Amino-3-(4-(3,4-difluorophenoxy)phenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- Peptidin-1-yl]-2-bromoprop-2-en-1-one (compound 52);

(R)-1-[3-[4-Amino-3-(4-(2,3-difluorophenoxy)phenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- Methylpiperidin-1-yl]-2-bromoprop-2-en-1-one (compound 53);

(R)-1-[3-[4-Amino-3-(4-(2,4-difluorophenoxy)phenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- Piperidine-1- 2-bromoprop-2-en-1-one (compound 54);

Its specific structure is as follows:

In the present invention, the term "alkyl" is a branched or straight-chain saturated aliphatic hydrocarbon group having the specified number of carbon atoms; as defined in "C ₁ -C ₁₀ alkyl" as being included in a straight chain or A group having 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 carbon atoms in the branched structure. For example, "C ₁ -C ₁₀ alkyl" specifically includes methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl, isobutyl, pentyl, hexyl, heptyl, octyl,壬基和癸基, etc.

In the present invention, the term "alkoxy" means a group formed by linking an alkyl group to an oxygen atom, that is, "RO-", and R is an alkyl group.

In the present invention, the term "alkylthio" means a group formed by linking an alkyl group to a sulfur atom, that is, "RS-", and R is an alkyl group.

In the present invention, the term "alkylamino" means an amino group in which one hydrogen in "NH ₃ " is substituted with an alkyl group.

In the present invention, the term "alkylene" (including when used alone or in another group) means a branched and straight-chain subsaturated aliphatic hydrocarbon group having 1 to 20 carbon atoms, preferably 1 ～10 carbon atoms, more preferably 1-8 carbon atoms, such as methylene, ethylene, propylene, isopropylidene, n-butylene, tert-butyl, isobutylene, pentylene Base, hexylene group, heptylene group, octylene group, fluorenylene group, fluorenylene group, arylene (4,4-dimethylpentyl), sub (2,2,4-trimethylpentyl), sub Undecyl, dodecylene, and various isomers thereof.

In the present invention, the term "cycloalkyl" means an all-carbon monocyclic or polycyclic group, preferably 1 to 3 cyclic cycloalkyl groups formed by 3 to 20 carbons, more preferably 3 to 10 carbons. , for example: cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, Cycloheptyl, cyclooctyl, cyclodecane or cyclododecyl.

In the present invention, the term "heterocycloalkyl" as used herein alone or as part of another group, refers to one or more of 1 to 4 heteroatoms (such as nitrogen, oxygen and sulfur). A 4 to 12 membered monocyclic or polycyclic group wherein each ring may contain one or more double bonds, but none of the rings have a fully conjugated pi-electron system. Furthermore, any heterocycloalkyl ring can be fused to a cycloalkyl, aryl, heteroaryl or heterocycloalkyl ring. Heterocycloalkyl groups within the scope of this definition include, but are not limited to, oxazoline, oxocyclobutyl, pyranyl, tetrahydropyranyl, azetidinyl, 1,4-dioxyl, Hexahydropyrazine, piperazinyl, piperidinyl, pyrrolidinyl, morpholinyl, thiomorpholinyl, dihydrofuranyl, dihydroimidazolyl, indanyl, dihydroisoxazole , dihydroisothiazolyl, dihydrooxadiazolyl, dihydrooxazolyl, dihydropyrazinyl, dihydropyrazolyl, dihydropyridyl, dihydropyrimidinyl, dihydropyrrolyl, dihydrogen Tetrazolyl, dihydrothiadiazolyl, dihydrothiazolyl, dihydrothienyl, dihydrotriazolyl, dihydroazetidinyl, tetrahydrofuranyl and tetrahydrothiophenyl and their N-oxides. The heterocycloalkyl group may be bonded to other groups via a carbon atom or a hetero atom therein.

In the present invention, the term "alkenyl" means a straight-chain, branched or cyclic non-aromatic hydrocarbon group containing a specified number of carbon atoms and at least one carbon-carbon double bond. Preferably there is one carbon-carbon double bond and up to four non-aromatic carbon-carbon double bonds may be present. Thus, "C ₂ -C ₁₂ alkenyl" means an alkenyl group having 2 to 12 carbon atoms. The "C ₂ -C ₄ alkenyl group" means an alkenyl group having 2 to 4 carbon atoms, and includes a vinyl group, a propenyl group, a 2-methyl-propenyl group, a 1-butenyl group, and a 2-butenyl group. The linear, branched or cyclic moiety of the alkenyl group may contain a double bond.

In the present invention, the term "alkynyl" means a straight-chain, branched or cyclic hydrocarbon group containing a specified number of carbon atoms and at least one carbon-carbon triple bond. There may be up to three carbon-carbon triple bonds. Thus, "C ₂ -C ₁₂ alkynyl" means an alkynyl group having 2 to 12 carbon atoms. The "C ₂ -C ₄ alkynyl group" means an alkynyl group having 2 to 4 carbon atoms, and includes an ethynyl group, a propynyl group, a 1-butynyl group, a 2-butynyl group and the like.

In the present invention, the term "aryl" means any stable monocyclic or bicyclic carbon ring which may be up to 7 atoms in each ring, wherein at least one ring is an aromatic ring; examples of the above aryl unit include benzene Base, naphthyl, tetrahydronaphthyl, indanyl, biphenyl, phenanthryl, anthracenyl or acenaphthyl. It will be understood that in the case where the aryl substituent is a bicyclic substituent and one of the rings is a non-aromatic ring, the linkage is carried out through an aromatic ring.

In the present invention, the term "heteroaryl" means a stable monocyclic or bicyclic ring of up to 7 atoms in each ring, wherein at least one ring is an aromatic ring and contains from 1 to 4 selected from O, N, and Heteroatoms of S; heterocyclic aryl groups within the scope of this definition include, but are not limited to, acridinyl, oxazolyl, porphyrinyl, quinoxalinyl, pyrazolyl, indolyl, benzotriazolyl , furyl, thienyl, benzothienyl, benzofuranyl, quinolyl, isoquinolyl, oxazolyl, isoxazolyl, indolyl, pyrazinyl, pyridazinyl, pyridyl, Pyrimidinyl, pyrrolyl, tetrahydroquinoline. As defined by the following heterocyclic ring, "heterocyclic aryl" is also understood to include any nitrogen-containing heteroaryl N-oxide derivative. In its In the case where the heterocyclic aryl substituent is a bicyclic substituent and one ring is a non-aromatic ring or does not contain a hetero atom, it is understood that the linking is carried out through an aromatic ring or through a hetero atom containing a ring, respectively.

In the present invention, the determination of the carbon number range "C _x1 - _y1" alkyl (x1 and y1 is an integer), cycloalkyl, heterocycloalkyl, aryl and heteroaryl, such as "C _{1 -12} alkyl", each means a C _1-12 alkyl group which does not contain a substituent.

In the present invention, the term "halogen" means fluorine, chlorine, bromine, iodine or hydrazine.

In the present invention, the term "cyano" means

Any combination of the above groups is also contemplated herein for each variable. It will be appreciated that the substituents and substitution patterns on the compounds provided herein can be selected by one of ordinary skill in the art to provide compounds that are chemically stable and that can be synthesized using techniques known in the art and the techniques set forth herein. .

Compounds of any of the pyrazolopyrimidine derivatives of Formula I can be synthesized using standard synthetic techniques known to those skilled in the art or using synthetic methods known in the art in combination with the synthetic methods described herein. Additionally, the solvents, temperatures, and other reaction conditions presented herein can vary depending on the skill of the art. As a further guide, the following synthesis methods can also be utilized.

In certain embodiments, provided herein are methods of making pyrazolopyrimidine derivatives of Formula I and methods of use thereof. In certain embodiments, the compounds described herein can be synthesized using the schemes synthesized below, and the target compounds can be synthesized by selecting appropriate starting materials using methods analogous to those described below.

Starting materials for the synthesis of the compounds described herein can be synthesized or can be obtained from commercial sources. The compounds described herein, and other related compounds having different substituents, can be synthesized using techniques and starting materials known to those skilled in the art, for example, in Encyclopedia of Reagents for Organic Synthesis (Wiley 2009) by Paquette, "Advanced Organic Chemistry" by March. Fourth Edition, (Wiley 1992); Carey and Sundberg, Advanced Organic Chemistry, Fourth Edition, Volumes A and B (Plenum 2000, 2001); Green and Wuts, "Protective Groups in Organic Synthesis," Third Edition, ( Wiley 1999); Fieser and Fieser, Reagents for Organic Synthesis, Volumes 1-17 (John Wiley and Sons, 1991); Organic Reactions, Volumes 1-40 (John Wiley and Sons, 1991), and Larock's "Comprehensive" Organic Transformations (VCH Publishers Inc., 1999) (all of which are incorporated herein by reference). The general methods of preparing the compounds disclosed herein can be derived from reactions known in the art, and the reactions can be modified to synthesize the target compound molecules by reagents and conditions deemed appropriate by those skilled in the art. The following synthesis methods can be used as a basic guide.

After the end of each reaction, the reaction product can be worked up (isolated and purified) using conventional techniques including, but not limited to, filtration, distillation, crystallization, chromatography, and the like. These products can be characterized using conventional analytical methods, including Physical constants and map data (m.p, HPLC, LC-MS, NMR, optical rotation, etc.). The compounds described herein can be prepared as a single isomer using the synthetic methods described herein.

The present invention also provides a process for the preparation of the pyrazolopyrimidine derivative of the formula I, which comprises the following steps: coupling a compound II with an acid ZOH to obtain a pyrazole as shown in the formula I. a pyrimidine derivative;

Wherein, the definitions of Ar, L, Y, Z and V are the same as described above; the coupling reaction can employ conventional methods and conditions for such reactions in the art.

In the present invention, the method for preparing a pyrazolopyrimidine derivative of the formula I, further comprising the step of: deprotecting the compound III with an acid to obtain the compound II. ;

Wherein, Ar, L, Y and V are as defined above; PG is a protecting group and may be Boc (tert-butoxycarbonyl), CBz (benzyloxycarbonyl) or Troc (trichloroethoxycarbonyl), preferably Boc. (tert-butoxycarbonyl). The deprotection group can be subjected to conventional methods and conditions for such reactions in the art.

In the present invention, the method for producing a pyrazolopyrimidine derivative of the formula I, further comprising the steps of: subjecting the compound IV to the compound V by a Suzuki coupling reaction under palladium catalysis to obtain the Compound III can be used;

或

Wherein, the definitions of Ar, L, Y, V and PG are the same as defined above; R' and R" are each independently H or alkyl (for example, C ₁ -C ₄ alkyl, said "C ₁ -C" An alkyl group of ₄ , for example, methyl, ethyl, propyl, isopropyl, butyl, isobutyl or t-butyl; or R', R" and an oxygen atom or a boron atom therebetween form a ring Structure (for example

or

The Suzuki coupling can employ conventional methods and conditions for such reactions in the art.

In the present invention, the method for producing a pyrazolopyrimidine derivative of the formula I, further comprising the steps of: performing a Mitsunobu reaction of the compound VI with the compound V to obtain the compound IV;

Wherein, the definition of Y is the same as described above; the Mitsunobu reaction described may employ conventional methods and conditions for such reactions in the art.

In the present invention, the preparation method of the pyrazolopyrimidine derivative represented by Formula I, for example, Method 1 or Method 2,

Method 1 comprises the steps of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (Compound VI-1) via (Mi) or (S)-N-Boc- Coupling of 3-hydroxypiperidine with (R) or (S)-N-Boc-3-hydroxypyrrole affords Boc protected intermediate compound IV-1. Then, the appropriately substituted phenylboronic acid is subjected to palladium metal-catalyzed cross-coupling under basic conditions to form an intermediate compound III-1. After deprotection with an acid, it is coupled with Z acid to complete the synthesis to obtain the target compound I-1.

Method 2 comprises the steps of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (Compound VI-1) via Mitsunobu reaction with (R) or (S)-N-Boc-oxime Amino alcohol coupling gave Boc protected intermediate IV-2. Then, the appropriately substituted phenylboronic acid is subjected to palladium metal-catalyzed cross-coupling under basic conditions to form an intermediate III-2. After deprotection with an acid, it is coupled with Z acid to complete the synthesis to obtain the target compound I-2.

The pyrazolopyrimidine derivatives of Formula I disclosed herein are obtained in good yield and purity using the synthetic methods described herein, as well as those known in the art. Compounds prepared according to the methods disclosed herein are purified by conventional methods known in the art, such as filtration, recrystallization, chromatography, distillation, and combinations thereof.

The present invention also provides the use of any of the compounds of Formula I, or a pharmaceutically acceptable salt thereof, or a stereoisomer, solvate, active metabolite, and prodrug thereof, for the preparation of a medicament, It can be administered alone or in combination with other therapeutic agents. The medicament for use in the treatment and/or prevention of Bruton's tyrosine kinase (Btk) activity or for the benefit of Bruton's tyrosine kinase (Btk) activity in a mammal (including a human) A disease, condition, or condition that is inhibited. The medicament may be administered alone or in combination with other therapeutic agents. Combination drugs include, but are not limited to, doxorubicin, dactinomycin, bleomycin, vinblastine, cisplatin, acevicin; arubicin; apodazole hydrochloride; acronin; Aldileukin; hexamethylene melamine; ampomycin; aramid acetate; aziridine; ampicillin; anastrozole; acitretin; asparaginase; triamcinol; azacitidine Azatto; azamycin; bamastat; benzozide; bicalutamide; chlorinated hydrochloride; dimethyl sulfamate; diazepam; bleomycin sulfate; Buquina sodium; bromopyrimidine; busulfan; actinomycin C; carrazine; calamine; carbemide; carboplatin; carmustine; carbofuran hydrochloride; New; Westfinoco; chlorambucil; sirromycin; cladribine; clinidine methanesulfonate; cyclophosphamide; cytarabine; dacarbazine; daunorubicin hydrochloride; Hebin; right omalim; ligated sulphonic acid; sulphonic acid sulphonic acid; mantle; doxorubicin; doxorubicin hydrochloride; droloxifene; droloxifene citrate; Thomsonone Isomycin; edoxacin hydrochloride; effluranzine hydrochloride; elsamitrucin; enroplatin; enpyl urethane; epipirin; erbazole; ebisopic hydrochloride; estramustine; Estrostatin sodium phosphate; etalazol; etoposide; etoposide phosphate; chlorophenethylamine; carbazole hydrochloride; fazaradine; fenretinide; fluorouridine; Fluorouracil; fluoxetine; phosphaquinone; fisostatin sodium; gemcitabine; gemcitabine hydrochloride; hydroxyurea; idarubicin hydrochloride; ifosfamide; imofosin; interleukin II (including recombinant white blood cells) Interleukin II or rIL2), interferon alpha-2a; interferon alpha-2b; interferon alpha-n1; interferon alpha-n3; interferon beta-1a; interferon gamma-1b; isopropaplatin; irinotene hydrochloride Kang; lanreotide acetate; letrozole; leuprolide acetate; liazol hydrochloride; lometrozine sodium; lomustine; loxosporine hydrochloride; masobolol; maytansin; Mustard; megestrol acetate; melengestrol acetate; melphalan; menolide; guanidine; methotrexate; methotrexate sodium; chloramphenicol; metopril; Amine; mitoxin; lysin; mitoxanthin; mitoxanthin; mitomycin; mitoxantrone; mitoxantrone; mitoxantrone hydrochloride; mycophenolic acid; Azole; noramycin; omacil; oxicilin; pease; pelimycin; nemustine; pilomycin sulfate; phosphatidylamide; piperob bromide; Pyridoxine hydrochloride; pucamycin; promethate; porphyrin sodium; porphyrin; prednistatin; procarbazine hydrochloride; puromycin; puromycin hydrochloride; pyrazole furophenone; Lipo adenosine; 洛米米特; Safingo; Safingo hydrochloride; semustine; octretonide; sodium phosphatidyl aspartate; saponin; Spirostatin; streptavidin; streptozin; sulfochlorophenylurea; talimycin; tegacarb sodium; tegafur; tiloxime hydrochloride; temoporfin; teniposide; Teloxicillin; testosterone; thiomethamate, thioguanine, thiotepa; thiazolidine; tirapazamine; toremifene citrate; trodonone acetate, 7-norronone acetate; Trifluralin phosphate; trimethoate; trimethoate glucuronide; triptorelin; tobracil hydrochloride; uracil mustard; uridine; depeptin; verteporfin; vinblastine sulfate Vincristine sulfate; vindesine; vindesine sulfate; vinapidine sulfate; vinorelbine sulfate; vinpocet sulfate; vinorelbine; vinorelbine sulfate; vinorelbine sulfate; Platinum; statin; zorubicin hydrochloride, mitoxantrone, paclitaxel, propanol, thiotepa, angiogenesis inhibitor, camptothecin, dexamethasone, aspirin, acetaminophen, comparable Xin, ibuprofen, ketoprofen, meloxicam, corticosteroids and adrenocortical steroids.

