CN111646995A

CN111646995A - 4-amino-pyrimidoazenitrogen heterocycle-phenylurea derivative and preparation method and application thereof

Info

Publication number: CN111646995A
Application number: CN202010123847.4A
Authority: CN
Inventors: 陈俐娟
Original assignee: Sichuan University
Current assignee: Sichuan University
Priority date: 2019-03-04
Filing date: 2020-02-27
Publication date: 2020-09-11
Anticipated expiration: 2040-02-27
Also published as: CN111646995B

Abstract

The invention belongs to the field of chemical medicine, and particularly relates to a 4-amino-pyrimidoazepine-phenylurea derivative, and a preparation method and application thereof. The invention provides a 4-amino-pyrimidoazepine-phenylurea derivative, the structural formula of which is shown in formula I. In addition, the invention also provides a preparation method and application of the 4-amino-pyrimidoazepine-phenylurea derivative. The 4-amino-pyrimidoazepine-phenylurea derivative provided by the invention can be used as a FLT3 kinase inhibitor, has a good effect, and provides a new choice for preparing antitumor drugs.

Description

4-amino-pyrimidoazenitrogen heterocycle-phenylurea derivative and preparation method and application thereof

Technical Field

The invention belongs to the field of chemical medicine, and particularly relates to a 4-amino-pyrimidoazepine-phenylurea derivative, and a preparation method and application thereof.

Background

Leukemia is an abnormal clonal disease of hematopoietic stem cells, is arrested in different stages of cell development and malignant proliferation because of the loss of the ability of the leukemia cells to differentiate into mature functional blood cells, and the leukemia cells are proliferated and accumulated in large quantity in bone marrow and other hematopoietic tissues and infiltrate into other organs and tissues, so that normal hematopoiesis is inhibited, and symptoms such as anemia, hemorrhage, infection, infiltration of various organs and the like are clinically shown. Leukemia belongs to cell heterogeneous malignant tumor, has various types, complex etiology and different clinical manifestations, and some leukemias have the characteristics of quick onset, high mortality, short survival period, easy relapse, poor prognosis and extremely difficult cure.

Protein kinases, the family of receptor protein tyrosine kinases in particular, which catalyze the phosphorylation of hydroxyl groups on tyrosine, serine and threonine residues of proteins, play an important role as growth factor receptors in the control of many signal transduction pathways responsible for cellular functions, such as cell cycle, cell growth, cell differentiation and cell death. Dysregulation of receptor tyrosine kinases is often found in diseases such as proliferative disorders, inflammatory disorders, and immune system disorders.

FLT3(Fms-like tyrosine kinase 3) is an Fms-like tyrosine kinase 3, which belongs to a member of the type III receptor tyrosine kinase (RTK III) family with c-Kit, c-Fms and PDGFR, and the structure of FLT3 includes an extracellular domain consisting of 5 immunoglobulin-like molecules, a transmembrane domain and an intracellular tyrosine kinase domain. FLT3 is expressed predominantly on the cell surface of normal hematopoietic stem and progenitor cells, with its ligand predominantly expressed in bone marrow stromal cells. When ligand is combined with the extracellular domain of FLT3 cell membrane, FLT3 receptor is promoted to dimerize, and tyrosine kinase domain in cell membrane is self-phosphorylated, so that a series of downstream signal transduction pathways, such as Ras/MAPK, PI3K/Akt/mTOR and STAT5, are activated to regulate the proliferation and differentiation of cells. FLT3 mutation usually leads to its abnormal activation, and in the absence of ligand binding, autophosphorylation activates downstream signaling pathways, leading to abnormal proliferation of hematopoietic and lymphoid cells, leading to a variety of hematological malignancies.

Activating mutations for FLT3 have been demonstrated to be mainly two: point mutations in the activation loop in the internal tandem repeat (ITD) and kinase domain (TKD). The FLT3-ITD mutation occurred in approximately 25% of acute myelocytic leukemia patients and was associated with some adverse prognosis, while the FLT3-TKD mutation occurred in approximately 5% of acute myelocytic leukemia patients.

Numerous studies have shown that FLT3 mutation is one of the most common molecular genetic abnormalities and poor prognosis factors in AML, and its transduction involves multiple signaling pathways, making FLT3 an ideal drug target. In previous studies, more than 20 compounds have shown inhibition of FLT3 tyrosine kinase. Many have entered clinical trials. These small molecule tyrosinase inhibitors are mostly heterocyclic purine analogs, are ATP analogs, or intermediates with a structure similar to tyrosine covalently bound to ATP. The FLT3 receptor is still expressed, but the ATP pathway is blocked, thus autophosphorylation and sustained phosphorylation of the substrate is terminated, thereby blocking the signaling pathway of FLT3-ITD dependent cell lines and exerting cytotoxic effects.

The current research on FLT3-ITD and FLT3-TKD inhibitors belongs to the hot spot of drug development, and the single-target FLT3 inhibitor AC220 is currently subjected to clinical three-phase tests and is expected to be on the market, but the drug resistance in the later treatment period becomes an urgent problem to be solved. Therefore, the development of inhibitors effective against both mutations is the direction of research and development.

Disclosure of Invention

In order to solve the problems, the invention provides a 4-amino-pyrimidoazepine-phenylurea derivative, the structural formula of which is shown as formula I:

wherein X is N or C;

R₁is-H,-OH, halogen, C1-C10 alkoxy, C1-C10 haloalkoxy,

C2-C10 alkenyl, C2-C10 alkynyl, C3-C10 cycloalkyl,

Or substituted or unsubstituted C1-C10 alkyl; the substituent of the substituted C1-C10 alkyl is-H,

-CN, -OH, phenyl, halogen, C1-C8 alkoxy, C3-C8 cycloalkyl, C1-C8 carbonyl or substituted or unsubstituted 3-6 membered heterocycloalkyl; the heteroatom of the 3-6 membered heterocycloalkyl is N, O or S, and the number of the heteroatoms is 1-3; the substituent for substituting the 3-to 6-membered heterocycloalkyl is-H, -OH, halogen, C1-C8 oxycarbonyl, C1-C8 alkyl, C1-C8 alkoxy or

R₄is-H, -OH, halogen, C1-C8 alkyl, C1-C8 alkoxy or

R₅～R₁₀Independently is-H or C1-C8 alkyl;

R₂is-H, -OH, halogen, C1-C10 alkyl, C3-C10 cycloalkyl, C1-C10 haloalkyl, amino-substituted C1-C10 alkyl, benzyl, substituted or unsubstituted C5-C10 aryl or substituted or unsubstituted 5-10 membered heteroaryl; the 5-to 10-membered heteroaryl has N, O or S as heteroatoms, and the number of the heteroatoms is 1-3; the substituent of the substituted C5-C10 aryl or 5-10-membered heteroaryl is-H, halogen or-NH₂C1-C8 alkyl, C3-C8 cycloalkyl, C1-C8 haloalkyl, C1-C8 alkoxy, C1-C8 haloalkoxy, -OH or C1-C8 carbonyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C10 alkyl.

As a preferred embodiment of the present inventionX is N or C; r₁is-H, -OH, halogen, C1-C8 alkoxy, C1-C8 haloalkoxy,

C2-C8 alkenyl, C2-C8 alkynyl, C3-C8 cycloalkyl,

Or substituted or unsubstituted C1-C8 alkyl; the substituent of the substituted C1-C8 alkyl is-H,

-CN, -OH, phenyl, halogen, C1-C6 alkoxy, C3-C6 cycloalkyl, C1-C6 carbonyl or substituted or unsubstituted 3-6 membered heterocycloalkyl; the heteroatom of the 3-6 membered heterocycloalkyl is N, O or S, and the number of the heteroatoms is 1-3; the substituent for substituting the 3-to 6-membered heterocycloalkyl is-H, -OH, halogen, C1-C6 oxycarbonyl, C1-C6 alkyl, C1-C6 alkoxy or

R₄is-H, -OH, halogen, C1-C6 alkyl, C1-C6 alkoxy or

R₅～R₁₀Independently is-H or C1-C6 alkyl;

R₂is-H, -OH, halogen, C1-C8 alkyl, C3-C8 cycloalkyl, C1-C8 haloalkyl, amino-substituted C1-C8 alkyl, benzyl, substituted or unsubstituted C5-C8 aryl or substituted or unsubstituted 5-8 membered heteroaryl; the 5-8 membered heteroaryl has N, O or S as heteroatoms, and the number of the heteroatoms is 1-3; the substituent of the substituted C5-C8 aryl or 5-8-membered heteroaryl is-H, halogen or-NH₂C1-C6 alkyl, C3-C6 cycloalkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 haloalkoxy, -OH or C1-C6 carbonyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C8 alkyl.

Further, the method comprisesIn a preferred embodiment of the present invention, X is N or C; r₁is-H, -OH, halogen, C1-C8 alkyl, C1-C6 alkoxy, C1-C6 halogenated alkoxy,

C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl,

Or substituted C1-C6 alkyl; the substituent of the substituted C1-C6 alkyl is-H,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl or substituted or unsubstituted 3-6 membered heterocycloalkyl; the heteroatom of the 3-6 membered heterocycloalkyl is N, O or S, and the number of the heteroatoms is 1-3; the substituent for substituting the 3-to 6-membered heterocycloalkyl is-H, -OH, halogen, C1-C4 oxycarbonyl, C1-C4 alkyl, C1-C4 alkoxy or

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₂is-H, -OH, halogen, C1-C6 alkyl, C3-C6 cycloalkyl, C1-C6 haloalkyl, amino-substituted C1-C6 alkyl, benzyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstituted 5-6 membered heteroaryl; the 5-6 membered heteroaryl has N, O or S as heteroatoms, and the number of the heteroatoms is 1-3; the substituent of the substituted C5-C6 aryl or 5-6-membered heteroaryl is-H, halogen or-NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

Preferably, the 4-amino-pyrimidoazepine-phenylurea derivative has X being N or C; r₁is-H, C1-C8 alkyl,

C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl or substituted C1-C4 alkyl; the substituent of the substituted C1-C4 alkyl is-H,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl or substituted or unsubstituted 5-to 6-membered heterocycloalkyl; the heteroatom of the 5-6 membered heterocycloalkyl is N or O, and the number of the heteroatoms is 1-2; the substituent for substituting the 5-to 6-membered heterocycloalkyl is-H, C1-C4 oxycarbonyl, C1-C4 alkyl, C1-C4 alkoxy or

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

Further, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl,

Or

R₄is-H, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl; r₁₁is-H, C1-C4 oxycarbonyl, C1-C4 alkyl, C1-C4 alkoxy or

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

Further, X is N or C; r₁is-H, C1-C8 alkyl,

C2-C6 alkenyl, C2-C6 alkynylC3-C6 cycloalkyl or substituted C1-C4 alkyl; the substituent of the substituted C1-C4 alkyl is-H,

-CN, -OH, phenyl, halogen, methoxy, ethoxy, N-acetyl, N,

Formyl, acetyl,

R₄is-H, C1-C4 alkyl, tert-butyloxy or

R₅～R₁₀Independently is-H, methyl or ethyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

Most preferably, X is N or C; r₁is-H, C1-C8 alkyl,

T-butyloxycarbonyl, C2-C6 alkenyl, C2-C6 alkynyl,

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

Preferably, the 4-amino-pyrimidoazepine-phenylurea derivative has X being N or C; r₂Is benzyl, C3-C6 cycloalkyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstituted 5-6 membered heteroaryl; the 5-6 membered heteroaryl has 1-2 heteroatoms and N or O as heteroatoms; the substituent of the substituted C5-C6 aryl or 5-6-membered heteroaryl is-H, halogen or-NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl;

R₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl or substituted or unsubstituted 5-to 6-membered heterocycloalkyl; the hetero atom of the 5-to 6-membered heterocycloalkyl group is N or OThe number of the active ingredients is 1-2; the substituent for substituting the 5-to 6-membered heterocycloalkyl is-H, C1-C4 oxycarbonyl, C1-C4 alkyl, C1-C4 alkoxy or

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

Further, X is N or C; r₂Is benzyl, C3-C6 cycloalkyl,

R₁₂～R₁₉Independently is-H, halogen, -NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl;

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

Further, X is N or C; r₂Is benzyl, C3-C6 cycloalkyl,

R₁₆～R₁₉Independently is-H, halogen, -NH₂C1-C4 alkyl, C1-C4 haloalkyl, -OH or C1-C4 carbonyl;

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

Still further, X is N or C; r₂Is benzyl, C3-C6 cycloalkyl,

R₁₆～R₁₉Independently is-H, -F, -Cl, -Br, -CF₃Methyl or acetyl;

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

Most preferably, X is N or C; r₂Is composed of

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

Preferably, the 4-amino-pyrimidoazepine-phenylurea derivative has X being N or C; r₃is-H, halogen, -OH or C1-C4 alkyl; r₂Is benzyl, C3-C6 cycloalkyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstituted 5-6 membered heteroaryl; the 5-6 membered heteroaryl has 1-2 heteroatoms and N or O as heteroatoms; the substituent of the substituted C5-C6 aryl or 5-6-membered heteroaryl is-H, halogen or-NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH orC1-C4 carbonyl;

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl.

Further, X is N or C; r₃is-H, halogen or C1-C4 alkyl; r₂Is benzyl, C3-C6 cycloalkyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstituted 5-6 membered heteroaryl; the 5-6 membered heteroaryl has 1-2 heteroatoms and N or O as heteroatoms; the substituent of the substituted C5-C6 aryl or 5-6-membered heteroaryl is-H, halogen or-NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl;

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl.

Further, X is N or C; r₃is-H or halogen; r₂Is benzyl, C3-C6 cycloalkyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstituted 5-6 membered heteroaryl; the 5-6 membered heteroaryl has 1-2 heteroatoms and N or O as heteroatoms; the substituent of the substituted C5-C6 aryl or 5-6-membered heteroaryl is-H, halogen or-NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl;

R₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl or substituted or unsubstituted 5-to 6-membered heterocycloalkyl; the heteroatom of the 5-6 membered heterocycloalkyl is N or O, and the number of the heteroatoms is 1-2; the substituent for substituting the 5-to 6-membered heterocycloalkyl is-H,C1-C4 oxycarbonyl, C1-C4 alkyl, C1-C4 alkoxy or

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl.

Most preferably, X is N or C; r₃is-H, -F, -Cl or-Br; r₂Is benzyl, C3-C6 cycloalkyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstituted 5-6 membered heteroaryl; the 5-6 membered heteroaryl has 1-2 heteroatoms and N or O as heteroatoms; the substituent of the substituted C5-C6 aryl or 5-6-membered heteroaryl is-H, halogen or-NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl;

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl.

As a preferred technical scheme of the invention, the 4-amino-pyrimidoazepine-phenylurea derivative has the structure that X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl,

R₄is-H, C1-C4 alkyl, C1-C4 alkoxy or

R₂Is benzyl, C3-C6 cycloalkyl,

R₃is-H, halogen or C1-C4 alkyl.

Preferably, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, methoxy, ethoxy, N-acetyl, N,

Formyl, acetyl,

R₄is-H, C1-C4 alkyl, tert-butyloxy or

R₅～R₁₀Independently is-H, methyl or ethyl;

R₂is benzyl, C3-C6 cycloalkyl,

R₃is-H or halogen.

