CN112442028A

CN112442028A - RET selective inhibitor and preparation method and application thereof

Info

Publication number: CN112442028A
Application number: CN201910825867.3A
Authority: CN
Inventors: 刘志华; 袁德闯; 陈坤成; 袁保昆; 任仁; 闵汪洋; 刘希杰; 张凯; 孙永亮; 路畅; 孙颖慧
Original assignee: Shouyao Holdings Beijing Co Ltd
Current assignee: Shouyao Holdings Beijing Co Ltd
Priority date: 2019-09-03
Filing date: 2019-09-03
Publication date: 2021-03-05

Abstract

The invention provides RET selective inhibitors, methods of preparation and uses thereof; the present invention provides compounds of formula (I) and pharmaceutically acceptable salts, solvates, polymorphs, or isomers thereof, pharmaceutical compositions comprising these compounds, and the use of such compounds in the treatment of RET related diseases.

Description

RET selective inhibitor and preparation method and application thereof

Technical Field

The present invention relates to compounds which selectively inhibit the activity of RET proteins, to processes for their preparation and to their use in pharmaceutical compositions.

Background

RET is a protooncogene, located on chromosome 10. The RET protein encoded by the RET gene is a Receptor Tyrosine Kinase (RTK) present in cell membranes, belongs to a member of the cadherin superfamily, and plays an important role in the normal development of the peripheral sympathetic and parasympathetic nervous systems of the brain, the production of calcitonin by thyroid C cells, and is also essential for the thyroid and lung, as well as hematopoietic progenitor cells and other tissues (Nat. Rev. cancer 2014; 14 (3): 173-86.).

The domain composition of RET proteins is identical to that of other protein tyrosine kinases, and contains a typical extracellular domain, a transmembrane segment, followed by an intracellular membrane proximal segment, a kinase domain, and a carboxy-terminal tail. The extracellular domain contains four cadherin-like domain repeats of about 110 amino acid residues (CD1/2/3/4) and a cysteine-rich segment of about 150 amino acid residues immediately adjacent to the transmembrane segment, with extracellular Ca present between CD2 and CD3²+ binding site (J.biol.CHEm.2001; 276 (38): 35808-35817).

RET is the only receptor protein tyrosine kinase with a cadherin-like domain. Although having the same domain composition as other protein tyrosine kinases, the activation mechanism of RET differs, and its activation requires an additional GDNF family receptor (GDNF/NTRN/ARTN/PSP) -GFR co-receptor (GFR 1/2/3/4). Members of the GDNF family typically exist as disulfide-linked homodimers that interact with two GFR complexes to form binary complexes that interact with two RET receptors, eventually forming a hexameric complex consisting of two receptors, two co-receptors, and a ligand dimer (nat. rev. cancer 2014; 14 (3): 173-86). RET is in a non-phosphorylated, inactive state in the absence of ligand-mediated stimulation. When the RET protein binds to its ligand, GDNF, it causes phosphorylation of the RET protein receptor and brings RET into an activated state. Activated RET will phosphorylate its substrate, resulting in activation of downstream signaling pathways. The signaling pathways involved in the RET protein include the PI3K-AKT-mTOR pathway and the RAS-RAF-MEK-ERK pathway. The PI3K-AKT-mTOR pathway is involved in cell survival, while the RAS-RAF-MEK-ERK pathway is involved in cell proliferation. Therefore, RET protein has a certain role in Cell survival, migration and proliferation (Cell 2010; 141 (7): 1117-1134; Cell. Signal. 2014; 26 (8): 1743-1752).

The mutations occurring in the RET gene encoding RET protein kinase include activation point mutation and gene fusion, which lead to excessive activation of RET signaling pathway and uncontrolled proliferation of cells, resulting in diseases such as malignant tumors. Chromosomal rearrangement of ret gene is associated with the occurrence of thyroid papillary carcinoma, and the chromosomal rearrangement may lead to a break in the middle of the ret gene, and the kinase domain of the ret gene is fused with more than ten different genes, such as kif5b, cccc 6, ncoa4, etc., to form a fusion gene that drives tumor proliferation (Transl Lung Cancer Res.2015; 4 (2): 156-. In 5% -40% of Papillary Thyroid Carcinomas (PTCs) of the thyroid, ret gene fusion is found, and ret gene rearrangement in papillary thyroid carcinomas is related to ionizing radiation, namely, the cancers occur due to nuclear radiation in the environment and the like. The fusion of ret genes resulting from chromosomal rearrangements also occurs in non-small cell lung cancer (NSCLC), with the frequency of ret gene fusions in NSCLC being approximately 1% -2%, and fusion partner genes for the four ret genes are currently identified as kif5b, cccd 6, ncoa4, and trim33, respectively, with the most common form of fusion kif5b-ret (Oncoloist.2013; 18 (7): 865-875). Point mutations in the ret gene, such as those caused by activation of the ret gene by mutations at the V804M, M918T alleles, also lead to tumorigenesis, mainly in multiple endocrine tumors (MEN2A, MEN2B) and Medullary Thyroid Carcinoma (MTC) (Ann Oncol.2016; 27 (7): 1286-1291). Germ cell ret gene mutations, occurring in almost all hereditary medullary thyroid cancers, comprise multiple endocrine adenomatosis type 2. In acquired medullary thyroid carcinoma, ret gene mutations also occur in 50% of patients (Ann Oncol.2016; 27 (7): 1286-1291).