In the present invention, the disease, disorder or condition which benefits from inhibition of Bruton's tyrosine kinase (Btk) activity includes, but is not limited to, cancer, inflammatory disease or disease state, immune disease or disease state, hyperplasia a disease or disease state and a degenerative disease or disease state.

In the present invention, the inflammatory disease or disease state, immune disease or disease state includes, but is not limited to, asthma, inflammatory bowel disease (including Crohn's disease and ulcerative colitis), appendicitis, orbital inflammation, bronchioles Inflammation, bronchitis, bursitis, cervicitis, cholangitis, cholecystitis, colitis, conjunctivitis, cystitis, lacrimal gland inflammation, dermatitis, dermatomyositis, encephalitis, endocarditis, endometritis, enteritis , enterocolitis, epicondylitis, epididymitis, fasciitis, fibrinitis, gastritis, gastroenteritis, hepatitis, suppurative sweat gland inflammation, laryngitis, mastitis, meningitis, myelitis, myocarditis, myositis, Nephritis, oophoritis, orchitis, osteitis, otitis, pancreatitis, mumps, pericarditis, peritonitis, pharyngitis, pleurisy, phlebitis, pneumonia or pneumonia, proctitis, prostatitis, pyelonephritis, rhinitis, Salpingitis, sinusitis, stomatitis, synovitis, tendonitis, tonsillitis, uveitis, vaginitis, vasculitis, vulvitis, HIV, adult respiratory distress syndrome, bone resorption, chronic obstructive pulmonary disease Chronic pneumonia, inflammatory skin disease, specific dermatitis, cystic fibrosis, septic shock, sepsis, endotoxic shock, hemodynamic shock, sepsis syndrome, reperfusion injury after ischemia, silver Psoriasis, fibrotic disease, cachexia, transplant rejection of graft versus host disease, autoimmune disease, xenogenic immune path or disease, radiation damage, asthma, oxygen-rich lung injury, microbial infection, and microbial infection syndrome.

The autoimmune diseases include, but are not limited to, rheumatoid arthritis, psoriatic arthritis, osteoarthritis, striata, juvenile arthritis, lupus, diabetes, myasthenia gravis, Hashimoto's thyroiditis, Des thyroiditis, Graves' disease, Sjogren's syndrome, multiple sclerosis, Guillain-Barré syndrome, acute disseminated encephalomyelitis, Addison's disease, strabismus syndrome, ankylosing spondylitis, Antiphospholipid antibody syndrome, aplastic anemia, autoimmune hepatitis, abdominal disease, Goodpasch's syndrome, idiopathic thrombocytopenic purpura, optic neuritis, scleroderma, primary biliary cirrhosis , Lytle's syndrome, high arteritis, temporal arteritis, Warm-antibody autoimmune hemolytic anemia, Wegener's granulomatosis, psoriasis, systemic hair loss, Becht's disease, chronic fatigue, autonomic dysfunction, endometriosis, interstitial bladder Inflammation, neuromuscular rigidity, scleroderma and vulvar pain.

The xenon immune condition or disease, including but not limited to graft versus host disease, transplantation, blood transfusion, allergic reaction, allergies (eg, for plant pollen, latex, drugs, food, insect toxins, animal hair, animal dander) , dust mites or sputum allergies), type I hypersensitivity, allergic conjunctivitis, allergic rhinitis and atopic dermatitis.

In the present invention, the cancer includes, but is not limited to, head cancer, thyroid cancer, neck cancer, eye cancer, skin cancer, oral cancer, throat cancer, esophageal cancer, chest cancer, bone cancer, blood cancer, bone marrow cancer, lung cancer, Colon cancer, sigmoid colon cancer, rectal cancer, stomach cancer, prostate cancer, breast cancer, ovarian cancer, kidney cancer, liver cancer, pancreatic cancer, brain cancer, intestinal cancer, heart cancer, adrenal cancer, subcutaneous tissue cancer, lymph node cancer, pigmentoma, Malignant glioma, B cell proliferative disease. The B cell proliferative diseases such as diffuse large B-cell lymphoma, follicular lymphoma, chronic lymphocytic lymphoma, chronic lymphocytic leukemia, B-cell pro-lymphocytic leukemia, lymphoplasmacytic lymphoma/Walden Stryst macroglobulinemia, spleen marginal lymphoma, plasma cell myeloma, plasmacytoma, extranodal marginal zone B-cell lymphoma, lymph node marginal zone B-cell lymphoma, mantle cell lymphoma, mediastinum (thymus Large B-cell lymphoma, intravascular large B-cell lymphoma, primary exudative lymphoma, Burkitt's lymphoma/leukemia or lymphomatoid granulomatosis.

The invention also provides a pharmaceutical composition comprising an active compound and, optionally, one or more pharmaceutically acceptable carriers or excipients; said active compound being as described in Formula I One or more of a pyrazolopyrimidine derivative, a pharmaceutically acceptable salt thereof, a stereoisomer thereof, a solvate, an active metabolite, and a prodrug.

The invention also provides a disease, disorder or condition for treating and/or preventing a mammal and for inhibiting Bruton's tyrosine kinase activity or for treating inhibition of Bruton's tyrosine kinase activity A method comprising the steps of: administering an effective amount of said pharmaceutical composition to a population in need thereof.

The pharmaceutical compositions may be formulated in a conventional manner using one or more pharmaceutically acceptable carriers including excipients and excipients which facilitate processing the active compound into a pharmaceutically acceptable formulation. The appropriate formulation will depend on the route of administration chosen. Any of the well-known techniques, carriers and excipients can be suitably employed and as understood in the art.

The pharmaceutical composition described in the present invention refers to a compound of any one of the compounds described herein and other chemical components such as a carrier, a stabilizer, an antioxidant, a disintegrant, a diluent, a dispersant, a filler, and a flavor. Mixtures of suspending agents, suspending agents, glidants, solubilizers, surfactants, wetting agents, thickening agents and/or excipients. The pharmaceutical composition facilitates administration of the active compound to an organism. In practicing the methods of treatment or use provided herein, Compositions A therapeutically effective amount of an active compound described herein is administered to a mammal having a disease, disorder or condition to be treated. Preferably, the mammal is a human. The therapeutically effective amount can vary widely, depending on the severity of the disease, the age and relative health of the subject, the efficacy of the compound employed, and the like. The active compound may be used alone or in combination with one or more therapeutic agents as a component of the mixture.

The pharmaceutical composition will comprise at least one of the pyrazolopyrimidine derivatives of the formula I described herein, in the form of the free acid or the free base, or as a pharmaceutically acceptable salt. Active ingredient. Additionally, the methods and pharmaceutical compositions described herein include the use of N-oxides, crystalline forms (also known as polymorphs), and active metabolites of these same type of pyrazolopyrimidine derivatives of Formula I. . In some cases, a pyrazolopyrimidine derivative as shown in Formula I can exist as a tautomer. All tautomers are included within the scope of the compounds provided herein. Additionally, the pyrazolopyrimidine derivatives of Formula I described herein can exist in unsolvated as well as solvated forms with pharmaceutically acceptable solvents such as water, ethanol, and the like. The solvated forms of the pyrazolopyrimidine derivatives as shown herein, as provided herein, are also considered to be disclosed herein.

The pharmaceutical compositions described herein can be administered to a subject by a variety of routes of administration including, but not limited to, oral, parenteral (eg, intravenous, subcutaneous, intramuscular), intranasal, buccal, Local, rectal or transdermal routes of administration. Dosage forms of the pharmaceutical compositions described herein include, but are not limited to, aqueous dispersions, self-emulsifying dispersions, solid solutions, liposome dispersions, aerosols, solid dosage forms, powders, immediate release formulations, controlled release formulations, instant solutions. Formulations, tablets, capsules, pills, sustained release preparations, extended release preparations, pulsatile release preparations, multiparticulate preparations, and mixed immediate and controlled release preparations.

Pharmaceutical preparations for oral administration can be obtained by mixing one or more solid excipients with one or more of the compounds described herein, optionally grinding the resulting mixture, if necessary, after adding suitable excipients The mixture is granulated to obtain a tablet or tablet core. Suitable excipients include, for example, fillers such as sugars, including lactose, sucrose, mannitol or sorbitol; cellulose preparations such as corn starch, wheat starch, rice starch, potato starch, gelatin, tragacanth, methyl Cellulose, microcrystalline cellulose, hydroxypropyl methylcellulose, sodium carboxymethylcellulose; and the like, such as polyvinylpyrrolidone (PVP or povidone) or calcium phosphate. If necessary, a disintegrating agent such as cross-linked croscarmellose sodium, polyvinylpyrrolidone, agar or alginic acid or a salt thereof such as sodium alginate may be added.

Pharmaceutical preparations which can be used orally include push-in capsules made of gelatin, and soft, sealed capsules made of gelatin and a plasticizer such as glycerol or sorbitol. The push-in capsules can contain the active ingredient in admixture in a filler such as lactose, a binder such as a starch and/or a lubricant such as talc or magnesium stearate, and optionally a stabilizer. In soft capsules, the active compound can be dissolved or suspended in a suitable liquid, such as a fatty oil, liquid paraffin or liquid polyethylene glycol. In addition, a stabilizer may be added. All formulations for oral administration should be in a form suitable for such administration.

In some embodiments, the solid dosage forms disclosed herein can be in the form of tablets (including suspension tablets, instant tablets, Chew disintegrating tablets, fast disintegrating tablets, effervescent tablets or capsules), pills, powders (including aseptically packaged powders, non-essential powders or effervescent powders), capsules (including soft capsules or Hard capsules, such as capsules made from gelatin of animal origin or capsules or "dispersed capsules" made from plant-derived HPMC), solid dispersions, solid solutions, bioerodible formulations, controlled release formulations, pulsatile release dosage forms, Multiparticulate formulations, pellets, granules or aerosols. In other embodiments, the pharmaceutical formulation is in powder form. In still other embodiments, the pharmaceutical formulation is in the form of a tablet, including but not limited to a fast dissolving tablet. Additionally, the pharmaceutical formulations described herein can be administered as a single capsule or as a multi-capsule dosage form. In some embodiments, the pharmaceutical formulation is administered in two, or three, or four capsules or tablets.

The pharmaceutical composition will comprise a formulation of at least one of the compounds described herein as any one of the pyrazolopyrimidine derivatives of Formula I, suitable for intramuscular, subcutaneous or intravenous injection, and may include physiologically acceptable sterile water. Or a non-aqueous solution, dispersion, suspension or emulsion, and sterile powder injection for reconstituting a sterile injectable solution or dispersion. Examples of suitable aqueous and nonaqueous vehicles, diluents, solvents or vehicles include water, ethanol, polyols (propylene glycol, polyethylene glycol, glycerol, cremophor, etc.), suitable mixtures thereof, vegetable oils (eg Olive oil) and organic esters for injection such as ethyl oleate. Appropriate fluidity can be maintained, for example by the use of a coating such as lecithin; by the maintenance of the required particle size; and by the use of surfactants. Formulations suitable for subcutaneous injection may also contain additives such as preservatives, wetting agents, emulsifying agents and dispersing agents. Prevention of microbial growth can be ensured by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, and the like. An isotonic agent such as sugar, sodium chloride or the like may also be included as needed. Prolonged absorption of the injectable pharmaceutical form can be brought about by the use of delayed absorption agents such as aluminum monostearate and gelatin.

The present application also relates to various pharmaceutical compositions known in the pharmaceutical arts for use in intraocular, intranasal, and intra-oral delivery. The pharmaceutical formulation comprises an aqueous ophthalmic solution of the active compound, which may be present in a water-soluble form such as an eye drop, or as a gellan gum or hydrogel; an ophthalmic ointment; an ophthalmic suspension, such as a microparticle, suspension. Small polymeric particles comprising a drug, a fat-soluble formulation, and microspheres; and an ophthalmic inserting agent in a liquid carrier medium. For stability and comfort, these suitable pharmaceutical preparations are most often and preferably manufactured as sterile, isotonic and buffered preparations. Pharmaceutical compositions also include drops and sprays which often mimic nasal secretions in a number of ways to ensure maintenance of normal ciliary action. As is well known to those skilled in the art, suitable formulations are most often and preferably isotonic, are lightly buffered to maintain a pH of 5.5 to 6.5, and most often and preferably include antimicrobial preservatives and appropriate drugs. stabilizer. Pharmaceutical formulations for intra-oral delivery include suspensions and ointments for topical application in the ear. Common solvents for these otic preparations include glycerin and water. It can be administered to a mammal, such as a human, by oral, parenteral (intravenous, intramuscular, subcutaneous, etc.), pulmonary, topical, dermal, and the like. The human dose of the compounds of the invention may range from about 0.1 mg to about 1000 mg.

The above preferred conditions can be arbitrarily combined without departing from the ordinary knowledge in the art, that is, preferred embodiments of the present invention.

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

In the present invention, room temperature means an ambient temperature of 10 ° C to 25 ° C.

The positive progress of the present invention is that the pyrazolopyrimidine derivative of the present invention has a good inhibitory effect on Bruton's tyrosine kinase and has good in vivo and in vitro inhibitory activity against tumor cell growth. Have a good market prospects.

Detailed ways

The invention is further illustrated by the following specific examples of the preparation of the compounds and the biological examples, which are intended to be used in more detail and are not intended to limit the invention in any way. The experimental methods in the following examples which do not specify the specific conditions are selected according to conventional methods and conditions, or according to the product specifications.

