CN114853730A

CN114853730A - Compound used as HPK1 kinase inhibitor and preparation method and application thereof

Info

Publication number: CN114853730A
Application number: CN202210590072.0A
Authority: CN
Inventors: 蒋晟; 肖易倍; 周立昕; 郝海平; 姚和权; 叶秀全; 张阔军; 王天雨; 王凯振; 章翔宇
Original assignee: China Pharmaceutical University
Current assignee: China Pharmaceutical University
Priority date: 2022-05-27
Filing date: 2022-05-27
Publication date: 2022-08-05
Anticipated expiration: 2042-05-27
Also published as: CN114853730B

Abstract

The invention discloses a compound used as an HPK1 kinase inhibitor, a preparation method and application thereof. Specifically, the invention provides a compound shown as a formula I, wherein the definition of each group is described in the specification. The compound has excellent HPK1 inhibition activity, so that the compound can be used for treating cancers and other HPK activity-related diseases.

Description

Compound used as HPK1 kinase inhibitor and preparation method and application thereof

Technical Field

The invention relates to a kinase inhibitor and a preparation method and application thereof, in particular to a compound used as an HPK1 kinase inhibitor and a preparation method and application thereof.

Background

Surgical resection, radiation therapy, chemotherapy, small molecule targeted drugs are the primary means for treating cancer, unfortunately, surgical resection is often not a viable option for many forms of cancer or tumors, and radiation therapy and chemotherapy, while killing tumor cells, also damage healthy cells. In addition, the instability of tumor cell genome promotes the mutation of tumor cells, further resulting in the rapid change of cancer genome, which makes it resistant to drugs specifically targeting tumors, which makes the treatment of cancer difficult. In recent years, the killing of tumor cells by the cancer patient's own immune system and the improvement of the body's anti-tumor immunity have been a new strategy for cancer treatment. One approach is to suppress negative regulators of the immune response that maintain peripheral tolerance, allowing tumors to be recognized as non-self antigens, and thus overcoming the immune escape of tumor cells. Hematopoietic progenitor kinase (HPK1) is one of the mitogen-activated protein kinase (MAP4K) family members, which also includes GCK/MAP4K2, GLK/MAP4K3, HGK/MAP4K4, KHS/MAP4K5, MINK/MAP4K 6. HPK1 is a negative regulator of B cell, T cell and dendritic cell activation reaction, and its expression can be inhibited to improve the anti-tumor immunity of the organism, and it is mainly expressed in hematopoietic cells, such as T cell, B cell, dendritic cell, macrophage, mast cell and neutrophil. In T cells, HPK1 regulates a role in T cell activation through the TCR signaling pathway. Following TCR activation, HPK1 interacts with T cell receptor proteins, is phosphorylated by tyrosine kinases Zap70 and Lck, and at the same time phosphorylates SLP-76 receptor protein, negatively regulating TCR signaling, thereby inhibiting T cell activation and proliferation. Research has shown that HPK1 can participate in many signaling cascades, including the MAKP signaling pathway, the Fas-induced apoptosis pathway, and the NF-. kappa.B signaling pathway. In addition, HPK1 can also inhibit AP-1, and AP-1 can promote cell proliferation, inhibit differentiation, and promote tumor cell invasion and metastasis during tumor formation and development. HPK1 kinase was not expressed in major organs, suggesting that HPK1 kinase inhibitors may not cause any serious complications.

At present, no medicine on the market exists aiming at the target of the hematopoietic progenitor cell kinase (HPK 1).

Disclosure of Invention

The purpose of the invention is as follows: the present invention aims to provide a compound useful as an inhibitor of HPK1 kinase with selectivity and high activity; it is another object of the present invention to provide a process for the preparation of compounds useful as inhibitors of HPK1 kinase; it is another object of the present invention to provide the use of a compound that acts as an inhibitor of HPK1 kinase for the preparation of a pharmaceutical composition for the prevention or treatment of a disease that responds to inhibition of HPK1 activity in a subject.

The technical scheme is as follows: the invention relates to a compound shown as the following formula I, or pharmaceutically acceptable salt, isomer or hydrate thereof:

wherein:

X ₁ selected from the group consisting of: none, NR, O, S (O) _t 、CHR、NRC(O)、C(O)NR、NRC(O)C(O)NR、NRC(O)NR、NRC(S)NR、NRC(O)NRCH ₂ 、NRC(S)NRCH ₂ Wherein R is selected from the group consisting of: H. c _1-6 Alkyl radical, C _1-6 Deuterated alkyl, C _3-8 Cycloalkyl or 3-12 membered heterocyclic group having 1-3 heteroatoms selected from the group consisting of N, S and O;

X ₂ or X ₃ Each independently is CH or N; when said X is ₂ Or X ₃ When it is CH, it may be substituted by R _a Substitution (i.e. X) ₂ 、X ₃ Is C);

ring a and ring B are each independently selected from the group consisting of: H. c _3-8 Cycloalkyl, 3-12 membered heterocyclyl, C _6-10 An aromatic ring, a 5-to 10-membered aromatic heterocycle having 1-3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered aromatic ring and a 5-to 7-membered heterocyclic ring having 1-3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered aromatic ring and a 5-to 7-membered aromatic heterocycle having 1-3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered heteroaromatic ring having 1-3 heteroatoms selected from the group consisting of N, S, O below and a 5-to 7-membered heterocyclic ring having 1-3 heteroatoms selected from the group consisting of N, S, O below, a 6-membered heteroaromatic ring having 1-3 heteroatoms selected from the group N, S, O below, and a fused ring having 1-3 heteroatoms selected from the group consisting of N, S, O belowN, S, O, the 5-7 membered heteroaromatic rings of the heteroatoms taken together;

R _a 、R ₁ 、R ₂ and R ₃ Each independently selected from the group consisting of: H. deuterium, halogen, OH, CN, NO ₂ 、C _1-6 Deuterated alkyl, C _1-6 Alkyl radical, C _3-8 Cycloalkyl radical, C _2-6 Alkenyl radical, C _2-6 Alkynyl, C _1-6 Alkoxy radical, C _6-10 Aryl, P (O) R ₆ R ₇ 、S(O) ₂ R ₆ 、S(O) ₂ NR ₆ R ₇ 、NR ₆ R ₇ 、C(O)NR ₆ R ₇ 、C(O)NR ₆ S(O) ₂ R ₇ 、NR ₆ S(O) ₂ R ₇ 、C(O)R ₆ 、NR ₆ C(O)R ₇ 5-12 membered heteroaryl having 1-3 heteroatoms selected from the group consisting of N, S, O, 3-12 membered heterocyclyl having 1-3 heteroatoms selected from the group consisting of N, S, O, wherein R is substituted with R _a 、R ₁ 、R ₂ And R ₃ The alkyl, alkenyl, alkynyl, alkoxy, phenyl, heteroaryl, cycloalkyl or heterocyclyl group in the group represented may be substituted with 1 to 3 substituents each independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _1-6 Haloalkoxy, NR ₆ R ₇ 、C(＝O)NR ₆ R ₇ 、C(＝O)NR ₆ S(＝O) ₂ R ₇ 、P(＝O)R ₆ R ₇ 、S(＝O) ₂ R ₆ 、S(＝O) ₂ NR ₆ R ₇ 、NR ₆ S(＝O) ₂ R ₇ ；

p and q are each independently 0, 1,2,3,4 or 5.

Further, the compound of formula I has any one of the structures shown in the following formulas:

wherein:

X ₁ selected from the group consisting of: is free of,NR、O、S(O) _t 、CHR、NRC(O)、C(O)NR、NRC(O)C(O)NR、NRC(O)NR、NRC(S)NR、NRC(O)NRCH ₂ 、NRC(S)NRCH ₂ Wherein R is selected from the group consisting of: H. c _1-6 Alkyl radical, C _1-6 Deuterated alkyl, C _3-8 Cycloalkyl or a 3-12 membered heterocyclic group having 1-3 heteroatoms selected from the group consisting of N, S and O;

X ₅ or X ₆ Each independently selected from the group consisting of none, NR _b 、O、S(O) _t 、C(O)、CR ₄ R ₅ ；

Y is selected from NR _b 、O、S(O) _t Or CR ₄ R ₅ ；

M ¹ 、M ² Or M ³ Each independently is CH or N; when said M is ¹ 、M ² Or M ³ When is CH, the

Can be located at said M ¹ 、M ² Or M ³ Upper (i.e. M) ¹ 、M ² Or M ³ Is C);

ring A is selected from the group consisting of: H. c _3-8 Cycloalkyl, 3-12 membered heterocyclyl, C _6-10 An aromatic ring, a 5-to 10-membered aromatic heterocycle having 1 to 3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered aromatic ring and a 5-to 7-membered heterocyclic ring having 1 to 3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered aromatic ring and a 5-to 7-membered aromatic heterocycle having 1 to 3 heteroatoms selected from the group consisting of N, S, O below, a 6-membered heteroaryl ring having 1-3 heteroatoms selected from the group consisting of the lower group N, S, O and a fused ring jointly composed of a 5-7-membered heterocyclic ring having 1-3 heteroatoms selected from the group consisting of the lower group N, S, O, a 6-membered heteroaryl ring having 1-3 heteroatoms selected from the group consisting of the lower group N, S, O and a 5-7-membered heteroaryl ring having 1-3 heteroatoms selected from the group consisting of the lower group N, S, O;

R _a 、R ₁ 、R ₂ 、R ₃ and R ₅ Each independently selected from the group consisting of: H. deuterium, halogen, OH, CN, NO ₂ 、C _1-6 Deuterated alkyl, C _1-6 Alkyl radical, C _3-8 Cycloalkyl radical, C _2-6 Alkenyl radical, C _2-6 Alkynyl, C _1-6 Alkoxy radical, C _6-10 Aryl, P (O) R ₆ R ₇ 、S(O) ₂ R ₆ 、S(O) ₂ NR ₆ R ₇ 、NR ₆ R ₇ 、C(O)NR ₆ R ₇ 、C(O)NR ₆ S(O) ₂ R ₇ 、NR ₆ S(O) ₂ R ₇ 、C(O)R ₆ 、NR ₆ C(O)R ₇ 5-12 membered heteroaryl having 1-3 heteroatoms selected from the group consisting of N, S, O below, 3-12 membered heterocyclyl having 1-3 heteroatoms selected from the group consisting of N, S, O below, wherein R is substituted with R _a 、R ₁ 、R ₂ 、R ₃ And R ₅ The alkyl, alkenyl, alkynyl, alkoxy, phenyl, heteroaryl, cycloalkyl or heterocyclyl group in the group represented may be substituted with 1 to 3 substituents each independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _1-6 Haloalkoxy, NR ₆ R ₇ 、C(＝O)NR ₆ R ₇ 、C(＝O)NR ₆ S(＝O) ₂ R ₇ 、P(＝O)R ₆ R ₇ 、S(＝O) ₂ R ₆ 、S(＝O) ₂ NR ₆ R ₇ 、NR ₆ S(＝O) ₂ R ₇ ；

R _b Selected from the group consisting of H, deuterium, C (O) C _1-6 Alkyl radical, C _1-6 Deuterated alkyl, C _1-6 Alkyl radical, C _3-8 Cycloalkyl radical, C _2-6 Alkenyl radical, C _2-6 Alkynyl or 3-12 membered heterocyclyl having 1-3 heteroatoms selected from the group N, S, O, wherein R is _b The alkyl, alkenyl, alkynyl, cycloalkyl or heterocyclyl group in the group represented may be substituted with 1 to 3 substituents each independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _1-6 Haloalkoxy, C _3-8 Cycloalkyl, 5-10 membered heterocyclic group and NR ₆ R ₇ ；

R ₄ Selected from the group consisting of: H. deuterium, C (O) C _1-6 Alkyl radical, C _1-6 Alkyl radical, C _1-6 Deuterated alkyl, C _3-8 Cycloalkyl radical, C _2-6 Alkenyl radical, C _2-6 Alkynyl or C having 1-3 heteroatoms selected from the group N, S, O _3-12 Heterocyclyl, wherein R is ₄ The alkyl, alkenyl, alkynyl, cycloalkyl or heterocyclyl group in the group represented may be substituted with 1 to 3 substituents each independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _1-6 Haloalkoxy, C _3-8 Cycloalkyl, C _5-10 A heterocyclic radical and NR ₆ R ₇ ；

R ₆ 、R ₇ Each independently is H or C _1-6 Alkyl radical, C _1-6 Deuterated alkyl, C _3-8 Cycloalkyl, C having 1-3 hetero atoms selected from the group consisting of N, S and O _3-12 Heterocyclyl, wherein R is ₆ Or R ₇ The alkyl, cycloalkyl and heterocyclic groups represented by the formula (I) may be substituted by 1 to 3 substituents independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _3-8 Cycloalkyl radical, C _3-12 A heterocyclic group.

R ₆ And R ₇ Together with the same nitrogen or phosphorus atom to which they are attached may form C _3-12 Heterocyclyl, which may be substituted with 1 to 3 substituents independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _1-6 Deuterated alkyl, C _1-6 Haloalkoxy, C _3-8 Cycloalkyl radical, C _3-12 A heterocyclic group;

n is 1,2 or 3;

m is 1,2 or 3; and n + m is less than or equal to 4;

p and q are each independently 0, 1,2,3,4 or 5;

t is 0, 1 or 2;

X ₁ selected from the group consisting of: none, NR, O, S (O) _t 、CHR、NRC(O)、C(O)NR、NRC(O)C(O)NR、NRC(O)NR、NRC(S)NR、NRC(O)NRCH ₂ 、NRC(S)NRCH ₂ Wherein R is selected from the group consisting of: H. c _1-6 Alkyl radical, C _1-6 Deuterated alkyl, C _3-8 Cycloalkyl or a 3-12 membered heterocyclic group having 1-3 heteroatoms selected from the group consisting of N, S and O;

M ¹ 、M ² 、M ³ 、M ⁴ or M ⁵ Each independently is CH or N; when said M is ¹ 、M ² 、M ³ 、M ⁴ Or M ⁵ When is CH, the

Can be located at said M ¹ 、M ² 、M ³ 、M ⁴ Or M ⁵ Upper (i.e. M) ¹ 、M ² 、M ³ 、M ⁴ Or M ⁵ Is C); when said M is ⁴ And M ⁵ When substituted at the same time, may form a bicyclic ring together with the atoms to which they are attached; wherein the bicyclic ring can contain one or more heteroatoms selected from N, S and O; and wherein said bicyclic ring is optionally substituted with one, two, three, four or five R ₃ ；

Ring A is selected from the group consisting of: H. c _3-8 Cycloalkyl, 3-12 membered heterocyclyl, C _6-10 An aromatic ring, a 5-to 10-membered aromatic heterocycle having 1 to 3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered aromatic ring and a 5-to 7-membered heterocyclic ring having 1 to 3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered aromatic ring and a 5-to 7-membered aromatic heterocycle having 1 to 3 heteroatoms selected from the group consisting of N, S, O below, a fused ring having 1 to 3 heteroatoms selected from the group consisting of N, S, O belowN, S, O with a 5-7 membered heterocyclic ring having 1-3 heteroatoms selected from the group consisting of group N, S, O, a 6-membered heteroaromatic ring having 1-3 heteroatoms selected from the group N, S, O, and a 5-7 membered heteroaromatic ring having 1-3 heteroatoms selected from the group N, S, O;

R _a 、R ₁ 、R ₂ and R ₃ Each independently selected from the group consisting of: H. deuterium, halogen, OH, CN, NO ₂ 、C _1-6 Deuterated alkyl, C _1-6 Alkyl radical, C _3-8 Cycloalkyl radical, C _2-6 Alkenyl radical, C _2-6 Alkynyl, C _1-6 Alkoxy radical, C _6-10 Aryl, P (O) R ₆ R ₇ 、S(O) ₂ R ₆ 、S(O) ₂ NR ₆ R ₇ 、NR ₆ R ₇ 、C(O)NR ₆ R ₇ 、C(O)NR ₆ S(O) ₂ R ₇ 、NR ₆ S(O) ₂ R ₇ 、C(O)R ₆ 、NR ₆ C(O)R ₇ 5-12 membered heteroaryl having 1-3 heteroatoms selected from the group consisting of N, S, O below, 3-12 membered heterocyclyl having 1-3 heteroatoms selected from the group consisting of N, S, O below, wherein R is substituted with R _a 、R ₁ 、R ₂ And R ₃ The alkyl, alkenyl, alkynyl, alkoxy, phenyl, heteroaryl, cycloalkyl or heterocyclyl group in the group represented may be substituted with 1 to 3 substituents each independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _1-6 Haloalkoxy, NR ₆ R ₇ 、C(＝O)NR ₆ R ₇ 、C(＝O)NR ₆ S(＝O) ₂ R ₇ 、P(＝O)R ₆ R ₇ 、S(＝O) ₂ R ₆ 、S(＝O) ₂ NR ₆ R ₇ 、NR ₆ S(＝O) ₂ R ₇ ；