Most preferably, X is N or C; r₁is-H, C1-C8 alkyl,

T-butyloxycarbonyl, C2-C6 alkenyl, C2-C6 alkynyl,

R₂Is composed of

R₃is-H, -F, -Cl or-Br.

As a preferred embodiment of the present invention, when R is the above-mentioned 4-amino-pyrimidoazepine-phenylurea derivative₂Is composed of

And the structure is shown as formula II:

wherein X is N or C; r₁is-H, -OH, halogen, C1-C10 alkoxy, C1-C10 haloalkoxy,

C2-C10 alkenyl, C2-C10 alkynyl, C3-C10 cycloalkyl,

R₄is-H,-OH, halogen, C1-C8 alkyl, C1-C8 alkoxy or

R₅～R₁₀Independently is-H or C1-C8 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C10 alkyl.

Preferably, X is N or C; r₁is-H, -OH, halogen, C1-C8 alkoxy, C1-C8 haloalkoxy,

C2-C8 alkenyl, C2-C8 alkynyl, C3-C8 cycloalkyl,

R₄is-H, -OH, halogen, C1-C6 alkyl, C1-C6 alkoxy or

R₅～R₁₀Independently is-H or C1-C6 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C8 alkyl.

Further preferably, X is N or C; r₁is-H, -OH, halogen, C1-C8 alkyl, C1-C6 alkoxy, C1-C6 halogenated alkoxy,

C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

Further, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl or substituted or unsubstituted 5-to 6-membered heterocycloalkyl; the heteroatom of the 5-6 membered heterocycloalkyl is N or O, and the number of the heteroatoms is 1-2; the substituted 5-to 6-membered heteroThe substituent of the cycloalkyl is-H, C1-C4 oxycarbonyl, C1-C4 alkyl, C1-C4 alkoxy or

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen or C1-C4 alkyl.

Further, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl,

R₄is-H, C1-C4 alkyl, C1-C4 alkoxy or

R₃is-H, -OH or halogen.

More preferably, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, methoxy, ethoxy, N-acetyl, N,

Formyl, acetyl,

R₄is-H, C1-C4 alkyl, tert-butyloxy or

R₅～R₁₀Independently is-H, methyl or ethyl;

R₃is-H or halogen.

Most preferably, X is N or C; r₁is-H, C1-C8 alkyl,

T-butyloxycarbonyl, C2-C6 alkenyl, C2-C6 alkynyl,

R₃is-H, -F, -Cl or-Br.

R₃When the structure is-H, the structure is shown as formula III:

wherein X is N or C; r₁is-H, -OH, halogen, C1-C10 alkoxy, C1-C10 haloalkoxy,

C2-C10 alkenyl, C2-C10 alkynyl, C3-C10 cycloalkyl,

R₄is-H, -OH, halogen, C1-C8 alkyl, C1-C8 alkoxy or

R₅～R₁₀Independently is-H or C1-C8 alkyl.

C2-C8 alkenyl, C2-C8 alkynyl, C3EC8 cycloalkyl,

R₄is-H, -OH, halogen, C1-C6 alkyl, C1-C6 alkoxy or

R₅～R₁₀Independently is-H or C1-C6 alkyl.

C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl or substituted or unsubstituted 3-6 membered heterocycloalkyl; the heteroatom of the 3-6 membered heterocycloalkyl is N, O or S, and the number of the heteroatoms is 1-3; the substituent for substituting the 3-to 6-membered heterocycloalkyl is-H, -OH, halogenElements, C1-C4 oxycarbonyl, C1-C4 alkyl, C1-C4 alkoxy or

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl.

Further, X is N or C; r₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl.

Further, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl,

R₄is-H, C1-C4 alkyl, C1-C4 alkoxy or

More preferably, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, methoxy, ethoxy, N-acetyl, N,

Formyl, acetyl,

R₄is-H, C1-C4 alkyl, tert-butyloxy or

R₅～R₁₀Independently is-H, methyl or ethyl.

Most preferably, X is N or C; r₁is-H, C1-C8 alkyl,

T-butyloxycarbonyl, C2-C6 alkenyl, C2-C6 alkynyl,

R₃is-H, X is C, R₁Is composed of

And when the structure is shown as formula IV:

wherein R is₁₆～R₁₉Independently is-H, halogen, -NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl.

Preferably, R₁₆～R₁₉Independently is-H, halogen, -NH₂C1-C4 alkylC1-C4 haloalkyl, -OH or C1-C4 carbonyl.

Most preferably, R₁₆～R₁₉Independently is-H, -F, -Cl, -Br, -CF₃Methyl or acetyl.

The 4-amino-pyrimidoazenitrogen heterocycle-phenylurea derivative has the following structural formula:

the invention also provides a synthesis method of the 4-amino-pyrimidoazenitrogen heterocycle-phenylurea derivative, which mainly adopts the following synthesis route:

the synthesis method of the compound comprises the following steps: the structure of the compound of the general formula (I) is divided into a part A and a part B, wherein the part A is an amine intermediate (formula V) compound, and the part B is an active urea intermediate (formula VI) compound or an isocyanate intermediate.

The first scheme is as follows: the synthesis method of the part A amine intermediate (formula V) compound is as follows:

the synthesis method of the pyrimido pyrrole intermediate is shown as the following scheme:

step a: ammoniation: taking 4-chloro-pyrrolopyrimidine (commercially available) as a starting material SM1, adding ammonia water (g/mL is approximately 5-7 times) into an autoclave, reacting for 4 hours under the pressure of 10-15 atm, cooling, filtering to obtain an intermediate M1, and drying without further purification;

step b: iodination: dissolving the intermediate M1 in a solvent at room temperature, adding N-iodosuccinimide (NIS) (1.5 times equivalent) for reacting for 1 hour, concentrating the solution, adding 30-40 times of water for pulping, adding 0.5 equivalent of sodium thiosulfate, filtering to obtain an intermediate M2, and drying without further purification; the solvent is tetrahydrofuran, DMF or acetonitrile, etc.;

step c: and (4) Boc protection: dissolving the intermediate M2 in a solvent, adding 2 equivalents of organic base, adding 1 equivalent of Boc anhydride (such as di-tert-butyl dicarbonate) and reacting at 50-60 ℃ for 2 hours, concentrating the reaction solution after the reaction is finished, pulping with 30-40 times of water, filtering to obtain an intermediate M3, and drying without further purification; the solvent is acetonitrile, DMF or dioxane, etc.; the organic base is triethylamine, DIEA or pyridine and the like;

step d: suzuki reaction: mixing the intermediate M3 with boric acid/boric acid pinacol ester, alkali, a catalyst and the like, adding a solvent, reacting under the protection of nitrogen at the reaction temperature of 70-90 ℃ for 2-4 hours, treating, and obtaining an intermediate M4 after simple treatment, wherein the intermediate M4 needs to be further purified; the alkali is inorganic alkali such as potassium carbonate, sodium carbonate, potassium bicarbonate or sodium bicarbonate; the catalyst is (dppf) PdCl₂Or tetrakis (triphenylphosphine) palladium; the solvent is a mixed solvent of dioxane, ethanol and water orA mixed solvent of toluene, ethanol and water;

step e: removing Boc: dissolving the intermediate M4 in a solvent at room temperature, adding strong acid, adjusting the pH of the system to about 1, adjusting the pH to be alkaline by using inorganic base after the reaction is finished, adjusting the pH to about 13, extracting by using dichloromethane, concentrating an organic phase, pulping by using diethyl ether, filtering to obtain an intermediate M5, and drying without further purification; the solvent is water or ethanol and the like; the strong acid is inorganic strong acid such as sulfuric acid, methanesulfonic acid or hydrochloric acid; the inorganic alkali is potassium hydroxide or sodium hydroxide and the like;

step f: and (3) substitution: dissolving the intermediate M5 in a solvent, adding 1.2 equivalents of halide, reacting at 70-80 ℃ for 1 hour, filtering the reaction solution to remove inorganic salts to obtain a solution of the intermediate M6 (the compound of formula V), and not requiring further purification; the solvent is acetonitrile, toluene, dioxane or DMF.

The synthesis method of the part B amine intermediate (formula VI) compound is as follows:

dissolving triphosgene in a solvent, adding 1 equivalent of amine raw material, adding organic base, reacting at 60-65 ℃ for 4-5 hours, filtering to remove solid residues after the reaction is finished, and concentrating the filtrate to obtain an intermediate (a compound shown in a formula VI); the solvent is tetrahydrofuran and the like; the organic base is triethylamine or DIEA and the like.

Then, dissolving the intermediate compound (formula V) in a solvent, adding 1 equivalent of the intermediate compound (formula VI), reacting at 70-80 ℃ for 1 hour, precipitating a large amount of solid, and filtering to obtain a filter cake, namely the final urea compound, namely the compound of the general formula (I); the solvent is acetonitrile, dioxane, toluene or DMF, etc.;

wherein X is N or C; r₁is-H, -OH, halogen, C1-C10 alkoxy, C1-C10 haloalkoxy,

C2-C10 alkenyl, C2-C10 alkynyl, C3-C10 cycloalkyl,

R₄is-H, -OH, halogen, C1-C8 alkyl, C1-C8 alkoxy or

R₅～R₁₀Independently is-H or C1-C8 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C10 alkyl.

Scheme II:

the synthesis of the intermediate compound (formula V) is the same as that of the first scheme, the intermediate compound (formula V) is dissolved in a solvent, 1 equivalent of isocyanate intermediate is added, the reaction is carried out for 1 hour at 70-80 ℃, a large amount of solids are separated out, and a filter cake after filtration is the final urea compound, namely the compound of the general formula (I); the solvent is acetonitrile, dioxane, toluene or DMF, etc.;

wherein X is N or C; r₁is-H, -OH, halogen, C1-C10 alkoxy, C1-C10 haloalkoxy,

C2-C10 alkenyl, C2-C10 alkynyl, C3-C10 cycloalkyl,

R₄is-H, -OH, halogen, C1-C8 alkyl, C1-C8 alkoxy or

R₅～R₁₀Independently is-H or C1-C8 alkyl;

R₂is-H, -OH, halogen, C1-C10 alkyl, C3-C10 cycloalkyl, C1-C10 haloalkyl, amino-substituted C1-C10 alkyl, benzyl, substituted or unsubstitutedA C5-C10 aryl or a substituted or unsubstituted 5-to 10-membered heteroaryl; the 5-to 10-membered heteroaryl has N, O or S as heteroatoms, and the number of the heteroatoms is 1-3; the substituent of the substituted C5-C10 aryl or 5-10-membered heteroaryl is-H, halogen or-NH₂C1-C8 alkyl, C3-C8 cycloalkyl, C1-C8 haloalkyl, C1-C8 alkoxy, C1-C8 haloalkoxy, -OH or C1-C8 carbonyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C10 alkyl.

The invention also provides the 4-amino-pyrimidoazenitrogen heterocyclic-phenylurea derivative, including tautomers, stereoisomers, mixtures of all proportions thereof and isotopically substituted compounds thereof.

The invention also provides pharmaceutically acceptable salts of the 4-amino-pyrimidoazepine-phenylurea derivatives. Wherein the salt with an acid is obtained by reacting the free base of the parent compound with an inorganic or organic acid. The inorganic acid includes hydrochloric acid, hydrobromic acid, nitric acid, phosphoric acid, metaphosphoric acid, sulfuric acid, sulfurous acid, perchloric acid and the like. The organic acid includes acetic acid, propionic acid, acrylic acid, oxalic acid, (D) or (L) malic acid, fumaric acid, maleic acid, hydroxybenzoic acid, γ -hydroxybutyric acid, methoxybenzoic acid, phthalic acid, methanesulfonic acid, ethanesulfonic acid, naphthalene-1-sulfonic acid, naphthalene-2-sulfonic acid, p-toluenesulfonic acid, salicylic acid, tartaric acid, citric acid, lactic acid, mandelic acid, succinic acid, malonic acid, or the like.

The term "pharmaceutically acceptable" as used herein, means that which, within the scope of sound medical judgment, is suitable for use in contact with the tissues of human beings and other mammals without undue toxicity, irritation, allergic response and the like, and which, when administered to a recipient, provides, directly or indirectly, a compound of the invention or a prodrug of the compound.

The invention also provides pharmaceutically acceptable hydrates of the 4-amino-pyrimidoazenitrogen heterocyclic-phenylurea derivatives. The term "hydrate" refers to a compound that further binds stoichiometric or non-stoichiometric water by non-covalent intermolecular forces.

The invention also provides pharmaceutically acceptable polymorphic substances of the 4-amino-pyrimidoazepine-phenylurea derivatives. The term "polymorph" denotes a solid crystalline form of a compound or a complex thereof, which can be characterized by physical means, such as x-ray powder diffraction patterns or infrared spectroscopy.

The invention also provides a pharmaceutically acceptable pharmaceutical composition of the 4-amino-pyrimidoazepine-phenylurea derivative, which is prepared by adding pharmaceutically acceptable auxiliary components into the 4-amino-pyrimidoazepine-phenylurea derivative shown in formulas I to IV or salt or hydrate thereof, wherein the auxiliary components are cyclodextrin, arginine or meglumine, the cyclodextrin is selected from α -cyclodextrin, β -cyclodextrin, gamma-cyclodextrin, (C) and (D)_1-4Alkyl) - α -cyclodextrins (C)_1-4Alkyl) - β -cyclodextrins (C)_1-4Alkyl) -gamma-cyclodextrin, (hydroxy-C)_1-4Alkyl) - α -cyclodextrins, (hydroxy-C_1-4Alkyl) - β -cyclodextrins, (hydroxy-C_1-4Alkyl) -gamma-cyclodextrin, (carboxy-C)_1-4Alkyl) - α -cyclodextrins, (carboxy-C_1-4Alkyl) - β -cyclodextrins, (carboxy-C_1-4Alkyl) -gamma-cyclodextrin, α -cyclodextrin saccharide ether, β -cyclodextrin saccharide ether, gamma-cyclodextrin saccharide ether, α -cyclodextrin sulfobutyl ether, β -cyclodextrin sulfobutyl ether and gamma-cyclodextrin sulfobutyl ether.

The pharmaceutical composition may be in liquid form or solid form. Wherein the liquid form may be an aqueous solution. The solid form may be in the form of a powder, granules, tablets or lyophilized powder. The pharmaceutical composition further comprises water for injection, saline solution, aqueous glucose solution, saline for injection/infusion, glucose for injection/infusion, Grignard solution or Grignard solution containing lactate.

The invention also provides application of the 4-amino-pyrimidoazepine-phenylurea derivatives, salts, hydrates or pharmaceutical compositions shown in the formulas I to IV in preparation of FLT3 kinase inhibitors; in particular mutant FLT3 kinase; especially FLT3/ITD mutant kinase.

The invention also provides application of the 4-amino-pyrimidoazepine-phenylurea derivatives, salts, hydrates or pharmaceutical compositions shown in the formulas I to IV in preparing medicines for treating diseases in the aspect of cell proliferation disorder.

The invention also provides application of the 4-amino-pyrimidoazepine-phenylurea derivatives, salts, hydrates or pharmaceutical compositions shown in the formulas I to IV in preparing medicaments for treating tumors.

Further, the tumor is a solid tumor and/or a hematological tumor.