1. Compound preparation part:

Example 1: (R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine Synthesis of 1-yl]-2-bromoprop-2-en-1-one (Compound 1)

Synthesis of (R)-1-Boc-3-(4-amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidin-1-yl)piperidine

4-Amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidine (10 g, 38 mmol), (S)-1-Boc-3-hydroxypiperidine (17 g, 85 mmol), triphenyl Phosphine (20 g, 76 mmol) was added to a three-necked flask, THF (120 ml) was added, and the mixture was cooled to 0 ° C, and diisopropyl azodicarboxylate (DIAD) (15.2 g, 76 mmol) and tetrahydrofuran (THF) (30 ml) were added dropwise. The solution was added dropwise after about 1 h, and slowly warmed to room temperature overnight. The reaction solution was sparged, water was added, ethyl acetate was evaporated, dried, and concentrated, and then purified by column chromatography to afford product (R)-1-Boc-3-(4-amino-3-iodo-1H-pyrazolo[3, 4-D]pyrimidin-1-yl)piperidine (13.5 g, yield 80%). ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.38 (s, 1H), 6.02 (bs, 2H), 4.82-4.64 (m, 1H), 4.31-4.02 (m, 2H), 3.44-3.20 (m) , 1H), 2.95-2.65 (m, 1H), 2.25-2.08 (m, 2H), 1.95-1.58 (m, 2H), 1.42 (s, 9H).

b. Synthesis of 4-phenoxyphenylboronic acid

Dissolve 4-bromodiphenyl ether (100 g, 0.402 mol) in dry THF (800 mL), under nitrogen atmosphere, cooled to -78 ° C, and then add n-butyllithium (n-BuLi) (177 mL, 0.442 mol) The temperature was kept below -65 ° C. After stirring for 1 h, the temperature was maintained at -60 ° C, triisopropyl borate (90 g, 0.482 mol) was added dropwise, and the mixture was slowly warmed to 0 ° C and stirred for 3 h. Water (300 mL) was added to the ice bath and stirred overnight. The reaction mixture was concentrated to remove the organic phase, which was adjusted to pH 1-2 with 12N concentrated hydrochloric acid, and solids were isolated and filtered to afford product (77 g, yield 90%). ¹ H-NMR (d-DMSO, 400 MHz, δ ppm): 8.0 (s, 2H), 7.80 (d, J = 8.4 Hz, 2H), 7.41 (t, J = 7.6 Hz, 2H), 7.16 (t, J) = 7.2 Hz, 1H), 7.03 (d, J = 8.4 Hz, 2H), 6.94 (d, J = 8.4 Hz, 2H).

c. Synthesis of (R)-1-Boc-3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)piperidine

(R)-1-Boc-3-(4-Amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidin-1-yl)piperidine (12.8 g, 29 mmol) was added to a three-neck bottle 4-phenoxyphenylboronic acid (6.8 g, 32 mmol), PdCl ₂ (dppf) (0.5 g, 0.69 mmol), sodium carbonate (6.1 g, 58 mmol), 1,4-dioxane (160 ml) and After water (40 ml) was replaced with nitrogen, the mixture was heated to 80 ° C overnight. After confirming the completion of the reaction, the mixture was filtered, dried, evaporated, evaporated, evaporated, evaporated, evaporated. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.38 (s, 1H), 7.64 (d, J = 8.4 Hz, 2H), 7.42-7.38 (m, 2H), 7.24-7.04 (m, 5H), 6.18(br,2H),4.86-4.81(m,1H),4.18-4.04(m,1H),3.72-3.68(m,1H),3.18-3.01(m,1H),2.84-2.78(m,1H) ), 2.36-2.18 (m, 1.5H), 2.08-1.84 (m, 1.5H), 1.78-1.68 (m, 1H), 1.44 (s, 9H).

Synthesis of (R)-3-(4-phenoxyphenyl)-1-(piperidin-3-yl)-1H-pyrazolo[3,4-D]pyrimidine-4-ammonium

In the reaction flask, (R)-1-Boc-3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)piperidin The pyridine (8 g, 16.4 mmol) was added to a mixed solution of trifluoroacetic acid (40 ml) and dichloromethane (40 ml), and was allowed to react overnight at room temperature. After confirming the completion of the reaction, the mixture was neutralized with a saturated Na ₂ CO ₃ solution to pH 7-8, and then extracted with dichloromethane (DCM). The purified product (5.4 g, yield 85%) was analyzed. ¹ H-NMR (d-DMSO, 400 MHz, δ ppm): 8.25 (s, 1H), 7.67 (d, J = 8.4 Hz, 2H), 7.44 (t, J = 8.4 Hz, 2H), 7.17 to 7.30 (m) , 5H), 4.78 (m, 1H), 3.31 to 3.40 (m, 1H), 3.22 to 3.30 (m, 1H), 3.17 to 3.21 (m, 1H), 3.00 to 3.10 (m, 1H), 2.60 to 2.75 (m, 1H), 1.99 to 2.18 (m, 2H), 1.59 to 1.83 (m, 2H).

(R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl ]-2-bromoprop-2-en-1-one (compound 1)

(R)-3-(4-Phenoxyphenyl)-1-(piperidin-3-yl)-1H-pyrazolo[3,4-D]pyrimidine-4-ammonium (193 mg, 0.5 mmol) , 2-bromoacrylic acid (88 mg, 0.58 mmol), 1-hydroxybenzotriazole (HOBT) (95 mg, 0.7 mmol) and 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride The salt (EDCI) (134 mg, 0.7 mmol) eluted elute After adding ethyl acetate, the mixture was diluted with water and the mixture was washed twice with ethyl acetate. The organic phase was combined, dried over anhydrous sodium sulfate, and then purified by column chromatography to afford compound 1 (195 mg, yield 75%).

¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.38 (s, 1H), 7.64 (d, J = 8.4 Hz, 2H), 7.46-7.38 (m, 2H), 7.22 - 7.16 (m, 3H), 7.08 (d, J = 8.4 Hz, 2H), 6.08-6.02 (s, 1H), 5.92-5.86 (m, 1H), 4.94-4.98 (m, 1H), 4.66-4.64 (m, 0.5H), 4.48 -4.52 (m, 0.5H), 4.14 - 4.11 (m, 0.5H), 3.96-3.99 (m, 0.5H), 3.78-3.76 (m, 0.5H), 3.54-3.50 (m, 0.5H), 3.24 - 3.21 (m, 0.5H), 2.94-2.90 (m, 0.5H), 2.38-2.34 (m, 2H), 2.08-2.04 (m, 1H), 1.86-1.82 (m, 1H).

Example 2: (R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine Synthesis of 1-yl]-2-chloroprop-2-en-1-one (Compound 2)

In a similar manner to the preparation of Compound 1, Compound 2 (72% yield) was obtained using 2-chloroacrylic acid as a starting material. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.28 (s, 1H), 7.62 (d, J = 8 Hz, 2H), 7.41 (t, J = 8 Hz, 2H), 7.24-7.08 (m, 5H) , 5.71-5.64 (m, 2H), 4.98-4.90 (m, 1H), 4.72-4.66 (m, 0.5H), 4.58-4.52 (m, 0.5H), 4.18-4.12 (m, 0.5H), 3.99 -3.95 (m, 0.5H), 3.80-3.78 (m, 0.5H), 3.57-3.52 (m, 0.5H), 3.33-3.28 (m, 0.5H), 2.96-2.90 (m, 0.5H), 2.42 -2.25 (m, 2H), 2.09-2.01 (m, 1H), 1.85-1.77 (m, 1H).

Example 3: (R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine Synthesis of 1-yl]-2-fluoroprop-2-en-1-one (Compound 3)

In a similar manner to the preparation of Compound 1, Compound 3 (68% yield) was obtained using 2-fluoroacrylic acid as a starting material. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.28 (s, 1H), 7.62 (d, J = 8 Hz, 2H), 7.41 (t, J = 8 Hz, 2H), 7.24-7.08 (m, 5H) , 5.71-5.64 (m, 2H), 4.98-4.90 (m, 1H), 4.72-4.66 (m, 0.5H), 4.58-4.52 (m, 0.5H), 4.18-4.12 (m, 0.5H), 3.99 -3.95 (m, 0.5H), 3.80-3.78 (m, 0.5H), 3.57-3.52 (m, 0.5H), 3.33-3.28 (m, 0.5H), 2.96-2.90 (m, 0.5H), 2.42 -2.25 (m, 2H), 2.09-2.01 (m, 1H), 1.85-1.77 (m, 1H).

Example 4: (R,E)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl] Synthesis of piperidin-1-yl]-4,4,4-trifluoro-but-2-en-1-one (Compound 4)

In a similar manner to the preparation of Compound 1, Compound 4 (69% yield) was obtained by using trans-4,4,4-trifluorobutenoic acid as a starting material. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.38 (s, 1H), 7.64 (d, J = 8.4 Hz, 2H), 7.38-7.34 (m, 2H), 7.21-7.14 (m, 3H), 7.08 (d, J=8.4 Hz, 2H), 7.03-6.95 (m, 1H), 6.75-6.64 (m, 1H), 5.53-5.51 (br, 2H), 4.91-4.78 (m, 1.5H), 4.41 -4.36(m,0.5H),4.14-4.12(m,0.5H),3.96-3.92(m,0.5H),3.74-3.70(m,0.5H),3.52-3.48(m,0.5H),3.34 - 3.29 (m, 0.5H), 3.26-3.18 (m, 0.5H), 2.52 - 2.46 (m, 1H), 2.32 - 2.22 (m, 1H), 2.16 - 2.02 (m, 2H).

Example 5: (R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine Synthesis of 1-yl]-2-trifluoromethylprop-2-en-1-one (Compound 5)

In a similar manner to the preparation of Compound 1, Compound 5 (58% yield) was obtained from 2-(trifluoromethyl)acrylic acid. _{^{1 H-NMR (CDCl 3,}} 400MHz, δppm): 8.34 (s, 1H), 7.61 (d, J = 8.4Hz, 2H), 7.41-7.39 (m, 2H), 7.20-7.14 (m, 3H), 7.08 (d, J=8.4 Hz, 2H), 6.06-6.03 (m, 1H), 5.73-5.69 (m, 1H), 4.95-4.86 (m, 1H), 4.80-4.61 (m, 1H), 4.12 4.06 (m, 0.5H), 3.92-3.84 (m, 0.5H), 3.78-3.71 (m, 0.5H), 3.61-3.52 (m, 0.5H), 3.34-3.26 (m, 0.5H), 2.98- 2.88 (m, 0.5H), 2.42 - 2.26 (m, 2H), 2.08 - 2.01 (m, 1H), 1.86-1.72 (m, 1H).

Example 6: (R,Z)-4-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl] Synthesis of piperidin-1-yl]-4-oxobut-2-enyl cyanide (compound 6)

a.(R,Z)-4-(3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl]-1 Synthesis of -piperidinyl]-4-oxo-2-butenyl cyanide

(R)-3-(4-Phenoxyphenyl)-1-(piperidin-3-yl)-1H-pyrazolo[3,4-D]pyrimidine-4-ammonium (193 mg, 0.5 mmol) , maleamic acid (67 mg, 0.58 mmol), 1-hydroxybenzotriazole (HOBT) (95 mg, 0.7 mmol) and 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride The salt (EDCI) (134 mg, 0.7 mmol) eluted elute After it was diluted with ethyl acetate, it was washed with water, and the aqueous phase was extracted twice with ethyl acetate. The organic phase was combined, dried over anhydrous sodium sulfate, and purified by column chromatography to afford compound 6 (140 mg, yield 60%). ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.34 (s, 1H), 7.61 (d, J = 8.0 Hz, 2H), 7.41-7.37 (m, 2H), 7.20-7.14 (m, 3H), 7.08 (d, J=8.0 Hz, 2H), 6.99-6.93 (t, 1H, J = 11.6 Hz), 5.77 (d, J ₁ = 11.6 Hz, 0.5 H), 5.70 (d, J ₁ = 11.6 Hz, 0.5H), 4.93-4.89 (m, 1.5H), 4.46-4.42 (m, 0.5H), 4.00-3.86 (m, 1H), 3.52-3.51 (m, 0.5H), 3.50-3.48 (m, 0.5) H), 3.33-3.31 (m, 0.5H), 3.18-3.12 (m, 0.5H), 2.48-2.36 (m, 2H), 2.12-2.07 (m, 1H), 1.86-1.78 (m, 1H). ESI-MS (m/z): 466.2 [M+H] ⁺ .

Example 7: (R,E)-4-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl] Synthesis of Methyl Piperidin-1-yl]-4-oxobut-2-enoate (Compound 7)

In a similar manner to the preparation of Compound 1, Compound 7 (66% yield) was obtained by using monomethyl fumarate as a starting material. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.38 (s, 1H), 7.62 (d, J = 8.4 Hz, 2H), 7.41 - 7.24 (m, 3H), 7.21-7.17 (m, 3H), 7.09 (d, J = 8.4 Hz, 2H), 6.95 (d, J = 15.6 Hz, 0.5H), 6.75 (d, J = 15.6 Hz, 0.5H), 4.92-4.79 (m, 1.5H), 4.43 4.37 (m, 0.5H), 4.14-4.10 (m, 0.5H), 4.03-3.95 (m, 0.5H), 3.92-3.88 (m, 0.5H), 3.81 (s, 3H), 3.49-3.46 (m , 0.5H), 3.27-3.24 (m, 0.5H), 3.11-3.06 (m, 0.5H), 2.42-2.21 (m, 2H), 2.10-2.02 (m, 1H), 1.81-1.68 (m, 1H) ).

Example 8: (R,E)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl] Synthesis of piperidin-1-yl]-3-chloroprop-2-en-1-one (compound 8)

In a similar manner to the preparation of Compound 1, compound 8 (67% yield) was obtained using trans-3-chloroacrylic acid as a starting material. ¹ H-NMR (d-DMSO, 400 MHz, δ ppm): 8.42 (s, 1H), 7.63 (d, J = 8.4 Hz, 2H), 7.42-7.36 (m, 2H), 7.29-7.12 (m, 4H) , 7.08 (d, J = 8.4 Hz, 2H), 6.76-6.66 (m, 1H), 5.92-5.88 (br, 2H), 4.87-4.84 (m, 1.5H), 4.48-4.45 (m, 0.5H) , 4.12-4.08 (m, 0.5H), 4.02-3.98 (m, 0.5H), 3.87-3.84 (m, 0.5H), 3.42-3.38 (m, 0.5H), 3.28-3.12 (m, 0.5H) , 3.06-2.98 (m, 0.5H), 2.42 - 2.33 (m, 2H), 2.12-1.98 (m, 2H).

Example 9: (R,Z)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl] Synthesis of piperidin-1-yl]-3-bromoprop-2-en-1-one (compound 9)

In a similar manner to the preparation of Compound 1, Compound 9 (63% yield) was obtained from cis-3-bromoacrylic acid. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.39 (s, 1H), 7.64 (d, J = 8.4 Hz, 2H), 7.42 - 7.36 (m, 2H), 7.20-7.16 (m, 3H), 7.06 (d, J = 8.4 Hz, 2H), 6.84 - 6.78 (m, 1H), 6.60 (d, J = 8.0 Hz, 0.5H), 6.53 (d, J = 8.0 Hz, 0.5H), 5.61-5.59 (br, 2H), 4.95-4.92 (m, 1.5H), 4.89-4.85 (m, 0.5H), 4.18-4.06 (m, 0.5H), 3.96-3.82 (m, 1H), 3.48-3.42 (m , 0.5H), 3.24 - 3.18 (m, 0.5H), 2.98-2.94 (m, 0.5H), 2.42 - 2.22 (m, 2H), 2.06-1.98 (m, 2H).

Example 10: (R,E)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl] Synthesis of piperidin-1-yl]-3-bromoprop-2-en-1-one (compound 10)

In a similar manner to the preparation of Compound 1, Compound 10 (62% yield) was obtained using trans-3-bromoacrylic acid as a starting material. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.40 (s, 1H), 7.64 (d, J = 8.4 Hz, 2H), 7.58-7.54 (m, 1H), 7.46-7.38 (m, 2H), 7.19-7.14(m,3H),7.08(d,J=8.4Hz,2H), 7.02-6.90(m,1H),5.52-5.51(br,2H),4.87-4.68(m,1.5H),4.45 -4.41 (m, 0.5H), 4.06-4.02 (m, 0.5H), 3.97-3.93 (m, 0.5H), 3.87-3.84 (m, 0.5H), 3.46-3.42 (m, 0.5H), 3.32 - 3.21 (m, 0.5H), 3.18-3.06 (m, 0.5H), 2.44-2.41 (m, 1H), 2.36-2.26 (m, 1H), 2.03-1.96 (m, 2H).