R ₆ 、R ₇ Each independently is H or C _1-6 Alkyl radical, C _1-6 Deuterated alkyl, C _3-8 Cycloalkyl, C having 1-3 hetero atoms selected from the group consisting of N, S and O _3-12 Heterocyclyl, wherein R is ₆ Or R ₇ Is represented byThe alkyl, cycloalkyl and heterocyclyl groups may be substituted with 1 to 3 substituents independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _3-8 Cycloalkyl radical, C _3-12 A heterocyclic group.

p and q are each independently 0, 1,2,3,4 or 5;

M ⁷ is CR ⁸ 、NR ⁸ N, O or S;

M ⁸ is CR ⁸ Or N;

M ⁹ and M ¹⁰ Each independently is C or N, provided that M ⁹ And M ¹⁰ Is C;

ring A is selected from the group consisting of: H. c _3-8 Cycloalkyl, 3-12 membered heterocyclyl, C _6-10 Fragrance compositionA ring, a 5-to 10-membered aromatic heterocyclic ring having 1 to 3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered aromatic ring and a 5-to 7-membered heterocyclic ring having 1 to 3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered aromatic ring and a 5-to 7-membered aromatic heterocyclic ring having 1 to 3 heteroatoms selected from the group consisting of N, S, O below, a 6-membered heteroaryl ring having 1-3 heteroatoms selected from the group consisting of the lower group N, S, O and a fused ring jointly composed of a 5-7-membered heterocyclic ring having 1-3 heteroatoms selected from the group consisting of the lower group N, S, O, a 6-membered heteroaryl ring having 1-3 heteroatoms selected from the group consisting of the lower group N, S, O and a 5-7-membered heteroaryl ring having 1-3 heteroatoms selected from the group consisting of the lower group N, S, O;

ring C is selected from the group consisting of: c _5-8 Cycloalkyl or a 5-to 8-membered heterocyclic ring having at least 3 ring-forming carbon atoms and 1,2, or 3 ring-forming heteroatoms independently selected from the group consisting of N, P, O and S; wherein C is _5-8 Cycloalkyl and 5-to 8-membered heterocycle are independently optionally selected from R by 1,2,3,4 or 5 independently ⁸ Substituted with the substituent(s); and wherein C _5-8 Two substituents of cycloalkyl or 5-to 8-membered heterocycle, when present, optionally together form a ring optionally substituted with 1,2,3 or 4 independently selected from R ⁸ Substituted spiro, fused or bridged cycloalkyl (e.g., C) _3-6 Cycloalkyl) or optionally substituted with 1,2,3 or 4 independently selected from R ⁸ A spiro, fused, or bridged heterocyclic group (e.g., a 3-to 6-membered heterocyclic group) substituted with the substituent(s) of (a);

R ₁ 、R ₂ and R ₃ Each independently selected from the group consisting of: H. deuterium, halogen, OH, CN, NO ₂ 、C _1-6 Deuterated alkyl, C _1-6 Alkyl radical, C _3-8 Cycloalkyl radical, C _2-6 Alkenyl radical, C _2-6 Alkynyl, C _1-6 Alkoxy radical, C _6-10 Aryl, P (O) R ₆ R ₇ 、S(O) ₂ R ₆ 、S(O) ₂ NR ₆ R ₇ 、NR ₆ R ₇ 、C(O)NR ₆ R ₇ 、C(O)NR ₆ S(O) ₂ R ₇ 、NR ₆ S(O) ₂ R ₇ 、C(O)R ₆ 、NR ₆ C(O)R ₇ Having 1-3 options5-12 membered heteroaryl of a heteroatom selected from the group consisting of N, S, O, 3-12 membered heterocyclyl having 1-3 heteroatoms selected from the group consisting of N, S, O, wherein R is ₂ And R ₃ The alkyl, alkenyl, alkynyl, alkoxy, phenyl, heteroaryl, cycloalkyl or heterocyclyl group in the group represented may be substituted with 1 to 3 substituents each independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _1-6 Haloalkoxy, NR ₆ R ₇ 、C(＝O)NR ₆ R ₇ 、C(＝O)NR ₆ S(＝O) ₂ R ₇ 、P(＝O)R ₆ R ₇ 、S(＝O) ₂ R ₆ 、S(＝O) ₂ NR ₆ R ₇ 、NR ₆ S(＝O) ₂ R ₇ ；

R ⁸ Selected from the group consisting of: H. halogen, cyano, C _1-6 Alkyl radical, C _3-8 Cycloalkyl radical, C _2-6 Alkenyl radical, C _2-6 Alkynyl, C _1-6 Alkoxy radical, C _6-10 Aryl, 5-10 membered heteroaryl having 1-3 heteroatoms selected from the group consisting of N, S, O below, 5-12 membered heteroaryl having 1-3 heteroatoms selected from the group consisting of N, S, O below, C (O) R ₆ 、C(O)OR ₆ 、C(O)NR ₆ R ₇ 、OC(O)R ₆ 、OC(O)NR ₆ R ₇ 、SR ⁶ 、S(O) ₂ R ₆ 、P(O)R ₆ R ₇ 、S(O) ₂ NR ₆ R ₇ 、NR ₆ R ₇ 、C(O)NR ₆ S(O) ₂ R ₇ 、NR ₆ S(O) ₂ R ₇ 、NR ₆ C(O)R ₇ 、NR ₆ C(O)OR ₇ And a 3-12 membered heterocyclic group having 1-3 heteroatoms selected from N, S, O below, wherein R is represented by ₁ 、R ₂ And R ₃ The alkyl, alkenyl, alkynyl, alkoxy, phenyl, heteroaryl, cycloalkyl or heterocyclyl group in the group represented may be substituted with 1 to 4 substituents each independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _1-6 Haloalkoxy, NR ₆ R ₇ 、C(＝O)NR ₆ R ₇ 、C(＝O)NR ₆ S(＝O) ₂ R ₇ 、P(＝O)R ₆ R ₇ 、S(＝O) ₂ R ₆ 、S(＝O) ₂ NR ₆ R ₇ 、NR ₆ S(＝O) ₂ R ₇ ；

q is 0, 1,2,3,4 or 5;

M ⁷ is CR ⁸ 、NR ⁸ N, O or S;

M ⁸ is CR ⁸ Or N;

ring A is selected from the group consisting of: H. c _3-8 Cycloalkyl, 3-12 membered heterocyclyl, C _6-10 An aromatic ring, a 5-to 10-membered aromatic heterocycle having 1-3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered aromatic ring and a 5-to 7-membered heterocyclic ring having 1-3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered aromatic ring and a 5-to 7-membered aromatic heterocycle having 1-3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered heteroaromatic ring having 1-3 heteroatoms selected from the group consisting of N, S, O below and a 5-to 7-membered heterocyclic ring having 1-3 heteroatoms selected from the group consisting of N, S, O below, a fused ring having 1-3 heteroatoms selected from the group N, S, O below, a fused ring composed of a 5-to 7-membered heterocyclic ring having 1-3 heteroatoms selected from the group N, S, O below, and a method for producing the sameA fused ring composed of the 6-membered heteroaromatic ring of (a) and a 5-7-membered heteroaromatic ring having 1-3 heteroatoms selected from group N, S, O below;

R ₁ 、R ₂ and R ₃ Each independently selected from the group consisting of: H. deuterium, halogen, OH, CN, NO ₂ 、C _1-6 Deuterated alkyl, C _1-6 Alkyl radical, C _3-8 Cycloalkyl radical, C _2-6 Alkenyl radical, C _2-6 Alkynyl, C _1-6 Alkoxy radical, C _6-10 Aryl, P (O) R ₆ R ₇ 、S(O) ₂ R ₆ 、S(O) ₂ NR ₆ R ₇ 、NR ₆ R ₇ 、C(O)NR ₆ R ₇ 、C(O)NR ₆ S(O) ₂ R ₇ 、NR ₆ S(O) ₂ R ₇ 、C(O)R ₆ 、NR ₆ C(O)R ₇ 5-12 membered heteroaryl having 1-3 heteroatoms selected from the group consisting of N, S, O below, 3-12 membered heterocyclyl having 1-3 heteroatoms selected from the group consisting of N, S, O below, wherein R is ₂ And R ₃ The alkyl, alkenyl, alkynyl, alkoxy, phenyl, heteroaryl, cycloalkyl or heterocyclyl group in the group represented may be substituted with 1 to 3 substituents each independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _1-6 Haloalkoxy, NR ₆ R ₇ 、C(＝O)NR ₆ R ₇ 、C(＝O)NR ₆ S(＝O) ₂ R ₇ 、P(＝O)R ₆ R ₇ 、S(＝O) ₂ R ₆ 、S(＝O) ₂ NR ₆ R ₇ 、NR ₆ S(＝O) ₂ R ₇ ；

q is 0, 1,2,3,4 or 5.

Still further, the compound of formula II-a has any one of the structures shown below:

the compound of formula II-b has any structure shown as the following formula:

the compound of formula II-c has any structure shown as the following formula:

the compound of formula II-d has any structure as shown in the following formula:

in another aspect, the present invention provides a method for preparing the above compound, or a pharmaceutically acceptable salt, isomer, or hydrate thereof, characterized in that: the method is any one of the following methods:

the method A comprises the following steps:

the method B comprises the following steps:

the compound of formula I is obtained by coupling and deprotection reactions of benzo 1,2, 4-triazine or pyrido 1,2, 4-triazine compounds.

In another aspect, the present invention provides a pharmaceutical composition comprising a therapeutically effective amount of one or more of the compounds described above, or a pharmaceutically acceptable salt, isomer, or hydrate thereof, and a pharmaceutically acceptable excipient.

In another aspect, the present invention provides a use of the above-mentioned compound, or a pharmaceutically acceptable salt, isomer or hydrate thereof for the preparation of a pharmaceutical composition for preventing or treating a disease responsive to inhibition of HPK1 activity in a subject.

A combination of the above compound, or a pharmaceutically acceptable salt, isomer or hydrate thereof, with other tumor immunotherapeutic agents selected from the group consisting of: small molecule compounds and antibodies (including but not limited to PD-1, PD-L1, CTLA-4, STING agonists, LAG3 antagonists, etc.), tumor targeting drugs, tumor vaccines, and radiation therapy regimens.

The use of the above compounds and their pharmaceutically acceptable salts, stereoisomers, prodrugs, solvates, esters and deuterated compounds in conjunction with CAR-T immunotherapy in the immunotherapy of cancer.

Further, the disease is cancer.

In another aspect, the present invention provides a use of the above compound, or a pharmaceutically acceptable salt, isomer or hydrate thereof, in an HPK1 kinase inhibitor.

Since the compound of the present invention has a very good HPK1 kinase inhibitory activity, the compound of the present invention and various crystalline forms, pharmaceutically acceptable organic or inorganic salts, solvates or hydrates thereof, and a pharmaceutical composition containing the compound of the present invention as a main active ingredient can be used for the prevention and/or treatment of diseases and the like (e.g., cancer) associated with the activity or expression level of HPK1 kinase.

The pharmaceutical composition of the present invention comprises the compound of the present invention in a safe and effective amount range and a pharmaceutically acceptable carrier or excipient. Wherein the safe and effective amount refers to: the amount of the compound is sufficient to significantly improve the condition without causing serious side effects. Typically, the pharmaceutical composition contains 1-2000mg of a compound of the invention per dose, more preferably, 10-200mg of a compound of the invention per dose. Preferably, said "dose" is a capsule or tablet.

By "pharmaceutically acceptable carrier" is meant: one or more compatible liquid or solid fillers or gel substances, which are suitable for human use and must be of sufficient purity and sufficiently low toxicity. By "compatible" is meant herein that the components of the composition are capable of intermixing with and between the compounds of the invention without significantly diminishing the efficacy of the compounds. Examples of pharmaceutically acceptable carrier moieties are cellulose and its derivatives (e.g., sodium ethylcellulose, sodium carboxymethylcellulose, cellulose acetate, etc.), gelatin, talc, calcium sulfate, solid lubricants (e.g., stearic acid, magnesium stearate), vegetable oils (e.g., soybean oil, sesame oil, peanut oil, olive oil, etc.), colorants, polyols (e.g., propylene glycol, glycerin, mannitol, sorbitol, etc.), emulsifiers (e.g., tween R, wetting agents (e.g., sodium lauryl sulfate), flavoring agents, preservatives, stabilizers, antioxidants, pyrogen-free water, etc.

The mode of administration of the compounds or pharmaceutical compositions of the present invention is not particularly limited, and representative modes of administration include (but are not limited to): oral, parenteral (intravenous, intramuscular or subcutaneous).

Solid dosage forms for oral administration include capsules, pills, tablets, powders and granules. In these solid dosage forms, the active compound is mixed with at least one conventional inert excipient (or carrier), such as dicalcium phosphate or sodium citrate, or with the following ingredients: (a) fillers or compatibilizers, for example: lactose, sucrose, starch, glucose, mannitol, and silicic acid; (b) binders, for example: hydroxymethyl cellulose, gelatin, alginates, polyvinylpyrrolidone, gum arabic, and sucrose; (c) humectants, such as: glycerol; (d) disintegrants, for example: calcium carbonate, agar-agar, potato or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (e) slow solvents, such as paraffin; (f) absorption accelerators, e.g., quaternary ammonium compounds; (g) wetting agents, such as cetyl alcohol and glycerol monostearate; (h) adsorbents, for example: kaolin; (i) lubricants, for example, talc, solid polyethylene glycol, calcium stearate, magnesium stearate, sodium lauryl sulfate or mixtures thereof. In capsules, tablets and pills, the dosage forms may also comprise buffering agents.

Solid dosage forms such as tablets, capsules, dragees, pills and granules can be prepared using shells and coatings such as enteric coatings and other materials well known in the art. They may contain opacifying agents and the release of the active compound or compounds in such compositions may be delayed in release in a certain part of the digestive tract. Examples of embedding components which can be used are polymeric substances and wax-like substances. If desired, the active compound may also be in microencapsulated form with one or more of the above-mentioned excipients.

Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, suspensions, solutions, syrups or tinctures. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly employed in the art, such as water or other solvents, solubilizing agents and emulsifiers, as exemplified by: ethanol, isopropanol, propylene glycol, ethyl acetate, ethyl carbonate, 1, 3-butylene glycol, dimethylformamide, and oils, especially peanut oil, cottonseed oil, corn germ oil, olive oil, sesame oil, and castor oil or mixtures thereof, and the like.

In addition to these inert diluents, the compositions can also contain adjuvants such as emulsifying, wetting and suspending agents, flavoring, sweetening and perfuming agents.

Suspensions, in addition to the active compounds, may contain suspending agents, for example: polyoxyethylene sorbitol and sorbitan esters, ethoxylated isostearyl alcohols, microcrystalline cellulose, agar and aluminum methoxide or mixtures of these and the like.

Compositions for parenteral injection may comprise physiologically acceptable sterile aqueous or dispersion, anhydrous solutions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Suitable aqueous and diluents, non-aqueous carriers, solvents or vehicles include water, polyols, ethanol and suitable mixtures thereof.

The compounds of the present invention may be administered alone or in combination with other pharmaceutically acceptable compounds.

When administered in combination, the pharmaceutical composition further comprises one or more other pharmaceutically acceptable compounds. One or more of the other pharmaceutically acceptable compounds may be administered simultaneously, separately or sequentially with a compound of the invention.

When the pharmaceutical composition is used, a safe and effective amount of the compound of the present invention is applied to a mammal (such as a human) in need of treatment, wherein the administration dose is a pharmaceutically acceptable effective dose, and for a human with a weight of 60kg, the daily administration dose is usually 1 to 2000mg, preferably 20 to 500 mg. Of course, the particular dosage will also take into account factors such as the health of the patient, the route of administration, and the like, which are within the skill of the skilled practitioner.

The invention will be further illustrated with reference to the following specific examples. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. Experimental procedures without specific conditions noted in the following examples, generally according to conventional conditions, or according to conditions recommended by the manufacturer. Unless otherwise indicated, percentages and parts are by weight.

Has the advantages that: compared with the prior art, the invention has the following remarkable advantages: has excellent HPK1 inhibiting activity, and can be used for preparing pharmaceutical composition for treating cancer and other diseases related to HPK activity.