Currently, in the United states, the number of new cases of tumors bearing RET mutations or RET fusion proteins is estimated to be about 10,000 per year, which is about the same as the incidence of chronic myelogenous leukemia (PHarmalogic ResearcH H2018; 128: 1-17). Several drugs against RET have been FDA approved for the treatment of cancer: cabozantinib, Vandertinib, Lenvatinib, Sorafenib, Alectonib, etc., which are multi-kinase inhibitors with anti-RET activity, have relatively poor clinical effects for RET targeted therapy and are accompanied by toxic side effects due to significant inhibition of other kinases such as vascular endothelial growth factor receptor 2(VEGFR2) (Nat Rev Clin Oncol.2018; 15 (3): 151-167). Therefore, the development of highly selective RET inhibitors has attracted research interest from numerous pharmaceutical companies, and the development of specific RET inhibitors has become an emerging therapeutic approach to improve the therapeutic efficacy of RET-driven cancers.

Disclosure of Invention

The invention provides a selective inhibitor of RET, which is a compound represented by a general formula (I) or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof. The invention also provides a series of compounds represented by the general formula (I) and pharmaceutically acceptable salts, solvates, polymorphs or isomers thereof, pharmaceutical compositions containing the compounds, and methods of treating diseases using the compounds.

In one aspect, the present invention provides a compound of formula (I) or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof:

(I)

wherein,

x and Y are each independently selected from CH and N;

a is selected from 5-6 membered heteroaromaticRadical, 3-to 12-membered heterocyclyl, -O-C_1-6Alkyl, -S-C_1-6Alkyl and-SO₂-C_1-6Alkyl, said heteroaryl and heterocyclyl being each optionally substituted by C_1-6Alkyl, -CN, -OH, -NR⁵R⁶Halogen, -O-C_1-6Alkyl, -S-C_1-6Alkyl, or-SO₂-C_1-6Alkyl substitution, said C_1-6Alkyl may optionally be substituted by-OH, halogen, or- (CO) N (CH)₃)₂Is substituted, and R⁵And R⁶Each independently selected from H and C_1-6An alkyl group;

R¹、R¹’、R²and R²' independently of one another are H or C_1-6Alkyl, or R¹And R¹' one of the compounds with R²And R²One of' may be taken together to form a bond or- (CH)₂)_m-；

Z is selected from- (CH)₂)_m-S-C_1-6Alkyl, - (CS) -N (R)⁸)₂、

And- (CH)₂)_n-Ar；

R⁸Each independently selected from H, C_1-6Alkyl, -O-C_1-6Alkyl and- (CH)₂)_m-O-C_1-6An alkyl group;

ar is selected from the group consisting of phenyl, pyridyl and pyrimidinyl, each of which may optionally be substituted by-O-C_1-6Alkyl, -S-C_1-6Alkyl, -SO-C_1-6Alkyl, or-SO₂-C_1-6Alkyl substitution;

each M is independently selected from 1 and 2;

n is selected from 0, 1 and 2;

in some embodiments of the invention, A is selected from 5-6 membered heteroaryl and 3-12 membered heterocyclyl, each of which may be optionally substituted with C_1-6Alkyl, -CN, -OH, -NR⁵R⁶Halogen, -O-C_1-6Alkyl, -S-C_1-6Alkyl, or-SO₂-C_1-6Alkyl substitution, said C_1-6Alkyl may optionally be substituted by-OH, halogen, or- (CO) N (CH)₃)₂Is substituted, and R⁵And R⁶Each independently selected from H and C_1-6An alkyl group;

in some embodiments of the invention, A is selected from 5-6 membered heteroaryl and 4-9 membered heterocyclyl, each of which may be optionally substituted with C_1-6Alkyl, -CN, -OH, -NR⁵R⁶Halogen, -O-C_1-6Alkyl, -S-C_1-6Alkyl, or-SO₂-C_1-6Alkyl substitution, said C_1-6Alkyl may optionally be substituted by-OH, halogen, or- (CO) N (CH)₃)₂Is substituted, and R⁵And R⁶Each independently selected from H and C_1-6An alkyl group;

in some embodiments of the invention, the compounds of the invention are selected from:

or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof.