Example 11: (R,E)-4-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl] Synthesis of piperidin-1-yl]-4-oxobut-2-enamide (Compound 11)

Synthesis of (E)-4-Amino-4-oxo-2-butenoic acid methyl ester

Monomethyl fumarate (15 g, 115 mmol) was dissolved in 20 ml of toluene, dichloromethane (20 ml) was added, and the mixture was evaporated. Under ice-cooling, a solution of NH ₃ in THF (80 ml) was added dropwise to the acid chloride prepared above, and the mixture was stirred and allowed to react at room temperature for 1 h. Washed with aqueous citric acid pH equal to 6, extracted with ethyl acetate, the organic phase was washed with saturated NaHCO _3, dried and concentrated to give (E) -4- Amino-4-oxo-2-butenoate (8.9 g of , yield 60%). ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 6.95 (d, J = 15.2 Hz, 1H), 6.84 (d, J = 15.6 Hz, 1H), 5.88-5.84 (m, 1H), 5.83-5.81 ( m, 1H), 3.82 (s, 3H).

a. Synthesis of (E)-4-amino-4-oxo-2-butenoic acid

Methyl (E)-4-amino-4-oxo-2-butenoate (4 g, 31 mmol) was dissolved in MeOH (40 mL). Concentration, 2M HCl was added to adjust the pH to 3, ethyl acetate was extracted twice, the organic phase was combined, dried and concentrated to give (E)-4-amino-4-oxo-2-butenoic acid (2.7 g, yield 75 %). ESI-MS (m/z): 116.1 [M+H] ⁺ .

c. (R,E)-4-(3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl]-1 Synthesis of -piperidinyl]-4-oxo-2-butenamide

In a similar manner to the preparation of Compound 1, compound 11 (60% yield) was obtained using (E)-4-amino-4-oxo-2-butenoic acid as a starting material. ¹ H-NMR (d-DMSO, 400 MHz, δ ppm): 8.36 (s, 1H), 7.62 (d, J = 8.4 Hz, 2H), 7.48-7.36 (m, 3H), 7.18-7.12 (m, 3H) , 7.07 (d, J = 8.4 Hz, 2H), 7.02-6.88 (m, 1H), 5.83-5.81 (br, 2H), 4.87-4.81 (m, 1.5H), 4.51-4.48 (m, 0.5H) , 4.21-4.19 (m, 0.5H), 4.17-4.15 (m, 0.5H), 3.87-3.84 (m, 0.5H), 3.38-3.34 (m, 0.5H), 3.26-3.19 (m, 0.5H) , 2.98-2.96 (m, 0.5H), 2.42 - 2.38 (m, 1H), 2.34 - 2.28 (m, 1H), 2.06-1.98 (m, 1H), 1.78-1.72 (m, 1H). ESI-MS (m/z): 484.7 [M+H] ⁺ .

Example 12: (R)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)- Synthesis of 4-tetrahydropyrrol-1-yl]-2-fluoroprop-2-en-1-one (Compound 12)

Synthesis of (R)-1-Boc-2-((4-amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidin-1-yl)methyl)tetrahydropyrrole

4-Amino-3-iodo-1H-oxazolo[3,4-D]pyrimidine (10 g, 38 mmol), Boc-D-prolinol (17.1 g, 85 mmol), triphenylphosphine (20 g, 76 mmol) Into a three-necked flask, THF (120 ml) was added, and the mixture was cooled to 0 ° C, and a mixture of DIAD (15.2 g, 76 mmol) and THF (30 ml) was added dropwise, and the mixture was added dropwise over 1 hour, and the mixture was slowly allowed to react to room temperature overnight. The reaction solution was sparged, water was added, ethyl acetate was evaporated, dried, concentrated, and then purified by column chromatography to give product (R)-1-Boc-2-((4-amino-3-iodo-1H-pyrazolo[3] 4-D]pyrimidin-1-yl)methyl)tetrahydropyrrole (11 g, yield 65%). ESI-MS (m/z): 445.2 [M+H] ⁺ .

b. (R)-1-Boc-2-((4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)methyl) Synthesis of tetrahydropyrrole

(R)-1-Boc-2-((4-amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidin-1-yl)methyl)tetrahydropyrrole (in a three-necked flask) 12.9g, 29mmol), 4-phenoxyphenylboronic acid (6.8g, 32mmol), Pd(dppf)Cl ₂ (0.5g, 0.69mmol), sodium carbonate (6.1g, 58mmol), 1,4-dioxane Hexacyclic (160 ml) and water (40 ml) were replaced with nitrogen and then warmed to 80 ° C overnight. After confirming the completion of the reaction, the mixture was filtered, dried, evaporated, evaporated, evaporated, evaporated. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.34 (s, 1H), 7.64 (d, J = 8.4 Hz, 2H), 7.41-7.37 (m, 2H), 7.17-7.13 (m, 3H), 7.09-7.07 (m, 2H), 4.61-4.34 (m, 3H), 3.40-3.26 (m, 2H), 1.84-1.78 (m, 4H), 1.42 (s, 9H).

c. Synthesis of (R)-3-(4-phenoxyphenyl)-1-(tetrahydropyrrole-2-methyl)-1H-pyrazolo[3,4-D]pyrimidine-4-amino

In the reaction flask, (R)-1-Boc-2-((4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl) Methyl)tetrahydropyrrole (7.98 g, 16.4 mmol) was added to a mixed solution of trifluoroacetic acid (40 ml) and dichloromethane (40 ml). After confirming the completion of the reaction, the mixture was neutralized with a saturated Na ₂ CO ₃ solution to pH 7-8, and then extracted with DCM. Yield 80%). ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.26 (s, 1H), 7.58 (d, J = 8.4 Hz, 2H), 7.36 (m, 2H), 7.16 (m, 1H), 7.06 (m, 4H), 4.86-4.77 (m, 2H), 4.32-4.30 (m, 1H), 3.52-3.46 (m, 1H), 3.36-3.32 (m, 1H), 2.22-2.18 (m, 1H), 2.06- 1.96 (m, 2H), 1.86-1.78 (m, 1H).

d.(R)-1-(2-((4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)methyl)tetra Synthesis of Hydropyrrole-1-yl)-2-fluoropropen-1-one

(R)-3-(4-Phenoxyphenyl)-1-(tetrahydropyrrole-2-methyl)-1H-pyrazolo[3,4-D]pyrimidine-4-amino (193 mg, 0.5 mmol), 2-fluoroacrylic acid (52 mg, 0.58 mmol), HOBT (95 mg, 0.7 mmol) and 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDCI) (134 mg) , 0.7 mmol), dissolved in anhydrous DCM (10 mL). After diluting with ethyl acetate, the mixture was washed with water, and the aqueous phase was extracted twice with ethyl acetate. The organic phase was combined, dried over anhydrous sodium sulfate, and then purified by column chromatography to afford compound 12 (160 mg, yield 70%). ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.36 (s, 1H), 7.62 (d, J = 8.4 Hz, 2H), 7.46-7.42 (m, 2H), 7.28-7.22 (m, 3H), 7.16 (d, J = 8.4 Hz, 2H), 5.46 (dd, J ₁ = 46.4 Hz, J ₂ = 3.2 Hz, 1H), 5.12 (dd, J ₁ = 15.6 Hz, J ₂ = 2.8 Hz, 1H), 4.75-4.71 (m, 3H), 3.67-3.65 (m, 1H), 3.58-3.56 (m, 1H), 1.86-1.82 (m, 2H), 1.78-1.68 (m, 2H).

Example 13: (R)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)- Synthesis of 4-tetrahydropyrrol-1-yl]-2-bromoprop-2-en-1-one (Compound 13)

In a similar manner to the preparation of compound 12, compound 13 (yield 58%) was obtained from 2-bromoacrylic acid. _{^{1 H-NMR (CDCl 3,}} 400MHz, δppm): 8.30 (s, 1H), 7.57 (d, J = 8.4Hz, 2H), 7.40-7.38 (m, 2H), 7.20-7.13 (m, 3H), 7.08 (d, J = 8.4 Hz, 2H), 5.98 (d, J = 2 Hz, 1H), 5.81 (d, J = 2 Hz, 1H), 4.72-4.68 (m, 3H), 3.68-3.62 (m, 1H) ), 3.52-3.48 (m, 1H), 2.02-1.94 (m, 2H), 1.92-1.86 (m, 2H).

Example 14: (R)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)- Synthesis of 4-tetrahydropyrrol-1-yl]-2-chloroprop-2-en-1-one (Compound 14)

In a similar manner to the preparation of compound 14, compound 13 (52% yield) was obtained from 2-chloroprop. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.31 (s, 1H), 7.61 (d, J = 8.4 Hz, 2H), 7.39 (t, J = 8 Hz, 2H), 7.22-7.08 (m, 5H) ), 5.68 (d, J = 2 Hz, 1H), 5.63 (d, J = 2 Hz, 1H), 4.77 - 4.66 (m, 3H), 3.58 - 3.57 (m, 1H), 3.45 - 3.43 (m, 1H) , 2.11-2.04 (m, 1H), 1.92-1.88 (m, 3H).

Example 15: (R,E)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl Synthesis of methyl]tetrahydropyrrol-1-yl]-4,4,4-trifluorobut-2-en-1-one (Compound 15)

In a similar manner to the preparation of compound 12, compound 15 (67% yield) was obtained using trans-4,4,4-trifluorobutenoic acid as a starting material. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.38 (s, 1H), 7.66 (d, J = 8.4 Hz, 2H), 7.38-7.32 (m, 2H), 7.19-7.15 (m, 3H), 7.08 (d, J=8.4 Hz, 2H), 6.82-6.78 (m, 1H), 6.69-6.67 (m, 1H), 5.52-5.50 (br, 2H), 4.78-4.67 (m, 2H), 4.64 4.59 (m, 1H), 3.76-3.69 (m, 1H), 3.56-3.48 (m, 1H), 2.10-2.05 (m, 2H), 1.78-1.66 (m, 2H).

Example 16: (R,E)-4-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl Synthesis of Methyl]Methyltetrahydropyrrol-1-yl]-4-oxobut-2-enoate (Compound 16)

In a similar manner to the preparation of compound 12, compound 16 (70% yield) was obtained using monomethyl fumarate as a starting material. ¹ H-NMR (d-DMSO, 400 MHz, δ ppm): 8.28 (s, 1H), 7.60 (d, J = 8.4 Hz, 2H), 7.40-7.36 (m, 2H), 7.22-7.08 (m, 5H) , 6.98 (d, J = 15.2 Hz, 0.5H), 6.83 (d, J = 15.2 Hz, 1H), 6.60 (d, J = 15.2 Hz, 0.5H), 4.71-4.68 (m, 2H), 4.39- 4.36 (m, 1H), 3.79 (s, 3H), 3.72-3.67 (m, 1H), 3.59-3.56 (m, 1H), 2.16-2.04 (m, 2H), 2.01-1.94 (m, 2H).

Example 17: (R,Z)-4-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl Synthesis of methyl]tetrahydropyrrol-1-yl]-4-oxobut-2-enyl cyanide (Compound 17)

In a similar manner to the preparation of compound 12, compound 17 (65% yield) was obtained using maleamic acid as a starting material. ¹ H-NMR (d-DMSO, 400 MHz, δ ppm): 8.36 (s, 1H), 7.62 (d, J = 8.4 Hz, 2H), 7.42-7.38 (m, 2H), 7.20-7.16 (m, 5H) , 6.95 (d, J = 11.6 Hz, 0.5 H), 6.82 (d, J = 11.2 Hz, 0.5 H), 5.76 (d, J = 11.2 Hz, 0.5 H), 5.62 - 5.69 (m, 2.5 H), 4.81-4.78 (m, 1H), 4.77-4.68 (m, 1H), 4.56-4.52 (m, 1H), 3.78-3.72 (m, 1H), 3.48-3.42 (m, 1H), 2.02-1.98 (m) , 2H), 1.86-1.78 (m, 2H).

Example 18: (S)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl) A Synthesis of 4-tetrahydropyrrol-1-yl]-2-fluoroprop-2-en-1-one (Compound 18)

In a similar manner to the preparation of Compound 12, Compound 18 (68% yield) was obtained from Boc-L-prolinol and 2-fluoroacrylic acid. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.42 (s, 1H), 7.62 (d, J = 8.4 Hz, 2H), 7.44 - 7.38 (m, 2H), 7.18-7.12 (m, 3H), 7.06 (d, J = 8.4 Hz, 2H), 5.48 (d, J = 48 Hz, 1H), 5.14 (d, J = 2.4 Hz, 1H), 4.75 - 4.67 (m, 2.5H), 4.52-4.48 (m , 0.5H), 3.65-3.60 (m, 1H), 3.50-3.48 (m, 1H), 1.96-1.88 (m, 2H), 1.86-1.68 (m, 2H).

Example 19: (S)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)- Synthesis of 4-tetrahydropyrrol-1-yl]-2-chloroprop-2-en-1-one (Compound 19)

In a similar manner to the preparation of compound 18, compound 19 (64% yield) was obtained using 2-chloroprop. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.34 (s, 1H), 7.62 (d, J = 8.4 Hz, 2H), 7.44 - 7.38 (m, 2H), 7.28-7.20 (m, 3H), 7.10 (d, J = 8.4 Hz, 2H), 5.68 (d, J = 2.4 Hz, 1H), 5.64 (d, J = 2.4 Hz, 1H), 4.82-4.68 (m, 3H), 3.58-3.52 (m , 1H), 3.48-3.42 (m, 1H), 2.14-2.10 (m, 1H), 1.95-1.78 (m, 3H).

Example 20: (S)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)- Synthesis of 4-tetrahydropyrrol-1-yl]-2-bromoprop-2-en-1-one (Compound 20)

In a similar manner to the preparation of compound 18, compound 20 (64% yield) was obtained using 2-bromoacrylic acid as a starting material. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.29 (s, 1H), 7.59 (d, J = 8.4 Hz, 2H), 7.44 - 7.40 (m, 2H), 7.23-7.19 (m, 3H), 7.09 (d, J = 8.4 Hz, 2H), 6.00 (d, J = 2 Hz, 1H), 5.83 (d, J = 2 Hz, 1H), 4.77 - 4.64 (m, 3H), 3.58 - 3.56 (m, 1H) ), 3.50-3.46 (m, 1H), 2.12-2.04 (m, 1H), 1.94-1.86 (m, 3H).

Example 21: (S,E)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl Synthesis of methyl]tetrahydropyrrol-1-yl]-4,4,4-trifluorobut-2-en-1-one (Compound 21)

In a similar manner to the preparation of compound 18, compound 21 (64% yield) was obtained using trans-4,4,4-trifluorobutenoic acid as a starting material. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.41 (s, 1H), 7.68 (d, J = 8.4 Hz, 2H), 7.46-7.38 (m, 2H), 7.24-7.09 (m, 5H), 6.88-6.82 (m, 1H), 6.78-6.67 (m, 1H), 5.54-5.52 (br, 2H), 4.78-4.68 (m, 2H), 4.44-4.42 (m, 1H), 3.76-3.67 (m , 1H), 3.52-3.45 (m, 1H), 2.16-2.08 (m, 2H), 1.78-1.62 (m, 2H).