Still further, the solid tumor includes lymphoma, B-cell lymphoma, diffuse large B-cell lymphoma, chronic lymphocytic lymphoma, lymphoplasmacytic lymphoma, ovarian cancer, breast cancer, prostate cancer, bladder cancer, kidney cancer, esophageal cancer, neck cancer, pancreatic cancer, colorectal cancer, gastric cancer, non-small cell lung cancer, thyroid cancer, brain cancer, lymphatic cancer, epidermal hyperproliferation, psoriasis, and/or prostatic hyperplasia.

Still further, the hematologic neoplasm comprises: acute myeloid leukemia, chronic myeloid leukemia, myeloma, acute lymphocytic leukemia, acute myelogenous leukemia, acute promyelocytic leukemia, chronic lymphocytic leukemia, chronic neutrophilic leukemia, acute undifferentiated cell leukemia, myelodysplastic syndrome, myelodysplasia, multiple myeloma, and/or myelosarcoma.

The invention also provides application of the 4-amino-pyrimidoazenitrogen heterocycle-phenylurea derivatives, salts, hydrates or pharmaceutical compositions shown in the formulas I to IV in preparation of oral or intravenous injection preparations. The oral or intravenous injection preparation at least comprises a 4-amino-pyrimidoazepine-phenylurea derivative, salt, hydrate or pharmaceutical composition shown in formulas I-IV and any excipient and/or adjuvant.

The 4-amino-pyrimidoazepine-phenylurea derivative provided by the invention can be used as a FLT3 kinase inhibitor, has a good effect, and provides a new choice for preparing antitumor drugs.

Drawings

FIG. 1MV4-11 verifies target pathways;

FIG. 2Molm-13 verifies the target pathway;

FIG. 3Molm-13 in vivo pharmacodynamic model;

FIG. 4MV4-11 in vivo pharmacodynamic model;

FIG. 5 survival study of NCG mice.

Detailed Description

The present invention will be described in further detail with reference to the following specific embodiments in the form of examples, but the present invention is not limited thereto.

In the examples, the reaction temperature is, without particular mention, room temperature, i.e.from 20 to 30 ℃.

Example 1: 1- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (4-fluorophenyl) urea compound CLJ-1

Step a: preparation of M1 intermediate

Weighing raw materials SM1(20g,130mmol) and ammonia water (150mL, 25% -28% industrial ammonia water), adding into a high-pressure reaction kettle, sealing the system, inserting a thermometer, setting the temperature to 130 ℃, reacting for 4 hours, and cooling to room temperature. The reaction suspension was directly filtered and the filter cake was washed with diethyl ether to give intermediate M1.¹H NMR(400MHz,DMSO):11.45(s,1H),8.03(s,1H),7.06(d,J＝3.2Hz,1H),6.88(s,2H),6.51(d,J＝3.3Hz,1H).

Step b: preparation of M2 intermediate

Intermediate M1(16g,112mmol) from the previous step was added to 200mL tetrahydrofuran at room temperature and NIS (37.8g,168mmol) was added in portions and reacted for 1 h. The reaction suspension was evaporated under reduced pressure to give a solid mixture, 300mL of water was added, sodium thiosulfate (8.8g,56mmol) was added with stirring, the mixture turned from brown to yellow, and stirring was continued for 0.After 5h, filtration was carried out and the filter cake was washed with diethyl ether to give intermediate M2.¹H NMR(400MHz,DMSO):11.97(s,1H),8.07(s,1H),7.36(s,1H),6.56(s,2H).

Step c: preparation of M3 intermediate

Intermediate M2(28g,107.7mmol), potassium carbonate (29.7g,215.4mmol), Boc anhydride (25.8g,118.5mmol) from the previous step was added to 500mL acetonitrile and the resulting mixture was heated to 60 ℃ for 2 h. And filtering the reaction liquid, discarding a filter cake, distilling the mother liquor under reduced pressure to obtain a mixture solid, adding 500mL of water for pulping, continuously stirring for 0.5h, filtering, draining the filter cake, transferring into a vacuum drying oven, and drying at 60 ℃ for 5h to obtain an intermediate M3.¹H NMR(400MHz,DMSO):8.28(s,1H),7.73(s,1H),6.87(s,2H),1.62(s,10H).

Step d: preparation of M4 intermediate

Intermediate M3(9.56g,26.6mmol), p-aminobenzoate (5.5g,31.8mmol), potassium carbonate (7.3g,53.1mmol) and dppf (Pd)₂Cl₂) The mixture was charged into a 250mL three-necked flask, dioxane/ethanol/water (7: 3:4 in total 120mL) was added as a solvent, nitrogen was replaced three times, and the mixture was put into an oil bath at 80 ℃ for reaction for 2 hours. After the reaction is finished, concentrating the reaction solution to be dry, mixing the sample, and separating the mixture by a column to obtain an intermediate M4.¹H NMR(400MHz,DMSO):9.57(s,1H),9.02(s,1H),8.28(s,1H),7.59(d,J＝8.6Hz,2H),7.47(s,1H),7.44(d,J＝8.5Hz,2H),6.52(s,1H),6.23(s,2H),1.61(s,9H),1.31(s,9H).

Step e: preparation of M5 intermediate

Intermediate M4(5g,15.4mmol) from the previous step was added to 50mL of dichloromethane, concentrated hydrochloric acid (10mL, 36-38% concentrated hydrochloric acid) was added, and the reaction was complete after 1 h. 200mL of dichloromethane and 100mL of water were added to adjust the pH of the mixture to basic pH (about 10), the organic phase was retained by extraction and dried to afford intermediate M5.¹H NMR(400MHz,DMSO):11.57(s,1H),8.07(s,1H),7.12(d,J＝8.3Hz,2H),7.03(s,1H),6.66(d,J＝8.3Hz,2H),5.94(s,2H),5.16(s,2H).

Step f: preparation of intermediate M6

Intermediate M5(225mg,1mmol) from the previous step, cesium carbonate (650mg,2mmol), N- (2-chloroethyl) morpholine saltThe acid salt (223mg,1.2mmol) was added to 20mL acetonitrile and heated to 80 ℃ for 1 h. The reaction was spin dried and extracted with dichloromethane to afford the purer intermediate M6.¹H NMR(400MHz,CDCl₃) 8.68(s,1H),7.25(d, J ═ 8.4Hz,4H),7.13(s,1H),6.80(d, J ═ 8.4Hz,2H),4.44(s,2H),3.72(s,4H),2.87(s,2H),2.59(s,4H). Preparation of the end product CLJ-1

Adding the intermediate M6(338mg,1mmol) in the previous step into 20mL of dichloromethane, adding 4-fluoroisocyanate (137mg,1mmol), reacting for 0.5h, separating out a solid, filtering, and rinsing the filter cake with diethyl ether to obtain the high-purity final product CLJ-1.¹H NMR(400MHz,DMSO):9.56(s,1H),9.47(s,1H),8.54(s,1H),7.71(s,1H),7.62(d,J＝8.6Hz,2H),7.52–7.45(m,2H),7.40(d,J＝8.6Hz,2H),7.16–7.10(m,2H),4.75(t,J＝6.5Hz,2H),3.89(s,4H),3.65(t,J＝6.4Hz,4H).HRMS(ESI),m/z:476.2214[M+H]⁺.

EXAMPLE 21- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (2-chlorophenyl) urea Compound CLJ-2

The phenyl 4-fluoroisocyanate was replaced with 2-chlorophenyl isocyanate in step g in the same manner as in the synthesis of example 1 to give CLJ-2 as a final product.¹H NMR(400MHz,DMSO):11.47(s,1H),10.10(s,1H),8.59(s,1H),8.55(s,1H),8.15(dd,J＝8.3,1.5Hz,1H),7.72(s,1H),7.66(d,J＝8.6Hz,2H),7.46(dd,J＝8.0,1.5Hz,1H),7.42(d,J＝8.6Hz,2H),7.34–7.28(m,1H),7.07–7.01(m,1H),4.75(t,J＝6.5Hz,2H),3.89(s,4H),3.65(t,J＝6.4Hz,3H).HRMS(ESI),m/z:492.1922[M+H]⁺.

EXAMPLE 31- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3-benzylurea Compound CLJ-3

In step g, benzyl isocyanate was used as in the synthesis of example 1Replacing 4-phenyl fluoroisocyanate to obtain the final product CLJ-3.¹H NMR(400MHz,DMSO):11.58(s,1H),9.31(s,1H),8.55(s,1H),7.70(s,1H),7.59(d,J＝8.6Hz,2H),7.37–7.30(m,6H),7.27–7.20(m,1H),7.05(s,1H),4.75(t,J＝6.5Hz,2H),4.32(s,2H),3.87(m,9H),3.65(t,J＝6.4Hz,3H).HRMS(ESI),m/z:472.2455[M+H]⁺.

EXAMPLE 41- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (4-chlorophenyl) urea Compound CLJ-4

The phenyl 4-fluoroisocyanate was replaced with phenyl 4-chloroisocyanate in step g in the same manner as in the synthesis of example 1 to give CLJ-4 as a final product.¹H NMR(400MHz,DMSO):8.86(s,1H),8.82(s,1H),8.14(s,1H),7.56(d,J＝8.5Hz,2H),7.53–7.49(m,2H),7.38(d,J＝8.5Hz,2H),7.35–7.30(m,3H),6.05(s,2H),4.28(t,J＝6.5Hz,2H),3.56–3.50(m,4H),2.71(t,J＝6.4Hz,2H),2.46(s,4H).HRMS(ESI),m/z:492.1913[M+H]⁺.

EXAMPLE 51- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (3-chlorophenyl) urea Compound CLJ-5

The phenyl 4-fluoroisocyanate was replaced with 3-chlorophenyl isocyanate in step g in the same manner as in the synthesis of example 1 to give CLJ-5 as a final product.¹H NMR(400MHz,DMSO):9.44(s,1H),9.33(s,1H),8.22(s,1H),7.73(s,1H),7.59(d,J＝8.5Hz,2H),7.43–7.35(m,3H),7.32–7.27(m,2H),7.05–6.98(m,1H),6.39(s,2H),4.49(s,2H),3.73(s,4H),2.98(s,4H).HRMS(ESI),m/z:492.1915[M+H]⁺.

EXAMPLE 61- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (2-fluorophenyl) urea Compound CLJ-6

The phenyl 4-fluoroisocyanate was replaced with phenyl 2-fluoroisocyanate in step g in the same manner as in the synthesis of example 1 to give CLJ-6 as a final product.¹H NMR(400MHz,DMSO):9.23(s,1H),8.62(s,1H),8.16(dd,J＝15.3,6.0Hz,2H),7.57(d,J＝8.4Hz,2H),7.39(d,J＝8.4Hz,2H),7.32(s,1H),7.29–7.20(m,1H),7.15(t,J＝7.6Hz,1H),7.01(dd,J＝12.7,6.5Hz,1H),6.04(s,2H),4.28(t,J＝6.4Hz,2H),3.54(s,4H),2.70(t,J＝6.5Hz,2H),2.45(s,4H).HRMS(ESI),m/z:476.2207[M+H]⁺.

EXAMPLE 71- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (4-chloro-3- (trifluoromethyl) phenyl) urea Compound CLJ-7

The phenyl 4-fluoroisocyanate was replaced in step g with 4-chloro-3-trifluoromethyl-phenyl isocyanate in the same manner as the synthesis of example 1 to give the final product CLJ-7.¹H NMR(400MHz,DMSO):11.11(s,1H),9.99(s,1H),9.66(s,1H),8.52(s,1H),8.14(d,J＝1.8Hz,1H),7.70(s,1H),7.66–7.59(m,3H),7.41(d,J＝8.6Hz,2H),4.73(t,J＝6.3Hz,2H),3.87(s,4H),3.64(m,3H),3.41–3.13(m,4H).HRMS(ESI),m/z:560.1786[M+H]⁺.

EXAMPLE 81- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (p-tolyl) urea Compound CLJ-8

The phenyl 4-fluoroisocyanate was replaced with p-tolylisocyanate in step g in the same manner as in the synthesis of example 1 to give the final product CLJ-8.¹H NMR(400MHz,DMSO):9.46(s,1H),9.20(s,1H),8.52(s,1H),7.69(s,1H),7.62(d,J＝8.6Hz,2H),7.39(d,J＝8.6Hz,2H),7.36(d,J＝8.5Hz,2H),7.09(d,J＝8.3Hz,2H),4.73(t,J＝6.5Hz,2H),3.88(s,4H),3.64(t,J＝6.1Hz,3H),3.44–3.11(m,4H),2.25(s,3H).HRMS(ESI),m/z:472.2458[M+H]⁺.

EXAMPLE 91- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (3-chloro-4-methylphenyl) urea Compound CLJ-9

The phenyl 4-fluoroisocyanate was replaced with 3-chloro-4-methyl-phenyl isocyanate in step g in the same manner as in the synthesis of example 1 to give the final product CLJ-9.¹H NMR(400MHz,DMSO):11.40(s,1H),9.66(d,J＝4.5Hz,2H),8.55(s,1H),7.72(s,1H),7.71(d,J＝1.9Hz,1H),7.62(d,J＝8.6Hz,2H),7.40(d,J＝8.6Hz,2H),7.27–7.19(m,2H),4.75(t,J＝6.5Hz,2H),3.89(s,4H),3.66(t,J＝6.5Hz,3H),2.26(s,3H).HRMS(ESI),m/z:506.2066[M+H]⁺.

EXAMPLE 101- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (3-acetylphenyl) urea Compound CLJ-10

The phenyl 4-fluoroisocyanate was replaced with 3-acetylphenyl isocyanate in step g in the same manner as in the synthesis of example 1 to give the final product CLJ-10.¹H NMR(400MHz,DMSO):11.39(s,1H),9.70(d,J＝8.8Hz,2H),8.54(s,1H),8.11(t,J＝1.8Hz,1H),7.73–7.68(m,2H),7.65(d,J＝8.6Hz,2H),7.61–7.57(m,1H),7.48–7.42(m,1H),7.41(d,J＝8.6Hz,2H),4.75(t,J＝6.5Hz,2H),3.89(s,4H),3.66(t,J＝6.5Hz,2H),2.57(s,3H).HRMS(ESI),m/z:500.2406[M+H]⁺.

EXAMPLE 111- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3-cyclohexylurea compound CLJ-11

In the same manner as in the synthesis of example 1, phenyl 4-fluoroisocyanate was replaced with cyclohexyl isocyanate in step g to giveThe final product CLJ-11.¹H NMR(400MHz,DMSO):8.38(s,1H),8.12(s,1H),7.47(d,J＝8.6Hz,2H),7.30(d,J＝8.6Hz,2H),7.28(s,1H),6.09(d,J＝7.9Hz,1H),4.27(t,J＝6.6Hz,2H),3.56–3.51(m,4H),2.69(t,J＝6.6Hz,2H),2.45(d,J＝4.1Hz,4H),1.81(dd,J＝8.5,3.9Hz,2H),1.67(dd,J＝9.0,4.0Hz,2H),1.59–1.49(m,1H),1.38–1.25(m,2H),1.24–1.12(m,4H).HRMS(ESI),m/z:464.2773[M+H]⁺.

EXAMPLE 121- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (4-bromophenyl) urea Compound CLJ-12

The phenyl 4-fluoroisocyanate was replaced in step g with 4-bromobenzene isocyanate in the same manner as the synthesis of example 1 to give the final product CLJ-12.¹H NMR(400MHz,DMSO):8.86(s,1H),8.82(s,1H),8.14(s,1H),7.56(d,J＝8.5Hz,2H),7.53–7.49(m,2H),7.38(d,J＝8.5Hz,2H),7.35–7.30(m,3H),6.05(s,2H),4.28(t,J＝6.5Hz,2H),3.56–3.50(m,4H),2.71(t,J＝6.4Hz,2H),2.46(s,4H).HRMS(ESI),m/z:536.1408,538.1390[M+H]⁺.