Detailed Description

The invention provides a compound shown as the following formula I:

wherein:

ring a and ring B are each independently selected from the group consisting of: H. c _3-8 Cycloalkyl, 3-12 membered heterocyclyl, C _6-10 An aromatic ring, a 5-to 10-membered aromatic heterocycle having 1 to 3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered aromatic ring and a 5-to 7-membered heterocyclic ring having 1 to 3 heteroatoms selected from the group consisting of N, S, O below, a fused ring composed of a 6-membered aromatic ring and a 5-to 7-membered aromatic heterocycle having 1 to 3 heteroatoms selected from the group consisting of N, S, O below, a 6-membered heteroaryl ring having 1-3 heteroatoms selected from the group consisting of the lower group N, S, O and a fused ring jointly composed of a 5-7-membered heterocyclic ring having 1-3 heteroatoms selected from the group consisting of the lower group N, S, O, a 6-membered heteroaryl ring having 1-3 heteroatoms selected from the group consisting of the lower group N, S, O and a 5-7-membered heteroaryl ring having 1-3 heteroatoms selected from the group consisting of the lower group N, S, O;

R _a 、R ₁ 、R ₂ and R ₃ Each independently selected from the group consisting of: H. deuterium, halogen, OH, CN, NO ₂ 、C _1-6 Deuterated alkyl, C _1-6 Alkyl radical, C _3-8 Cycloalkyl radical, C _2-6 Alkenyl radical, C _2-6 Alkynyl, C _1-6 Alkoxy radical, C _6-10 Aryl, P (O) R ₆ R ₇ 、S(O) ₂ R ₆ 、S(O) ₂ NR ₆ R ₇ 、NR ₆ R ₇ 、C(O)NR ₆ R ₇ 、C(O)NR ₆ S(O) ₂ R ₇ 、NR ₆ S(O) ₂ R ₇ 、C(O)R ₆ 、NR ₆ C(O)R ₇ 5-12 membered heteroaryl having 1-3 heteroatoms selected from the group consisting of N, S, O below, 3-12 membered heterocyclyl having 1-3 heteroatoms selected from the group consisting of N, S, O below, wherein R is substituted with R _a 、R ₁ 、R ₂ And R ₃ Alkyl, alkenyl, alkynyl, alkoxy, phenyl, heteroaryl, cycloalkane of the groupThe group or the heterocyclic group may be substituted with 1 to 3 substituents each independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _1-6 Haloalkoxy, NR ₆ R ₇ 、C(＝O)NR ₆ R ₇ 、C(＝O)NR ₆ S(＝O) ₂ R ₇ 、P(＝O)R ₆ R ₇ 、S(＝O) ₂ R ₆ 、S(＝O) ₂ NR ₆ R ₇ 、NR ₆ S(＝O) ₂ R ₇ ；

p and q are each independently 0, 1,2,3,4 or 5;

in the compound of the formula I, each chiral center is in an R configuration or an S configuration.

Preferably, said X ₁ 、X ₂ 、X ₃ 、Y、M ¹ 、M ² 、M ³ 、M ⁴ 、M ⁵ 、R、R ₁ 、R ₂ 、R ₃ 、R ₄ 、R ₅ 、R ₆ 、R ₇ 、R ₈ 、R _a 、R _b N, m, p, q, t, are each independently the corresponding group in the specific compound in each example.

The compounds of the invention may be useful as inhibitors of HPK1 kinase, and in a preferred embodiment, are selective inhibitors of HPK1 kinase.

Preparation of Compounds of formula I

The compounds of formula I of the present invention may be prepared by the following exemplary method a or method B:

the method A comprises the following steps:

the method B comprises the following steps:

The invention will be further illustrated with reference to the following specific examples. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. The experimental procedures, in which specific conditions are not noted in the following examples, are generally carried out under conventional conditions or conditions recommended by the manufacturers. Unless otherwise indicated, percentages and parts are by weight.

Preparation of key intermediate 1 (3-amino-6-chlorobenzo [ e ] [1,2,4] triazin-8-yl) carbamic acid tert-butyl ester

The method comprises the following steps: boc at room temperature ₂ O (53.8g, 247mmol) was slowly added to a solution of 5-chloro-2-nitroaniline (10g, 57.95mmol) and DMAP (10.6g, 86.76mmol) in THF (150ml) and the temperature was raised to 70 ℃ for 3 h. Cooling to room temperature, decompressing, concentrating, and purifying by column chromatography to obtain a light yellow compound (5-chloro-2-nitrophenyl) di-tert-butyl carbamate. MS (ESI) M/Z373.8 [ M + H ]] ⁺ 。

Step two: slowly dropwise adding a tetrahydrofuran (20ml) solution of potassium tert-butoxide (5.2g, 46.4mmol) into a tetrahydrofuran solution (40ml) of di-tert-butyl (5-chloro-2-nitrophenyl) carbamate (2.17g, 5.8mmol) and guanidine hydrochloride (1.11g, 11.6mmol) at room temperature, heating to 65 ℃ for overnight reaction, cooling to room temperature after the reaction is finished, adding a saturated ammonium chloride solution, extracting with ethyl acetate, drying an organic phase anhydrous sodium sulfate, concentrating under reduced pressure, and purifying by column chromatography to obtain a bright yellow target compound (3-amino-6-chlorobenzo [ e ]][1,2,4]Triazin-8-yl) carbamic acid tert-butyl ester. MS (ESI) 296.7[ M + H ]] ⁺ . Key intermediate 2.7-iodo-6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinoline preparation

The method comprises the following steps: at 0 ℃ in advanceAdding 7-methoxy-1, 2,3, 4-tetrahydroisoquinoline (6g, 33.85mmol) and concentrated nitric acid (3.46g) in sequence into cold trifluoroacetic acid (46ml), stirring at 0 ℃ for 1h, concentrating under reduced pressure, extracting with DCM, drying the organic phase with anhydrous sodium sulfate, suction filtering, spin drying, and purifying by column chromatography to obtain 6-methoxy-7-nitro-1, 2,3, 4-tetrahydroisoquinoline (1.85g, 24.6%). MS (ESI) M/Z209.2 [ M + H ]] ⁺ 。

Step two: at 0 deg.C, in 7-methoxy-6-nitro-1, 2,3, 4-four hydrogen isoquinoline (7.58g,46.44mmo1) methanol (82mL) solution is added 37% formaldehyde solution (23mL), at 0 deg.C stirring for 15 minutes, add NaBH batch wise ₄ (6.15g), the mixture at room temperature stirring for 3h, using ice water (20mL) quenching, using DCM extraction, organic phase with anhydrous sodium sulfate drying, suction filtration, spin drying, column chromatography purification to obtain 6-methoxy-2-methyl-7-nitro-1, 2,3, 4-four hydrogen isoquinoline. MS (ESI) M/Z223.2 [ M + H ]] ⁺ 。

Step three: adding Fe powder (13.2g,237.60mmol) and hydrochloric acid (1mL) into a solution of 7-methoxy-2-methyl-6-nitro-1, 2,3, 4-tetrahydroisoquinoline (8.8g,39.6 mmol 1) in water (10mL) and EtOH (90mL), stirring at 60 ℃ for 3 hours, cooling to room temperature, filtering, washing the filter cake with methanol, washing the filtrate with saturated sodium bicarbonate, concentrating by organic phase decompression, and purifying by column chromatography to obtain 6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinoline-7-amine. MS (ESI) M/Z193.3 [ M + H ]] ⁺ 。

Step four: at room temperature, p-toluenesulfonic acid monohydrate (1.71g, 9mmol) was added to a solution of 7-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinolin-6-amine (576.8mg, 3mmol) in acetonitrile (14ml), cooled to 0 ℃, and KI (1.25g, 7.5 mmol) and NaNO were slowly added dropwise ₂ (414mg, 6mmol) in water (6ml), reacted at this temperature for 2h and then slowly warmed to room temperature. Adding saturated sodium bicarbonate water solution, extracting with ethyl acetate, concentrating organic phase under reduced pressure, and purifying by column chromatography to obtain target product 7-iodo-6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinoline. MS (ESI) 304.1[ M + H ]] ⁺ . The key intermediate 3. N-tert-butoxycarbonyl-N- [ 4-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -3-pyridyl]Preparation of carbamates

The method comprises the following steps: the same procedure as the first step of the key intermediate 1 is adopted to prepare the N-tert-butoxycarbonyl-N- (4-methyl-5-bromo-3-pyridyl) carbamate. MS (ESI) M/Z388.3 [ M + H ]] ⁺ 。

Step two: N-tert-Butoxycarbonyl-N- (4-methyl-5-bromo-3-pyridyl) carbamate (2.383g, 6.16mmol) was dissolved in N, N-dimethylacetamide (24 mL). To this solution were added bis (pinacol) diboron (7.823g, 30.8mmol), KOAc (1.813g, 18.5mmol) and Pd (dppf) Cl ₂ (454mg, 0.62 mmol). The mixture was heated to 90 ℃ under nitrogen and stirred for 2 hours. Cooled to room temperature, quenched with water (20mL) and extracted with ethyl acetate (3X 20 mL). The organic layers were combined, dried over sodium sulfate, filtered and evaporated. Purifying the residue with silica gel column to obtain tert-butyl N-tert-butoxycarbonyl-N- [ 4-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -3-pyridyl]Carbamate (2.077g, 78%). MS (ESI) M/Z353.2 [ M (boronic acid) + H] ⁺ 。

Preparation of key intermediate 4.8-methyl-7- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -2, 3-dihydropyrido [2,3-b ] [1,4] oxazine-1-carboxylic acid tert-butyl ester

The method comprises the following steps: ethyl glycolate (13.17mL,139.2mmol) was dissolved in THF (5mL) and cooled to 0 ℃. Potassium tert-butoxide (13.39g, 119.3mmol) was added portionwise to the solution. After stirring at 0 ℃ for 5 minutes, 5-bromo-2-chloro-4-methyl-3-nitropyridine (5.0g,19.88mmol) was added. After stirring for 2 min at 0 ℃, 15ml of saturated ammonium chloride was added, followed by extraction with EtOAc, and the organic layer was dried over sodium sulfate and concentrated in vacuo. The crude product was purified by column chromatography to give ethyl 2- ((5-bromo-4-methyl-3-nitropyridin-2-yl) oxy) acetate. MS (ESI) M/Z320.1 [ M + H ]] ⁺ 。

Step two: ethyl 2- ((5-bromo-4-methyl-3-nitropyridin-2-yl) oxy) acetate (1.3g, 4.07mmol) and Fe (1.14g, 20.4mmol) were dissolved in EtOAc (10mL) and AcOH (5 mL). After heating to 85 ℃ for 16H, the reaction mixture was cooled to room temperature and basified with saturated sodium carbonate solution, extracted with EtOAc, and the combined organic layers were dried over sodium sulfate and concentrated in vacuo to give 7-bromo-8-methyl-1H-pyrido [2,3-b ]][1,4]Oxazin-2 (3H) -ones, which were used without further purification by MS (ESI) M/Z244.1 [ M + H ]] ⁺ 。

Step three: under the protection of nitrogen, 7-bromo-8-methyl-1H-pyrido [2,3-b ] is reacted][1,4]Oxazin-2 (3H) -one (331mg,1.36mmol) was added to the flask, cooled to 0 deg.C, borane in THF (4.54mL, 0.9M, 3equiv.) was slowly added, heated to 50 deg.C for 30min, cooled to room temperature, and saturated NaHCO was added dropwise ₃ Extracted with EtOAc, and the organic layer dried over sodium sulfate and concentrated in vacuo to give 7-bromo-8-methyl-2, 3-dihydro-1H-pyrido [2,3-b ]][1,4]Oxazines, used without further purification. MS (ESI) 244.1[ M + H ]] ⁺ 。

Step four: to 7-bromo-8-methyl-2, 3-dihydro-1H-pyrido [2,3-b ] at 0 deg.C][1,4]LiHMDS (8.73mL,8.73mmol, lmol/L) was added dropwise to a solution of oxazine (1g,4.37mmol) in tetrahydrofuran (2 mL). The resulting solution was stirred at 0 ℃ under nitrogen for 0.5 hour. Then, Boc anhydride (2.85g,13.07mmol) was added, and the reaction was stirred at room temperature for 2 hours. The reaction was quenched with methanol (50 mL). The solvent was concentrated under vacuum. Purifying the residue by silica gel flash chromatography to obtain 7-bromo-8-methyl-2, 3-dihydropyrido [2,3-b ]][1,4]Oxazine-1-carboxylic acid tert-butyl ester. MS (ESI) M/Z230.1 [ M + H ]] ⁺ 。

Step five: reacting 7-bromo-8-methyl-2, 3-dihydropyrido [2,3-b ]][1,4]Oxazine-1-carboxylic acid tert-butyl ester (6.2g,18.83mmol), dipinaconodiboron (23.93g,94.22mmol), Pd (dppf) Cl ₂ (II) (2.76g,3.77mmol) and potassium acetate (5.55g,56.62mmol) were added to 1, 4-dioxane (2mL) and stirred at 90 ℃ for 2.5 h under nitrogen. The solvent was concentrated under vacuum. The residue is purified by flash chromatography on silica gel to give 8-methyl-7- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaboronHeterocyclopent-2-yl) -2, 3-dihydropyrido [2,3-b][1,4]Oxazine-1-carboxylic acid tert-butyl ester. MS (ESI) M/Z376.3 [ M + H ]] ⁺ 。

Preparation of key intermediate 5. tert-butyl (3-fluoro-5-methyl-4- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) benzyl (methyl) carbamate

The method comprises the following steps: n-bromosuccinimide (15.8g,89mmol) was added to a solution of 3-fluoro-5-methylaniline (11g,88mmol) in DMF (80mL) at 0 ℃. The reaction mixture was stirred at 0 ℃ for 30 minutes. After warming to room temperature, the reaction was stirred for an additional 1 hour. Water and EtOAc were then added and the organic phase was washed with saturated aqueous NaHCO3 solution. Then the organic phase is dried by magnesium sulfate, the solvent is evaporated under reduced pressure, and the 4-bromo-3-fluoro-5-methylaniline is obtained after column chromatography purification. MS (ESI) 204.0[ M + H ]] ⁺ 。

Step two: an aqueous solution (10mL) of sulfuric acid (4.75mL,89mmol) was slowly added to a solution of 4-bromo-3-fluoro-5-methylaniline (7.28g,36mmol) in acetonitrile (190mL) in an ice bath, after stirring for 5 minutes, a solution of sodium nitrite (4.92g, 71.4mmol) in water (10mL) was added dropwise and the reaction mixture was stirred at 0 ℃ for a further 15 minutes. Then, a solution of potassium iodide (23.7g,143mmol) in water (20mL) was added, the reaction was stirred for 20 minutes after warming to room temperature, then a saturated aqueous solution of sodium thiosulfate was added to quench the reaction, the mixture was extracted with ethyl acetate, the combined organic phases were washed with brine, dried over magnesium sulfate, and concentrated under reduced pressure. And purifying the crude product by column chromatography to obtain 2-bromo-1-fluoro-5-iodo-3-methylbenzene. MS (ESI) 315.9[ M + H ]] ⁺ 。

Step three: vinyl boronic acid pinacol ester (6.16mL,34.5mmol), Pd (dppf) Cl ₂ (2.40g,3.3mmol) and tripotassium phosphate (13.9g,65.7mmol) were added to a mixed solution of dioxane (80ml) and water (13.3 ml). The mixture is heated to 70 ℃ under the protection of nitrogen and reacted for 1 hour. After cooling to room temperature, the reaction mixture was filtered through celite, diluted with water, extracted with ethyl acetate, the combined organic phases washed with brine and dried over magnesium sulfateAnd concentrating under reduced pressure. The crude product is purified by column chromatography to obtain 2-bromo-1-fluoro-3-methyl-5-vinylbenzene. MS (ESI) 216.1[ M + H ]] ⁺ 。

Step four: to a mixed solution of 2-bromo-1-fluoro-3-methyl-5-vinylbenzene (5.46g,25.4mmol) in acetone (46mL) and water (4.6mL) were added sodium periodate (21.7g,102mmol) and a 4% aqueous osmium tetroxide solution (8.07mL, 1.27mmol) in that order. The reaction was stirred at room temperature for 2 hours. The reaction mixture was then filtered through celite, diluted with water and extracted with ethyl acetate, the combined organic phases washed with brine, the organic phase dried over magnesium sulfate and concentrated under reduced pressure. And (5) purifying the crude product by column chromatography. MS (ESI) M/Z218.0 [ M + H ]] ⁺ 。