Example 22: (S,E)-4-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl Synthesis of Methyl]Methyltetrahydropyrrol-1-yl]-4-oxobut-2-enoate (Compound 22)

In a similar manner to the preparation of compound 18, compound 22 (70% yield) was obtained starting from monomethyl fumarate. ¹ H-NMR (d-DMSO, 400 MHz, δ ppm): 8.23 (s, 1H), 7.64 (dd, J1 = 8.4 Hz, J2 = 22.4 Hz, 2H), 7.46-7.42 (m, 2H), 7.21 - 7.12 (m, 5H), 6.69-6.63 (m, 1H), 6.11 (d, J = 15.2 Hz, 1H), 4.63-4.61 (m, 1H), 4.47-4.43 (m, 1H), 4.29-4.24 (m , 1H), 3.58 (s, 3H), 3.52-3.48 (m, 2H), 2.06-1.98 (m, 3H), 1.86-1.82 (m, 1H).

Example 23: (S,Z)-4-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl Synthesis of methyl]tetrahydropyrrol-1-yl]-4-oxobut-2-enyl cyanide (Compound 23)

In a similar manner to the preparation of compound 18, compound 23 (64% yield) was obtained from maleamic acid. _{^{1 H-NMR (CDCl 3,}} 400MHz, δppm): 8.32 (s, 1H), 7.61 (d, J = 6.4Hz, 2H), 7.39-7.36 (m, 2H), 7.15-7.06 (m, 5H), 6.92 (d, J = 11.2 Hz, 0.5H), 6.82 (d, J = 11.2 Hz, 0.5H), 5.76 (d, J = 11.2 Hz, 0.5H), 5.59 (d, J = 11.2 Hz, 0.5H) ), 4.77-4.72 (m, 1H), 4.66-4.50 (m, 1H), 4.42-4.38 (m, 1H), 3.72-3.64 (m, 1H), 3.48-3.44 (m, 1H), 2.15. (m, 4H).

Example 24: (S,E)-1-[2-[(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl Synthesis of methyl]tetrahydropyrrol-1-yl]-4-(dimethylamino)but-2-en-1-one (Compound 24)

Synthesis of a. 4-dimethylamino croton hydrochloride

The compound methyl crotonate (20 g, 200 mmol) was added to a 500 ml single-mouth bottle, dissolved in carbon tetrachloride (200 ml), and azobisisobutyronitrile (AIBN) (66 mg, 0.4 mmol) and N-bromine were added with stirring. Desuccinimide (NBS) (39.1 g, 220 mmol), the reaction mixture was reacted with heating under reflux for 6 h, cooled, filtered, and the solid was filtered, washed with water (20 ml × 3), dried over anhydrous Na ₂ SO ₄ , filtered This was triturated to give a yellow oil, methyl 4-bromocrotonate (34 g).

The compound 4-bromocrotonate methyl ester (34 g, 190 mmol) was added to a 500 ml single-necked flask, dissolved in THF, and the hydrochloride salt of dimethylamine (18.5 g, 230 mmol) and Et ₃ N (74.2 ml, 530 mmol) were added with stirring. After stirring at room temperature overnight, methyl 4-bromocrotonate was completely reacted, and the solvent was evaporated. The remaining yellow oil was added to isopropyl alcohol (200 ml), and 4 M HCl/dioxane solution was added dropwise to adjust pH ≤2. A white solid was isolated, filtered, and then filtered and washed with EtOAc EtOAc EtOAc EtOAc ¹ H-NMR (d-DMSO, 400 MHz, δ ppm): 6.89-6.92 (m, 1H), 6.30 (d, J = 16 Hz, 1H), 3.91 (d, J = 8 Hz, 2H), 3.70 (s, 3H) ), 2.72 (s, 6H).

The compound 4-dimethylamino crotonate methyl ester hydrochloride (10 g, 56 mmol) was added to a 250 ml single-mouth bottle, and 6N aqueous hydrochloric acid (100 ml) was added thereto, and the mixture was refluxed for 6 hours, and the reaction mixture was dried to give a white solid. Amino croton hydrochloride (9.0 g, 98% yield). ¹ H-NMR (d-DMSO, 400 MHz, δ ppm): 6.75-6.81 (m, 1H), 6.18 (d, J = 16 Hz, 1H), 3.89 (d, J = 8 Hz, 2H), 2.74 (s, 6H) ).

b.(S,E)-1-(2-((4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)) Synthesis of tetrahydropyrrol-1-yl)-4-(dimethylamino)-2-buten-1-one

In a similar manner to the preparation of compound 18, compound 24 (64% yield) was obtained using 4-dimethylamino crotonate as a starting material. _{^{1 H-NMR (CDCl 3,}} 400MHz, δppm): 8.32 (s, 1H), 7.78-7.67 (m, 2H), 7.36-7.28 (m, 2H), 7.18-7.05 (m, 5H), 6.83-6.77 (m, 1H), 6.55-6.32 (m, 1H), 4.64-4.58 (m, 2H), 4.38-4.32 (m, 1H), 3.64-3.52 (m, 2H), 3.32-3.18 (m, 1H) , 3.02-2.98 (m, 1H), 2.48 (s, 6H), 2.02-1.74 (m, 4H).

Example 25: (R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine Synthesis of 1-yl]prop-2-en-1-one (Compound 25)

Synthesis of 2-(4-bromobenzene)-2-benzene-1,3-dithiocyclohexane

4-bromobenzoylbenzene (10g, 38mmol), propylenedithiol (5g, 46mmol), p-toluenesulfonic acid (1.1g, 6.4mmol) was dissolved in toluene (50ml), refluxed for 12h, dried organic The solvent was beaten with petroleum ether and filtered to give the product (11 g, 82% yield). ESI-MS (m/z): 350.9 [M+H] ⁺ .

b. Synthesis of 1-bromo-4-(difluoro(phenyl)methyl)benzene

2-(4-Bromobenzene)-2-benzene-1,3-dithiocyclohexane (5 g, 14 mmol) was dissolved in anhydrous dichloromethane (50 ml). After the completion of the dropwise addition, the reaction was stirred, and the mixture was evaporated. EtOAc (EtOAc) _{^{1 H-NMR (CDCl 3,}} 400MHz, δppm): 7.55 (d, J = 8.8Hz, 2H), 7.48-7.46 (m, 2H), 7.44-7.41 (m, 3H), 7.37 (d, J = 8.8 Hz, 2H). ¹⁹ F-NMR: (CDCl ₃ , 400 MHz, δ ppm): -88.9.

c. Synthesis of 4-benzoylbenzeneboronic acid

1-Bromo-4-(difluoro(phenyl)methyl)benzene (2 g, 7.1 mmol) was dissolved in anhydrous THF (20 mL), cooled to -78 ° C under nitrogen, and n-butyl lithium was added dropwise. n-BuLi) (3.1 mL, 7.8 mmol), maintaining the temperature below -65 ° C, stirring for 1 h, maintaining the temperature at -60 ° C, adding triisopropyl borate (1.5 g, 7.8 mmol), slowly rising to 0 ° C Stir for 3 h. Water (10 mL) was added to the ice bath and stirred overnight. The reaction mixture was concentrated to remove the organic phase, which was adjusted to pH 1-2 with 12N concentrated hydrochloric acid, and solids were isolated and filtered to give the product (1.5 g, yield 95%). ESI-MS (m/z): 227.1 [M+H] ⁺ .

d. (R)-1-(3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)tetrahydropyrrole- Synthesis of 1-yl)-2-propen-1-one

In a similar manner to the preparation of Compound 1, compound 25 (62% yield) was obtained from 4-benzoylbenzeneboronic acid and acrylic acid. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.40 (s, 1H), 7.98 (d, J = 8.4 Hz, 2H), 7.84-7.8 (m, 4H), 7.65-7.61 (m, 1H), 7.54-7.51(m,2H),6.62-6.58(m,1H),6.34-6.28(m,1H),5.68-5.62(m,3H),4.96-4.94(m,1H),4.86-4.84(m , 0.5H), 4.68-4.64 (m, 0.5H), 4.24-4.20 (m, 0.5H), 4.12-4.08 (m, 0.5H), 3.74-3.71 (m, 0.5H), 3.41-3.38 (m , 0.5H), 3.22-3.19 (m, 0.5H), 2.98-2.96 (m, 0.5H), 2.42-2.38 (m, 2H), 2.18-2.02 (m, 2H). ESI-MS (m/z): 453.2 [M+H] ⁺ .

Example 26: (R)-[4-[4-Amino-1-(1-(vinylsulfone)piperidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl Synthesis of phenyl](phenyl)methanone (Compound 26)

In a similar manner to the preparation of compound 25, compound 26 (53% yield) was obtained using vinylsulfonyl chloride as a starting material. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.40 (s, 1H), 7.98 (d, J = 8.4 Hz, 2H), 7.91 - 7.84 (m, 4H), 7.64 - 7.60 (m, 1H), 7.54-7.50 (m, 2H), 6.51-6.44 (m, 1H), 6.28-6.24 (d, J = 16.8 Hz, 1H), 6.08-6.03 (m, 1H), 5.72 (br, 2H), 5.06- 5.04 (m, 1H), 4.03-4.00 (m, 1H), 3.84-3.81 (m, 1H), 3.34-3.31 (m, 1H), 2.72-2.68 (m, 1H), 2.28-2.22 (m, 2H) ), 2.20-2.18 (m, 2H).

Example 27: (R,E)-4-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl] Synthesis of piperidin-1-yl]-4-oxobut-2-enyl cyanide (Compound 27)

a.(E)-Synthesis of methyl cyanoacrylate

Methyl (E)-4-amino-4-oxo-2-butenoate (3.18 g, 25 mmol) was dissolved in pyridine (27 mL). EtOAc (3. After 1 h, the mixture was stirred at room temperature for 1 h. After TLC showed that the reaction was completed, ice cubes were added, stirred for 0.5 h, extracted with DCM three times, and the organic phase was washed with 2M HCl, and then washed with NaHCO ₃ , Product (1 g, 36% yield). ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 6.71 (d,J = 16.4 Hz, 1H), 6.48 (d,J = 16.4 Hz, 1H), 3.83 (s, 3H).

b. Synthesis of (E)-3-cyano-acrylic acid

(E)-Methyl cyanoacrylate (1.4 g, 13 mmol) was dissolved in THF (5 mL), MeOH (EtOAc) (EtOAc) After stirring at 10 ° C for 3 h, the reaction was completed, 2M HCl was added to adjust pH to 3, ethyl acetate was extracted, and the organic phase was dried and concentrated to give product (945 mg, yield 75%). ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 9.29 (s, 1H), 6.72 (d, J = 16.4 Hz, 1H), 6.56 (d, J = 16.4 Hz, 1H).

c. (R,E)-4-(3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)tetrahydrogen Synthesis of pyrrol-1-yl)-4-oxo-2-butenyl cyanide

In a similar manner to the preparation of Compound 1, Compound 27 (56% yield) was obtained from (E)-3-cyano-acrylic acid. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.39 (s, 1H), 7.61 (d, J = 8.4 Hz, 2H), 7.41 - 7.37 (m, 2H), 7.31 - 7.19 (dd, J1 = 16 Hz , J2=42Hz, 1H), 7.17-7.14(m,3H), 7.08(d, J=8.4Hz, 2H), 6.49(d, J=16Hz, 0.5H), 6.43(d, J=16Hz, 0.5 H), 5.79-5.72 (br, 2H), 4.90-4.86 (m, 1H), 4.82-4.78 (m, 0.5H), 4.38-4.35 (m, 0.5H), 4.12-4.06 (m, 0.5H) , 3.93-3.89 (m, 1H), 3.52-3.46 (m, 0.5H), 3.32-3.28 (m, 0.5H), 3.18-3.12 (m, 0.5H), 2.52-2.38 (m, 1H), 2.32 -2.22 (m, 1H), 2.12 - 2.01 (m, 1H), 1.82-1.68 (m, 1H). ESI-MS (m/z): 466.6 [M+H] ⁺ .

Example 28: (R)-1-[3-[4-Amino-3-(4-(difluoro(phenyl)methyl)phenyl)-1H-pyrazolo[3,4-d]pyrimidine Synthesis of 1-yl]piperidin-1-yl]prop-2-en-1-one (Compound 28)

Synthesis of 2-(4-(difluoro(phenyl)methyl)phenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborane

1-Bromo-4-(difluoro(phenyl)methyl)benzene (3.76 g, 13 mmol) was dissolved in dimethoxyethane (DME) (100 mL) and dimethyl sulfoxide (DMSO) (5 mL) Under nitrogen protection, [1,1'-bis(diphenylphosphino)ferrocene]palladium(II) chloride (Pd(dppf)Cl ₂ ) (960 mg, 1.3 mmol) was added and the reaction was carried out at 80 ° C. After overnight, TLC was monitored. After the reaction was completed, dichloromethane and water were added, and the organic phase was dried over anhydrous sodium sulfate and purified by column chromatography to afford product (2.1 g, yield 47.9%). ¹⁹ F-NMR (400 MHz, CDCl ₃ , δ ppm): -89.67. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 7.85 (d, J = 8.0 Hz, 2H), 7.52-7.47 (m, 4H), 7.43-7.39 (m, 3H), 1.34 (s, 12H).

b.(R)-1-(3-(4-Amino-3-(4-(difluoro(phenyl)methyl)phenyl)-1H-pyrazolo[3,4-D]pyrimidine-1 Synthesis of -yl)piperidin-1-yl)-2-propen-1-one

A method similar to the preparation of Compound 1, 2-(4-(difluoro(phenyl)methyl)phenyl)-4,4,5,5-tetramethyl-1,3,2-dioxa Starting from borane and acrylic acid, compound 28 (61% yield) was obtained.

¹⁹ F-NMR (400 MHz, CDCl ₃ , δ ppm): -89.3. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.38 (s, 1H), 7.78-7.68 (m, 4H), 7.58-7.54 (m, 2H), 7.45-7.39 (m, 3H), 6.68-6. (m, 1H), 6.36-6.28 (m, 1H), 5.73-5.53 (m, 3H), 4.88-4.85 (m, 1.5H), 4.61-4.52 (m, 0.5H), 4.28-4.22 (m, 0.5H), 4.08-3.96 (m, 0.5H), 3.76-3.68 (m, 0.5H), 3.36-3.32 (m, 0.5H), 3.21-3.18 (m, 0.5H), 2.94-2.88 (m, 0.5H), 2.41-2.28 (m, 2H), 2.04-1.92 (m, 1H), 1.88-1.78 (m, 1H). ESI-MS (m/z): 475.5 [M+H] ⁺ .

Example 29: (R)-3-[4-(Difluoro(phenyl)methyl)phenyl]-1-(1-(vinylsulfone)piperidin-3-yl)-1H-pyrazolo[ Synthesis of 3,4-d]pyrimidine-4-ammonia (Compound 29)

In a similar manner to the preparation of compound 28, compound 29 (53% yield) was obtained from methylenesulfonyl chloride. ¹⁹ F-NMR (400 MHz, CDCl ₃ , δ ppm): -89.41. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.38 (s, 1H), 7.70 (dd, J ₁ = 8.0 Hz, J ₂ = 20.4 Hz, 4H), 7.54-7.52 (m, 2H), 7.46- 7.44 (m, 3H), 6.45-6.42 (dd, J ₁ = 16.8 Hz, J ₂ = 10.0 Hz, 1H), 6.24 (d, J = 16.8 Hz, 1H), 6.03 (d, J = 10.0 Hz, 1H) ), 5.52-5.49 (br, 2H), 5.04-5.01 (m, 1H), 4.00-3.96 (m, 1H), 3.82-3.79 (m, 1H), 3.28-3.23 (m, 1H), 2.75-2.69 (m, 1H), 2.23-2.19 (m, 2H), 1.98-1.88 (m, 2H). ESI-MS (m/z): 511.5 [M+H] ⁺ .