EXAMPLE 131- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-13

Preparation of active urea intermediate 1, 3-bis (5- (tert-butyl) isoxazol-3-yl) urea

Triphosgene (47.07g,156.9mmol) was added to 300mL of tetrahydrofuran, and 3-amino-5-tert-butylisoxazole (20g,142.65mmol) was dissolved in 100mL of tetrahydrofuran under an ice bath, then added dropwise to the triphosgene solution, and finally triethylamine (39.8mL,285.3mmol) was added dropwise. The reaction was transferred to a 60 ℃ oil bath and reacted for 5 h. After the reaction is completed, the reaction mixture is filtered, the filtrate is retained, and the filtrate is concentrated under reduced pressure to obtain a solid, namely the active urea intermediate.¹H NMR(400MHz,DMSO):6.71(s,1H),6.35(s,1H),4.90(s,2H),1.36(s,9H),1.35(s,9H).

Combining steps f and h into a one-pot reaction process

Intermediate M5(225mg,1mmol) from the previous step, cesium carbonate (650mg,2mmol), N- (2-chloroethyl) morpholine hydrochloride (223mg,1.2mmol) were added to 20mL acetonitrile and heated to 80 ℃ for 1 h. The reaction solution was filtered, and the mother liquor was retained. Transferring the mother liquor to 80 ℃, adding an active urea intermediate (306mg,1mmol), reacting for 0.5h, separating out a large amount of solid, filtering, and leaching with diethyl ether to obtain the high-purity final product CLJ-13.¹H NMR(400MHz,DMSO):9.53(s,1H),8.91(s,1H),8.14(s,1H),7.56(d,J＝8.3Hz,2H),7.40(d,J＝8.2Hz,2H),7.33(s,1H),6.52(s,1H),6.06(s,2H),4.29(t,J＝6.3Hz,2H),3.54(s,4H),2.71(t,J＝6.4Hz,2H),2.46(s,4H),1.31(s,9H).HRMS(ESI),m/z:505.2672[M+H]⁺.

Example 141- (4- (4-amino-7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-14

And adding the intermediate M5(225mg,1mmol) in the last step into 20mL of acetonitrile, heating to 80 ℃ for reaction, adding an active urea intermediate (306mg,1mmol), reacting for 0.5h, precipitating a large amount of solid, filtering, and leaching with diethyl ether to obtain the high-purity final product CLJ-14.¹H NMR(400MHz,DMSO):11.74(s,1H),9.53(s,1H),8.93(s,1H),8.11(s,1H),7.55(d,J＝8.5Hz,2H),7.40(d,J＝8.5Hz,2H),7.19(d,J＝2.3Hz,1H),6.52(s,1H),5.99(s,2H),1.31(s,9H).HRMS(ESI),m/z:392.1831[M+H]⁺.

EXAMPLE 151- (4- (4-amino-7-tert-butoxycarbonyl-7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea compound CLJ-15

Intermediate M4(325mg,1mmol) from the previous step was added to 20mL acetonitrileHeating to 80 ℃ for reaction, adding an active urea intermediate (306mg,1mmol), reacting for 0.5h, separating out a large amount of solid, filtering, and leaching with diethyl ether to obtain the high-purity final product CLJ-15.¹H NMR(400MHz,DMSO):9.57(s,1H),9.02(s,1H),8.28(s,1H),7.59(d,J＝8.6Hz,2H),7.47(s,1H),7.44(d,J＝8.5Hz,2H),6.52(s,1H),6.23(s,2H),1.61(s,9H),1.31(s,9H).HRMS(ESI),m/z:592.2353[M+H]⁺.

EXAMPLE 164- (2- (4-amino-5- (4- (3- (5- (tert-butyl) isoxazol-3-yl) ureido) phenyl) -7H-pyrrolo [2,3-d ] tert-butylpyrimidin-7-yl) ethyl) piperazine-1-carboxylic acid tert-butyl ester Compound CLJ-16

In the same manner as in example 13, N-Boc-2-ethylpiperazine was used in place of N- (2-chloroethyl) morpholine hydrochloride, to give CLJ-16 as a highly pure final product.¹H NMR(400MHz,DMSO):9.53(s,1H),8.91(s,1H),8.14(s,1H),7.56(d,J＝8.5Hz,2H),7.39(d,J＝8.5Hz,2H),7.33(s,1H),6.52(s,1H),6.06(s,2H),4.29(t,J＝6.5Hz,2H),3.28(s,4H),2.73(t,J＝6.5Hz,2H),2.46–2.37(m,4H),1.39(s,9H),1.31(s,9H).HRMS(ESI),m/z:604.3353[M+H]⁺.

Example 171- (4- (4-amino-7- (2-piperazinoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-17

Compound CLJ-16(252mg,0.5mmol) was added to 20mL of methylene chloride, concentrated hydrochloric acid (10mL, 36-38% concentrated hydrochloric acid) was added, and the reaction was completed after 1 h. 200mL of methylene chloride and 100mL of water were added to adjust the pH of the mixture to basic pH (about 10), the organic phase was retained by extraction, and the final product, CLJ-17, was obtained after concentration.¹H NMR(400MHz,DMSO):9.58(s,1H),9.01(s,1H),8.14(s,1H),7.57(d,J＝8.5Hz,2H),7.39(d,J＝8.5Hz,2H),7.33(s,1H),6.52(s,1H),6.05(s,2H),4.27(t,J＝6.6Hz,2H),2.75–2.63(m,6H),2.41(s,4H),1.31(s,9H).HRMS(ESI),m/z:504.2829[M+H]⁺.

Example 181- (4- (4-amino-7- (3-morpholinopropyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-18

In the same manner as in example 13, high-purity CLJ-18 was obtained by substituting N- (2-chloroethyl) morpholine hydrochloride with N- (3-chloropropyl) morpholine.¹H NMR(400MHz,DMSO):9.53(s,1H),8.90(s,1H),8.14(s,1H),7.56(d,J＝8.6Hz,2H),7.40(d,J＝8.6Hz,2H),7.31(s,1H),6.52(s,1H),6.05(s,2H),4.20(t,J＝7.0Hz,2H),3.59–3.51(m,4H),2.29(dd,J＝14.3,7.0Hz,6H),2.02–1.91(m,2H),1.31(s,9H).HRMS(ESI),m/z:519.2825[M+H]⁺.

EXAMPLE 191- (4- (4-amino-7- (2- (dimethylamino) ethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea compound CLJ-19

In the same manner as in example 13, high-purity CLJ-19 was obtained by substituting N, N-dimethyl-2-chloroethyl for N- (2-chloroethyl) morpholine hydrochloride.¹H NMR(400MHz,DMSO):9.54(s,1H),8.93(s,1H),8.14(s,1H),7.56(d,J＝8.3Hz,2H),7.39(d,J＝8.3Hz,2H),7.33(s,1H),6.52(s,1H),6.05(s,2H),4.26(t,J＝6.6Hz,2H),2.67(t,J＝6.5Hz,2H),2.20(s,6H),1.31(s,9H).HRMS(ESI),m/z:463.2563[M+H]⁺.

EXAMPLE 201- (4- (4-amino-7-allyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-20

In the same manner as in the synthesis of example 13, bromopropene was used in place of N- (2-chloroethyl) morpholine hydrochlorideThus obtaining the high-purity final product CLJ-20.¹H NMR(400MHz,DMSO):9.53(s,1H),8.91(s,1H),8.15(s,1H),7.56(d,J＝8.5Hz,2H),7.40(d,J＝8.4Hz,2H),7.25(s,1H),6.52(s,1H),6.06(m,3H),5.18(d,J＝10.1Hz,1H),5.09(d,J＝17.1Hz,1H),4.81(d,J＝5.4Hz,2H),1.31(s,9H).HRMS(ESI),m/z:432.2149[M+H]⁺.

Example 211- (4- (4-amino-7- (cyanomethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-21

High purity final product CLJ-21 was obtained by substituting chloroacetonitrile for N- (2-chloroethyl) morpholine hydrochloride in the same manner as in example 13.¹H NMR(400MHz,DMSO):9.54(s,1H),8.94(s,1H),8.24(s,1H),7.58(d,J＝7.4Hz,2H),7.42(d,J＝7.6Hz,2H),7.38(s,1H),6.53(s,1H),6.26(s,2H),5.41(s,2H),1.31(s,9H).HRMS(ESI),m/z:431.1937[M+H]⁺.

Example 221- (4- (4-amino-7-isopropyl-7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-22

In the same manner as in example 13, high-purity CLJ-22 was obtained by substituting 2-iodopropane for N- (2-chloroethyl) morpholine hydrochloride.¹H NMR(400MHz,DMSO):9.55(s,1H),8.97(s,1H),8.33(s,1H),7.65(s,1H),7.59(d,J＝8.6Hz,2H),7.43(d,J＝8.5Hz,2H),6.52(s,1H),5.10–4.93(m,1H),1.49(d,J＝6.8Hz,6H),1.31(s,9H).HRMS(ESI),m/z:434.2299[M+H]⁺.

EXAMPLE 231- (4- (4-amino-7- (4-hydroxybutyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-23

In the same manner as in example 13, 4-iodobutanol was used in place of N- (2-chloroethyl) morpholine hydrochloride to give high purity CLJ-23.¹H NMR(400MHz,DMSO):9.53(s,1H),8.91(s,1H),8.14(s,1H),7.56(d,J＝8.4Hz,2H),7.40(d,J＝8.4Hz,2H),7.32(s,1H),6.52(s,1H),6.05(s,2H),4.41(t,J＝5.2Hz,1H),4.17(t,J＝7.0Hz,2H),4.09(q,J＝5.2Hz,1H),3.41(dd,J＝11.8,6.2Hz,2H),3.18(d,J＝5.2Hz,2H),1.88–1.76(m,2H),1.46–1.36(m,2H),1.31(s,9H).HRMS(ESI),m/z:464.2409[M+H]⁺.

Example 241- (4- (4-amino-7- (N, N-diethylformyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-24

In the same manner as in example 13, N-diethylchloroformamide was used in place of N- (2-chloroethyl) morpholine hydrochloride to obtain high-purity CLJ-24.¹H NMR(400MHz,DMSO):9.55(s,1H),8.96(s,1H),8.20(s,1H),7.58(d,J＝8.6Hz,2H),7.44(d,J＝8.5Hz,2H),7.41(s,1H),6.53(s,1H),6.25(s,2H),3.39(m,4H),1.31(s,9H),1.21–1.06(m,6H).HRMS(ESI),m/z:491.2493[M+H]⁺.

EXAMPLE 251- (4- (4-amino-7- (2- (methylsulfonyl) piperazin-1-yl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-25

Compound CLJ-17(252mg,0.5mmol) was dissolved in 20mL dichloromethane, triethylamine (61mg,0.6mmol) was added, methanesulfonyl chloride (57mg,0.5mmol) diluted with dichloromethane was added under ice bath, after 1h reaction, extraction was performed with dichloromethane and water, and the organic phase was dried by spin drying to give the purer final product CLJ-25.¹H NMR(400MHz,DMSO):9.53(s,1H),8.92(s,1H),8.15(s,1H),7.56(d,J＝8.6Hz,2H),7.40(d,J＝8.5Hz,2H),7.34(s,1H),6.52(s,1H),6.05(s,2H),4.30(t,J＝6.4Hz,2H),3.12–3.03(m,4H),2.85(s,3H),2.79(t,J＝6.5Hz,2H),2.62–2.54(m,4H),1.31(s,9H).HRMS(ESI),m/z:582.2610[M+H]⁺.

EXAMPLE 261- (4- (4-amino-7- (2-methoxyethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-26

In the same manner as the synthesis method in example 13, N- (2-chloroethyl) morpholine hydrochloride was replaced with chloroethyl methyl ether to obtain high-purity final product CLJ-26.¹H NMR(400MHz,DMSO):9.56(s,1H),8.95(s,1H),8.15(s,1H),7.56(d,J＝8.4Hz,2H),7.39(d,J＝8.4Hz,2H),7.29(s,1H),6.53(s,1H),6.09(s,2H),4.33(t,J＝5.3Hz,2H),3.71(t,J＝5.4Hz,2H),3.26(s,3H),1.31(s,9H).HRMS(ESI),m/z:450.2249[M+H]⁺.

EXAMPLE 271- (4- (4-amino-7- (but-2-yn-1-yl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-27

In the same manner as in example 13, 1-bromo-2-butyne was used in place of N- (2-chloroethyl) morpholine hydrochloride to give high purity final product CLJ-27.¹H NMR(400MHz,DMSO):9.55(s,1H),8.97(s,1H),8.17(s,1H),7.57(d,J＝8.5Hz,2H),7.41(d,J＝8.4Hz,2H),7.32(s,1H),6.53(s,1H),6.12(s,1H),4.97(d,J＝2.3Hz,2H),1.81(dd,J＝5.3,3.1Hz,3H),1.31(s,9H).HRMS(ESI),m/z:444.2147[M+H]⁺.

Example 281- (4- (4-amino-7- (pent-2-yn-1-yl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-28

1-bromo was reacted as in the synthesis of example 13The N- (2-chloroethyl) morpholine hydrochloride is replaced by the-2-pentyne, and the high-purity final product CLJ-28 can be obtained.¹H NMR(400MHz,DMSO):9.53(s,1H),8.93(s,1H),8.16(s,1H),7.56(d,J＝8.6Hz,2H),7.40(d,J＝8.5Hz,2H),7.31(s,1H),6.52(s,1H),6.12(s,2H),4.98(t,J＝2.1Hz,2H),2.25–2.16(m,2H),1.30(s,9H),1.06(t,J＝7.5Hz,3H).HRMS(ESI),m/z:458.2299[M+H]⁺.

EXAMPLE 291- (4- (4-amino-7- (but-3-yn-1-yl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-29

In the same manner as in example 13, 1-bromo-2-pentyne was substituted for N- (2-chloroethyl) morpholine hydrochloride to give high purity final product CLJ-29.¹H NMR(400MHz,DMSO):9.57(s,1H),8.99(s,1H),8.15(s,1H),7.56(d,J＝8.4Hz,2H),7.39(d,J＝8.3Hz,2H),7.32(s,1H),6.52(s,1H),6.05(s,2H),5.82(ddd,J＝23.8,10.3,6.7Hz,1H),5.14–4.93(m,2H),4.24(t,J＝7.0Hz,2H),2.56(dd,J＝13.3,6.5Hz,2H),1.31(s,9H).HRMS(ESI),m/z:446.2302[M+H]⁺.

EXAMPLE 301- (4- (4-amino-7- (2-ethoxyethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea compound CLJ-30

The same synthesis method as that in example 13 is adopted, and bromoethyl ether is used to replace N- (2-chloroethyl) morpholine hydrochloride to obtain the high-purity final product CLJ-30.¹H NMR(400MHz,DMSO):9.53(s,1H),8.92(s,1H),8.15(s,1H),7.56(d,J＝8.4Hz,2H),7.39(d,J＝8.4Hz,2H),7.29(s,1H),6.52(s,1H),6.06(s,2H),4.32(t,J＝5.4Hz,2H),3.74(t,J＝5.5Hz,2H),3.46(q,J＝6.9Hz,2H),1.31(s,9H).HRMS(ESI),m/z:464.2406[M+H]⁺.