Step five: 4-bromo-3-fluoro-5-methylbenzaldehyde (1.46g, 6.70mmol) was added to MeOH (6.70mL), a 33% methylamine solution in ethanol (3.15g, 33.5mmol) and tetraisopropyl titanate (0.982mL, 3.35mmol) were slowly added under nitrogen, and after stirring at room temperature for 3 hours, sodium borohydride (1.01g, 26.8mmol) was added to the reaction in portions and stirring at room temperature was continued for 1.5 hours. At this point, ammonia was added to the reaction mixture and stirring was continued for 15 minutes. The reaction was then acidified with 1N HCl and extracted with ethyl acetate. The aqueous phase was made basic and extracted with ethyl acetate and concentrated under reduced pressure to give 1- (4-bromo-3-fluoro-5-methylphenyl) -N-methyl-amine as a pale yellow oil. MS (ESI) M/Z231.9 [ M + H ]] ⁺ 。

Step six: to a solution of 1- (4-bromo-3-fluoro-5-methylphenyl) -N-methylmethanamine (1.32g, 5.67mmol) and triethylamine (1.58mL, 11.34mmol) in THF (18.9mL) was added di-tert-butyl dicarbonate (1.58mL, 6.80mmol), and the mixture was stirred at room temperature for 1 hour. The reaction mixture was then diluted with water and extracted with ethyl acetate. The combined organic phases were dried over magnesium sulfate and concentrated under reduced pressure. Purifying by column chromatography to obtain (4-bromo-3-fluoro-5-methylbenzyl) (methyl) carbamic acid tert-butyl ester. MS (ESI) M/Z276.0 [ M + H ]] ⁺ 。

Step seven: tert-butyl (4-bromo-3-fluoro-5-methylbenzyl) (methyl) carbamate (573mg,1.73mmol) was dissolved in THF (11.5mL), cooled to-78 deg.C, and n-butyllithium (1.6M in hexane, 1.19mL, 1) was slowly added dropwise.90 mmol). The reaction mixture was stirred for 3 minutes, then pinacol 2-isopropylborate (427uL,2.25mmol) was added slowly. The reaction was warmed to room temperature and stirred for 5 hours. Quenching the reaction with water, acidifying to pH 5-6 with 1N HCl, extracting with ethyl acetate, drying the organic phase over magnesium sulfate, and concentrating to give tert-butyl (3-fluoro 5-methyl-4- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) benzyl (methyl) carbamate the crude product was used in the next step without further purification ms (esi) M/Z324.2 [ M + H ═ M] ⁺ 。

Preparation of the Key intermediate 6. tert-butyl (tert-butoxycarbonyl) (2-methyl-3- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) phenyl) carbamate

The method comprises the following steps: the operation process of the key intermediate 3, step one, is to prepare the (3-bromo-2-methylphenyl) (tert-butyloxycarbonyl) carbamic acid tert-butyl ester. MS (ESI) M/Z387.3 [ M + H ]] ⁺ 。

Step two: the operation process of the key intermediate 3, step two is used for preparing the tert-butyl (tert-butyloxycarbonyl) (2-methyl-3- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) phenyl) carbamate. MS (ESI) M/Z434.4 [ M + H ]] ⁺ 。

Preparation of the key intermediate 7. tert-butyl (tert-butoxycarbonyl) (2, 4-dimethyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-3-yl) carbamate

The method comprises the following steps: the same procedure as in step one of key intermediate 3 was used to prepare tert-butyl (5-bromo-2, 4-dimethylpyridin-3-yl) (tert-butoxycarbonyl) carbamate. MS (ESI) M/Z402.3 [ M + H ]] ⁺ 。

Step two: the operation process of the key intermediate 3 step two is adopted to prepare the tert-butyl (tert-butoxycarbonyl)) (2, 4-dimethyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-3-yl) carbamate. MS (ESI) M/Z449.4 [ M + H ]] ⁺ 。

Preparation of key intermediate, tert-butyl 88-methyl-7- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -3, 4-dihydro-2H-1, 5-naphthyridine-1-carboxylate

The method comprises the following steps: a solution of 5-bromo-4-methyl-pyridin-3-amine (60.0g, 320.79mmol) and N-iodosuccinimide (72.2g, 320.92mmol) in acetic acid (1000mL) was stirred at 25 deg.C for 12 hours. The reaction mixture was concentrated under vacuum. The residue was purified by flash chromatography on silica gel to give 5-bromo-2-iodo-4-methyl-pyridin-3-amine (72g, 177.16mmol, 55.2% yield) as a yellow solid. MS (ESI) M/Z414.1 [ M + H ]] ⁺ 。

Step two: a solution of 5-bromo-2-iodo-4-methyl-pyridin-3-amine (55g, 175.76mmol) and NaHMDS (360mL, 720mmol) in tetrahydrofuran (500mL) was stirred at 0 deg.C for 30 minutes. Then adding Boc ₂ O (43g, 197.25mmol), and stirred at 25 ℃ for 1.5 h. The resulting solution was diluted with water and then extracted with ethyl acetate. The organic layers were combined and concentrated under vacuum. The residue was purified by flash chromatography on silica gel to give tert-butyl N- (5-bromo-2-iodo-4-methyl-3-pyridinyl) carbamate (58g, 79.9%). MS (ESI) 374.3[ M + H ═ M/Z] ⁺ 。

Step three: n- (5-bromo-2-iodo-4-methyl-3-pyridinyl) carbamic acid tert-butyl ester (500.0mg, 1.21mmol), bromo- (3-methoxy-3-oxo-propyl) zinc (15mL, 6.89mmol) and Pd (PPh) under nitrogen ₃ ) ₄ A mixture of (115.0mg, 0.10mmol) and tetrahydrofuran (10mL) was warmed to 70 ℃ and stirred for 2 hours. The reaction mixture was diluted with water. The resulting solution was extracted with ethyl acetate, and the organic layers were combined. The organic layer was dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by flash chromatography on silica gel to give 3- [ 5-bromo-3- (tert-butoxycarbonylamino) -4-methyl-2-pyridineBase of]Methyl propionate (313mg, 65.1%). MS (ESI) M/Z373.0 [ M + H ]] ⁺ 。

Step four: 3- [ 5-bromo-3- (tert-butoxycarbonylamino) -4-methyl-2-pyridinyl ] at room temperature]A solution of methyl propionate (313.0mg, 0.79mmol) in trifluoroacetic acid (1mL) and dichloromethane (4mL) was stirred for 1 hour. The reaction mixture was concentrated under vacuum. The residue was purified by flash chromatography on silica gel to give 7-bromo-8-methyl-3, 4-dihydro-1H-1, 5-naphthyridin-2-one (140mg, 70%). MS (ESI) M/Z241.0 [ M + H ]] ⁺ 。

Step five: 7-bromo-8-methyl-3, 4-dihydro-1H-1, 5-naphthyridin-2-one (120.0mg, 0.47mmol) in tetrahydrofuran (10mL) and 1M BH at 60 deg.C ₃ In THF (1.4mL, 1.42mmol) for 2 hours. The reaction was quenched with methanol and dilute hydrochloric acid. The mixture was concentrated under vacuum. The resulting 7-bromo-8-methyl-1, 2,3, 4-tetrahydro-1, 5-naphthyridine (70mg, 61.9%) was purified by column chromatography. MS (ESI) M/Z227.0 [ M + H ]] ⁺ 。

Step six: NaHDMS (0.5mL, 0.26mmol) was added to a solution of 7-bromo-8-methyl-1, 2,3, 4-tetrahydro-1, 5-naphthyridine (60.0mg, 0.26mmol) in tetrahydrofuran (10mL) at 0 deg.C, and the mixture was stirred at this temperature for 0.5 h. Then add (Boc) ₂ O (100.0mg, 0.46mmol), and stirred at room temperature for 1 hour. The reaction was quenched with methanol (1 mL). The reaction mixture was concentrated under vacuum. Purification by column chromatography gave 7-bromo-8-methyl-3, 4-dihydro-2H-1, 5-naphthyridine-1-carboxylic acid tert-butyl ester (70mg, 76.9%). MS (ESI) M/Z327.0 [ M + H ]] ⁺ 。

Step seven: 7-bromo-8-methyl-3, 4-dihydro-2H-1, 5-naphthyridine-1-carboxylic acid tert-butyl ester (30.0g, 91.68mmol), 4,4,4',4',5,5,5',5' -octamethyl-2, 2' -bis (1,3, 2-dioxaborolane) (117.0g, 460.63mmol), Pd (dppf) Cl at 100 ℃ under nitrogen ₂ A mixture of (6.7g, 9.17mmol) and KOAc (27.0g, 275.51mmol) in 1, 4-dioxane (500mL) was stirred for 3 hours. The solids were removed by filtration. The mixture was then concentrated under vacuum. Purification by column chromatography gave 8-methyl-7- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -3, 4-dihydro-2H-1, 5-naphthyridine-1-carboxylic acid tert-butyl ester (30g, 87.4%). MS (ESI) M-Z＝375.0[M+H] ⁺ 。

Preparation of key intermediate 9. tert-butyl (tert-butoxycarbonyl) (4-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridazin-3-yl) carbamate

The method comprises the following steps: following the procedure of step one of key intermediate 3, tert-butyl (5-bromo-4-methylpyridazin-3-yl) (tert-butoxycarbonyl) carbamate was prepared. MS (ESI) M/Z389.3 [ M + H ]] ⁺ 。

Step two: the operation process of the key intermediate 3, step two is used for preparing tert-butyl (tert-butoxycarbonyl) (4-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridazin-3-yl) carbamate. MS (ESI) M/Z436.3 [ M + H ]] ⁺ 。

Key intermediate 10 preparation of tert-butyl (tert-butoxycarbonyl) (5-fluoro-2-methyl-3- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) phenyl) carbamate

The method comprises the following steps: the same procedure as in step one of key intermediate 3 was used to prepare tert-butyl (3-bromo-5-fluoro-2-methylphenyl) (tert-butoxycarbonyl) carbamate. MS (ESI) 405.3[ M + H ]] ⁺ 。

Step two: the operation process of the key intermediate 3 step two is used for preparing tert-butyl (tert-butoxycarbonyl) (5-fluoro-2-methyl-3- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) phenyl) carbamate. MS (ESI) M/Z452.3 [ M + H ]] ⁺ 。

Preparation of key intermediate 11, tert-butyl (tert-butoxycarbonyl) (5-methyl-6- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridazin-4-yl) carbamate

The method comprises the following steps: following the procedure of step one of key intermediate 3, tert-butyl (6-bromo-5-methylpyridazin-4-yl) (tert-butoxycarbonyl) carbamate was prepared. MS (ESI) M/Z389.3 [ M + H ]] ⁺ 。

Step two: and the operation process of the key intermediate 3, step two is used for preparing tert-butyl (tert-butyloxycarbonyl) (5-methyl-6- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridazin-4-yl) carbamate. MS (ESI) M/Z436.3 [ M + H ]] ⁺ 。

Preparation of the Key intermediate, tert-butyloxycarbonyl) (3- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -2- (trifluoromethyl) phenyl) carbamic acid tert-butyl ester

The method comprises the following steps: the same procedure as in step one of key intermediate 3 was used to prepare tert-butyl (3-bromo-2- (trifluoromethyl) phenyl) (tert-butoxycarbonyl) carbamate. MS (ESI) M/Z441.3 [ M + H ]] ⁺ 。

Step two: the same operation process of the key intermediate 3 step two is adopted to prepare (tert-butoxycarbonyl) (3- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -2- (trifluoromethyl) phenyl) carbamic acid tert-butyl ester. MS (ESI) M/Z488.3 [ M + H ]] ⁺ 。

Key intermediate 13 preparation of tert-butyl (tert-butoxycarbonyl) (6- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyrazin-2-yl) carbamate

The method comprises the following steps: the same operation process as the first step of the key intermediate 3 is adopted to prepare the (6-bromopyrazin-2-yl) (tert-butoxycarbonyl) carbamic acid tert-butyl ester. MS (ESI) M/Z375.2 [ M + H ]] ⁺ 。

Step two: the operation process of the key intermediate 3 step two is used for preparing tert-butyl (tert-butoxycarbonyl) (6- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyrazin-2-yl) carbamate. MS (ESI) 422.3[ M + H ]] ⁺ 。

Preparation of key intermediate 14, tert-butyl (tert-butoxycarbonyl) (4-ethyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-3-yl) carbamate

The method comprises the following steps: the operation process of the key intermediate 3 step one is used for preparing the (5-bromo-4-ethylpyridin-3-yl) (tert-butyloxycarbonyl) carbamic acid tert-butyl ester. MS (ESI) M/Z402.3 [ M + H ]] ⁺ 。

Step two: and the operation process of the key intermediate 3, step two is used for preparing tert-butyl (tert-butyloxycarbonyl) (4-ethyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridine-3-yl) carbamate. MS (ESI) M/Z449.4 [ M + H ]] ⁺ 。

Preparation of key intermediate 15. methyl (4-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-3-yl) carbamic acid tert-butyl ester

The method comprises the following steps: the same operation process of the key intermediate 4 step four is adopted to prepare the (5-bromo-4-methylpyridin-3-yl) (methyl) carbamic acid tert-butyl ester. MS (ESI) M/Z302.3 [ M + H ]] ⁺ 。

Step two: and the operation process of the key intermediate 4 step five is used for preparing methyl (4-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridine-3-yl) carbamic acid tert-butyl ester. MS (ESI) 349.3[ M + H ]] ⁺ . Key intermediate 16.4- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -2, 3-dihydro-1H-pyrrolo [2, 3-c)]Preparation of pyridine-1-carboxylic acid tert-butyl ester

The method comprises the following steps: the same operation process of the key intermediate 4 step four is adopted to prepare the 4-bromo-2, 3-dihydro-1H-pyrrolo [2,3-c ]]Pyridine-1-carboxylic acid tert-butyl ester. MS (ESI) M/Z300.2 [ M + H ]] ⁺ 。

Step two: the same operation process of the key intermediate 4 step five is adopted to prepare 4- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -2, 3-dihydro-1H-pyrrolo [2, 3-c)]Pyridine-1-carboxylic acid tert-butyl ester. MS (ESI) 349.3[ M + H ]] ⁺ . Key intermediate 17.7-bromo-8-methyl-1H-pyrido [2,3-b ]][1,4]Preparation of oxazin-2 (3H) -ones

Step two: ethyl 2- ((5-bromo-4-methyl-3-nitropyridin-2-yl) oxy) acetate (1.3g, 4.07mmol) and Fe (1.14g, 20.4mmol) were dissolved in EtOAc (10mL) and AcOH (5 mL). After heating to 85 ℃ for 16H, the reaction mixture was cooled to room temperature and basified with saturated sodium carbonate solution, extracted with EtOAc, and the combined organic layers were dried over sodium sulfate and concentrated in vacuo to give 7-bromo-8-methyl-1H-pyrido [2,3-b ]][1,4]Oxazin-2 (3H) -one, which was used without further purification. MS (ESI) 244.1[ M + H ]] ⁺ 。

Preparation of key intermediate 18 (7-bromo-3, 4-dihydro-1, 5-naphthyridine-1 (2H) -carboxylic acid tert-butyl ester

The method comprises the following steps: 60% sodium hydride (5g, 126mmol) was added to 100ml of N-dimethylformamide, and diethyl malonate (20g,126mmol) was added dropwise at 0 ℃ and reacted at this temperature for 30 min. After the reaction system is clarified, a solution of intermediate 5-bromo-2-chloro-3-nitropyridine (10g, 42.1mmol) in N, N-dimethylformamide (20mL) is slowly added dropwise to the reaction solution, and the reaction is carried out overnight at room temperature. And after the reaction is completed, spin-drying the N, N-dimethylformamide, dissolving the oily substance in water, extracting with ethyl acetate, and spin-drying the organic phase under reduced pressure to obtain a crude yellow oily intermediate which is directly used for the next reaction without purification. MS (ESI) M/Z362.2 [ M + H ]] ⁺ 。

Step two: 60% sodium hydride (2.2g, 53.2mmo1) was added to 40mL of N, N-dimethylformamide, and diethyl 2- (5-bromo-3-nitropyridin-2-yl) malonate (9.68,26.2mmol) was added dropwise at 0 ℃ and reacted at this temperature for 1 h. A solution of ethyl bromoacetate (6.68,40mmol) in N, N-dimethylformamide (10mL) was then slowly added dropwise to the reaction solution and allowed to react overnight at room temperature. After completion of the reaction, N-dimethylformamide was spin-dried, a saturated sodium bicarbonate solution (50mL) was added, and extraction was performed with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, filtered and the filtrate was spin-dried. Purifying by column chromatography to obtain yellow oily intermediate (5-bromo-3-nitropyridine-2-yl) triethyl methanetricarboxylate. MS (ESI) M/Z434.2 [ M + H ]] ⁺ 。

Step three: triethyl (5-bromo-3-nitropyridin-2-yl) methanetricarboxylate (2.58,5.6mmol) was added to 10ml 6N HCl and reacted overnight at 80 ℃ under nitrogen. Cooled to room temperature, adjusted to pH 9 with saturated sodium bicarbonate solution and extracted with dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, filtered and the filtrate was spin-dried. And purifying by column chromatography to obtain a yellow oily intermediate 5-bromo-3-nitropyridine-2-ethyl propionate. MS (ESI) 304.1[ M + H ]] ⁺ 。