Example 30: [(R)-3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidine-1 Synthesis of -()-((R)-oxiran-2-yl)methanone (Compound 30)

a. Synthesis of (R)-Ethylene oxide-2-carboxylate

Methyl (R)-glycidylate (1.0 g, 9.8 mmol) was dissolved in ethanol (8 ml) and H ₂ O (1 ml), and KOH (0.66 g, 12 mmol). After stirring at room temperature for 2 hours, the material of the plate was completely reacted, and the solution was directly dried to give a pale yellow solid, which was directly used for the next step.

b.((R)-3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)piperidin-1-yl Synthesis of ((R)-oxiran-2-yl)methanone (Compound 30)

(R)-3-(4-Phenoxyphenyl)-1-(piperidin-3-yl)-1H-pyrazolo[3,4-D]pyrimidine-4-ammonium (307 mg, 0.79 mmol) And (R)-oxirane-2-carboxylate (200 mg, 1.6 mmol), 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDCI) (167 mg, 0.87 mmol) and 1-hydroxybenzotriazole (HOBT) (118 mg, 0.87 mmol) were dissolved in DMF (5 mL) solvent, then N,N-diisopropylethylamine (DIPEA) (307 mg, 2.38 mmol) . The reaction was carried out at 42 ° C for 2 hours, and the material of the spot plate disappeared. After cooling, it was washed with water, ethyl acetate (5 ml × 3), and then dried and evaporated. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.37 (s, 1H), 7.64 (d, J = 8.4 Hz, 2H), 7.41-7.37 (m, 2H), 7.17-7.14 (m, 3H), 7.08 (d, J=8.4 Hz, 2H), 5.50-5.48 (br, 2H), 4.89-4.71 (m, 1H), 4.53-4.37 (m, 0.5H), 4.34-4.30 (m, 0.5H), 4.22-4.16 (m, 0.5H), 3.90-3.80 (m, 0.5H), 3.68-3.64 (m, 1H), 3.47-3.41 (m, 0.5H), 3.31-3.20 (m, 0.5H), 3.02 -2.98 (m, 2H), 2.91-2.86 (m, 1H), 2.47-2.34 (m, 1H), 2.28-2.25 (m, 1H), 2.02-1.98 (m, 1H), 1.80-1.74 (m, 1H).

Example 31: [(R)-3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidine-1 Synthesis of -()-(()-oxiran-2-yl)methanone (Compound 31)

a. Synthesis of (S)-2-bromo-3-hydroxypropionic acid

D-serine (1.0 g, 9.5 mmol) and KBr (4.0 g, 33 mmol) were stirred in an aqueous solution (8 ml) to a total solution, and HBr (4.1 g, 21 mmol, a 47% by mass aqueous solution) was added at room temperature. The mass concentration refers to the mass of hydrogen bromide as a percentage of the total mass of the aqueous hydrogen bromide solution. The solution is cooled to -12 ° C, then NaNO ₂ (0.72 g, 11 mmol) is slowly added in portions, each time the solution is added to a tan The reaction was stirred until fading was added again, and after 2 hours of addition, the mixture was returned to 0 ° C, and the reaction was stirred overnight. The reaction mixture was extracted with EtOAc (EtOAc m. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 4.24 (dd, J ₁ = 5.6 Hz, J ₂ = 8.0 Hz, 1H), 3.80 (dd, J ₁ = 8.0 Hz, J ₂ = 11.2 Hz, 1H) , 3.67 (dd, J ₁ = 5.6 Hz, J ₂ = 11.2 Hz, 1H).

b. Synthesis of (S)-Ethylene oxide-2-carboxylate

(S)-2-Bromo-3-hydroxypropionic acid (1.0 g, 5.9 mmol) was dissolved in absolute ethanol (8 ml), the reaction mixture was dropped to -20 ° C, and KOH (0.63 g, 11.3) was slowly added under a nitrogen atmosphere. A solution of mmol) in absolute ethanol, and the reaction was allowed to react at -20 ° C to 0 ° C overnight. Most of the ethanol was removed by rotary evaporation. EtOAc (3 mL) was evaporated.

c.((R)-3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)piperidin-1-yl Synthesis of ((S)-oxiran-2-yl)methanone

In a similar manner to the preparation of compound 30, compound 31 (36% yield) was obtained from (S)-ethyl oxy-2-carboxylate. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.37 (s, 1H), 7.66-7.63 (m, 2H), 7.41-7.37 (m, 2H), 7.18-7.14 (m, 3H), 7.09-7.07 (m, 2H), 5.57-5.54 (br, 2H), 4.89-4.78 (m, 1.5H), 4.58-4.55 (m, 0.5H), 4.05-4.00 (m, 0.5H), 3.88-3.84 (m , 0.5H), 3.74-3.68 (m, 1H), 3.32-3.26 (m, 0.5H), 3.22-3.15 (m, 0.5H), 2.81-2.72 (m, 0.5H), 2.44-2.40 (m, 0.5H), 2.38-2.22 (m, 2H), 2.00-1.92 (m, 1H), 1.78-1.72 (m, 3H).

Example 32: (R)-1-[3-[4-Amino-3-(4-phenylthiophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine Synthesis of 1-yl]-2-bromoprop-2-en-1-one (Compound 32)

Synthesis of (4-bromophenyl)(phenyl)sulfane

Benzyl bromide (5.0 g, 26 mmol) and iodobenzene (2.7 g, 13 mmol), tetrabutylammonium bromide (TBAB) (4.3 g, 13 mmol), KOH (1.1 g, 20 mmol) in water (50 ml) CuI (0.05 g, 0.26 mmol) was added to the solution, and the reaction was carried out at 85 ° C overnight. The reaction of the iodophenyl group was carried out, washed with water, and extracted with ethyl acetate (30 ml × 3). The organic phase was dried and then evaporated to dryness. The product was purified by column chromatography (2.8 g, yield 80%). ESI-MS (m/z): 264.9 [M+H] ⁺ .

b. Synthesis of 4-(phenylthio)benzeneboronic acid

Under a nitrogen atmosphere, a solution of (4-bromophenyl)(phenyl)sulfane (1.0 g, 3.8 mmol) in anhydrous THF (10 mL) was reduced to -70 ° C, and n-BuLi (2.1 ml, 4.2) was slowly added dropwise. Mm), after the completion of the dropwise addition, the reaction was stirred for 15 min, isopropyl borate (0.86 g, 4.6 mmol) was added, the dry ice bath was removed, and the reaction was carried out at 0 ° C for 30 min. The solution became cloudy and white solids were separated. After 3 h of reaction, water (6 ml) was added and the reaction was allowed to stand overnight. Water (8 ml) was added again, and the mixture was extracted with HCl (pH 1-2), EA (8 ml×3), and the organic phase was dried and then evaporated to dryness, washed with petroleum ether and collected by filtration (0.75 g, yield 86%) .

¹ H-NMR (d-DMSO, 400 MHz, δ ppm): 8.11 (s, 2H), 7.74 (d, J = 8.0 Hz, 2H), 7.39-7.35 (m, 4H), 7.23 (d, J = 8.0 Hz) , 2H), 6.56 (s, 1H).

c. 1-[(3R)-3-[4-Amino-3-(4-phenylthiophenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl]-1-piper Synthesis of pyridyl]-2-bromopropen-1-one

In a similar manner to the preparation of Compound 1, compound 32 (45% yield) was obtained using 4-(phenylthio)benzeneboronic acid as a starting material. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.37 (s, 1H), 7.60 (d, J = 8.4 Hz, 2H), 7.48-7.46 (m, 2H), 7.42 - 7.34 (m, 5H), 6.04-6.01(s,1H),5.85-5.78(m,1H),5.46-5.38(br,2H),4.96-4.91(m,1H),4.68-4.62(m,0.5H),4.53-4.47( m, 0.5H), 4.17-4.11 (m, 0.5H), 3.99-3.94 (m, 0.5H), 3.79-3.71 (m, 0.5H), 3.52-3.45 (m, 0.5H), 3.22-3.17 ( m, 0.5H), 2.93-2.87 (m, 0.5H), 2.35-2.22 (m, 2H), 2.02-1.98 (m, 1H), 1.80-1.75 (m, 1H).

Example 33: (R)-1-[3-[4-Amino-3-(4-benzylphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine- Synthesis of 1-yl]-2-bromoprop-2-en-1-one (Compound 33)

Synthesis of a 4-benzylbenzeneboronic acid sterol ester

In a 50 ml single-necked flask, 4-bromodiphenylmethane (2.5 g, 10 mmol), boranoic acid pinacol ester (3.885 g, 15 mmol), KOAc (3.1 g, 31 mmol), DME (65 ml), DMSO (3.3 ml) ). The N _{2 was} replaced 3 times, and a catalyst Pd(dppf)Cl ₂ (740 mg, 0.5 mmol) was added. The N _{2 was} replaced 3 times, heated to 80 ° C, refluxed overnight, and the starting material was completely reacted. The reaction mixture was passed through celite, EtOAc (EtOAc)EtOAc ESI-MS (m/z): 295.2.

b. Synthesis of (R)-1-Boc-3-(4-amino-3-(4-benzylphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)piperidine

(R)-1-Boc-3-(4-Amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidin-1-yl)piperidine (2.6 g, 5.9) was added to a 500 ml vial Ment), 4-benzylphenylboronic acid sterol ester (1.2 g, 5.9 mmol), Na ₂ CO ₃ (2.31 g, 31 mmol), dimethoxyethane (DME) (150 ml), H ₂ O (30 ml) ). The N _{2 was} replaced 3 times, and the catalyst tetrakistriphenylphosphine palladium (Pd(PPh ₃ ) ₄ ) (0.23 g, 0.3 mmol) was added, and N _{2 was} substituted 3 times. Heat to 80 ° C, reflux overnight, the reaction of the starting material. The reaction system was an orange clear solution. Most of the solvent was evaporated, water was added, and the mixture was evaporated. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.28 (s, 1H), 7.62-7.57 (m, 2H), 7.54-7.52 (m, 2H), 7.30-7.22 (m, 2H), 7.19-7.14 (m, 3H), 5.66-5.55 (br, 2H), 4.78-4.73 (m, 1H), 4.32-4.05 (m, 2H), 3.98-4.01 (m, 2H), 3.40-3.26 (m, 1H) , 2.79-2.71 (m, 1H), 2.18-2.11 (m, 2H), 1.82-1.81 (m, 1H), 1.63-1.60 (m, 1H), 1.36 (s, 9H).

c. (R)-1-[3-[4-Amino-3-(4-benzylphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine-1- Synthesis of 2-bromoprop-2-en-1-one (Compound 33)

A method similar to the preparation of Compound 1, with (R)-1-Boc-3-(4-amino-3-(4-benzylphenyl)-1H-pyrazolo[3,4-D]pyrimidine- Starting from 1-yl)piperidine, Compound 33 (40% yield) was obtained.

¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 8.36 (s, 1H), 7.60 (d, J = 7.6 Hz 2H), 7.37-7.31 (m, 4H), 7.23-7.22 (m, 3H), 6.03 -6. , 0.5H), 4.17-4.11 (m, 0.5H), 4.05 (s, 2H), 4.01-3.94 (m, 0.5H), 3.79-3.70 (m, 0.5H), 3.48-3.45 (m, 0.5H) ), 3.18-3.18 (m, 0.5H), 2.89-2.85 (m, 0.5H), 2.35-2.23 (m, 2H), 1.80-1.59 (m, 2H).

Example 34: (R)-1-[3-[4-Amino-3-(4-(4-fluorophenoxy)phenyl)-1H-pyrazolo[3,4-d]pyrimidine-1 Synthesis of 1-ylpiperidin-1-yl]-2-bromoprop-2-en-1-one (Compound 34)

Synthesis of a. 1-(4-Bromophenoxy)-4-fluorobenzene

4-Fluorodiphenyl ether (30 g, 0.16 mol) and a few iodine were dissolved in CS ₂ (300 ml), and bromine (10 ml, 0.19 mol) was slowly added dropwise under ice-cooling. The purified product (28 g, 64%) was obtained. ¹ H-NMR (CDCl ₃ , 400 MHz, δ ppm): 7.46-7.42 (m, 2H), 7.10 - 6.97 (m, 4H), 6.88 - 6.84 (m, 2H).

b. Synthesis of 4-(4-fluorophenoxy)phenylboronic acid

1-(4-Bromophenoxy)-4-fluorobenzene (10 g, 37 mmol) was dissolved in anhydrous THF (EtOAc) (EtOAc) 41 mmol), the temperature was kept below -65 ° C, and after stirring for 1 h, the temperature was maintained at -60 ° C, triisopropyl borate (8.3 g, 44.4 mmol) was added dropwise, and the mixture was slowly warmed to 0 ° C and stirred for 3 h. Water (30 mL) was added to the ice bath and stirred overnight. The reaction mixture was concentrated to remove the organic phase, which was adjusted to pH 1-2 with 12N concentrated hydrochloric acid, and solids were separated and filtered to afford product (6.4 g, yield 75%). ESI-MS (m/z): 233.1 [M+H] ⁺ .

c. (R)-1-(3-(4-Amino-3-(4-(4-fluorophenoxy)phenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl Synthesis of 1-(piperidinyl)-2-bromopropen-1-one

In a similar manner to the preparation of Compound 1, compound 34 (41% yield) was obtained from 4-phenoxyphenylboronic acid. ESI-MS (m/z): 537.1 [M+H] ⁺ .

Example 35: [(R)-3-(4-Amino-3-(3-fluoro-4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl) Synthesis of Tetrahydropyrrol-1-yl]((S)-oxiran-2-yl)methanone (Compound 35)

Synthesis of (R)-1-Boc-3-(4-amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidin-1-yl)tetrahydropyrrole

4-Amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidine (10 g, 38 mmol), (R)-1-Boc-3-hydroxypyrrole (16 g, 85 mmol), triphenylphosphine (20g, 76mmol) was added to a three-necked flask, THF (120ml) was added, the temperature was lowered to 0 ° C, and a mixture of diisopropyl azodicarboxylate (DIAD) (15.2 g, 76 mmol) and THF (30 ml) was added dropwise for about 1 h. After the dropwise addition was completed, the mixture was slowly warmed to room temperature and allowed to react overnight. The reaction mixture was evaporated to dryness. ESI-MS (m/z): 431.1 [M+H] ⁺ .

b. Synthesis of 3-fluoro-4-phenoxyphenylboronic acid

In a similar manner to the preparation of compound 34, 4-fluoro-4-phenoxyphenylboronic acid can be obtained by using 4-bromo-2-fluoro-1-phenoxybenzene as a starting material. ESI-MS (m/z): 233.1 [M+H] ⁺ .

b.(3R)-1-Boc-3-(4-Amino-3-(3-fluoro-4-phenyloxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl) Synthesis of tetrahydropyrrole

(R)-1-Boc-3-(4-Amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidin-1-yl)tetrahydropyrrole (12.5 g, 29 mmol) was added to a three-necked flask. , 3-fluoro-4-phenoxyphenylboronic acid (7.4 g, 32 mmol), Pd(dppf)Cl ₂ (0.5 g, 0.69 mmol), sodium carbonate (6.1 g, 58 mmol), 1,4-dioxane Hexacyclic (160 ml) and water (40 ml) were replaced with nitrogen and then warmed to 80 ° C overnight. After confirming the completion of the reaction, the mixture was filtered, dried, evaporated, evaporated, evaporated, evaporated. ESI-MS (m/z): 4921. [M+H] ⁺ .