EXAMPLE 311- (4- (4-amino-7- (3-methylbut-2-en-1-yl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-31

In the same manner as in example 13, 3-dimethylallyl bromide was used in place of N- (2-chloroethyl) morpholine hydrochloride to give high-purity CLJ-31 as a final product.¹H NMR(400MHz,DMSO):9.53(s,1H),8.92(s,1H),8.15(s,1H),7.56(d,J＝8.4Hz,2H),7.39(d,J＝8.4Hz,2H),7.23(s,1H),6.52(s,1H),6.05(s,2H),5.41(t,J＝6.8Hz,1H),4.76(d,J＝7.0Hz,2H),1.81(s,3H),1.72(s,3H),1.31(s,9H).HRMS(ESI),m/z:460.2457[M+H]⁺.

Example 321- (4- (4-amino-7- (2-methylallyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-32

In the same manner as in example 13, 2-methylallyl bromide was used in place of N- (2-chloroethyl) morpholine hydrochloride to give high-purity CLJ-32 as a final product.¹H NMR(400MHz,DMSO):9.53(s,1H),8.92(s,1H),8.15(s,1H),7.56(d,J＝8.6Hz,2H),7.40(d,J＝8.6Hz,2H),7.20(s,1H),6.52(s,1H),6.09(s,2H),4.87(s,1H),4.73(s,2H),4.61(s,1H),1.68(s,3H),1.31(s,9H).HRMS(ESI),m/z:446.2298[M+H]⁺.

EXAMPLE 331- (4- (4-amino-7- (pent-4-en-1-yl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-33

In the same manner as in example 13, high-purity CLJ-33 was obtained by substituting 5-bromo-1-pentene for N- (2-chloroethyl) morpholine hydrochloride.¹H NMR(400MHz,DMSO):9.77(s,1H),9.58(s,1H),8.48(s,1H),7.68(s,1H),7.61(d,J＝8.6Hz,2H),7.42(d,J＝8.6Hz,2H),6.53(s,1H),5.83(ddt,J＝16.8,10.2,6.4Hz,1H),5.09–4.95(m,2H),4.27(t,J＝7.0Hz,2H),2.05(dd,J＝13.7,6.7Hz,2H),1.99–1.87(m,2H),1.31(s,9H).HRMS(ESI),m/z:460.2458[M+H]⁺.

EXAMPLE 341- (4- (4-amino-7- (hex-5-en-1-yl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-34

In the same manner as in example 13, high-purity CLJ-34 was obtained by substituting 5-bromo-1-pentene for N- (2-chloroethyl) morpholine hydrochloride.¹H NMR(400MHz,DMSO):9.78(s,1H),9.60(s,1H),8.48(s,1H),7.68(s,1H),7.61(d,J＝8.6Hz,2H),7.41(d,J＝8.6Hz,2H),6.53(s,1H),5.78(ddt,J＝16.9,10.2,6.7Hz,1H),5.05–4.90(m,2H),4.27(t,J＝7.0Hz,2H),2.06(q,J＝7.2Hz,2H),1.83(dt,J＝14.9,7.2Hz,2H),1.41–1.31(m,2H),1.31(s,9H).HRMS(ESI),m/z:474.2618[M+H]⁺.

Example 351- (4- (4-amino-7-methyl-7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-35

The same synthesis method as that in example 13 was used to replace N- (2-chloroethyl) morpholine hydrochloride with methyl iodide to obtain high purity final product CLJ-35.¹H NMR(400MHz,DMSO):9.53(s,1H),8.93(s,1H),8.11(s,1H),7.55(d,J＝8.5Hz,2H),7.40(d,J＝8.5Hz,2H),7.19(d,J＝2.3Hz,1H),6.52(s,1H),5.99(s,2H),3.7(s,3H),1.31(s,9H).HRMS(ESI),m/z:406.1992[M+H]⁺.

EXAMPLE 361- (4- (4-amino-7-ethyl-7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-36

Same as the synthesis method of example 13And simultaneously, the iodoethane is used for replacing N- (2-chloroethyl) morpholine hydrochloride to obtain the high-purity final product CLJ-36.¹H NMR(400MHz,DMSO):9.78(s,1H),9.61(s,1H),8.48(s,1H),7.69(s,1H),7.61(d,J＝8.6Hz,2H),7.42(d,J＝8.5Hz,2H),6.53(s,1H),4.30(q,J＝7.2Hz,2H),1.43(t,J＝7.2Hz,3H),1.31(s,9H).HRMS(ESI),m/z:420.2142[M+H]⁺.

Example 371- (4- (4-amino-7-propyl-7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-37

High purity final product CLJ-37 was obtained by substituting iodopropane for N- (2-chloroethyl) morpholine hydrochloride in the same manner as in the synthesis of example 13.¹H NMR(400MHz,DMSO):9.80(s,1H),9.66(s,1H),8.48(s,1H),7.68(s,1H),7.61(d,J＝8.5Hz,2H),7.41(d,J＝8.5Hz,2H),6.53(s,1H),4.23(t,J＝7.0Hz,2H),1.90–1.77(m,2H),1.31(s,9H),0.87(t,J＝7.4Hz,3H).HRMS(ESI),m/z:434.2303[M+H]⁺.

Example 381- (4- (4-amino-7-butyl-7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-38

High purity final product CLJ-38 was obtained by substituting N- (2-chloroethyl) morpholine hydrochloride with bromobutane in the same manner as in the synthesis of example 13.¹H NMR(400MHz,DMSO):9.78(s,1H),9.61(s,1H),8.48(s,1H),7.68(s,1H),7.61(d,J＝8.6Hz,2H),7.41(d,J＝8.5Hz,2H),6.53(s,1H),4.26(t,J＝7.1Hz,2H),1.87–1.75(m,2H),1.31(s,9H),1.26(dd,J＝14.9,7.5Hz,2H),0.91(t,J＝7.4Hz,3H).HRMS(ESI),m/z:448.2460[M+H]⁺.

Example 391- (4- (4-amino-7-cyclopentyl-7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-39

The same synthesis method as that in example 13 is adopted, and bromocyclopentane is used to replace N- (2-chloroethyl) morpholine hydrochloride to obtain high-purity final product CLJ-39.¹H NMR(400MHz,DMSO):9.78(s,1H),9.62(s,1H),8.47(s,1H),7.73(s,1H),7.61(d,J＝8.6Hz,2H),7.44(d,J＝8.5Hz,2H),6.53(s,1H),5.22–5.08(m,1H),2.16(d,J＝7.9Hz,2H),2.02–1.83(m,4H),1.71(d,J＝7.0Hz,2H),1.31(s,9H).HRMS(ESI),m/z:460.2457[M+H]⁺.

EXAMPLE 401- (4- (4-amino-7-hexyl-7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-40

The same synthesis method as that in example 13 was used to replace N- (2-chloroethyl) morpholine hydrochloride with N-bromo-hexane to obtain high-purity final product CLJ-40.¹H NMR(400MHz,DMSO):9.71(s,1H),9.40(s,1H),8.46(s,1H),7.67(s,1H),7.61(d,J＝8.6Hz,2H),7.41(d,J＝8.5Hz,2H),6.52(s,1H),4.25(t,J＝7.1Hz,2H),1.88–1.77(m,2H),1.31(s,9H),1.28(m,6H),0.86(t,J＝6.8Hz,3H).HRMS(ESI),m/z:476.2770[M+H]⁺.

EXAMPLE 411- (4- (4-amino-7-benzyl-7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-41

High purity final product CLJ-41 was obtained by substituting benzyl bromide for N- (2-chloroethyl) morpholine hydrochloride in the same manner as in the synthesis of example 13.¹H NMR(400MHz,DMSO):9.71(s,1H),8.49(s,1H),7.74(s,1H),7.60(d,J＝8.6Hz,2H),7.41(d,J＝8.5Hz,2H),7.38–7.27(m,5H),6.52(s,1H),5.49(s,2H),1.31(s,9H).HRMS(ESI),m/z:482.2303[M+H]⁺.

Example 421- (4- (4-amino-7-propynyl-7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-42

The same synthesis method as that in example 13 is adopted, and N- (2-chloroethyl) morpholine hydrochloride is replaced by propargyl bromide to obtain the high-purity final product CLJ-42.¹H NMR(400MHz,DMSO):9.66(s,1H),9.28(s,1H),8.49(s,1H),7.66(s,1H),7.61(d,J＝8.5Hz,2H),7.43(d,J＝8.5Hz,2H),6.52(s,1H),5.15(d,J＝2.3Hz,2H),3.52(t,J＝2.4Hz,2H),1.31(s,9H).HRMS(ESI),m/z:430.1985[M+H]⁺.

Example 431- (4- (4-amino-7- (2-cyanoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-43

The high purity final product CLJ-43 can be obtained by replacing N- (2-chloroethyl) morpholine hydrochloride with bromopropionitrile in the same way as the synthesis method of the embodiment 13.¹H NMR(400MHz,DMSO):9.71(s,1H),9.42(s,1H),8.50(s,1H),7.70(s,1H),7.63(d,J＝8.6Hz,2H),7.42(d,J＝8.5Hz,2H),6.53(s,1H),4.56(t,J＝6.4Hz,2H),3.19(t,J＝6.4Hz,2H),1.31(s,9H).HRMS(ESI),m/z:445.2097[M+H]⁺.

Example 441- (4- (4-amino-7H-pyrrolo [2,3-d ] pyrimidin-5-) -2-fluorophenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-44

Step i: preparation of M7 intermediate

Intermediate M3(9g,25mmol), pinacol 3-fluoro-4-aminophenylborate (6.5g,27.5mmol), potassium carbonate (6.9g,50mmol) and dppf (PdCl)₂) The mixture was charged into a 250mL three-necked flask, dioxane/ethanol/water (7: 3:4 in total 120mL) was added as a solvent, nitrogen was replaced three times, and the mixture was put into an oil bath at 80 ℃ for reaction for 2 hours. Inverse directionAfter the reaction is finished, the reaction solution is concentrated and then is mixed with a sample and is separated by a column, and an intermediate M7 is obtained.

Step j: preparation of M8 intermediate

Intermediate M7(4g,11.6mmol) from the previous step was added to 50mL of dichloromethane, concentrated hydrochloric acid (10mL, 36-38% concentrated hydrochloric acid) was added, and the reaction was complete after 1 h. 200mL of dichloromethane and 100mL of water were added to adjust the pH of the mixture to basic pH (about 10), the organic phase was retained by extraction and dried to afford intermediate M8.¹H NMR(400MHz,DMSO):11.66(s,1H),8.08(s,1H),7.11(d,J＝2.1Hz,1H),7.07(dd,J＝12.5,1.8Hz,1H),6.98(dd,J＝8.1,1.8Hz,1H),6.85(dd,J＝9.5,8.2Hz,1H),5.99(s,2H),5.21(s,2H).

Step k: preparation of CLJ-44

Adding the intermediate M8(243mg,1mmol) in the previous step into 20mL of acetonitrile, adding an active urea intermediate (306mg,1mmol), reacting for 0.5h, separating out a solid, filtering, and rinsing the filter cake with diethyl ether to obtain the high-purity final product CLJ-44.¹H NMR(400MHz,DMSO):13.04(s,1H),10.13(s,1H),9.20(s,1H),8.46(s,1H),8.23(t,J＝8.5Hz,1H),7.61(d,J＝2.3Hz,1H),7.37(dd,J＝12.0,1.7Hz,1H),7.27(d,J＝8.4Hz,1H),6.51(s,2H),1.31(s,9H).HRMS(ESI),m/z:410.1734[M+H]⁺.

EXAMPLE 451- (4- (4-amino-7-propynyl-7H-pyrrolo [2,3-d ] pyrimidin-5-) -2-fluorophenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-45

Combining steps l and m into a reaction process of a' one-pot method

Intermediate M8(243mg,1mmol), cesium carbonate (650mg,2mmol), and bromopropyne (143mg,1.2mmol) from the previous step were added to 20mL of acetonitrile, and heated to 80 ℃ for 1 h. The reaction solution was filtered, and the mother liquor was retained. The mother liquor was transferred to 80 ℃ and the active urea intermediate (306mg,1mmol) was added, reaction 0.After 5h, a large amount of solid is separated out, filtered and rinsed by ether, and the high-purity final product CLJ-45 can be obtained.¹H NMR(400MHz,DMSO):10.01(s,1H),9.08(s,1H),8.52(s,1H),8.26(t,J＝8.5Hz,1H),7.73(s,1H),7.39(dd,J＝12.0,1.8Hz,1H),7.30–7.25(m,1H),6.51(s,1H),5.16(d,J＝2.4Hz,2H),1.31(s,9H).HRMS(ESI),m/z:448.1891[M+H]⁺.

Example 461- (4- (4-amino-7-allyl-7H-pyrrolo [2,3-d ] pyrimidin-5) -2-fluorophenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-46

The high purity final product, CLJ-46, was obtained by substituting bromopropyne for bromopropene in the same manner as in example 45.¹H NMR(400MHz,DMSO):9.86(s,1H),8.88(s,1H),8.20(t,J＝8.5Hz,1H),8.16(s,1H),7.33(s,1H),7.33(dd,J＝12.1,1.8Hz,2H),7.26(dd,J＝8.5,1.3Hz,1H),6.51(s,1H),6.19(s,2H),6.05(ddd,J＝22.5,10.6,5.5Hz,1H),5.18(dd,J＝10.2,1.3Hz,1H),5.09(dd,J＝17.1,1.4Hz,1H),4.80(d,J＝5.5Hz,2H),1.31(s,9H).HRMS(ESI),m/z:450.2042[M+H]⁺.

EXAMPLE 471- (4- (4-amino-7- (but-3-en-1-yl) -7H-pyrrolo [2,3-d ] pyrimidin-5) -2-fluorophenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-47

In the same manner as in example 45, the bromopropyne was replaced with 4-bromo-1-butene, thereby obtaining high-purity final product CLJ-47.¹H NMR(400MHz,DMSO):10.05(s,1H),9.12(s,1H),8.49(s,1H),8.25(t,J＝8.5Hz,1H),7.73(s,1H),7.36(dd,J＝12.0,1.7Hz,1H),7.26(d,J＝8.4Hz,1H),6.51(s,1H),5.80(ddt,J＝17.0,10.3,6.7Hz,1H),5.10–4.99(m,2H),4.34(t,J＝7.0Hz,2H),2.61(q,J＝6.8Hz,2H),1.31(s,9H).HRMS(ESI),m/z:464.2212[M+H]⁺.

Example 481- (4- (4-amino-7-methoxyethyl-7H-pyrrolo [2,3-d ] pyrimidin-5) -2-fluorophenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-48

In the same manner as the synthesis method in example 45, bromopropyne was replaced with chloroethyl methyl ether, thereby obtaining high-purity final product CLJ-48.¹H NMR(400MHz,DMSO):10.10(s,1H),9.16(s,1H),8.51(s,1H),8.25(t,J＝8.5Hz,1H),7.70(s,1H),7.43–7.33(m,1H),7.26(m,1H),6.51(s,1H),4.43(t,J＝5.1Hz,2H),3.76(t,J＝5.3Hz,2H),1.31(s,9H).HRMS(ESI),m/z:468.2159[M+H]⁺.