Step four, adding 5-bromo-3-nitropyridine-2-ethyl propionateThe ester (420mg,14mmol) was dissolved in ethanol (10mL), stannous chloride dihydrate (1.3g,7mmol) was added in portions, warmed to 80 ℃ and stirred overnight. Cooling to room temperature, and filtering to obtain a white solid intermediate 7-bromo-3, 4-dihydro-1, 5-naphthyridin-2 (1H) -one. MS (ESI) M/Z208.1 [ M + H ]] ⁺ 。

Step five: 7-bromo-3, 4-dihydro-1, 5-naphthyridin-2 (1H) -one (200mg, 0.88mmol) was dissolved in 5mL of borane in tetrahydrofuran and heated to 80 ℃ for reaction for 2H. The reaction solution was cooled to room temperature, quenched with methanol, and 3N HCl (5mL) was added dropwise, the temperature was raised to 110 ℃ and the reaction was continued for 3 h. After cooling to room temperature, the pH was adjusted to 9 with saturated aqueous sodium bicarbonate solution, and extracted with dichloromethane (20 mL). And drying the organic phase by using anhydrous sodium sulfate, filtering, spin-drying the filtrate, and purifying by column chromatography to obtain a colorless oily intermediate 7-bromo-1, 2,3, 4-tetrahydro-1, 5-naphthyridine. MS (ESI) 214.1[ M + H ]] ⁺ 。

Step six: the same operation as the key intermediate 4, step four, to prepare (7-bromo-3, 4-dihydro-1, 5-naphthyridine-1 (2H) -carboxylic acid tert-butyl ester MS (ESI) M/Z314.2 [ M + H ]] ⁺ 。

Key intermediate 19 preparation of tert-butyl (tert-butoxycarbonyl) (5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-3-yl) carbamate

The method comprises the following steps: the same procedure as in step one of key intermediate 3 was used to prepare tert-butyl (5-bromopyridin-3-yl) (tert-butoxycarbonyl) carbamate. MS (ESI) 374.3[ M + H ═ M/Z] ⁺ 。

Step two: the operation process of the key intermediate 3 step two is used for preparing tert-butyl (tert-butoxycarbonyl) (5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridine-3-yl) carbamate. MS (ESI) M/Z421.3 [ M + H ]] ⁺ 。

Preparation of key intermediate, tert-butyl 20.7-methyl-6- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -1H-imidazo [4,5-b ] pyridine-1-carboxylate

The method comprises the following steps: the 6-bromo-7-methyl-1H-imidazo [4,5-b ] is prepared by the same operation process as the key intermediate 4 step four]Pyridine-1-carboxylic acid tert-butyl ester. MS (ESI) 313.2[ M + H ]] ⁺ 。

Step two: the same operation process of the key intermediate 4 step five is adopted to prepare the 7-methyl-6- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -1H-imidazo [4,5-b]Pyridine-1-carboxylic acid tert-butyl ester. MS (ESI) M/Z360.2 [ M + H ]] ⁺ . Preparation of key intermediate 21. tert-butyl (tert-butoxycarbonyl) (4-methoxy-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-3-yl) carbamate

The method comprises the following steps: the same procedure as in step one of key intermediate 3 was used to prepare tert-butyl (5-bromo-4-methoxypyridin-3-yl) (tert-butoxycarbonyl) carbamate. MS (ESI) M/Z404.3 [ M + H ]] ⁺ 。

Step two: the operation process of the key intermediate 3 step two is used for preparing tert-butyl (tert-butoxycarbonyl) (4-methoxy-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridine-3-yl) carbamate. MS (ESI) M/Z451.3 [ M + H ]] ⁺ 。

Preparation of key intermediate 22. tert-butyl (tert-butoxycarbonyl) (4-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-2-yl) carbamate

The method comprises the following steps: the operation process of the key intermediate 3 step one is used for preparing the (5-bromo-4-methylpyridin-2-yl) (tert-butyloxycarbonyl) carbamic acid tert-butyl ester. MS (ESI) M/Z388.3 [ M + H ]] ⁺ 。

Step two: the operation process of the key intermediate 3 step two is used for preparing tert-butyl (tert-butoxycarbonyl) (4-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridine-2-yl) carbamate. MS (ESI) M/Z435.3 [ M + H ]] ⁺ 。

Key intermediate 23 preparation of tert-butyl (tert-butoxycarbonyl) (5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-2-yl) carbamate

The method comprises the following steps: the same procedure as in step one of key intermediate 3 was used to prepare tert-butyl (5-bromopyridin-2-yl) (tert-butoxycarbonyl) carbamate. MS (ESI) 374.3[ M + H ═ M/Z] ⁺ 。

Step two: and the operation process of the key intermediate 3, step two is used for preparing tert-butyl (tert-butyloxycarbonyl) (5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridine-2-yl) carbamate. MS (ESI) M/Z421.3 [ M + H ]] ⁺ . Preparation of key intermediate 24.5-bromo-2- (oxazol-2-yl) pyridine

The method comprises the following steps: to a solution of 5-bromopicolinoyl chloride (0.5g,2.5mmol) in EtOH (30mL) was added 2, 2-dimethoxyethylamine (0.26g,2.5mmol) and TEA (1.1g,10 mmol). The reaction mixture was refluxed for 2 hours, concentrated and purified by column chromatography to give 5-bromo-N- (2, 2-dimethoxyethyl) picolinamide (0.58g, 80%) as a pale yellow solid. MS (ESI) M/Z290.1 [ M + H ]] ⁺ 。

Step two: to a solution of 5-bromo-N- (2, 2-dimethoxyethyl) picolinamide (0.58g, 2.0mmol) in DCM (30mL) was added concentrated HCl (2 mL). The reaction mixture was stirred at room temperature for 4h, saturated NaHCO was added ₃ The solution was brought to pH7.5 and the organic layer was washed with brine, dried over MgSO4 and concentrated to give 5-bromo-N- (2-oxoethyl) picolinamide as a white solid. MS (ESI) 2M/Z44.1[M+H] ⁺ 。

Step three: to a solution of 5-bromo-N- (2-oxoethyl) picolinamide (0.24g, 1mmol) in toluene (10mL) was added Ph ₃ PO (0.56g, 2 mmol). The reaction mixture was refluxed overnight, concentrated and purified by column chromatography to give 5-bromo-2- (oxazol-2-yl) pyridine as a white solid. MS (ESI) M/Z226.1 [ M + H ]] ⁺ 。

Preparation of key intermediate 25.6-bromo-5-methyl 3H-1, 3-benzoxazol-2-one

5-methyl-3H-1, 3-benzoxazol-2-one (500mg,3.35mmol) and NBS (656mg,3.69mmol) were stirred in acetic acid (10mL) at 20 ℃ for 16H. The reaction mixture was diluted with water (25mL) and stirring was continued for 15 min. The mixture was then filtered and washed with water to give 6-bromo-5-methyl 3H-1, 3-benzoxazol-2-one as a white solid. MS (ESI) M/Z229.1 [ M + H ]] ⁺ 。

Preparation of key intermediate 26.3-bromo-4-methyl-2-phenylpyridine

2, 3-dibromo-4-methylpyridine (500mg,2.00mmol), phenylboronic acid (257mg, 2.lmmol), tetrakis (triphenylphosphine) palladium (116mg, 0.10mmol), and anhydrous potassium carbonate (833mg, 6.03mmol) were added to a mixed solvent of toluene 20mL and water 2 mL. And reacting for 15h at 50 ℃ under the protection of nitrogen. Cooling to room temperature, filtering, distilling under reduced pressure, and purifying with column chromatography to obtain white solid 3-bromo-4-methyl-2-phenylpyridine MS (ESI) M/Z249.1 [ M + H ]] ⁺ 。

Preparation of key intermediate 27.5-iodo-1, 4-dimethyl-pyrazol-3-amine

The method comprises the following steps: 4-methyl-3-Nitro-1H-pyrazole (500mg, 3.93mmol), iodomethane (837.9mg, 5.9mmol) and potassium tert-butoxide (484.6mg, 4.33mmol) were added to a solution of tetrahydrofuran (20mL) and stirred at 25 ℃ for 1 hour. The solvent was concentrated under vacuum. The residue was purified by flash chromatography on silica gel to give 1, 4-dimethyl-3-nitro-pyrazole as a white solid. MS (ESI) M/Z142.1 [ M + H ]] ⁺ 。

Step two: 1, 4-dimethyl-3-nitro-pyrazole (530mg, 3.76mmol) and palladium on carbon (10%) (50mg, 3.76mmol) were added to methanol (20mL) at 20 deg.C and stirred for 1 hour. After filtration, the filtrate was concentrated under reduced pressure to give 1, 4-dimethylpyrazol-3-amine as a pale yellow oil. MS (ESI) M/Z112.2 [ M + H ]] ⁺ 。

Step three: a solution of 1, 4-dimethylpyrazol-3-amine (1.53g, 13.77mmol), 2, 5-hexanedione (3.14g, 27.53mmol) and p-toluenesulfonic acid (0.24g, 1.38mmol) in toluene (60mL) was stirred at 110 ℃ for 3 hours. The solvent was concentrated under vacuum. The residue was purified by flash chromatography on silica gel to give 3- (2, 5-dimethylpyrrol-1-yl) -1, 4-dimethyl-pyrazole as a grey solid. MS (ESI) M/Z190.3 [ M + H ]] ⁺ 。

Step four: to a solution of 3- (2, 5-dimethylpyrrol-1-yl) -1, 4-dimethyl-pyrazole (500mg, 2.64mmol) in tetrahydrofuran (20mL) was slowly added n-butyllithium (1.27mL,3.17mmol) at-78 ℃. Stirring was carried out at-78 ℃ for 1 hour. Iodine (671.03mg, 2.64mmol) was then added and stirred at-78 ℃ for 10 min. The mixture was stirred at room temperature for 1 hour. The reaction was quenched with saturated aqueous sodium thiosulfate. The resulting solution was extracted with ethyl acetate and dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated under vacuum. The residue was purified by flash chromatography on silica gel to give 3- (2, 5-dimethylpyrrol-1-yl) -5-iodo-1, 4-dimethyl-pyrazole as a pale yellow oil. MS (ESI) M/Z316.2 [ M + H ]] ⁺ 。

Step five: a solution of 3- (2, 5-dimethylpyrrol-1-yl) -5-iodo-1, 4-dimethyl-pyrazole (500mg, 1.59mmol), potassium hydroxide (444.23mg, 7.93mmol), hydroxylamine hydrochloride (1.09g, 15.87mmol) in ethanol (20mL) and water (20mL) was stirred at 90 ℃ for 12 hours. Concentrating under vacuum, and purifying the residue by flash chromatography on silica gel to obtain white solid 5-iodo-1, 4-dimethyl-pyrazole-3-amine. MS (ESI) M/Z238.0 [ M + H ]] ⁺ 。

Key intermediate 28. preparation of tert-butyl (tert-butoxycarbonyl) (4-methoxy-2-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-3-yl) carbamate

The method comprises the following steps: the same procedure as in step one of key intermediate 3 was used to prepare tert-butyl (5-bromo-4-methoxy-2-methylpyridin-3-yl) (tert-butoxycarbonyl) carbamate. MS (ESI) M/Z418.3 [ M + H ]] ⁺ 。

Step two: the operation process of the key intermediate 3 step two is used for preparing tert-butyl (tert-butoxycarbonyl) (4-methoxy-2-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridine-3-yl) carbamate. MS (ESI) M/Z465.4 [ M + H ]] ⁺ 。

Key intermediate 29 preparation of tert-butyl (tert-butoxycarbonyl) (6-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-3-yl) carbamate

The method comprises the following steps: the same procedure as in step one of key intermediate 3 gave tert-butyl (5-bromo-6-methylpyridin-3-yl) (tert-butoxycarbonyl) carbamate. MS (ESI) M/Z388.3 [ M + H ]] ⁺ 。

Step two: the operation process of the key intermediate 3 step two is used for preparing tert-butyl (tert-butoxycarbonyl) (6-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridine-3-yl) carbamate. MS (ESI) M/Z435.3 [ M + H ]] ⁺ 。

Key intermediate 30 preparation of tert-butyl (tert-butoxycarbonyl) (2-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-3-yl) carbamate

The method comprises the following steps: the same procedure as in step one of key intermediate 3 was used to prepare tert-butyl (5-bromo-2-methylpyridin-3-yl) (tert-butoxycarbonyl) carbamate. MS (ESI) M/Z388.3 [ M + H ]] ⁺ 。

Step two: the same procedure as in step two of key intermediate 3 was used to prepare tert-butyl (tert-butoxycarbonyl) (2-methyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-3-yl) carbamate. MS (ESI) M/Z435.3 [ M + H ]] ⁺ 。

Preparation of key intermediate 31.7-bromo-4- [ tert-butyl (dimethyl) silyl ] oxy-8-methyl-3, 4-dihydro-2H-1, 5-naphthyridine-1-carboxylic acid tert-butyl ester

The method comprises the following steps: a solution of 5-bromo-2-iodo-4-methyl-pyridin-3-amine (55g, 175.76mmol) and NaHMDS (360mL, 720mmol) in tetrahydrofuran (500mL) was stirred at 0 deg.C for 30 minutes. Then add (Boc) ₂ O (43g, 197.25mmol) and stirred at 25 ℃ for 1.5 h. The resulting solution was diluted with water and then extracted with ethyl acetate. The organic layers were combined and concentrated under vacuum. The residue was purified by flash chromatography on silica gel to give tert-butyl N- (5-bromo-2-iodo-4-methyl-3-pyridinyl) carbamate (58g, 79.9%) as a yellow oil. MS (ESI) M/Z414.1 [ M + H ]] ⁺ 。

Step two: a solution of tert-butyl N- (5-bromo-2-iodo-4-methyl-3-pyridinyl) carbamate (58g, 140.42mmol) in tetrahydrofuran (500mL) was stirred at room temperature for 1 hour. 4-bromo-1-butene (50g, 370.37mmol) and NaI (55g, 366.67mmol) were then added and stirred at 95 ℃ for 16 h. The reaction was then quenched by addition of water and extracted with ethyl acetate. The organic layers were combined, dried and concentrated in vacuo. Purification by flash chromatography on silica gel gave N- (5-bromo-2-iodo-4-methyl) as a yellow oil-3-pyridinyl) -N-but-3-enyl-carbamic acid tert-butyl ester (30g, 45.7%). MS (ESI) M/Z467.1 [ M + H ]] ⁺ 。

Step three: n- (5-bromo-2-iodo-4-methyl-3-pyridinyl) -N-but-3-enyl-carbamic acid tert-butyl ester (30g, 64.22mmol), Pd (dppf) Cl ₂ A solution of (2.4g, 3.28mmol) and TEA (13.1g, 128.43mmol) in N, N-dimethylformamide (1500mL) was stirred under nitrogen at 110 ℃ for 1 h. The resulting solution was diluted with water and extracted with ethyl acetate. The organic layers were combined, dried and concentrated in vacuo. Purification by flash chromatography on silica gel afforded tert-butyl 7-bromo-8-methyl-4-methylene-2, 3-dihydro-1, 5-naphthyridine-1-carboxylate (13g, 59.7%). MS (ESI) M/Z420.1 [ M + H ]] ⁺ 。

Step four: 7-bromo-8-methyl-4-methylene-2, 3-dihydro-1, 5-naphthyridine-1-carboxylic acid tert-butyl ester (13g, 38.32mmol), RuCl, was added at 25 deg.C ₃ (2.4g, 11.59mmol) and NaIO ₄ A solution of (20.5g, 95.79mmol) in acetonitrile (350mL), carbon tetrachloride (350mL) and water (350mL) was stirred for 1.5 hours. The reaction was then quenched by the addition of 300mL of sodium thiosulfate solution. The resulting solution was adjusted to pH6-7 with sodium bicarbonate and extracted with ethyl acetate. The organic phase is then concentrated under vacuum. Purification by flash chromatography on silica gel afforded tert-butyl 7-bromo-8-methyl-4-oxo-2, 3-dihydro-1, 5-naphthyridine-1-carboxylate (7g, 53.5%) as a yellow solid. MS (ESI) M/Z342.2 [ M + H ]] ⁺ 。

Step five: 7-bromo-8-methyl-4-oxo-2, 3-dihydro-1, 5-naphthyridine-1-carboxylic acid tert-butyl ester (500mg, 1.47mmol) and NaBH at room temperature ₄