Synthesis of (R)-3-(3-fluoro-4-phenoxyphenyl)-1-(pyrrol-3-yl)-1H-pyrazolo[3,4-D]pyrimidine-4-ammonia

In the reaction flask, (R)-1-Boc-3-(4-amino-3-(3-fluoro-4-phenyloxyphenyl)-1H-pyrazolo[3,4-D]pyrimidine-1 Tetrahydropyrrole (7.3 g, 14.8 mmol) was added to a mixed solution of trifluoroacetic acid (40 ml) and dichloromethane (40 ml) and allowed to react at room temperature overnight. After confirming the completion of the reaction, the mixture was neutralized with a saturated Na ₂ CO ₃ solution to pH 7-8, and then extracted with DCM. Yield 86%). ESI-MS (m/z): 391.2 [M+H] ⁺ .

d.((R)-3-(4-Amino-3-(3-fluoro-4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)-1 Synthesis of (tetrahydropyrrolyl)((S)-oxiran-2-yl)methanone (Compound 35)

A procedure similar to the preparation of compound 31, (R)-3-(3-fluoro-4-phenoxyphenyl)-1-(pyrrol-3-yl)-1H-pyrazolo[3,4-D Pyrimidine-4-ammonia was used as a starting material to give compound 35 (43% yield). ESI-MS (m/z): 461.2 [M+H] ⁺ .

Example 36: (R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]tetrahydro Synthesis of pyrrol-1-yl]-2-bromoprop-2-en-1-one (Compound 36)

a. (R)-1-Boc-3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)tetrahydropyrrole synthesis

(R)-1-Boc-3-(4-Amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidin-1-yl)tetrahydropyrrole (12.5 g, 29 mmol) was added to a three-necked flask. , 4-phenoxyphenylboronic acid (6.8 g, 32 mmol), Pd(dppf)Cl ₂ (0.5 g, 0.69 mmol), sodium carbonate (6.1 g, 58 mmol), 1,4-dioxane (160 ml) After cooling with water (40 ml), the mixture was heated to 80 ° C overnight. After confirming the completion of the reaction, the mixture was filtered, dried, evaporated, evaporated, evaporated, evaporated. ESI-MS (m/z): 473.2 [M+H] ⁺ .

b. Synthesis of (R)-3-(4-phenoxyphenyl)-1-(pyrrol-3-yl)-1H-pyrazolo[3,4-D]pyrimidine-4-ammonia

In the reaction flask, (R)-1-Boc-3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl)tetra Hydropyrrole (7 g, 14.8 mmol) was added to a mixed solution of trifluoroacetic acid (40 ml) and dichloromethane (40 ml) and allowed to react at room temperature overnight. After confirming the completion of the reaction, the mixture was neutralized with a saturated Na ₂ CO ₃ solution to pH 7-8, and then extracted with DCM. Yield 86%). ESI-MS (m/z): 373.2 [M+H] ⁺ .

c. (R)-1-(3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl]-1-tetra Synthesis of Hydropyrrolyl]-2-bromopropen-1-one (Compound 36)

(R)-3-(4-Phenoxyphenyl)-1-(pyrrol-3-yl)-1H-pyrazolo[3,4-D]pyrimidine-4-ammonium (184 mg, 0.5 mmol), 2-Bromoacrylic acid (88 mg, 0.58 mmol), HOBT (95 mg, 0.7 mmol), EtOAc (EtOAc, EtOAc, EtOAc (EtOAc) . After diluting with ethyl acetate, the mixture was washed with EtOAc (EtOAc). ESI-MS (m/z): 505.1 [M+H] ⁺ .

Example 37: (R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]tetrahydro Synthesis of pyrrol-1-yl]-2-chloroprop-2-en-1-one (Compound 37)

In a similar manner to the preparation of compound 36, compound 37 (73% yield) was obtained using 2-chloroprop. ESI-MS (m/z): 461.1 [M+H] ⁺ .

Example 38: (R)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]tetrahydro Synthesis of pyrrol-1-yl]-2-fluoroprop-2-en-1-one (Compound 38)

In a similar manner to the preparation of compound 36, compound 38 (73% yield) was obtained from 2-fluoroacrylic acid. ESI-MS (m/z): 445.2 [M+H] ⁺ .

Example 39: (R,Z)-4-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl] Synthesis of Tetrahydropyrrol-1-yl]-4-oxobut-2-enyl Cyanide (Compound 39)

In a similar manner to the preparation of compound 36, compound 39 (73% yield) was obtained from maleamic acid. ESI-MS (m/z): 452.2 [M+H] ⁺ .

Example 40: [(R)-3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)tetrahydropyrrole- Synthesis of 1-yl]((R)-oxiran-2-yl)methanone (Compound 40)

In a similar manner to the preparation of compound 36, compound 40 (72% yield) was obtained using (R)-e. ESI-MS (m/z): 443.2 [M+H] ⁺ .

Example 41: (S,E)-1-[3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl) Synthesis of Tetrahydropyrrol-1-yl]-3-chloroprop-2-en-1-one (Compound 41)

Synthesis of (S)-1-Boc-3-(4-amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidin-1-yl)pyrrole

4-Amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidine (12 g, 46 mmol), (S)-1-Boc-3-hydroxypyrrole (19 g, 101 mmol), triphenylphosphine (24g, 92mmol) was added to a three-necked flask, THF (120ml) was added, the temperature was lowered to 0 ° C, and a mixture of DIAD (18.6 g, 76 mmol) and THF (30 ml) was added dropwise, and the mixture was added dropwise over 1 h, and slowly allowed to react to room temperature overnight. . The reaction mixture was evaporated to dryness. ESI-MS (m/z): 431.1 [M+H] ⁺ .

b.(S,E)-1-(3-(4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-D]pyrimidin-1-yl]-1 Synthesis of -tetrahydropyrrolyl]-3-chloropropen-1-one

A procedure similar to the preparation of compound 36, with (S)-1-Boc-3-(4-amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidin-1-yl)pyrrole and E)-3-chloroacrylic acid was used as a starting material to give compound 41 (75% yield). ESI-MS (m/z): 461.2 [M+H] ⁺ .

Example 42: (S,Z)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl] Synthesis of Tetrahydropyrrol-1-yl]-3-bromoprop-2-en-1-one (Compound 42)

In a similar manner to the preparation of compound 41, compound 42 (71% yield) was obtained using (Z)-3-bromoacrylic acid as a starting material. ESI-MS (m/z): 505.2 [M+H] ⁺ .

Example 43: (R)-1-[3-[4-Amino-3-(4-(pyridin-4-yloxy)phenyl)-1H-pyrazolo[3,4-d]pyrimidine-1 Synthesis of 1-ylpiperidin-1-yl]-2-bromoprop-2-en-1-one (Compound 43)

Synthesis of 4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenoxy)pyridine

In a 50 ml single-necked flask, 4-(4-bromophenoxy)pyridine (2.5 g, 10 mmol), boranoic acid pinacol ester (3.9 g, 15 mmol), KOAc (3.1 g, 31 mmol), DME (65 ml), DMSO (3.3 ml). The N _{2 was} replaced 3 times, and a catalyst Pd(dppf)Cl ₂ (740 mg, 0.5 mmol) was added. The N _{2 was} replaced 3 times, heated to 80 ° C, refluxed overnight, and the starting material was completely reacted. The reaction solution was passed through celite, water was added, EA was evaporated, and the organic phase was dried and concentrated, and the product was purified by column chromatography (2.4 g, yield 80%). ESI-MS (m/z): 298.2.

A method similar to the preparation of Compound 1, 4-amino-3-iodo-1H-pyrazolo[3,4-D]pyrimidine, 4-(4-(4,4,5,5-tetramethyl-) Compound 43 (66% yield) was obtained by using 1,3,2-dioxaborolan-2-yl)phenoxy)pyridine and 2-bromoacrylic acid as starting materials. ESI-MS (m/z): 520.1 [M+H] ⁺ .

Example 44: (S)-1-[3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidine Synthesis of 1-yl]-2-bromoprop-2-en-1-one (Compound 44)

A method similar to the preparation of Compound 1, 4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidine, (R)-1-Boc-3-hydroxypiperidine and 2-bromoacrylic acid As a starting material, Compound 43 (70% yield) was obtained. ESI-MS (m/z): 519.1 [M+H] ⁺ .

Examples 45-54: Synthesis of Compound 45-54

A method similar to the preparation of Compound 1, 4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidine, (S)-1-Boc-3-hydroxypiperidine and 2-bromoacrylic acid As a starting material, compounds 45-54 can be obtained.

2. Biological activity test part:

Biological Example 1

Series of compounds in different tumor cell growth inhibition assays (CCK8 assay)

First, the cell line:

L1210: murine lymphocyte leukemia cell line, DMEM + 10% FBS;

WSU-DLCL2: human B lymphoma cell line, RPMI1640 + 10% FBS;

K562: human chronic myeloid leukemia cell line, IMDM + 10% FBS;

HL-60: human promyelocytic leukemia cell line, IMDM + 20% FBS;

Second, reagents and consumables:

CCK8 kit; anti-tumor compound; DMSO.

Third, the test method:

1. Cell culture

Collect logarithmic growth phase cells, count, resuspend cells with complete medium,

The cell concentration was adjusted to an appropriate concentration, and 96-well plates were seeded, and 100 μl of the cell suspension was inoculated per well.

The cells were incubated for 24 hours at 37 ° C in a 100% relative humidity, 5% CO ₂ incubator.

2. Relative inhibition rate experiment

The logarithmic growth phase cells were collected, counted, and the cells were resuspended in complete medium, and the cell concentration was adjusted to an appropriate concentration (determined according to the cell density optimization test results), and 96-well plates were seeded, and 100 μl of the cell suspension was added to each well. The cells were incubated for 24 hours at 37 ° C in a 100% relative humidity, 5% CO ₂ incubator.

The test compound was diluted with the medium to the corresponding concentration of action set, and the cells were added at 25 μl/well. The final concentration of the compound was diluted from 10 μM to 0 μM in 4 fold gradients for a total of 10 concentration points.

The cells were incubated for 72 hours at 37 ° C in a 100% relative humidity, 5% CO ₂ incubator.

The medium was aspirated, and the complete medium containing 10% CCK-8 was added and incubated in a 37 ° C incubator for 2-4 hours.

The absorbance at a wavelength of 450 nm was measured on a SpectraMax M5 Microplate Reader with gentle shaking, and the absorbance at 650 nm was used as a reference to calculate the inhibition rate.

Data processing and results

The inhibition rate of the drug on tumor cell growth was calculated as follows: tumor cell growth inhibition rate % = [(Ac-As) / (Ac-Ab)] × 100%

As: OA of the sample (cell + CCK-8 + test compound)

Ac: negative control OA (cell + CCK-8 + DMSO)

Ab: positive control OA (medium + CCK-8 + DMSO)

The IC ₅₀ curve was fitted and the IC ₅₀ value was calculated using the software Graphpad Prism 5 and using the calculation formula log(inhibitor) vs.response.

Table 1 shows the growth inhibitory activity (IC ₅₀ ) of some compounds on different tumor cells

Biological Example 2

The following experiment can be used to determine the inhibitory effect of the compounds of the invention on Bruton's tyrosine kinase (BTK) enzymatic activity.

The in vitro kinase assay was performed using Cisbio's HTRF kinEASE TK kit, and the procedure was followed by reference to the kit instructions, which examined the inhibitory effect of the test compound on BTK enzyme activity in vitro. The specific steps are as follows:

First, prepare a 2.5% DMSO solution using the configured 1X kinase buffer (the DMSO concentration will affect the reaction, control the final concentration of DMSO to 1%), and then dilute the test compound with the enzyme corresponding 2.5% DMSO solution. To the corresponding test concentration (4X). In addition to the control wells, 4 μL of the diluted test compound solution was added to the wells used, and 4 μL of the previously prepared BTK enzyme-containing 2.5% DMSO solution was added to the control wells.

To all wells, 2 μL of the previously prepared BTK enzyme corresponding substrate concentration of TK-biotin substrate solution was added (see Table 1 for the amount of substrate for enzyme screening).

To all wells except the negative wells, 2 μL of the previously prepared enzyme solution of the corresponding concentration (see Table 1 for the amount of enzyme) was added, and the negative wells were filled with 2 μL of enzyme corresponding to 1Xkinase buffer. The plate was sealed with a sealing film, and the mixture was incubated at room temperature for 10 minutes to allow the compound and the enzyme to fully bind.

The kinase reaction was initiated by adding 2 μL of ATP solution at the corresponding concentration of BTK enzyme to all wells. The enzymatic reaction time of BTK was 25 minutes (see Table 1 for the corresponding ATP concentration and reaction time for enzyme screening).

The BTK test solution was prepared 5 minutes before the end of the kinase reaction. Streptavidin-XL665 and TK antibody europium cryptate (1:100) solutions were prepared using the detection buffer in the kit. The concentrations of the corresponding detection reagents for enzyme screening are shown in Table 1.

After the completion of the kinase reaction, 5 μL of the diluted Streptavidin-XL665 was added to all wells, and the diluted TK antibody europium cryptate assay solution was added immediately after mixing.

The plate was mixed and reacted at room temperature for 1 h, and the fluorescence signal (320 nm stimulation, 665 nm, 615 nm emission) was detected using an ENVISION (Perkinelmer) instrument. Inhibition was calculated for each well by total active aperture hole and a background signal, duplicate wells were averaged for each test compound simultaneously fitting the median inhibitory activity (IC ₅₀₎ with specialized drawing software PRISM 5.0.

Table 2, IC ₅₀ inhibitory activity test of series compounds on BTK enzyme

Biological Example 3

In vivo pharmacodynamic evaluation

SCID Beige mice, female, 5-6 weeks old, weighing 18g ± 2g, purchased from Beijing Weitong Lihua Experimental Animal Co., Ltd., SPF-level environment.

Take WSU-DLCL2 cells in logarithmic growth phase, centrifuge at 1000 rpm for 5 min, resuspend in serum-free medium, and take 10 μL. The number of viable cells (cell survival rate > 90%) was counted by trypan blue staining, and the cell density was adjusted to 1 x 107 cells/0.2 mL. Four 5-6 week old SCID mice were injected, and 1×107/side/0.2 mL of WSU-DLCL2 cells were injected under the left and right sacs under sterile conditions. Apparent subcutaneous tumor formation was observed in about two weeks. When the tumor grows to 300-500 mm3, the mice are euthanized, the tumor is exfoliated under sterile conditions, placed in 1640 medium, and cut into 1-2 mm3 tumor blocks (each subcutaneous tumor can be divided into 20-30 Tumor block). Tumors were inoculated into the right subcutaneous of 80 5-6 week old SCID mice using a 12 gauge trocar.

When tumors of tumor-bearing mice grew to a measurable size, mice were randomly divided into 8 groups according to the mean tumor volume balance principle using SPSS 17.0 software. Compound Example 1 and Compound Example 2 were administered orally at 90, 30, 10 mg/kg per day; the positive control drug, ibrutinib, was administered orally at 30 mg/kg daily for a dose of 0.1 ml/10 g. . The drug was administered once a day for 23 days, and the negative control group was given an equal amount of solvent (1% DMSO in physiological saline solution). During the administration period and recovery period, the body weight and tumor diameter of the mice were measured 2 to 3 times per week. Tumor volume and relative tumor volume were calculated according to the measured data. Tumor Volume (TV) was calculated as: TV=1/2×a×b2, where a and b represent the long and short diameters of the tumor, respectively. According to the measurement results, the relative tumor volume (RTV) was calculated, and the formula was: RTV=Vt/V0, where V0 is the tumor volume at the start of the experiment, and Vt is the tumor volume measured each time. The anti-tumor activity evaluation index is the relative tumor growth rate T/C (%), and the formula is as follows: T/C (%) = TRTV / CRTV × 100%, TRTV is the treatment group RTV; CRTV is the negative control group RTV, relative The tumor growth inhibition rate (tumor inhibition rate)% = (1-T/C) × 100%, and the results are shown in Table 3.