Example 491- (4- (4-amino-7-ethoxyethyl-7H-pyrrolo [2,3-d ] pyrimidin-5) -2-fluorophenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-49

The synthesis method of example 45 was followed, and bromopropyne was replaced with bromoethyl ether to obtain high-purity final product CLJ-49.¹H NMR(400MHz,DMSO):10.02(s,1H),9.09(s,1H),8.48(s,1H),8.26(t,J＝8.5Hz,1H),7.68(s,1H),7.37(dd,J＝12.0,1.8Hz,1H),7.27(d,J＝8.5Hz,1H),6.51(s,1H),4.42(t,J＝5.3Hz,2H),3.78(t,J＝5.3Hz,2H),3.46(dd,J＝14.0,7.0Hz,2H),1.31(s,9H),1.06(t,J＝7.0Hz,3H).HRMS(ESI),m/z:482.2313[M+H]⁺.

Example 501- (4- (4-amino-7-cyanomethyl-7H-pyrrolo [2,3-d ] pyrimidin-5) -2-fluorophenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-50

In the same manner as in example 45, the bromopropyne was replaced with chloroacetonitrile, whereby CLJ-50, a high-purity final product, was obtained.¹H NMR(400MHz,DMSO):10.10(s,1H),9.16(s,1H),8.57(s,1H),8.26(t,J＝8.5Hz,1H),7.72(s,1H),7.39(dd,J＝11.9,1.8Hz,1H),7.28(d,J＝8.4Hz,1H),6.51(s,1H),5.55(s,2H),1.31(s,9H).HRMS(ESI),m/z:449.1848[M+H]⁺.

EXAMPLE 511- (4- (4-amino-7- (2-cyanoethyl) -7H-pyrrolo [2,3-d ] pyrimidin-5) -2-fluorophenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-51

In the same manner as in example 45, the bromopropyne was replaced with bromopropionitrile, whereby CLJ-51, a high-purity final product, was obtained.¹H NMR(400MHz,DMSO):10.05(s,1H),9.12(s,1H),8.54(s,1H),8.28(t,J＝8.5Hz,1H),7.79(s,1H),7.65(s,1H),7.37(dd,J＝11.9,1.8Hz,1H),7.28(d,J＝8.5Hz,1H),6.51(s,1H),4.60–4.51(t,J＝6.5Hz,2H),3.19(t,J＝6.6Hz,2H),1.31(s,9H).HRMS(ESI),m/z:463.2004[M+H]⁺.

Example 521- (4- (4-amino-7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-52

The synthetic route is shown as follows;

the synthetic route corresponds essentially to example 13, except that the starting material is replaced by 4-chloropyrazolo [2,3-d ]]Pyrimidine is reacted in 6 steps to obtain the final product CLJ-52.¹H NMR(400MHz,DMSO):13.80(s,1H),9.35(s,1H),9.14(s,1H),8.50(s,1H),7.53(s,2H),7.24–7.18(m,2H),6.73–6.67(m,2H),6.46(s,1H),1.29(s,9H).HRMS(ESI),m/z:392.1907[M+H]⁺.

Example 531- (4- (4-amino-7-allyl-7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea compound CLJ-53

In line with the route of example 52, except that bromopropene was added in g, and the reactive urea intermediate reacted with the next step was synthesized according to the "one-pot" synthesis to give final product CLJ-53.¹H NMR(400MHz,DMSO):9.56(s,1H),9.02(s,1H),8.26(s,1H),7.63(t,J＝7.0Hz,4H),7.00(dd,J＝114.0,63.4Hz,2H),6.53(s,1H),6.10–6.01(m,1H),5.19(d,J＝10.3Hz,1H),5.11(d,J＝17.1Hz,1H),4.97(d,J＝5.5Hz,2H),1.31(s,9H).HRMS(ESI),m/z:433.2099[M+H]⁺.

Example 541- (4- (4-amino-7- (2-morpholinoethyl) -7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea compound CLJ-54

This compound was synthesized in the same manner as in example 53 except that N- (2-chloroethylmorpholine) hydrochloride was used in place of bromopropene to give the desired end product CLJ-54.¹H NMR(400MHz,DMSO)9.57(s,1H),9.04(s,1H),8.25(s,1H),7.66–7.63(m,2H),7.63–7.61(m,2H),6.75(s,2H),6.53(s,1H),4.48–4.46(m,2H),3.52–3.48(m,4H),2.83–2.81(m,2H),2.49–2.46(m,4H),1.31(s,9H).HRMS(ESI),m/z:506.2628[M+H]⁺.

Example 551- (4- (4-amino-7- (2- (piperidin-1-yl) ethyl) -7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-55

This compound was synthesized in the same manner as in example 53 except that N- (2-chloroethylpiperidine) hydrochloride was used in place of bromopropene to obtain the objective end product CLJ-55.¹H NMR(400MHz,DMSO)：9.58(s,1H),9.05(s,1H),8.25(s,1H),7.66–7.63(m,2H),7.63–7.61(m,2H),6.75(s,2H),6.53(s,1H),4.52–4.48(m,2H),4.44–4.40(m,2H),2.78–2.74(m,2H),2.44–2.38(m,4H),1.46–1.40(m 4H),1.31(s,9H).HRMS(ESI),m/z:504.2829[M+H]⁺.

EXAMPLE 561- (4- (4-amino-7- (cyclopropylmethyl) -7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea compound CLJ-56

The synthesis of this compound was performed in the same manner as in example 53 except that 2-chlorocyclopropane was used instead of bromopropene to obtain the desired final product, CLJ-56.¹H NMR(400MHz,DMSO):9.56(s,1H),9.02(s,1H),8.25(s,1H),7.66–7.63(m,2H),7.63–7.61(m,2H),6.90(s,2H),6.53(s,1H),5.84–5.76(m,1H),5.05–4.97(m,2H),4.42–4.39(m,2H),2.66–2.61(m,2H),1.31(s,9H).HRMS(ESI),m/z:447.2257[M+H]⁺.

Example 571- (4- (4-amino-7-propargyl-7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-57

The synthesis of this compound was performed in the same manner as in example 53 except that bromopropyne was used in place of bromopropene to obtain the objective final product, CLJ-57.¹H NMR(400MHz,DMSO)9.63(s,1H),9.10(s,1H),8.34(s,1H),7.72–7.68(m,4H),6.59(s,1H),5.51(s,2H),5.25(s,2H),3.43(s,1H),1.37(s,9H).HRMS(ESI),m/z:431.1944[M+H]⁺.

Example 581- (4- (4-amino-7-methyl-7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-58

The synthesis of this compound was performed in the same manner as in example 53 except that methyl iodide was used instead of propylene bromide to obtain the desired final product, CLJ-58.¹H NMR(400MHz,DMSO):9.56(s,1H),9.05(s,1H),8.26(s,1H),7.66–7.61(m,4H),6.53(s,1H),5.41(s,2H),4.03–3.99(m,3H),1.31(s,9H).HRMS(ESI),m/z:407.1948[M+H]⁺.

Example 591- (4- (4-amino-7-ethyl-7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea compound CLJ-59

The synthesis of this compound was performed in the same manner as in example 53 except that iodoethane was used instead of bromopropene to obtain the desired final product, CLJ-59.¹H NMR(400MHz,DMSO):9.58(s,1H),9.05(s,1H),8.26(s,1H),7.66–7.61(m,4H),6.53(s,1H),5.41(s,2H),3.97–3.93(m,2H),1.31(s,9H),1.26–1.22(m,3H).HRMS(ESI),m/z:421.2100[M+H]⁺.

Example 601- (4- (4-amino-7- (2-cyanoethyl) -7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-60

This compound was synthesized in the same manner as in example 53 except that chloropropionitrile was used in place of bromopropene to give the desired final product CLJ-60.¹H NMR(400MHz,DMSO)9.62(s,1H),9.10(s,1H),8.34(s,1H),7.72–7.66(m,4H),6.59(s,1H),5.67(s,2H),4.66(s,2H),3.22–3.18(m,2H),1.36(s,9H).HRMS(ESI),m/z:446.2056[M+H]⁺.

Example 611- (4- (4-amino-7-hexyl-7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-61

This compound was synthesized in the same manner as in example 53 except that 1-bromohexane was used in place of bromopropene to obtain the objective final product CLJ-61.¹H NMR(400MHz,DMSO)9.56(s,1H),9.03(s,1H),8.25(s,1H),7.65–7.59(m,4H),6.54(s,1H),5.42(s,2H),4.34–4.31(m,2H),1.84–1.82(m,4H),1.31(s,9H),0.85–0.82(m,7H).HRMS(ESI),m/z:477.2396[M+H]⁺.

Example 621- (4- (4-amino-7-propyl-7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-62

The synthesis of this compound was performed in the same manner as in example 53 except that 1-bromopropyl was used instead of bromopropene to obtain the desired final product, CLJ-62.¹H NMR(400MHz,DMSO):9.56(s,1H),9.02(s,1H),8.25(s,1H),7.64–7.60(m,4H),6.53(s,1H),5.41(s,2H),4.32–4.28(m,2H),1.92–1.84(m,2H),1.31(s,9H),0.89–0.85(m,3H).HRMS(ESI),m/z:435.2256[M+H]⁺.

Example 631- (4- (4-amino-7-cyclopentyl-7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-63

The synthesis of this compound was the same as in example 53 except that bromocyclopentane was substituted for bromopropene to give the desired end product, CLJ-63.¹H NMR(400MHz,DMSO)9.55(s,1H),9.03(s,1H),8.23(s,1H),7.64–7.60(m,4H),6.72(s,2H),6.53(s,1H),5.28–5.19(m,1H),2.08–2.08(m,4H),1.92–1.87(m,2H),1.74–1.67(m,2H),1.31(s,9H).HRMS(ESI),m/z:461.2407[M+H]⁺.

Example 641- (4- (4-amino-7- (2-ethoxyethyl) -7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea compound CLJ-64

The synthesis of this compound was the same as in example 53 except that bromoethyl ether was used instead of bromopropene to give the desired end product, CLJ-64.¹H NMR(400MHz,DMSO):9.57(s,1H),9.05(s,1H),8.25(s,1H),7.67–7.61(m,4H),6.88(s,2H),6.54(s,1H),4.49–4.42(m,4H),1.31(s,9H),1.04–1.00(m,5H).HRMS(ESI),m/z:465.2361[M+H]⁺.

Example 651- (4- (4-amino-7- (3-methylbut-2-en-1-yl) -7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea Compound CLJ-65

The synthesis of this compound was the same as in example 53 except that 3, 3-dimethylbromopropene was used instead of bromopropene to give the desired end product, CLJ-65.¹H NMR(400MHz,DMSO):9.57(s,1H),9.05(s,1H),8.25(s,1H),7.65–7.59(m,4H),6.53(s,1H),5.42(s,2H),4.94–4.88(m,3H),1.82–1.79(m,5H),1.70(s,3H),1.31(s,9H).HRMS(ESI),m/z:461.2407[M+H]⁺.

EXAMPLE 661- (4- (4-amino-7- (2, 2-dimethoxyethyl) -7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea compound CLJ-66

The synthesis of this compound is the same as in example 53 except that 2, 2-dimethyl bromoethyl is substituted for bromopropene to give the desired end product, CLJ-66.¹H NMR(400MHz,DMSO):9.57(s,1H),9.04(s,1H),8.27(s,1H),7.66–7.60(m,4H),6.54(s,1H),5.46(s,2H),4.96–4.93(m,1H),4.44–4.42(m,2H),3.33(s,6H),1.31(s,9H).HRMS(ESI),m/z:481.2307[M+H]⁺.

EXAMPLE 671- (4- (4-amino-7- (4-oxopentyl) -7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (5-tert-butyl-isoxazol-3-yl) urea compound CLJ-67

The synthesis of this compound was the same as in example 53 except that 2, 2-dimethylbromoethyl was used instead of bromopropene to give the desired end product, CLJ-67.¹H NMR(400MHz,DMSO):9.57(s,1H),9.05(s,1H),8.21(s,1H),7.66–7.60(m,4H),6.54(s,1H),5.42(s,2H),4.30–4.25(m,2H),2.48–2.42(m,4H),2.05(s,3H),1.31(s,9H).HRMS(ESI),m/z:477.2396[M+H]⁺.

Example 681- (4- (4-amino-7H-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (3-tert-butyl-1H-pyrazol-5-yl) urea Compound CLJ-68

Preparation of active urea intermediate 1,3- (3-tert-butyl-1H-pyrazol-5-yl) urea

Triphosgene (4.71g,15.7mmol) was added to 50mL of tetrahydrofuran, and 3-amino-5-tert-butylpyrazole (1.99g,14.3mmol) was dissolved in 5mL of tetrahydrofuran under ice bath, then added dropwise to the triphosgene solution, and finally triethylamine (4.0mL,28.5mmol) was added dropwise. The reaction was transferred to a 60 ℃ oil bath and reacted for 5 h. And after the reaction is completed, filtering the reaction mixture, reserving filtrate, concentrating the filtrate under reduced pressure to obtain a solid, and separating the solid by a column to obtain the active urea intermediate.¹H NMR(400MHz,DMSO):12.66(s,2H),9.51(s,2H),6.30(s,2H),1.30(s,18H).

And adding the intermediate M5(225mg,1mmol) into 20mL of acetonitrile, heating to 80 ℃ for reaction, adding the active urea intermediate (390mg,1mmol) in the previous step, reacting for 0.5h, precipitating a large amount of solid, filtering, and leaching with diethyl ether to obtain the high-purity final product CLJ-68.¹H NMR(400MHz,DMSO):12.64(s,1H),11.54(s,1H),9.52(s,1H),8.86(s,1H),8.11(s,1H),7.80(d,J＝8.5Hz,2H),7.65(d,J＝8.5Hz,2H),7.19(d,J＝2.3Hz,1H),6.54(s,1H),6.01(s,2H),1.31(s,9H).HRMS(ESI),m/z:391.1924[M+H]⁺.

Example 691- (4- (4-amino-7- (2-morpholinoethyl) -pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (3-tert-butyl-1H-pyrazol-5-yl) urea Compound CLJ-69

The synthesis of this compound is the same as in example 54, except that the activity of the pyrazolylureasThe intermediate replaces an isoxazole active intermediate to obtain a target final product CLJ-69.¹H NMR(400MHz,DMSO)12.00(s,1H),9.46(s,1H),9.05(s,1H),8.24(s,1H),7.61(dd,J＝21.8,8.6Hz,4H),6.78(s,2H),6.02(s,1H),4.46(t,J＝6.6Hz,2H),3.54–3.47(m,4H),2.80(t,J＝6.7Hz,2H),2.46(d,J＝3.8Hz,4H),1.27(s,9H).HRMS(ESI),m/z:505.2811[M+H]⁺.

Example 701- (4- (4-amino-7-ethyl-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (3-tert-butyl-1H-pyrazol-5-yl) urea Compound CLJ-70

The synthesis of this compound is the same as in example 59 except that the isoxazole reactive intermediate is replaced by the reactive intermediate of pyrazolylurea to give the desired end product CLJ-70.¹H NMR(400MHz,DMSO):12.01(s,1H),9.42(s,1H),9.00(s,1H),8.25(s,1H),7.61(dd,J＝18.2,8.6Hz,4H),6.69(s,2H),6.02(s,1H),4.37(dd,J＝14.2,7.0Hz,2H),1.42(t,J＝7.1Hz,3H),1.19(s,9H).HRMS(ESI),m/z:420.2279[M+H]⁺.