A solution of (150mg, 4.05mmol) in ethanol (30mL) was stirred for 2 hours. The reaction mixture was diluted with water. The resulting mixture was extracted with ethyl acetate. The organic layers were combined, dried over anhydrous sodium sulfate and concentrated in vacuo to give 7-bromo-4-hydroxy-8-methyl-3, 4-dihydro-2H-1, 5-naphthyridine-1-carboxylic acid tert-butyl ester as a yellow solid (400mg, 71.6%). MS (ESI) M/Z344.2 [ M + H ]] ⁺ 。

Step six: to tert-butyl 7-bromo-4-hydroxy-8-methyl-3, 4-dihydro-2H-1, 5-naphthyridine-1-carboxylate (330mg, 0.96mmol) and imidazole (200mg,2.94mmol) in dichloromethane (10mL) TBDMSCl (293mg, 1.95mmol) was added and the resulting solution stirred at 25 ℃ for 2h under nitrogen. Purification by flash chromatography on silica gel to give 7-bromo-4- [ tert-butyl (dimethyl) silyl ] 7-bromo-4- [ as a colorless oil]Oxy-8-methyl-3, 4-dihydro-2H-1, 5-naphthyridine-1-carboxylic acid tert-butyl ester (400mg, 90.9%). MS (ESI) M/Z458.5 [ M + H ]] ⁺ 。

Preparation of key intermediate 32.6-bromo-4- ((tert-butyldimethylsilyl) oxy) -5-methyl-3, 4-dihydro-2H-pyranol [2,3-b ] pyridine

The method comprises the following steps: to a suspension of sodium acetate (9.86g,120mmol) and 2-methoxy-4-methylpyridine (5g,39mmol) in acetic acid (40mL) was added bromine (6.0mL,120 mmol). The mixture was then stirred at 80 ℃ for 12 hours. The reaction mixture was cooled to room temperature and concentrated under reduced pressure. The residue was then treated with 10% aqueous sodium hydroxide (28g,698mmol) and saturated sodium sulfite solution (15g,120 mmol). The resulting mixture was extracted with isopropyl ether. The combined organic layers were dried over anhydrous magnesium sulfate, filtered and concentrated, and purified by column chromatography to give 3, 5-dibromo-2-methoxy-4-methylpyridine as a white solid. MS (ESI) M/Z281.0 [ M + H ]] ⁺ 。

Step two: a solution of 3, 5-dibromo-2-methoxy-4-methylpyridine (7.6g,27mmol) in diethyl ether (100mL) was cooled to-65 ℃ under nitrogen. N-butyllithium (2.5M in hexane, 12mL, 29.7mmol) was added dropwise with constant stirring, while maintaining the temperature at-65 ℃. The reaction mixture was cooled to-70 ℃ and stirred under nitrogen for 1 hour. Acrolein (4.43g, 79.12mmol) was added dropwise at-65 ℃. The reaction was then allowed to warm to room temperature overnight. 30mL of saturated sodium bicarbonate solution was added to quench the reaction. The resulting mixture was extracted with dichloromethane, dried over magnesium sulfate, and then filtered. The filtrate was concentrated to give 1- (5-bromo-2-methoxy-4-methylpyridin-3-yl) prop-2-en-1-ol as a clear oil which was used directly in the next step without further work-upAnd (5) purifying. MS (ESI) M/Z259.1 [ M + H ]] ⁺ 。

Step three: to a solution of 1- (3-bromo-6-methoxy-2-methyl-phenyl) prop-2-en-1-ol (16.84g, 65.48mmol) in dichloromethane (150mL) was added dess-martin oxidant (41.7g, 98.21mmol) at room temperature. The reaction was stirred at room temperature overnight and then quenched by the addition of saturated sodium bicarbonate solution. The organic layer was collected. The aqueous layer was extracted with dichloromethane. The combined organic layers were dried over magnesium sulfate, filtered and concentrated, and purified by flash chromatography to give 1- (3-bromo-6-methoxy-2-methyl-phenyl) prop-2-en-1-one (4.81g, 28.6% over two steps) as a clear oil. MS (ESI) M/Z257.1 [ M + H ]] ⁺ 。

Step four: to a solution of 1- (5-bromo-2-methoxy-4-methyl-3-pyridinyl) prop-2-en-1-one (4.81g, 18.78mmol) in acetic acid (80mL) was added hydrobromic acid (9.5g, 56.35 mmol). The reaction was heated to 100 ℃ for 45 minutes, then cooled to room temperature and extracted with isopropyl ether. The combined organic layers were washed with saturated sodium bicarbonate solution. The organic layer was collected, dried over magnesium sulfate, filtered and concentrated under vacuum. Purifying by flash chromatography to obtain white solid 6-bromo-5-methyl-2, 3-dihydropyrano [2,3-b ]]Pyridin-4-one. MS (ESI) M/Z443.1 [ M + H ]] ⁺ 。

Step five: to 6-bromo-5-methyl-2, 3-dihydropyrano [2,3-b ]]To a solution of pyridin-4-one (1.81mg, 7.45mmol) in methanol (50ml) was added sodium borohydride (657mg, 17.35 mmol). The mixture was stirred at room temperature for 12 hours. The reaction was quenched with water, and the mixture was extracted with ethyl acetate. The combined organic layers were then dried over magnesium sulfate, filtered and concentrated to give 6-bromo-5-methyl-3, 4-dihydro-2H-pyrano [2,3-b ] as a white solid]Pyridin-4-ol, impure, was used directly in the next step. MS (ESI) M/Z245.1 [ M + H ]] ⁺ 。

Step six: to 6-bromo-5-methyl-3, 4-dihydro-2H-pyranol [2,3-b ]]Triethylamine (0.75mL,5.4mmol) was added to a solution of pyridin-4-ol (507.2mg,2.1mmol) in dichloromethane (12 mL). The mixture was cooled to 0 ℃ under a nitrogen atmosphere, and then tert-butyldimethylsilyl trifluoromethanesulfonate (0.58mL,2.47mmol) was added. The reaction mixture was allowed to stand at room temperatureStir for 45 minutes, then add concentrated ammonium chloride solution to quench. The mixture was extracted with dichloromethane and the organic layer was dried over sodium sulfate, filtered and concentrated, purified by column chromatography to give 6-bromo-4- ((tert-butyldimethylsilyl) oxy) -5-methyl-3, 4-dihydro-2H-pyranol [2,3-b ] as a white solid]Pyridine. MS (ESI) 359.4[ M + H ]] ⁺ 。

Preparation of key intermediate 33.6-bromo-5-methyl-3, 4-dihydro-2H-pyrano [2,3-b ] pyridine-4-carbonitrile

The method comprises the following steps: reacting 6-bromo-5-methyl-3, 4-dihydro-2H-pyrano [2,3-b ] at 0 deg.C]A solution of pyridin-4-ol (1.77g, 7.26mmol) and triethylamine (3.03ml, 21.77mmol) in dichloromethane (50ml) was stirred for 5 minutes. Methanesulfonyl chloride (0.84ml, 10.88mmol) was then added. The mixture was stirred at 0 ℃ for 2 hours. The reaction was quenched with water and extracted with DCM. The combined organic layers were dried over anhydrous sodium sulfate and concentrated in vacuo to give 6-bromo-4-chloro-5-methyl-3, 4-dihydro-2H-pyrano [2,3-b ] as a red oil]Pyridine (2.28g, 8.71mmol), was used in the next step without purification. MS (ESI) M/Z323.2 [ M + H ]] ⁺ 。

Step two: reacting 6-bromo-4-chloro-5-methyl-3, 4-dihydro-2H-pyrano [2,3-b ] at 70 deg.C]A solution of pyridine (2.29mg, 8.71mmol) and tetraethylammonium cyanide (4.08g, 26.11mmol) in toluene (90ml) was stirred for 3 hours. The reaction mixture was diluted with water and extracted with ethyl acetate. The organic layers were combined, dried over anhydrous sodium sulfate and concentrated in vacuo. Purifying by flash chromatography on silica gel to obtain 6-bromo-5-methyl-3, 4-dihydro-2H-pyrano [2,3-b ] as yellow solid]Pyridine-4-carbonitrile (800.0mg, two steps 42.42%). MS (ESI) M/Z254.1 [ M + H ]] ⁺ 。

Key intermediate 34. tert-butyl (tert-butoxycarbonyl) (4, 6-dimethyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-3-yl) carbamate

The method comprises the following steps: the same procedure as in step one of key intermediate 3 was used to prepare tert-butyl (5-bromo-4, 6-dimethylpyridin-3-yl) (tert-butoxycarbonyl) carbamate. MS (ESI) M/Z402.3 [ M + H ]] ⁺ 。

Step two: the operation process of the key intermediate 3 step two is used for preparing tert-butyl (tert-butoxycarbonyl) (4, 6-dimethyl-5- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridine-3-yl) carbamate. MS (ESI) M/Z449.4 [ M + H ]] ⁺ 。

Preparation of key intermediate 35.3-bromo-6, 7-dihydropyrazolo [1,5-a ] pyrazine-5 (4H) -carboxylic acid tert-butyl ester

The method comprises the following steps: 1H-pyrazole-5-carbaldehyde (800mg, 8.33mmol) and a solution of 2- (benzylamino) ethanol (1.182ml, 8.33mmol) in MeOH (40ml) were stirred at room temperature for 1 hour. Sodium triacetoxyborohydride (5.29g,24.98mmol) and AcOH (0.953ml,16.65mmol) were then added in portions. The reaction mixture was stirred at room temperature for an additional 4 hours. The reaction mixture was quenched with 2mL of water and saturated NaHCO was added ₃ The aqueous solution was extracted with ethyl acetate, the organic layer was dried over MgSO4, filtered and concentrated, and the crude product was used directly in the next step without further purification. MS (ESI) 232.0[ M + H ]] ⁺ 。

Step two: thionyl chloride (4.26mL,58.3mmol) was added slowly to a solution of 2- (((1H-pyrazol-5-yl) methyl) (benzyl) amino) ethanol (1.93g,8.33mmol) in DCM (40mL) at 0 ℃ and the mixture was stirred overnight at RT. The solvent was removed under reduced pressure. The residue was redissolved in DMF (25mL), 60% NaH (2.0g, 50.0mmol) was added in portions, and stirred at room temperature for 1 hour. The reaction mixture was quenched with saturated ammonium chloride, extracted with EtAOc, and the organic layer was MgSO ₄ Dried, filtered and concentrated. Purifying by column chromatography to obtain 5-benzyl-4, 5,6, 7-tetrahydropyrazolo [1,5-a ]]Pyrazine (0.876g, 50% yield). MS (ESI) M/Z214.0 [ M +H] ⁺ 。

Step three: pd (OH) at room temperature ₂ (346mg,0.493mmol) and Boc ₂ O (2.77ml,11.91 mmol) was added to 5-benzyl-4, 5,6, 7-tetrahydropyrazolo [1,5-a ]]Pyrazine (876mg,4.11mmol) in MeOH (20mL) and reaction mixture was taken up in H ₂ Stirred under atmosphere for 3 hours. More Pd (OH) added ₂ (346mg,0.493mmol), reacted in H ₂ And reacting for 3 hours in the atmosphere. The reaction mixture was filtered through celite, and the filtrate was concentrated. Purifying by column chromatography to obtain 6, 7-dihydropyrazolo [1,5-a]Pyrazine-5 (4H) -carboxylic acid tert-butyl ester (916mg, 100% yield). MS (ESI) M/Z224.1 [ M + H ]] ⁺ 。

Step four: reacting 6, 7-dihydropyrazolo [1,5-a ]]Pyrazine-5 (4H) -carboxylic acid tert-butyl ester (480mg,2.150mmol) and NBS (383mg,2.150mmol) were added to DCM (15mL) at room temperature and the reaction stirred for 2H. The mixture was diluted with EtOAc and washed with brine, and the organic layer was dried over anhydrous magnesium sulfate, filtered and concentrated to give 3-bromo-6, 7-dihydropyrazolo [1,5-a]Pyrazine-5 (4H) -carboxylic acid tert-butyl ester. The crude product was used without further purification. MS (ESI) M/Z301.9/303.9 [ M + H ]] ⁺ . Key intermediate 36.6-iodo-7-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinoline preparation

Step one, step two, step three, step four: the operation process of the key intermediate 2 is the same, and 6-iodine-7-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinoline is prepared. MS (ESI) M/Z193.3 [ M + H ]] ⁺ 。

Key intermediate 37.6-iodo-7-methoxy-2, 4-dimethyl-1, 2,3, 4-tetrahydroisoquinoline preparation

The method comprises the following steps: methyl iodide (0.33mL, 5.27mmol) was added to a solution of 6-methoxy-1-methyl-3, 4-dihydroisoquinoline (840mg, 4.79mmol) in acetone (20 mL). The reaction was stirred overnight, thenThen concentrating under vacuum to obtain 6-methoxy-l, 2-dimethyl-3, 4-dihydroisoquinoline-2-ammonium iodide. MS (ESI) M/Z191.3 [ M + H ]] ⁺ 。

Step two: under the condition of 0 ℃, add NaBH4(3.36mmol) in batches to a MeOH (8mL) solution containing 6-methoxy-l, 2-dimethyl-3, 4-dihydroisoquinoline-2-ammonium iodide (0.65mmol), react at room temperature for 3 hours by using saturated NaHCO ₃ Quenching the aqueous solution, extracting with DCM, drying the organic phase with anhydrous sodium sulfate, filtering, spin-drying, and purifying by column chromatography to obtain 6-methoxy-1, 2-dimethyl-1, 2,3, 4-tetrahydroisoquinoline. MS (ESI) M/Z192.3 [ M + H ]] ⁺ 。

Step three, step four and step five: the operation process of the key intermediate 2 is the same, and 6-iodine-7-methoxy-2, 4-dimethyl-1, 2,3, 4-tetrahydroisoquinoline is prepared. MS (ESI) M/Z318.2 [ M + H ]] ⁺ 。

Key intermediate 38.7-iodo-6-methoxy-2, 4, 4-trimethyl-1, 2,3, 4-tetrahydroisoquinoline preparation

The method comprises the following steps: 2- (3-methoxybenzene) -2-methylpropen-1-amine (lg, 5.58mmol) and formic acid (2.8mL) were added to paraformaldehyde (168mg, 5.59mmol) at 0 deg.C and stirred at 50 deg.C for 14 h. The resulting mixture was cooled to room temperature, diluted with water, extracted with DCM, the aqueous phase was separated, basified with 4N sodium hydroxide solution, extracted with DCM, the combined organic phases were dried over anhydrous sodium sulfate, filtered and concentrated to give 2- (3-methoxybenzene) -2-methylpropylamine. MS (ESI) M/Z192.27 [ M + H ] +.