Table 3: Therapeutic effects of the compounds of the invention on WSU-DLCL2 nude mice xenografts

While the invention has been described with respect to the preferred embodiments of the embodiments of the embodiments of the invention modify. Accordingly, the scope of the invention is defined by the appended claims.

Claims (17)

A pyrazolopyrimidine derivative of the formula I, a stereoisomer, solvate, pharmaceutically acceptable salt, active metabolite or prodrug thereof,

Where: L is O, S,

When L is O, S,

When Z is

When L is

When Z is

Wherein X is a halogen atom, a cyano group,

R is a C ₁ -C ₄ alkyl group; Ar is a substituted or unsubstituted aryl group or a substituted or unsubstituted heteroaryl group; the "substituted" as used in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means Substituted by one or more substituents selected from the group consisting of a halogen atom, a cyano group, a substituted or unsubstituted alkyl group, an alkoxy group, an alkenyl group, an alkynyl group, and an aryl group, said "substituted or unsubstituted alkyl group" The term "substituted" as used herein means substituted by one or more halogen atoms, and when a plurality of substituents are present, the substituents are the same or different; V is a hydrogen atom or a fluorine atom; Y is a substituted or unsubstituted alkylene group, cycloalkyl group or heterocycloalkyl group; the substitution described in the "substituted or unsubstituted alkylene group" means that the hetero atom is an oxygen, sulfur or nitrogen atom. a C ₄ -C ₇ heterocycloalkyl group having ₁ to 3 hetero atoms, a C ₁ -C ₆ alkoxy group, a C ₁ -C ₆ alkylthio group, and a C ₁ -C ₆ alkylamino group. Replaced by one or more. The pyrazolopyrimidine derivative of the formula I according to claim 1, which is a stereoisomer, a solvate thereof, A pharmaceutically acceptable salt, active metabolite or prodrug characterized by: V is a hydrogen atom; and / or, When X is a halogen atom, the halogen atom is a fluorine atom, a chlorine atom or a bromine atom; and / or, When R is a C ₁ -C ₄ alkyl group, the "C ₁ -C ₄ alkyl group" is methyl, ethyl, propyl, isopropyl, butyl, isobutyl or tert-butyl. ; and / or, The substitution described in the "substituted or unsubstituted alkylene group" described in the above Y means C ₄ to C in which the hetero atom is an oxygen, sulfur or nitrogen atom and the number of hetero atoms is 1-3. _When the heterocycloalkyl group of ₇ is substituted, the "hetero atom is an oxygen, sulfur or nitrogen atom, and the C ₄ - C ₇ heterocycloalkyl group having 1 to 3 hetero atoms" is a nitrogen atom of a hetero atom. a C ₄ -C ₅ heterocycloalkyl group having 1-2 hetero atoms; and / or, When the substitution described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means that it is substituted by one or more halogen atoms, the "halogen atom" is a fluorine atom. a chlorine atom or a bromine atom; and / or, When the substituent described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means substituted by one or more substituted or unsubstituted alkyl groups, the said "substituted" Or an unsubstituted alkyl group" is a substituted or unsubstituted C ₁ -C ₄ alkyl group; and / or, The substitution described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means substituted by one or more substituted or unsubstituted alkyl groups, and the said "substituted" The substitution described in the "unsubstituted alkyl group" means that when the halogen atom is substituted, the halogen atom is a fluorine atom, a chlorine atom or a bromine atom; and / or, When the substitution described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means that it is substituted by one or more alkoxy groups, the "alkoxy group" is a C ₁ -C ₄ alkoxy group; and / or, When the substitution described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means that it is substituted by one or more alkenyl groups, the "alkenyl group" is C _{2 .} ～C ₄ alkenyl; and / or, When the substitution described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means that it is substituted by one or more alkynyl groups, the "alkynyl group" is C _{2 .} Alkynyl group of -C ₄ ; and / or, When the substitution described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means that it is substituted by one or more aryl groups, the "aryl group" is C ₅ An aryl group of -C ₁₀ ; and / or, When the Ar is a substituted or unsubstituted aryl group, the "substituted or unsubstituted aryl group" is a "substituted or unsubstituted C ₅ - C ₁₀ aryl group"; and / or, When the Ar is a substituted or unsubstituted heteroaryl group, the "substituted or unsubstituted heteroaryl group" is a substituted or unsubstituted hetero atom which is a nitrogen atom and has a hetero atom number of 1-2. a heteroaryl group of C ₃ to C ₉ ; When Y is a substituted or unsubstituted alkylene group, the "substituted or unsubstituted alkylene group" is a substituted or unsubstituted C ₁ -C ₆ alkylene group; and / or, When Y is a cycloalkyl group, the cycloalkyl group is a C ₄ -C ₇ cycloalkyl group; and / or, When Y is a heterocycloalkyl group, the heterocycloalkyl group is a C ₄ - C ₇ heterocycloalkyl group in which the hetero atom is an oxygen, sulfur or nitrogen atom and the number of hetero atoms is 1-3. A pyrazolopyrimidine derivative of the formula I according to claim 2, which is a stereoisomer, a solvate, a pharmaceutically acceptable salt, an active metabolite or a prodrug, which is characterized in that: The substitution described in the "substituted or unsubstituted alkylene group" described in the above Y means a C ₄ - C ₅ heterocyclic ring having a hetero atom of a nitrogen atom and a hetero atomic number of 1-2. When the alkyl group is substituted, the "hetero atom is a nitrogen atom, and the C ₄ - C ₅ heterocycloalkyl group having 1-2 hetero atoms" is a pyrrolidinyl group; and / or, The substitution as described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means when substituted by one or more substituted or unsubstituted C ₁ -C ₄ alkyl groups , the "substituted C ₁ -C ₄ alkyl group" is a trifluoromethyl group; and / or, The substitution as described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means when substituted by one or more substituted or unsubstituted C ₁ -C ₄ alkyl groups The "unsubstituted C ₁ -C ₄ alkyl group" is a methyl group; and / or, When the substitution described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means that it is substituted by one or more C ₁ -C ₄ alkoxy groups, "C ₁ -C ₄ alkoxy" is methoxy; and / or, When the substitution described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means that it is substituted by one or more C ₂ -C ₄ alkenyl groups, The C ₂ -C ₄ alkenyl group" is a vinyl group; and / or, When the substitution described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means that it is substituted by one or more C ₂ -C ₄ alkynyl groups, the " An alkynyl group of C ₂ to C ₄ is an ethynyl group; and / or, When the substitution described in the "substituted or unsubstituted aryl group or substituted or unsubstituted heteroaryl group" means that it is substituted by one or more C ₅ -C ₁₀ aryl groups, the " The aryl group of C ₅ to C ₁₀ is a phenyl group; and / or, When the Ar is a substituted or unsubstituted C ₅ -C ₁₀ aryl group, the "substituted or unsubstituted C ₅ -C ₁₀ aryl group" is a substituted or unsubstituted phenyl group; and / or, When the Ar is a substituted or unsubstituted C ₃ -C ₉ heteroaryl group in which the hetero atom is a nitrogen atom and the number of hetero atoms is 1-2, the "hetero atom is a nitrogen atom and a hetero atom number. 1-2 substituted or unsubstituted C ₃ -C ₉ heteroaryl" is a substituted or unsubstituted C ₃ -C ₅ heteroaryl group having a nitrogen atom as a hetero atom and a hetero atom; and / or, When Y is a substituted or unsubstituted C ₁ -C ₆ alkylene group, the "substituted or unsubstituted C ₁ -C ₆ alkylene group" is a substituted or unsubstituted methylene group; When Y is a C ₄ -C ₇ cycloalkyl group, the "C ₄ -C ₇ cycloalkyl group" is a C ₅ -C ₆ cycloalkyl group; and / or, When Y is a C ₄ -C ₇ heterocycloalkyl group in which the hetero atom is an oxygen, sulfur or nitrogen atom and the number of hetero atoms is 1-3, the "hetero atom is an oxygen, sulfur or nitrogen atom, a hetero atom" The number of 1-3 C ₄ -C ₇ heterocycloalkyl groups" is a C ₄ -C ₅ heterocycloalkyl group in which the hetero atom is a nitrogen atom and the number of hetero atoms is 1-2. A pyrazolopyrimidine derivative of the formula I, a stereoisomer, a solvate, a pharmaceutically acceptable salt, an active metabolite or a prodrug thereof, according to claim 3, wherein: When the Ar is a substituted or unsubstituted phenyl group, the "substituted phenyl group" is a phenyl group substituted by one or more fluorine atoms, a phenyl group substituted by one or more methyl groups, One or more cyano substituted phenyl groups, phenyl substituted by one or more trifluoromethyl groups, phenyl substituted by one or more methoxy groups, taken from one or more vinyl groups a phenyl group, a phenyl group substituted by one or more ethynyl groups or a phenyl group substituted by one or more phenyl groups; and / or, When the Ar is a substituted or unsubstituted C ₃ -C ₅ heteroaryl group in which the hetero atom is a nitrogen atom and the number of hetero atoms is one, the "hetero atom is a nitrogen atom and the number of hetero atoms is One substituted or unsubstituted C ₃ -C ₅ heteroaryl" is a substituted or unsubstituted pyridyl group; and / or, When Y is a substituted or unsubstituted methylene group, the "substituted methylene group" is a pyrrolidinyl-substituted methylene group; When Y is a C ₅ -C ₆ cycloalkyl group, the "C ₆ cycloalkyl group" is a cyclohexyl group; and / or, When Y is a C ₄ -C ₅ heterocycloalkyl group in which the hetero atom is a nitrogen atom and the number of hetero atoms is 1-2, the "hetero atom is a nitrogen atom and the number of hetero atoms is 1-2". _{The 4-} to C ₅ heterocycloalkyl group is a pyrrolidinyl group or a piperidinyl group. A pyrazolopyrimidine derivative of the formula I, a stereoisomer, a solvate, a pharmaceutically acceptable salt, an active metabolite or a prodrug thereof, according to claim 4, wherein: When the Ar is a phenyl group substituted by one or more fluorine atoms, the "phenyl group substituted by one or more fluorine atoms" is

and / or, When the Ar is a phenyl group substituted by one or more methyl groups, the "phenyl group substituted by one or more methyl groups" is a 4-methylphenyl group; and / or, When the Ar is a phenyl group substituted by one or more cyano groups, the "phenyl group substituted by one or more cyano groups" is a 2-cyanophenyl group; and / or, When the Ar is a phenyl group substituted by one or more trifluoromethyl groups, the "phenyl group substituted by one or more trifluoromethyl groups" is 4-trifluoromethylphenyl; and / or, When the Ar is a phenyl group substituted by one or more methoxy groups, the "phenyl group substituted by one or more methoxy groups" is a 3-methoxyphenyl group; and / or, When the Ar is a phenyl group substituted by one or more vinyl groups, the "phenyl group substituted by one or more vinyl groups" is

and / or, When the Ar is a phenyl group substituted by one or more ethynyl groups, the "phenyl group substituted by one or more ethynyl groups" is

and / or, When the Ar is a phenyl group substituted by one or more phenyl groups, the "phenyl group substituted by one or more phenyl groups" is

and / or, When the Ar is a substituted or unsubstituted pyridyl group, the "unsubstituted pyridyl group" is

and / or, When Y is a pyrrolidinyl-substituted methylene group, the "pyrrolidinyl-substituted methylene group" is

and / or, When Y is a pyrrolidinyl group, the "pyrrolidinyl group" is

and / or, When Y is piperidinyl, the "piperidinyl" is

A pyrazolopyrimidine derivative of the formula I according to claim 1, which is a stereoisomer, a solvate, a pharmaceutically acceptable salt, an active metabolite or a prodrug, which is characterized in that: When L is O, S,

When Z is

Wherein X is Cl, Br or CN, and Y is pyrrolidinyl or piperidinyl; and / or, When L is

When Z is

Wherein X is Cl, Br or CN, and Y is pyrrolidinyl or piperidinyl. A pyrazolopyrimidine derivative of the formula I according to claim 1, which is a stereoisomer, a solvate, a pharmaceutically acceptable salt, an active metabolite or a prodrug, which is characterized in that: The pyrazolopyrimidine derivative of formula I is a compound of formula I-A:

Wherein, the definition of Ar is as defined in any one of claims 1 to 5, the definition of L is as defined in claim 1 or 6, the definition of Y is as defined in any one of claims 1 to 6, and the definition of Z is as claimed. As defined in any one of 1, 2 or 6, V is as defined in claim 1 or 2. A pyrazolopyrimidine derivative of the formula I, a stereoisomer, a solvate, a pharmaceutically acceptable salt, an active metabolite or a prodrug thereof, according to claim 7, wherein: In the present invention, the pyrazolopyrimidine derivative represented by Formula I is any one of the following compounds:

The method for producing a pyrazolopyrimidine derivative according to formula I according to any one of claims 1 to 8, which comprises the step of coupling a compound II with an acid ZOH to obtain a formula The pyrazolopyrimidine derivative shown by I can be used;

Wherein, the definition of Ar is as defined in any one of claims 1 to 5, the definition of L is as defined in claim 1 or 6, the definition of Y is as defined in any one of claims 1 to 6, and the definition of Z is as claimed. As defined in any one of 1, 2 or 6, V is as defined in claim 1 or 2. Any one of the compounds of formula I, or a pharmaceutically acceptable salt thereof, or a stereoisomer, solvate, active metabolite thereof, and prodrug thereof, according to any one of claims 1 to 8, for the preparation of a medicament For use, alone or in combination with other therapeutic agents. Any one of the compounds of formula I, or a pharmaceutically acceptable salt thereof, or a stereoisomer, solvate, active metabolite thereof, and prodrug thereof, according to any one of claims 1 to 8, for the preparation of a medicament Use of the medicament for the treatment and/or prevention of a disease in a mammal and a compound for inhibiting Bruton's tyrosine kinase activity or for treating a disease that benefits from the inhibition of Bruton's tyrosine kinase activity, The use of a condition or condition. The use according to claims 10-11, wherein said inhibition of Bruton's tyrosine kinase activity is for the treatment of cancer, including but not limited to solid tumors and hematological cancers. The use according to any one of claims 10 to 11, wherein said Bruton's tyrosine kinase activity is inhibited for treating an inflammatory disease or disease state, an immune disease or disease state, a hyperproliferative disease or disease state, and a denaturation Disease or disease state. A pharmaceutical composition comprising an active compound and one or more pharmaceutically acceptable carriers or excipients; said active compound being of formula I as described in any one of claims 1-8 A pyrazolopyrimidine derivative, one or more of a stereoisomer, a solvate, a pharmaceutically acceptable salt, an active metabolite, and a prodrug. The pharmaceutical composition according to claim 14, wherein the pharmaceutical composition comprises, but is not limited to, an aqueous liquid dispersion, a self-emulsified dispersion, a solid solution, a liposome dispersant, an aerosol, Solid dosage forms, powders, immediate release preparations, controlled release preparations, instant preparations, tablets, capsules, pills, sustained release preparations, extended release preparations, pulsatile release preparations, multiparticulate preparations, and mixed immediate and controlled release preparations. A method of treating and/or preventing a disease, condition or condition in a mammal for inhibiting Bruton's tyrosine kinase activity or for treating inhibition of Bruton's tyrosine kinase activity It consists in the steps comprising the use of an effective amount of the pharmaceutical composition according to claim 14 or 15 in a population in need thereof. The method of claim 16 wherein said disease, disorder or condition that benefits from inhibition of Bruton's tyrosine kinase activity includes, but is not limited to, cancer, inflammatory disease or disease state, immune disease Or a disease state, a hyperproliferative disease or disease state, and a degenerative disease or disease state.