Example 711- (4- (4-amino-7- (cyclopropylmethyl) -pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (3-tert-butyl-1H-pyrazol-5-yl) urea Compound CLJ-71

The synthesis of this compound is the same as in example 56 except that the active intermediate of pyrazolylurea is substituted for the active intermediate of isoxazole to give the desired end product CLJ-71.¹H NMR(400MHz,DMSO):12.00(s,1H),9.49(s,1H),9.04(s,1H),8.25(s,1H),7.61(dd,J＝21.9,8.5Hz,4H),6.69(s,2H),6.03(s,1H),5.03(dd,J＝35.8,13.0Hz,2H),4.40(d,J＝7.1Hz,2H),2.63(dd,J＝13.6,6.8Hz,2H),1.19(s,9H),0.84(dd,J＝10.0,7.0Hz,1H).HRMS(ESI),m/z:446.2456[M+H]⁺.

Example 721- (4- (4-amino-7-propyl-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (3-tert-butyl-1H-pyrazol-5-yl) urea Compound CLJ-72

The synthesis of this compound is the same as in example 62 except that the active intermediate of pyrazolylurea is substituted for the active intermediate of isoxazole to give the desired end product CLJ-72.¹H NMR(400MHz,DMSO):12.00(s,1H),9.43(s,1H),8.98(s,1H),8.25(s,1H),7.61(dd,J＝21.9,8.5Hz,4H),6.02(s,1H),4.29(t,J＝6.2Hz,2H),1.90–1.84(m,2H),1.27(s,9H),0.86(m,3H).HRMS(ESI),m/z:434.2419[M+H]⁺.

Example 731- (4- (4-amino-7-cyclopentyl-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (3-tert-butyl-1H-pyrazol-5-yl) urea Compound CLJ-73

The synthesis of this compound is the same as in example 63 except that the active intermediate of pyrazolylurea is substituted for the active intermediate of isoxazole to give the desired end product CLJ-73.¹H NMR(400MHz,DMSO)12.01(s,1H),9.40(s,1H),8.98(s,1H),8.23(s,1H),7.61(dd,J＝19.4,8.4Hz,4H),6.72(s,2H),6.02(s,1H),5.23(dt,J＝14.8,7.5Hz,1H),2.12–2.03(m,4H),1.90(d,J＝8.5Hz,2H),1.73–1.65(m,2H),1.27(s,9H).HRMS(ESI),m/z:460.2624[M+H]⁺.

Example 741- (4- (4-amino-7-allyl-pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (3-tert-butyl-1H-pyrazol-5-yl) urea Compound CLJ-74

The synthesis of this compound is the same as in example 53 except that the active intermediate of pyrazolylurea is substituted for the active intermediate of isoxazole to give the desired end product CLJ-74.¹H NMR(400MHz,DMSO):12.04(s,1H),9.41(s,1H),9.00(s,1H),8.25(s,1H),7.61(dd,J＝16.8,7.2Hz,4H),,6.72(s,2H),6.11–6.04(m,1H),6.02(s,1H),5.19(d,J＝10.4Hz,1H),5.11(d,J＝17.5Hz,1H),4.97(s,2H),1.27(s,9H).HRMS(ESI),m/z:432.2307[M+H]⁺.

Example 751- (4- (4-amino-7- (2-cyanoethyl) -pyrazolo [2,3-d ] pyrimidin-5-yl) phenyl) -3- (3-tert-butyl-1H-pyrazol-5-yl) urea Compound CLJ-75

The synthesis method of the compound is the same as that of example 60, except that the active intermediate of the pyrazole urea is used for replacing the active intermediate of the isoxazole to obtain the target final product CLJ-75.¹H NMR(400MHz,DMSO):12.00(s,1H),9.52(s,1H),9.05(s,1H),8.28(s,1H),7.63(dd,J＝19.1,8.1Hz,4H),6.73(s,2H),6.03(s,1H),4.61(t,J＝6.1Hz,2H),3.17(t,J＝6.0Hz,2H),1.27(s,9H).HRMS(ESI),m/z:445.2244[M+H]⁺.

Pharmacodynamic test section

The following representative experiments, without limitation, were used to analyze the biological activity of the compounds of the present invention.

MTT method for measuring MV4-11, Molm-13 and RS 4; 11 cell proliferation inhibition assay

The test compounds of the invention were tested for their effect on cancer cell viability on MV4-11 and Molm-13 cells, a human leukemia cell line, expressing the constitutive FLT3 receptor and containing the FLT3-ITD mutation. If the compound has strong growth inhibition activity on FLT3-ITD expression cells, the compound has obvious effect on FLT3-ITD mutant strains. If the growth inhibition activity on cells expressing FLT3-WT is poor, the compound has poor effect on FLT3-WT wild type, and the selectivity is better. The invention selects FLT3-ITD high specificity compound AC220 (CAS: 950769-58-1) as a positive control, and is synthesized by the laboratory according to the preparation method of the literature (J.Med.chem.2009,52, 7808-.

MV4-11 cells (from the American type culture center, by Sichuan university biotherapy national Key laboratory cell Bank cultures) were plated in 96-well plates in a medium of 100. mu.L IMDM, and Molm-13 cells (from the American type culture center, by Sichuan university biotherapy national Key laboratory cell Bank cultures) in 100. mu.L RPMI1640 containing 10% fetal bovine serum per 96-well plate10000-15000 cells in the well, test compounds in 100% DMSO, added to the cells, concentration of 100nM to 0.032nM (according to 5 times the dilution of the concentration of 6 concentration points) culture dish at 37 degrees C5% CO₂And culturing for 72 h. RS 4; 11 cells (from the American type culture center, from Sichuan university biotherapy national emphasis laboratory cell bank culture stock) were plated in 96-well plates in 100. mu.L RPMI1640 containing 10% fetal bovine serum, 10000-15000 cells per well, test compounds were prepared in 100% DMSO, added to the cells to a concentration of 1000nM, and plates were plated at 37 ℃ with 5% CO₂And culturing for 72 h. At the end point, 20 μ LMTT (5mg/mL) was added to each well and the cells were incubated for an additional 1-4 hours. After treatment with 20% SDS overnight, an absorbance value at a wavelength of 570nM was obtained on a spectrophotometer (Molecular Devices, Sunnyvale, USA). Calculation of IC Using percent growth compared to untreated control₅₀The values and measurement results are shown in Table 1.

TABLE 1 IC inhibition of leukemia cell proliferation by test Compounds₅₀Value of

******:0.01-0.1nM；*****:0.1-1nM；****:1-10nM；***:10-100nM；**:100-1000nM；*:>1000nM

The results show that the test compounds of the present invention have better inhibitory activity against MV4-11 and Molm-13 cell proliferation, some of which show better antiproliferative activity than AC220, and against RS 4; 11 is poor in inhibitory activity and is a novel, potential and potential inhibitor for treating FLT3-ITD related diseases.

In vitro kinase inhibition assay

In a reaction tube, buffer (8mM) MOPS, pH 7.0,0.2mM EDTA,10mM MnCl was added thereto₂) A kinase to be tested, a substrate for the kinase to be tested, 10mM magnesium acetate and gamma^33PATP solution, and compounds of different concentrations, then MgATP was added to the reaction to start the enzymatic process, and incubated at room temperature for 40 min. Finally, stopping the reaction by using 5 microliter of 3% phosphate buffer solution, titrating 10 microliter of reaction solution onto a Filtermat A membrane, washing three times by using 75mM phosphate solution, washing for 5 minutes each time by using methanol, finally drying the Filtermat A membrane and carrying out scintillation counting on the Filtermat A membrane, wherein the scintillation counting value reflects the phosphorylation degree of a substrate, so that the kinase activity inhibition condition can be represented.

TABLE 2 partial compound FLT3 kinase inhibitor Activity of the present invention

Test compounds	FLT3(IC₅₀,nM)
		CLJ-13	20
CLJ-14	6
		CLJ-20	7
CLJ-21	9
		CLJ-22	20
CLJ-42	13
		CLJ-44	4

The results show that the compound has better in-vitro enzymology inhibitory activity on FLT 3.

TABLE 3 dissociation constants of the inventive compound CLJ-20 for FLT3 mutant kinase

Type of mutation	Kd,nM
		FLT3 WT	4.37
FLT3(D835V)	7.04
		FLT3(ITD)	20.52
FLT3(ITD,D835V)	41.26
		FLT3(ITD,F691L)	48.47
FLT3(N841I)	2.37
		FLT3(R834Q)	10.15
FLT3(D835H)	5.14
		FLT3(D835Y)	7.78
FLT3(K663Q)	2.48

The results show that CLJ-20 also has good combination effect on various mutations of FLT3 kinase.

Part of tested compounds verify the target effect on Western Blot

The test method comprises the following steps: MV4-11 cells, Molm-13 cells, were treated with the indicated concentrations of the compounds. The cells were then harvested and total protein extracted with RIPA lysis buffer (beyond Co. P0013B, fraction: 50mM Tris, pH 7.4,150mM NaCl, 1% Triton X-100, 1% sodium deoxycholate, 0.1% SDS, 1). 1mM sodium orthovanadate, sodium fluoride, EDTA and leupeptin). The protein concentration was measured by BCA protein assay (ThermoScientific, USA). An equivalent sample (30. mu.g protein) was subjected to SDS-PAGE, and then the protein was transferred onto a PVDF membrane (Millipore, USA). After blocking with 5% skim milk for 1 hour at room temperature, the membranes were incubated with the indicated primary antibodies (FLT3(Cell signaling technology,3462S), p-FLT3(Cell signaling technology,3464S), STAT5(Cell signaling technology,9363S), p-STAT5(Abcam, AB32364), ERK (Zen Bioscience,220003), p-ERK (Zen Bioscience,340767), β -Actin (Abways, AB0035)) at 4 ℃ overnight, followed by detection with an appropriate secondary antibody (Abways, F300409) coupled to horseradish peroxidase for 1 hour. The immunoreactive bands were visualized using enhanced chemiluminescence (Millipore, USA). The molecular size of the detected protein was determined by comparison with a relevant protein marker (ThermoScientific, USA).

The experimental results are shown in figures 1 and 2, and the results show that the tested compound CLJ-20 can significantly reduce p-FLT3, p-STAT5 and p-ERK in a dose-dependent manner, and AC220 also shows the same effect, which indicates that the tested compound really influences the phosphorylation of a downstream signal channel by inhibiting the activity of FLT 3.

In vivo pharmacodynamic experiment of compound CLJ-20 on NOD/SCID nude mice

The purpose of this experiment was to examine the in vivo anti-tumor effect of the compounds of the invention using a mouse subcutaneous tumor model using MV4-11 and Molm-13 cells as cell lines and AC220, a developing clinical drug, as a positive control.

The test method comprises the following steps: 6-8 weeks NOD/SCID mice (purchased from Beijing Huafukang Biotech GmbH) were used, as 10⁷0.1mL of Molm-13 and MV4-11 cells are inoculated in the subcutaneous posterior costal region of the mouse, and the tumor is expanded to 200-³Then, the mice are divided into groups (6 mice in each group) and administered, the drug dissolution mode is 2% DMSO + 2% PEG-400+ 96% normal saline, the experimental group is a drug solvent control group, the oral gavage is carried out by 200 microliters every day, the compound CLJ-20 is orally gavage every day according to the dose of 3mg/kg, the compound CLJ-20 is orally gavage every day according to the dose of 10mg/kg, and the oral gavage is carried out by the positive drug AC220 according to the dose of 3mg/kg every day. The observation indexes are that the body weight of the mice and the long diameter and the short diameter of the tumor are measured once every two days, the volume of the tumor is calculated, and the physiological state of each group of mice is observed.

The experimental results are as follows: the Molm-13 model experiment results are shown in FIG. 3, and the MV4-11 model results are shown in FIG. 4. Experimental results show that the compound CLJ-20 has an obvious in-vivo anti-tumor effect on a Molm-13 model, can obviously inhibit tumor growth under the oral dosage of 3mg/kg, has the tumor inhibition rate of 94 percent, has the tumor inhibition rate of 96 percent at the time of 10mg/kg, has a better anti-tumor effect on AC220, has the tumor inhibition rate of 98 percent, and shows that the CLJ-20 and the AC220 have equivalent in-vivo effects. In the MV4-11 model, the tested drug CLJ-20 showed a tumor suppression rate of 98% at 3mg/kg, and two mice achieved tumor regression, 10mg/kg achieved tumor regression on the eighth day of administration, and the AC 2203 mg/kg group also achieved tumor regression in all mice on the eighth day. No adverse reactions such as weight reduction, rash, diarrhea and the like of the mice are found in the administration process. While a slight decrease in body weight was observed in the AC220 group, indicating that CLJ-20 was less toxic over the range of doses administered at the doses tested.

In vivo survival study of compound CLJ-20 in NCG mouse model

The test method comprises the following steps: NCG mice (purchased from Jiangsu Jiejiaokang Biotech Co., Ltd.) for 7-8 weeks were used according to 10⁶When the Molm-13 cells are inoculated in the tail vein of the mouse at a concentration, the cells can be massively proliferated in the blood circulation system of the mouse, so that the mouse has the symptoms of hind limb paralysis and even death. The mice are divided into groups (8 mice in each group) and are administrated, the drug dissolution mode is 2% DMSO + 2% PEG-400+ 96% normal saline, the experimental group is a drug solvent control group, the gavage is carried out by oral administration for 200 microliter every day, the compound CLJ-20 is carried out by oral administration for 30 days according to the dose of 10mg/kg every day. The therapeutic effect of CLJ-20 was examined by observing and recording the survival time of mice after the model was established.

The results of the experiment show that the Median Survival Time (MST) of the mice in the solvent control group is 20 days, as shown in fig. 5. The median survival time of the 10mg/kg dose group reaches 46.5 days, is prolonged by 26.5 days compared with the control group, and is sufficiently prolonged by more than one time. This indicates that CLJ-20 has significant tumor growth inhibition effects in AML xenograft in situ models, greatly prolonging survival.

Although the present invention has been described in detail hereinabove by way of general description, specific examples and experiments, it will be apparent to those skilled in the art that modifications and improvements can be made thereto based on the present invention. Accordingly, it is intended that all such modifications and variations be included within the scope of the invention as claimed and not departing from the spirit thereof.

Claims

1.4-amino-pyrimidoazepine-phenylurea derivatives having the structural formula shown in formula I:

wherein X is N or C;

R₁is-H, -OH, halogen, C1-C10 alkoxy, C1-C10 haloalkoxy,

C2-C10 alkenyl, C2-C10 alkynyl, C3-C10 cycloalkyl,

R₄is-H, -OH, halogen, C1-C8 alkyl, C1-C8 alkoxy or

R₅～R₁₀Independently is-H or C1-C8 alkyl;

R₂is-H, -OH, halogen, C1-C10 alkyl, C3-C10 cycloalkyl, C1-C10 haloalkyl, amino-substituted C1-C10 alkyl, benzyl, substituted or unsubstituted C5-C10 aryl or substituted or unsubstituted 5-10 membered heteroaryl; the 5-to 10-membered heteroaryl has N, O or S as heteroatoms, and the number of the heteroatoms is 1-3; the substituent of the substituted C5-C10 aryl or 5-10-membered heteroaryl is-H, halogen or-NH₂C1-C8 alkyl, C3-C8 cycloalkyl, C1-C8 haloalkyl, C1-C8 alkoxyC1-C8 haloalkoxy, -OH or C1-C8 carbonyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C10 alkyl.