Step two, step three, step four, step five: the operation process of the key intermediate 2 is the same, and 7-iodine-6-methoxy-2, 4, 4-trimethyl-1, 2,3, 4-tetrahydroisoquinoline is prepared. MS (ESI) M/Z332.2 [ M + H ]] ⁺ 。

Key intermediate 39.8-iodo-9-methoxy-1, 3,4,6,11,11 a-hexahydro-2H-pyrido [1,2-b ]]Preparation of isoquinolines

The method comprises the following steps: the 9-methoxy-1, 3,4,6,11,11 a-hexahydro-2H-pyrido [1,2-b ] is obtained by the same operation process of the key intermediate 38]An isoquinoline. MS (ESI) M/Z218.3 [ M + H ]] ⁺ 。

Step two, step three, step four: the 8-iodine-9-methoxyl-1, 3,4,6,11,11 a-hexahydro-2H-pyrido [1,2-b ] is prepared by the same operation process of the key intermediate 2]An isoquinoline. MS (ESI) M/Z344.2 [ M + H ]] ⁺ 。

Key intermediate 40.5-bromo-7-iodo-2-methyl-1, 2,3, 4-tetrahydroisoquinoline preparation

The method comprises the following steps: 2-methyl-7-nitro-1, 2,3, 4-tetrahydroisoquinoline (500.0mg, 2.60mg) was added to trifluoromethanesulfonic acid (6mL), cooled to 0 deg.C, and N-bromosuccinimide (926mg, 5.20mmol) was added. The reaction was heated to 60 ℃ and then stirred for 19 hours. More N-bromosuccinimide (926mg) was then added, the reaction was stirred at 60 ℃ for an additional 5 hours, the resulting mixture was poured into ice water (50mL), the mixture was diluted with 10% aqueous sodium thiosulfate (20mL), stirred for 5 minutes, and then treated with 4N aqueous NaOH until the pH was around 10. Extracting the obtained mixture with dichloromethane, washing with 10% sodium thiosulfate aqueous solution, saturated sodium bicarbonate aqueous solution and saturated brine, drying with anhydrous sodium sulfate, filtering, spin-drying, and purifying by column chromatography to obtain 8-bromo-2-methyl-7-nitro-1, 2,3, 4-tetrahydroisoquinoline. MS (ESI) M/Z272.1 [ M + H ]] ⁺ 。

Step two, step three: the 5-bromo-7-iodo-2-methyl-1, 2,3, 4-tetrahydroisoquinoline is prepared by the same operation process of the key intermediate 2. MS (ESI) M/Z353.0 [ M + H ]] ⁺ 。

Key intermediate 41.6-fluoro-7-iodo-2-methyl-1, 2,3, 4-tetrahydroisoquinoline preparation

The method comprises the following steps: potassium nitrate (578.47mg, 5.72mmol) was added portionwise to a solution of 6-fluoro-3, 4-dihydro-2H-isoquinolin-1-one (900mg, 5.45mmol) in H2SO4(9mL) at 0 deg.C, reacted at 0 deg.C for 2H and poured into ice water, the precipitated solid was collected by filtration and dried under vacuum to afford 6-fluoro-7-nitro-3, 4-dihydro-2H-isoquinolin-1-one (1.04g, 86%). MS (ESI) M/Z221.3 [ M + H ]] ⁺ 。

Step two: to a solution of 6-fluoro-7-nitro-3, 4-dihydro-2H-isoquinolin-1-one (2.02g,9.09mmol) in THF (200mL) was added 1M BH3/THF (45.46mmol,45.5mL), stirred under reflux for 20 hours, carefully quenched with methanol (30mL) under ice bath, the resulting solution was concentrated under vacuum, added 2N HC1(50mL), heated at 80 ℃ for 3 hours with ammonia water to adjust pH to 7-8, extracted with DCM, and the organic phase was dried over anhydrous sodium sulfate, filtered, and spin-dried to give 6-fluoro-7-nitro-1, 2,3, 4-tetrahydroisoquinoline (1.89g, 100.00%). MS (ESI) M/Z197.2 [ M + H ]] ⁺ 。

Step three, step four and step five: the operation process of the key intermediate 2 is the same, and the 6-fluoro-7-iodo-2-methyl-1, 2,3, 4-tetrahydroisoquinoline is prepared. MS (ESI) M/Z292.1 [ M + H ]] ⁺ 。

Preparation of key intermediate 42.2-bromo-5-methyl-4, 7-dihydropyrazolo [1,5-a ] pyrazin-6-one

The method comprises the following steps: a solution of 3-bromo-5-methyl-1H-pyrazole (500.0mg, 3.11mmol), methyl chloroacetate (505.53mg, 4.66mmol), tetrabutylammonium iodide (57.3mg, 0.16mmol), and potassium carbonate (771.43mg, 5.59mmol) in tetrahydrofuran (10mL) was stirred at 20 ℃ for 12 hours. The mixture was concentrated under vacuum. The residue was purified by flash chromatography on silica gel to give methyl 2- (3-bromo-5-methyl-pyrazol-1-yl) acetate as a white solid. MS (ESI) 234.1[ M + H ]] ⁺ 。

Step two: a solution of methyl 2- (3-bromo-5-methyl-pyrazol-1-yl) acetate (20g, 85.81mmol), AIBN (1.41g, 8.58mmol), NBS (16.80g, 94.4mmol) in carbon tetrachloride (400mL) was stirred at 80 ℃ for 2 hours. Concentrating the resulting mixture under vacuumA compound (I) is provided. Purifying the residue by flash chromatography on silica gel to obtain 2- [ 3-bromo-5- (bromomethyl) pyrazol-1-yl as yellow solid]Methyl acetate. MS (ESI) M/Z312.0 [ M + H ]] ⁺ 。

Step three: 2- [ 3-bromo-5- (bromomethyl) pyrazol-1-yl]A solution of methyl acetate (900mg, 2.88mmol) and methylamine (896.08mg, 28.85mmol) in tetrahydrofuran (10mL) was stirred at 25 ℃ for 2 h. The mixture was concentrated under vacuum. Purifying the residue by flash chromatography on silica gel to obtain 2-bromo-5-methyl-4, 7-dihydropyrazolo [1,5-a ] as a white solid]Pyrazin-6-one. MS (ESI) M/Z231.1 [ M + H ]] ⁺ 。

Preparation of key intermediate 43.4- (4-bromo-1H-pyrazol-1-yl) piperidine-1-carboxylic acid tert-butyl ester

Sodium hydride (8.88 mmol, 60% in oil) was added portionwise to a solution of 4-bromo-1H-pyrazole (8.07mmol) in DMF (10ml) cooled to 0 ℃, stirred for 1H, 1-Boc-4-methanesulfonyloxypiperidine (1.60g, 8.88mmol) was added to the mixture, and the resulting mixture was heated to 100 ℃.

Reaction for 10h, cool the mixture to room temperature, quench the reaction with water, extract with ethyl acetate, and concentrate the organic layer under vacuum. Purifying by silica gel column chromatography. MS (ESI) M/Z331.2 [ M + H ]] ⁺ 。

Preparation of key intermediate 44.4- (2- (3-bromo-1H-pyrazol-1-yl) ethyl) morpholine

3-bromopyrazole (6.63mmol), potassium carbonate (1.83g,13.3mmol) were added to acetonitrile (30 mL). The reaction was allowed to proceed overnight at 60 ℃ and poured onto ice and extracted with DCM. The organic phase was separated, washed with brine and Na ₂ SO ₄ And (5) drying. The solvent was removed in vacuo and purified by column chromatography to give 4- (2- (3-bromo-1H-pyrazol-1-yl) ethyl) morpholine as a yellow solid. MS (ESI) M/Z261.1 [ M + H ]] ⁺ 。

Key intermediate 45.2-bromo-4, 5,6, 8-tetrahydropyrazolo [1,5-d ] [1,4] diazepin-7-one preparation

The method comprises the following steps: 2- [ 3-bromo-5- (bromomethyl) pyrazol-1-yl]A solution of methyl acetate (35.4g, 113.48mmol) and NaCN (8.87g, 181.02mmol) in DMSO (550mL) was stirred at room temperature for 1 hour, and the NaCN was carefully quenched. The reaction solution was diluted with EA (2.5L). The solution was washed with water, and the organic layers were combined. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated in vacuo and purified by flash chromatography to give methyl 2- (3-bromo-5- (cyanomethyl) -1H-pyrazol-1-yl) acetate. MS (ESI) M/Z259.1 [ M + H ]] ⁺ 。

Step two: to 2- [ 3-bromo-5- (cyanomethyl) pyrazol-1-yl]To a solution of methyl acetate (3.0g, 11.62mmol) in methanol (600mL) was added PtO ₂ (600mg, 2.64 mmol). The mixture was stirred at 25 ℃ in hydrogen at 10atm for 15 hours. The mixture was filtered. The filtrate was used in the next step without further purification. MS (ESI) M/Z263.1 [ M + H ]] ⁺ 。

Step three: to 2- [5- (2-aminoethyl) -3-bromo-pyrazol-1-yl]TEA (70mL) was added to a solution of methyl acetate (600 mL). The mixture was stirred at 25 ℃ for 15 hours. The resulting solution was concentrated in vacuo. Purification of the residue by flash chromatography on silica gel gave 2-bromo-4, 5,6, 8-tetrahydropyrazolo [1,5-d][1,4]Diaza-7-ones. MS (ESI) M/Z245.1 [ M + H ]] ⁺ 。

Key intermediate 46.2' -bromo-6 ' -methyl-5 ',6' -dihydrospiro [ cyclopropane-1, 4' -pyrazolo [1,5-d ] [1,4] diazepin-7 ' (8' H) -one preparation

The method comprises the following steps: to a solution of 3, 5-dibromo-1H-pyrazole (137g, 607mmol) in MeCN (959mL) were added tert-butyl 2-chloroacetate (137g, 910mmol, 131mL), K ₂ CO ₃ (137g, 989mmol) and TBAI (11.0g, 29.7 mmol). The resulting solution was stirred at 25 ℃ for 12 hours. The combined filtrates were concentrated by filtration to give a residue. The residue was dissolved in EtOAc and washed with water. The organic phase is passed through anhydrous Na ₂ SO ₄ Drying, filtration and concentration in vacuo afforded the crude compound tert-butyl 2- (3, 5-dibromo-1H-pyrazol-1-yl) acetate.

Step two: to a solution of compound 2- (3, 5-dibromo-1H-pyrazol-1-yl) acetic acid tert-butyl ester (248g, 729mmol) in DCM (140mL) was added TFA (2.08kg,18.2mol), and the resulting solution was heated to 80 ℃ and stirred for 2 hours. The combined reaction mixture was concentrated under reduced pressure to give the crude product. The crude product was diluted with petroleum ether/ethyl acetate (4/1, 1.0L) and the resulting suspension was stirred at 25 ℃ for 1 hour and then filtered. The filter cake was collected and dried in vacuo to give the title compound 2- (3, 5-dibromo-1H-pyrazol-1-yl) acetic acid.

Step three: a solution of 2- (3, 5-dibromo-1H-pyrazol-1-yl) acetic acid (15g, 52.84mmol), N-methylpropenylamine (5.7g, 80.15mmol), N-diisopropylethylamine (27g, 208.9mmol), and HATU (30g, 78.9mmol) in N, N-dimethylformamide (500mL) was stirred at room temperature for 16 hours. The resulting mixture was diluted with ethyl acetate and then washed with sodium chloride solution. The organic phase was dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated under vacuum. The residue is purified by flash chromatography on silica gel to give 2- (3, 5-dibromo-1H-pyrazol-1-yl) -N-methyl-N-propenyl acetamide.

Step four: a mixture of 2- (3, 5-dibromo-1H-pyrazol-1-yl) -N-methyl-N-propenylacetamide (5g, 14.84mmol), palladium acetate (166mg, 0.74mmol), triphenylphosphine (388mg, 1.48mmol), TBAB (4.8g, 14.890mmol) and potassium acetate (4.2g, 42.80mmol) in N, N-dimethylformamide (100mL) was stirred at 80 ℃ for 10 hours. After filtration, the filtrate was concentrated in vacuo. Purifying the residue on silica gel column to obtain 2-bromo-6-methyl-4-methylene-4H, 5H,6H,7H, 8H-pyrazolo [1,5-d][1,4]Diaza-7-ones. MS (ESI) M/Z258.1 [ M + H ]] ⁺ 。

Step five: trimethyl sulphoxide iodide (1.29g, 5.86mmol) and potassium tert-butoxide (656mg, 5.85mmol) in dimethyl sulphoxide (3) at room temperature0mL) was stirred for 30 minutes. Then 2-bromo-6-methyl-4-methylene-4H, 5H,6H,7H, 8H-pyrazolo [1,5-d ] is added][1,4]Diaza-7-one (500mg, 1.95mmol) in dimethyl sulfoxide (3 mL). The mixture was then stirred at 50 ℃ for 12 hours. The reaction mixture was diluted with ethyl acetate and then washed with water. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under vacuum. Purifying the residue with silica gel column to obtain 2' -bromo-6 ' -methyl-5 ',6' -dihydrospiro [ cyclopropane-1, 4' -pyrazolo [1, 5-d)][1,4]Diaza derivatives]-7'(8' H) -one. MS (ESI) M/Z271.10 [ M + H ]] ⁺ 。

Preparation of the Key intermediate, tert-butyl (3-amino-6-chloro-5, 7-difluorobenzo [ e ] [1,2,4] triazin-8-yl) carbamate

Step one and step two: the (3-amino-6-chloro-5, 7-difluorobenzo [ e ] is prepared by the same operation process of the key intermediate 1][1,2,4]Triazin-8-yl) carbamic acid tert-butyl ester. MS (ESI) M/Z332.7 [ M + H ]] ⁺ 。

Preparation of the Key intermediate, tert-butyl (3-amino-6-chloro-7-difluorobenzo [ e ] [1,2,4] triazin-8-yl) carbamate

Step one and step two: the (3-amino-6-chloro-7-difluorobenzo [ e ] is prepared by the same operation process of the key intermediate 1][1,2,4]Triazin-8-yl) carbamic acid tert-butyl ester. MS (ESI) M/Z314.7 [ M + H ]] ⁺ 。

Preparation of the Key intermediate, tert-butyl (3-amino-6-chloro-5-fluorobenzo [ e ] [1,2,4] triazin-8-yl) carbamate

Step one and step two: preparing (3-amino) by the same operation process of the key intermediate 1-6-chloro-5-fluorobenzo [ e ]][1,2,4]Triazin-8-yl) carbamic acid tert-butyl ester. MS (ESI) M/Z314.7 [ M + H ]] ⁺ 。

Preparation of the Key intermediate, tert-butyl (3-amino-6-chloro-7-cyanobenzo [ e ] [1,2,4] triazin-8-yl) carbamate

Step one and step two: the (3-amino-6-chloro-7-cyanobenzo [ e ] is prepared by the same operation process of the key intermediate 1][1,2,4]Triazin-8-yl) carbamic acid tert-butyl ester. MS (ESI) 321.7[ M + H ]] ⁺ 。

Preparation of the Key intermediate, tert-butyl (3-amino-6-chloropyrido [4,3-e ] [1,2,4] triazin-8-yl) carbamate

Step one and step two: the (3-amino-6-chloropyrido [4,3-e ] is prepared by the same operation process of the key intermediate 1][1,2,4]Triazin-8-yl) carbamic acid tert-butyl ester. MS (ESI) M/Z297.7 [ M + H ]] ⁺ 。

Preparation of the Key intermediate, tert-butyl (3-amino-6-chloropyrido [2,3-e ] [1,2,4] triazin-8-yl) carbamate

Step one and step two: the (3-amino-6-chloropyrido [2,3-e ] is prepared by the same operation process of the key intermediate 1][1,2,4]Triazin-8-yl) carbamic acid tert-butyl ester. MS (ESI) M/Z297.7 [ M + H ]] ⁺ 。

Synthesis of key intermediate 53: preparation of tert-butyl (6-chloro-3- ((2-morpholinoethyl) amino) benzo [ e ] [1,2,4] triazin-8-yl) carbamate

Key intermediate 1(2mmol), 4- (2-chloroethyl) morpholine (2.2mmol) and potassium iodide (2mmol) were added to acetonitrile (20ml) and reacted at 110 ℃. Cooling to room temperature after the reaction is completed, adding water, extracting with ethyl acetate, concentrating the organic phase under reduced pressure, and purifying by column chromatography to obtain the target compound (6-chloro-3- ((2-morpholinoethyl) amino) benzo [ e ]][1,2,4]Triazin-8-yl) carbamic acid tert-butyl ester. MS (ESI) M/Z409.9 [ M + H ]] ⁺ 。

Synthesis of key intermediate 54: preparation of 7-bromo-8-methyl-4-oxo-2, 3-dihydro-1, 5-naphthyridine-1-carboxylic acid tert-butyl ester

The method comprises the following steps: a solution of 5-bromo-2-iodo-4-methyl-pyridin-3-amine (55g, 175.76mmol) and NaHMDS (360mL, 720mmol) in tetrahydrofuran (500mL) was stirred at 0 deg.C for 30 minutes. Then add (Boc) ₂ O (43g, 197.25mmol), and stirred at 25 ℃ for 1.5 h. The resulting solution was diluted with water and then extracted with ethyl acetate. The organic layers were combined and concentrated under vacuum. The residue was purified by flash chromatography on silica gel to give tert-butyl N- (5-bromo-2-iodo-4-methyl-3-pyridinyl) carbamate (58g, 79.9%) as a yellow oil. MS (ESI) M/Z414.1 [ M + H ]] ⁺ 。

Step two: a solution of tert-butyl N- (5-bromo-2-iodo-4-methyl-3-pyridinyl) carbamate (58g, 140.42mmol) in tetrahydrofuran (500mL) was stirred at room temperature for 1 hour. 4-bromo-1-butene (50g, 370.37mmol) and NaI (55g, 366.67mmol) were then added and stirred at 95 ℃ for 16 h. The reaction was then quenched by addition of water and extracted with ethyl acetate. The organic layers were combined, dried and concentrated in vacuo. Purification by flash chromatography on silica gel afforded N- (5-bromo-2-iodo-4-methyl-3-pyridinyl) -N-but-3-enyl-carbamic acid tert-butyl ester (30g, 45.7%) as a yellow oil. MS (ESI) M/Z467.1 [ M + H ]] ⁺ 。

Step three: reacting N- (5-bromo-2-iodo-4-methyl-3-pyridyl) -N-but-3-enyl-aminoTert-butyl formate (30g, 64.22mmol), Pd (dppf) Cl ₂ A solution of (2.4g, 3.28mmol) and TEA (13.1g, 128.43mmol) in N, N-dimethylformamide (1500mL) was stirred under nitrogen at 110 ℃ for 1 h. The resulting solution was diluted with water and extracted with ethyl acetate. The organic layers were combined, dried and concentrated in vacuo. Purification by flash chromatography on silica gel afforded tert-butyl 7-bromo-8-methyl-4-methylene-2, 3-dihydro-1, 5-naphthyridine-1-carboxylate (13g, 59.7%). MS (ESI) M/Z420.1 [ M + H ]] ⁺ 。

Example 1.N ³ - (6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinolin-7-yl) -6-phenylbenzo [ e][1,2,4]Triazine-3, 8-diamine method a:

the method comprises the following steps: key intermediate 1(400mg,1.35mmol) was dissolved in a mixed solution of dioxane (21ml) and water (3ml), and pinacol ester of phenylboronic acid (329mg,2.7mmol), Pd (dppf) Cl were added ₂ (197mg,0.27mmol), sodium carbonate (429mg,4.05mmol), refluxing at 90 deg.C for 3h under nitrogen protection, cooling to room temperature, filtering with diatomaceous earth, concentrating the filtrate under reduced pressure, and purifying by column chromatography to obtain (3-amino-6-phenylbenzo [ e ]][1,2,4]Triazin-8-yl) carbamic acid tert-butyl ester. MS (ESI) M/Z338.4 [ M + H ]] ⁺ 。

Step two: the (3-amino-6-phenylbenzo [ e ] obtained in the last step][1,2,4]Dissolving triazine-8-yl) carbamic acid tert-butyl ester (263.2mg,0.7mmol) in a mixed solution of toluene (4ml) and tetrahydrofuran (0.8ml), adding key intermediate 2(212.1mmol,0.7mmol), cuprous iodide (136.5mg,0.7mmol), N' -dimethylethylenediamine (61.6mg,0.7mmol) and potassium phosphate (446mg,2.1mmol), refluxing at 110 ℃ for 24h under the protection of nitrogen, cooling to room temperature, filtering with diatomaceous earth, spin-drying the filtrate under reduced pressure, and purifying by column chromatography to obtain (3- ((6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinolin-7-yl) amino) -6-phenylbenzo [ e ]][1,2,4]Triazin-8-yl) carbamic acid tert-butyl ester. MS (ESI) M/Z513.6 [ M + H ]] ⁺ 。

Step three: to (3- ((6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinolin-7-yl) amino) -6-phenylbenzo [ e][1,2,4]To a solution of t-butyl triazin-8-yl) carbamate (51.3mg,0.1mmol) in dichloromethane (5ml) was added trifluoroacetic acid (1ml) and the mixture was stirred at room temperature for 2 h. Adding saturated sodium bicarbonate to adjust pH to neutral, extracting with dichloromethane for three times, mixing organic phases, drying under reduced pressure, and purifying by column chromatography to obtain orange red target compound N ³ - (6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinolin-7-yl) -6-phenylbenzo [ e][1,2,4]Triazine-3, 8-diamine. (37mg, 89.7%). MS (ESI) M/Z413.5 [ M + H ]] ⁺ 。 ^1H ^NMR (400MHz,DMSO-d ₆ )δ8.88(s,1H),7.99(s,1H),7.77–7.70(m,2H),7.55–7.52(m,1H),7.51(d,J＝6.1Hz,1H),7.49–7.43(m,1H),6.94–6.86(m,5H),3.90(s,2H),3.86(s,3H),3.01(d,2H),2.95(d,J＝4.9Hz,2H),2.63(s,3H).

The method B comprises the following steps:

the method comprises the following steps: preparing tert-butyl (6-chloro-3- ((6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinolin-7-yl) amino) benzo [ e) by the operation of the second step of the reference method A][1,2,4]Triazin-8-yl) carbamates. MS (ESI) M/Z471.0 [ M + H ]] ⁺ 。

Step two: the tert-butyl (6-chloro-3- ((6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinolin-7-yl) amino) benzo [ e obtained in the last step][1,2,4]Triazin-8-yl) carbamates (1eq.) in solutionTo the dioxane solution, pinacol diboron (1.3eq.), Pd (dppf) Cl were added ₂ (0.2eq.), sodium carbonate (3eq.), and reflux at 90 ℃ for 3h under the protection of nitrogen, cooling to room temperature, suction-filtering with diatomite, concentrating the filtrate under reduced pressure, and purifying by column chromatography to obtain tert-butyl (3- ((6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinolin-7-yl) amino) -6- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) benzo [ e][1,2,4]Triazin-8-yl) carbamates. MS (ESI) M/Z563.5 [ M + H ]] ⁺ 。

Step three: tert-butyl (3- ((6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinolin-7-yl) amino) -6- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) benzo [ e][1,2,4]Triazin-8-yl) carbamates (1eq.), bromobenzenes (1eq.), Pd (dppf) Cl ₂ (0.2eq.) and sodium carbonate (3eq.) are dissolved in a mixed solution (7:1) of dioxane and water to react for 3h at 90 ℃ under the protection of nitrogen, the mixture is cooled to room temperature, diatomite is filtered and filtered, the filtrate is decompressed and concentrated, and column chromatography purification is carried out to obtain (3- ((6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinolin-7-yl) amino) -6-phenylbenzo [ e ]][1,2,4]Triazin-8-yl) carbamic acid tert-butyl ester. MS (ESI) M/Z513.6 [ M + H ]] ⁺ 。

Step four: preparing orange red target compound N by referring to the operation of step three of the method A ³ - (6-methoxy-2-methyl-1, 2,3, 4-tetrahydroisoquinolin-7-yl) -6-phenylbenzo [ e][1,2,4]Triazine-3, 8-diamine. MS (ESI) M/Z413.5 [ M + H ]] ⁺ 。

Biological Activity test example 1HPK1 ADP-Glo enzymatic Activity test

Preparing enzyme activity test buffer containing 40mM Tris, pH7.5; 20mM MgC 12; 0.1mg/ml BSA; 50uM DTT. The compounds were dissolved in pure DMSO at a stock concentration of 10 mM. The compound DMSO solution is continuously diluted three times of gradient from 100uM, 11 concentrations are counted, the diluted compound is diluted by enzyme activity test buffer solution in a ratio of 1:20, 1ul of the diluted compound is added into a working hole, and each concentration is two times of the diluted compound. Both negative and positive control wells were loaded with 1ul of DMSO solution diluted 1: 20. 2.5 Xsubstrate/ATP working solution was prepared as enzyme activity assay buffer containing 45uM ATP and 0.25ug/ul MBP protein, 2.5 Xsubstrate/ATP working solution 2ul was added to each well. 2.5 Xenzyme reaction working solution is prepared to be enzyme activity test buffer solution containing 0.5ng/ul HPK1 recombinant protein (Signalchem, the product number is M23-11G-10), 2.5 Xenzyme reaction working solution is added into each working hole, and only 2ul enzyme activity test buffer solution is added into a negative control hole. After the reaction is finished, 5ul of ADP-Glo reagent (Promega, product number V1901) is added into each hole, the reaction is continued for 40 minutes at room temperature, 10ul of kinase detection reagent (Promega, product number V1901) is added, the reaction is continued for 20 minutes at room temperature, and then the final luminescence signal is measured.

The positive and negative well mean values were calculated as positive control value (Signalpos) and negative control value (Signalneg), respectively. The Inhibition ratio was calculated by the formula (signaling) Inhibition rate ═ signaling-signaling)/(signaling-signaling) × 100% for the working well signal value (signaling). Calculated inhibition concentration-inhibition curves were plotted as a non-linear fit in GraphPad Prism software and IC50 values were calculated.

Biological Activity test example 2GLK ADP-Glo enzymatic Activity test

Preparing enzyme activity test buffer containing 40mM Tris, pH7.5; 20mM MgC 12; 0.1mg/ml BSA; 50uM DTT. The compounds were dissolved in pure DMSO at a stock concentration of 10 mM. The compound DMSO solution is continuously diluted three times of gradient from 100uM, 11 concentrations are counted, the diluted compound is diluted by enzyme activity test buffer solution in a ratio of 1:20, 1ul of the diluted compound is added into a working hole, and each concentration is two times of the diluted compound. Both negative and positive control wells were loaded with 1ul of DMSO solution diluted 1: 20. The 2.5 Xsubstrate/ATP working solution was prepared as enzyme activity assay buffer containing 105uM ATP and 0.5ug/ul PKA substrate polypeptide, 2.5 Xsubstrate/ATP working solution 2ul was added to each working well. 2.5 Xenzyme reaction working solution containing 2.5ng/ul enzyme activity test buffer solution of GLK recombinant protein (Signalchem, cat # M25-11G-10) was prepared, 2ul enzyme reaction working solution was added to each working well, and only 2ul enzyme activity test buffer solution was added to the negative control well. The plate-sealing membrane was attached and centrifuged, and the reaction mixture was reacted at room temperature for 1 hour, 5ul of ADP-Glo reagent (Promega, cat # V1901) was added to each well after the reaction was completed and reacted at room temperature for 40 minutes, followed by addition of 10ul of kinase detection reagent (Promega, cat # V1901) and reaction at room temperature for 20 minutes, and the final luminescence signal was measured.

The positive and negative well mean values were calculated as positive control value (Signalpos) and negative control value (Signalneg), respectively. The Inhibition ratio of the working well signal value (Signaltest) was calculated according to the formula (Signalpos-Signaltest)/(Signalpos-Signalneg) × 100%. Calculated inhibition concentration-inhibition curves were plotted as a non-linear fit in GraphPad Prism software and IC50 values were calculated. The results of the experiment are shown in the following table:

TABLE 1HPK1, GLK enzyme activity test results

Claims

1. A compound of formula I, or a pharmaceutically acceptable salt, isomer, or hydrate thereof:

wherein:

p and q are each independently 0, 1,2,3,4 or 5.

2. The compound of claim 1, or a pharmaceutically acceptable salt, isomer, or hydrate thereof, having any of the structures shown below:

wherein:

Y is selected from NR _b 、O、S(O) _t Or CR ₄ R ₅ ；

Can be located at said M ¹ 、M ² Or M ³ Above (Namely M ¹ 、M ² Or M ³ Is C);

R ₄ Selected from the group consisting of: H. deuterium, C (O) C _1-6 Alkyl radical, C _1-6 Alkyl radical, C _1-6 Deuterated alkyl, C _3-8 Cycloalkyl radical, C _2-6 Alkenyl radical, C _2-6 Alkynyl or C having 1-3 heteroatoms selected from the group N, S, O _3-12 Heterocyclyl, wherein R is ₄ The alkyl, alkenyl, alkynyl, cycloalkyl or heterocyclyl group in the group represented may be substituted with 1 to 3 substituents each independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _1-6 Haloalkoxy, C _3-8 Cycloalkyl radical, C _5-10 A heterocyclic radical and NR ₆ R ₇ ；

R ₆ 、R ₇ Each independently is H or C _1-6 Alkyl radical, C _1-6 Deuterated alkyl, C _3-8 Cycloalkyl, C having 1-3 hetero atoms selected from the group consisting of N, S and O _3-12 Heterocyclyl, wherein R is ₆ Or R ₇ The alkyl, cycloalkyl and heterocyclic groups represented by the formula (I) may be substituted by 1 to 3 substituents independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 HalogenatedAlkyl radical, C _1-6 Alkoxy radical, C _3-8 Cycloalkyl radical, C _3-12 A heterocyclic group.

n is 1,2 or 3;

m is 1,2 or 3; and n + m is less than or equal to 4;

p and q are each independently 0, 1,2,3,4 or 5;

t is 0, 1 or 2;

R _a 、R ₁ 、R ₂ and R ₃ Each independently selected from the group consisting of: H. deuterium, halogen, OH, CN, NO ₂ 、C _1-6 Deuterated alkyl, C _1-6 Alkyl radical, C _3-8 Cycloalkyl radical, C _2-6 Alkenyl radical, C _2-6 Alkynyl, C _1-6 Alkoxy radical, C _6-10 Aryl, P (O) R ₆ R ₇ 、S(O) ₂ R ₆ 、S(O) ₂ NR ₆ R ₇ 、NR ₆ R ₇ 、C(O)NR ₆ R ₇ 、C(O)NR ₆ S(O) ₂ R ₇ 、NR ₆ S(O) ₂ R ₇ 、C(O)R ₆ 、NR ₆ C(O)R ₇ 5-12 membered heteroaryl having 1-3 heteroatoms selected from the group consisting of N, S, O below, 3-12 membered heterocyclyl having 1-3 heteroatoms selected from the group consisting of N, S, O below, wherein R is substituted with R _a 、R ₁ 、R ₂ And R ₃ Alkyl, alkenyl, alkynyl, alkoxy in the group representedThe phenyl, heteroaryl, cycloalkyl or heterocyclyl group may be substituted with 1 to 3 substituents each independently selected from the group consisting of: halogen, OH, CN, NO ₂ 、C _1-6 Alkyl radical, C _1-6 Haloalkyl, C _1-6 Alkoxy radical, C _1-6 Haloalkoxy, NR ₆ R ₇ 、C(＝O)NR ₆ R ₇ 、C(＝O)NR ₆ S(＝O) ₂ R ₇ 、P(＝O)R ₆ R ₇ 、S(＝O) ₂ R ₆ 、S(＝O) ₂ NR ₆ R ₇ 、NR ₆ S(＝O) ₂ R ₇ ；

p and q are each independently 0, 1,2,3,4 or 5;

X ₁ selected from the group consisting of: none, NR, O, S (O) _t 、CHR、NRC(O)、C(O)NR、NRC(O)C(O)NR、NRC(O)NR、NRC(S)NR、NRC(O)NRCH ₂ 、NRC(S)NRCH ₂ Wherein, in the step (A),the R is selected from the following group: H. c _1-6 Alkyl radical, C _1-6 Deuterated alkyl, C _3-8 Cycloalkyl or a 3-12 membered heterocyclic group having 1-3 heteroatoms selected from the group consisting of N, S and O;

M ⁷ is CR ⁸ 、NR ⁸ N, O or S;

M ⁸ is CR ⁸ Or N;

ring C is selected from the group consisting of: c _5-8 Cycloalkyl or a 5-to 8-membered heterocyclic ring having at least 3 ring-forming carbon atoms and 1,2, or 3 ring-forming heteroatoms independently selected from the group consisting of N, P, O and S; wherein C is _5-8 Cycloalkyl and 5-to 8-membered heterocycle are independently optionally selected from R by 1,2,3,4 or 5 independently ⁸ Substituted with the substituent(s); and wherein C _5-8 Two substituents of cycloalkyl or 5-to 8-membered heterocycle, when present, optionally together form a ring optionally substituted with 1,2,3 or 4 independently selected from R ⁸ Substituted spiro, fused or bridged cycloalkyl (e.g., C) _3-6 Cycloalkyl) orOptionally substituted by 1,2,3 or 4 independently selected from R ⁸ A spiro, fused or bridged heterocyclic group (e.g., a 3-to 6-membered heterocyclic group) substituted with the substituent(s) of (a);

q is 0, 1,2,3,4 or 5;

M ⁷ is CR ⁸ 、NR ⁸ 、NO or S;

M ⁸ is CR ⁸ Or N;

q is 0, 1,2,3,4 or 5.

3. The compound of claim 2, or a pharmaceutically acceptable salt, isomer, or hydrate thereof, wherein the compound of formula II-a has any of the following structures:

the compound of formula II-b has any structure shown as the following formula:

the compound of formula II-c has any structure shown as the following formula:

4. the compound of claim 1, or a pharmaceutically acceptable salt, isomer, or hydrate thereof, having any of the structures shown below:

5. a process for preparing a compound according to any one of claims 1 to 4, or a pharmaceutically acceptable salt, isomer or hydrate thereof, characterized in that: the method is any one of the following methods:

the method A comprises the following steps:

the compound of the formula I is obtained by carrying out reactions such as Suzuki coupling, Ullmann coupling, deprotection and the like on benzo 1,2, 4-triazine or pyrido 1,2, 4-triazine compounds.

The method B comprises the following steps:

performing Ullmann coupling, Suzuki coupling, deprotection and other reactions on a benzo 1,2, 4-triazine or pyrido 1,2, 4-triazine compound to obtain a compound shown in the formula I.

6. A pharmaceutical composition comprising a therapeutically effective amount of one or more compounds of any one of claims 1-4, or a pharmaceutically acceptable salt, isomer, or hydrate thereof, and a pharmaceutically acceptable excipient.

7. Use of a compound according to any one of claims 1-4, or a pharmaceutically acceptable salt, isomer, or hydrate thereof, for the preparation of a pharmaceutical composition for the prevention or treatment of a disease responsive to inhibition of HPK1 activity in a subject.

8. The use according to claim 7, wherein the disease is cancer.

9. Use of a compound according to any one of claims 1-4, or a pharmaceutically acceptable salt, isomer, or hydrate thereof, in an HPK1 kinase inhibitor.