2. The 4-amino-pyrimidoazetidic-phenylurea derivative according to claim 1, characterized in that: x is N or C; r₁is-H, -OH, halogen, C1-C8 alkoxy, C1-C8 haloalkoxy,

C2-C8 alkenyl, C2-C8 alkynyl, C3-C8 cycloalkyl,

R₄is-H, -OH, halogen, C1-C6 alkyl, C1-C6 alkoxy or

R₅～R₁₀Independently is-H or C1-C6 alkyl;

R₂is-H, -OH, halogen, C1-C8 alkyl, C3-C8 cycloalkyl, C1-C8 haloalkyl, amino-substituted C1-C8 alkyl, benzyl, substituted or unsubstituted C5-C8 aryl or substituted or unsubstituted 5-8 membered heteroaryl; the hetero atom of the 5-to 8-membered heteroaryl is N,O or S, the number of heteroatoms is 1-3; the substituent of the substituted C5-C8 aryl or 5-8-membered heteroaryl is-H, halogen or-NH₂C1-C6 alkyl, C3-C6 cycloalkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 haloalkoxy, -OH or C1-C6 carbonyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C8 alkyl;

preferably, X is N or C; r₁is-H, -OH, halogen, C1-C8 alkyl, C1-C6 alkoxy, C1-C6 halogenated alkoxy,

C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₂is-H, -OH, halogen, C1-C6 alkyl, C3-C6 cycloalkyl, C1-C6 haloalkyl, amino substituted C1-C6 alkyl, benzyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstitutedA 5-6 membered heteroaryl; the 5-6 membered heteroaryl has N, O or S as heteroatoms, and the number of the heteroatoms is 1-3; the substituent of the substituted C5-C6 aryl or 5-6-membered heteroaryl is-H, halogen or-NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

3. The 4-amino-pyrimidoazetidic-phenylurea derivative according to claim 2, characterized in that: x is N or C; r₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₂is-H, -OH, halogen, C1-C6 alkyl, C3-C6 cycloalkyl, C1-C6 haloalkyl, amino-substituted C1-C6 alkyl, benzyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstituted 5-6 membered heteroaryl; what is needed isThe 5-6 membered heteroaryl has N, O or S as heteroatoms, and the number of the heteroatoms is 1-3; the substituent of the substituted C5-C6 aryl or 5-6-membered heteroaryl is-H, halogen or-NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl;

preferably, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl,

R₄is-H, C1-C4 alkyl, C1-C4 alkoxy or

R₂is-H, -OH, halogen, C1-C6 alkyl, C3-C6 cycloalkyl, C1-C6 haloalkyl, amino-substituted C1-C6 alkyl, benzyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstituted 5-6 membered heteroaryl; the hetero atom of the 5-to 6-membered heteroaryl is N,O or S, the number of heteroatoms is 1-3; the substituent of the substituted C5-C6 aryl or 5-6-membered heteroaryl is-H, halogen or-NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl;

more preferably, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, methoxy, ethoxy, N-acetyl, N,

Formyl, acetyl,

R₄is-H, C1-C4 alkyl, tert-butyloxy or

R₅～R₁₀Independently is-H, methyl or ethyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl;

most preferably, X is N or C; r₁is-H, C1-C8 alkyl,

T-butyloxycarbonyl, C2-C6 alkenyl, C2-C6 alkynyl,

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

4. 4-amino-pyrimidoazepine-phenylurea derivatives according to claim 2 or 3, characterized in that: x is N or C; r₂Is benzyl, C3-C6 cycloalkyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstituted 5-6 membered heteroaryl; the 5-6 membered heteroaryl has 1-2 heteroatoms and N or O as heteroatoms; the substituent of the substituted C5-C6 aryl or 5-6-membered heteroaryl is-H, halogen or-NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxyA radical, -OH or a C1-C4 carbonyl group;

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl;

preferably, X is N or C; r₂Is benzyl, C3-C6 cycloalkyl,

R₁is-H, C1-C8 alkyl,

C2-C6 alkenyl, C2-C6 alkynyl, C3EC6 cycloalkyl or substituted C1-C4 alkyl; the substituent of the substituted C1-C4 alkyl is-H,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl;

more preferably, X is N or C; r₂Is benzyl, C3-C6 cycloalkyl,

R₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl or substituted or unsubstituted 5-6 memberedA heterocycloalkyl group; the heteroatom of the 5-6 membered heterocycloalkyl is N or O, and the number of the heteroatoms is 1-2; the substituent for substituting the 5-to 6-membered heterocycloalkyl is-H, C1-C4 oxycarbonyl, C1-C4 alkyl, C1-C4 alkoxy or

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl;

further preferably, X is N or C; r₂Is benzyl, C3-C6 cycloalkyl,

R₁₆～R₁₉Independently is-H, -F, -Cl, -Br, -CF₃Methyl or acetyl;

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl;

most preferably, X is N or C; r₂Is composed of

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl.

5. The 4-amino-pyrimidoazetidine-phenylurea derivative according to any one of claims 2 to 4, wherein: x is N or C; r₃is-H, halogen, -OH or C1-C4 alkyl; r₂Is benzyl, C3-C6 cycloalkyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstituted 5-6 membered heteroaryl; the 5-6 membered heteroaryl has 1-2 heteroatoms and N or O as heteroatoms; the substituent of the substituted C5-C6 aryl or 5-6-membered heteroaryl is-H, halogen or-NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl;

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

preferably, X is N or C; r₃is-H, halogen or C1-C4 alkyl; r₂Is benzyl, C3-C6 cycloalkyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstituted 5-6 membered heteroaryl; the 5-6 membered heteroaryl has 1-2 heteroatoms and N or O as heteroatoms; the substituent of the substituted C5-C6 aryl or 5-6-membered heteroaryl is-H, halogen or-NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl;

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

more preferably, X is N or C; r₃is-H or halogen; r₂Is benzyl, C3-C6 cycloalkyl, substituted or unsubstituted C5-C6 aryl or substituted or unsubstituted 5-6 membered heteroaryl; the 5-6 membered heteroaryl has 1-2 heteroatoms and N or O as heteroatoms; the substituted C5-C6 aryl isThe substituent of the 5-to 6-membered heteroaryl is-H, halogen, -NH₂C1-C4 alkyl, C3-C6 cycloalkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl;

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl.

6. The 4-amino-pyrimidoazetidine-phenylurea derivative according to any one of claims 2 to 5, wherein: x is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl,

R₄is-H, C1-C4 alkyl, C1-C4 alkoxy or

R₂Is benzyl, C3-C6 cycloalkyl,

R₃is-H, halogen or C1-C4 alkyl;

further, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, methoxy, ethoxy, N-acetyl, N,

Formyl, acetyl,

R₄is-H, C1-C4 alkyl, tert-butyloxy or

R₅～R₁₀Independently is-H, methyl or ethyl;

R₂is benzyl, C3-C6 cycloalkyl,

R₃is-H or halogen;

most preferably, X is N or C; r₁is-H, C1-C8 alkyl,

T-butyloxycarbonyl, C2-C6 alkenyl, C2-C6 alkynyl,

R₃is-H, -F, -Cl or-Br.

7. The 4-amino-pyrimidoazetidic-phenylurea derivative according to claim 1, characterized in that: when R is₂Is composed of

And the structure is shown as formula II:

wherein X is N or C; r₁is-H, -OH, halogen, C1-C10 alkoxy,C1-C10 haloalkoxy,

C2-C10 alkenyl, C2-C10 alkynyl, C3-C10 cycloalkyl,

R₄is-H, -OH, halogen, C1-C8 alkyl, C1-C8 alkoxy or

R₅～R₁₀Independently is-H or C1-C8 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C10 alkyl.

8. The 4-amino-pyrimidoazetidic-phenylurea derivative according to claim 7, characterized in that: x is N or C; r₁is-H, -OH, halogen, C1-C8 alkoxy, C1-C8 haloalkoxy,

C2-C8 alkenyl, C2-C8 alkynyl, C3-C8 cycloalkyl,

R₄is-H, -OH, halogen, C1-C6 alkyl, C1-C6 alkoxy or

R₅～R₁₀Independently is-H or C1-C6 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C8 alkyl;

C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl or substituted or unsubstituted 3-6 membered heterocycloalkyl; the heteroatom of the 3-6 membered heterocycloalkyl is N, O or S, and the number of the heteroatoms is 1-3;the substituent for substituting the 3-to 6-membered heterocycloalkyl is-H, -OH, halogen, C1-C4 oxycarbonyl, C1-C4 alkyl, C1-C4 alkoxy or

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen, -NH₂Or C1-C6 alkyl;

further, X is N or C; r₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

R₃is-H, -OH, halogen or C1-C4 alkyl;

further, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl,

R₄is-H, C1-C4 alkyl, C1-C4 alkoxy or

R₃is-H, -OH or halogen;

more preferably, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, benzeneHalogen, methoxy, ethoxy,

Formyl, acetyl,

R₄is-H, C1-C4 alkyl, tert-butyloxy or

R₅～R₁₀Independently is-H, methyl or ethyl;

R₃is-H or halogen;

most preferably, X is N or C; r₁is-H, C1-C8 alkyl,

T-butyloxycarbonyl, C2-C6 alkenyl, C2-C6 alkynyl,

R₃is-H, -F, -Cl or-Br.

9. The 4-amino-pyrimidoazetidic-phenylurea derivative according to claim 1, characterized in that: when R is₂Is composed of

R₃When the structure is-H, the structure is shown as formula III:

wherein X is N or C; r₁is-H, -OH, halogen, C1-C10 alkoxy, C1-C10 haloalkoxy,

C2-C10 alkenyl, C2-C10 alkynyl, C3-C10 cycloalkyl,

R₄is-H, -OH, halogen, C1-C8 alkyl, C1-C8 alkoxy or

R₅～R₁₀Independently is-H or C1-C8 alkyl.

10. The 4-amino-pyrimidoazetidic-phenylurea derivative according to claim 9, characterized in that: x is N or C; r₁is-H, -OH, halogen, C1-C8 alkoxy, C1-C8 haloalkoxy,

C2-C8 alkenyl, C2-C8 alkynyl, C3-C8 cycloalkyl,

R₄is-H, -OH, halogen, C1-C6 alkyl, C1-C6 alkoxy or

R₅～R₁₀Independently is-H or C1-C6 alkyl;

C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl,

Or substituted or unsubstituted C1-C6 alkyl; the substituent of the substituted C1-C6 alkyl is-H,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl or substituted or unsubstituted 3-6 membered heterocycloalkyl; the heteroatom of the 3-6 membered heterocycloalkyl is N, O or S, and the number of the heteroatoms is 1-3; the substituent of the substituted 3-to 6-membered heterocycloalkyl is-H, -OH, halogen, C1-C4Oxycarbonyl, C1-C4 alkyl, C1-C4 alkoxy or

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

further, X is N or C; r₁is-H, C1-C8 alkyl,

R₄is-H, -OH, halogen, C1-C4 alkyl, C1-C4 alkoxy or

R₅～R₁₀Independently is-H or C1-C4 alkyl;

further, X is N or C; r₁is-H, C1-C8 alkyl,

C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl or substituted C1-C4 alkyl; the takingThe substituent of the C1-C4 alkyl is-H,

-CN, -OH, phenyl, halogen, C1-C4 alkoxy, C3-C6 cycloalkyl, C1-C4 carbonyl,

R₄is-H, C1-C4 alkyl, C1-C4 alkoxy or

More preferably, X is N or C; r₁is-H, C1-C8 alkyl,

-CN, -OH, phenyl, halogen, methoxy, ethoxy, N-acetyl, N,

Formyl radicalsA group, acetyl group,

R₄is-H, C1-C4 alkyl, tert-butyloxy or

R₅～R₁₀Independently is-H, methyl or ethyl;

most preferably, X is N or C; r₁is-H, C1-C8 alkyl,

T-butyloxycarbonyl, C2-C6 alkenyl, C2-C6 alkynyl,

11. The 4-amino-pyrimidoazetidic-phenylurea derivative according to claim 1, characterized in that: when R is₂Is composed of

R₃is-H, X is C, R₁Is composed of

And when the structure is shown as formula IV:

wherein R is₁₆～R₁₉Independently is-H, halogen, -NH₂C1-C4 alkyl, C3-C6 ringAlkyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, -OH or C1-C4 carbonyl.

12. The 4-amino-pyrimidoazetidic-phenylurea derivative according to claim 11, characterized in that: r₁₆～R₁₉Independently is-H, halogen, -NH₂C1-C4 alkyl, C1-C4 haloalkyl, -OH or C1-C4 carbonyl; preferably, R₁₆～R₁₉Independently is-H, -F, -Cl, -Br, -CF₃Methyl or acetyl.

4-amino-pyrimidoazepine-phenylureas derivatives of the formula:

14. a pharmaceutically acceptable salt of a 4-amino-pyrimidoazepine-phenylurea derivative according to any one of claims 1 to 13.

15. A pharmaceutically acceptable hydrate of a 4-amino-pyrimidoazepine-phenylurea derivative according to any one of claims 1 to 13.

16. A pharmaceutical composition comprising the 4-amino-pyrimidoazepine-phenylurea derivative according to any one of claims 1 to 13, the salt according to claim 14 or the hydrate according to claim 15 as an active ingredient, together with a pharmaceutically acceptable carrier.

17. Use of a 4-amino-pyrimidoazepin-phenylurea derivative according to any one of claims 1 to 13, a salt according to claim 14, a hydrate according to claim 15 or a pharmaceutical composition according to claim 16 for the preparation of an inhibitor of FLT3 kinase; preferably, the FLT3 kinase is a mutant FLT3 kinase; more preferably, the mutant FLT3 kinase is FLT3/ITD mutant kinase.

18. Use of a 4-amino-pyrimidoazepine-phenylurea derivative according to any one of claims 1 to 13, a salt according to claim 14, a hydrate according to claim 15 or a pharmaceutical composition according to claim 16 for the preparation of a medicament for the treatment of a tumor.

19. The use of claim 18, wherein the tumor comprises a solid tumor and/or a hematological tumor.

20. The use of claim 19, the solid tumor comprising: lymphoma, B-cell lymphoma, diffuse large B-cell lymphoma, chronic lymphocytic lymphoma, lymphoplasmacytic lymphoma, ovarian cancer, breast cancer, prostate cancer, bladder cancer, kidney cancer, esophageal cancer, neck cancer, pancreatic cancer, colorectal cancer, gastric cancer, non-small cell lung cancer, thyroid cancer, brain cancer, lymphatic cancer, epidermal hyperplasia, psoriasis and/or prostatic hyperplasia.

21. The use of claim 19, the hematological neoplasm comprising: acute myeloid leukemia, chronic myeloid leukemia, myeloma, acute lymphocytic leukemia, acute myelogenous leukemia, acute promyelocytic leukemia, chronic lymphocytic leukemia, chronic neutrophilic leukemia, acute undifferentiated cell leukemia, myelodysplastic syndrome, myelodysplasia, multiple myeloma, and/or myelosarcoma.