Yet another aspect of the present invention is directed to a pharmaceutical composition comprising a compound of formula (I) of the present invention, or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, and a pharmaceutically acceptable carrier;

in another aspect, the present invention provides a method of treating a RET-associated disease, said method comprising administering to a subject an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, or a combination thereof; in some embodiments, the RET-associated disease is lung cancer, papillary thyroid cancer, medullary thyroid cancer, differentiated thyroid cancer, recurrent thyroid cancer, refractory differentiated thyroid cancer, multiple endocrine tumors of type 2A or 2B (MEN2A or MEN2B, respectively), pheochromocytoma, parathyroid hyperplasia, breast cancer, colorectal cancer, papillary renal cell carcinoma, gastrointestinal mucosal ganglion cell tumor, and cervical cancer, preferably RET-fused body lung cancer or medullary thyroid cancer, more preferably small cell lung cancer, non-small cell lung cancer, bronchiolar lung cancer, or lung adenocarcinoma;

in some embodiments of the invention, the subject to which the invention relates is a mammal including a human;

in another aspect, the present invention provides the use of a compound of formula (I), or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, in the manufacture of a medicament for the treatment of a RET-associated disease; in some embodiments, the RET-associated disease is lung cancer, papillary thyroid cancer, medullary thyroid cancer, differentiated thyroid cancer, recurrent thyroid cancer, refractory differentiated thyroid cancer, multiple endocrine tumors of type 2A or 2B (MEN2A or MEN2B, respectively), pheochromocytoma, parathyroid hyperplasia, breast cancer, colorectal cancer, papillary renal cell carcinoma, gastrointestinal mucosal ganglion cell tumor, or cervical cancer, preferably RET fusogenic lung cancer or medullary thyroid cancer, more preferably small cell lung cancer, non-small cell lung cancer, or bronchiolar lung cancer or lung adenocarcinoma.

Detailed Description

Exemplary embodiments utilizing the principles of the present invention are set forth in the following detailed description of the invention. The features and advantages of the present invention may be better understood by reference to the following summary.

It should be understood that the scope of the various aspects of the invention is defined by the claims and that methods and structures within the scope of these claims and their equivalents are intended to be covered thereby.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the claimed subject matter belongs. All patents, patent applications, and publications cited herein are incorporated by reference in their entirety unless otherwise indicated.

It is to be understood that both the foregoing general description and the following detailed description are exemplary, explanatory and are not restrictive of any inventive subject matter. The use of the singular forms also includes the plural unless specifically stated otherwise. The use of "or", "or" means "and/or" unless stated otherwise. Furthermore, the term "comprising" as well as other forms, such as "includes," "including," and "containing," are not limiting.

Certain chemical terms

The terms "optional," "optional," or "optionally" mean that the subsequently described event or circumstance may or may not occur, and that the description includes instances where said event or circumstance occurs and instances where it does not. For example, "optionally substituted alkyl" means "unsubstituted alkyl" or "substituted alkyl". And, optionally substituted groups may be unsubstituted (e.g.: CH)₂CH₃) Fully substituted (e.g.: -CF₂CF₃) Monosubstituted (e.g.: -CH₂CH₂F) Or any level between mono-and fully substituted (e.g.: -CH₂CHF₂、-CF₂CH₃、-CFHCHF₂Etc.). It will be appreciated by those skilled in the art that any group containing one or more substituents will not incorporate any substitution or substitution pattern which is sterically impossible and/or cannot be synthesized.

Unless otherwise indicated, conventional methods within the skill of the art are employed, such as mass spectrometry, nuclear magnetism, high performance liquid chromatography, infrared and ultraviolet/visible spectroscopy, and pharmacological methods. Unless specific definitions are set forth, the nomenclature used herein in the analytical chemistry, organic synthetic chemistry, and pharmaceutical and medicinal chemistry, as well as the laboratory procedures and techniques, are those known in the art. Standard techniques can be used in chemical synthesis, chemical analysis, pharmaceutical preparation, formulation and delivery, and treatment of patients. For example, the reaction and purification can be carried out using the instructions of the kit from the manufacturer, or according to the methods known in the art or the instructions of the present invention. The techniques and methods described above can generally be practiced according to conventional methods well known in the art, as described in various general and more specific documents referred to and discussed in this specification. In the present specification, groups and substituents thereof may be selected by one skilled in the art to provide stable moieties and compounds.

When a substituent is described by a general formula written from left to right, the substituent also includes chemically equivalent substituents obtained when the formula is written from right to left. For example, -CH₂O-is equivalent to-OCH₂-。

As used herein, the terms "group", "chemical group" or "chemical group" refer to a particular portion or functional group of a molecule. Chemical groups are often considered as chemical entities embedded in or attached to a molecule.

Some of the chemical groups named herein may be referred to by a shorthand notation for the total number of carbon atoms. E.g. C₁-C₆Alkyl describes an alkyl group, as defined below, having a total of 1 to 6 carbon atoms. The total number of carbon atoms indicated by shorthand notation does not include carbon atoms on possible substituents.

The terms "halogen", "halo" or "halide" refer to bromine, chlorine, fluorine or iodine.

The terms "aromatic", "aromatic ring", "aromatic" and "aromatic-cyclic" as used herein refer to a planar ring portion of one or more rings having a delocalized electron-conjugated system of 4n +2 electrons, where n is an integer. The aromatic ring may be formed of 5, 6, 7, 8, 9 or more atoms. The aromatic compound may be optionally substituted and may be monocyclic or fused-ring polycyclic. The term aromatic compound includes all carbocyclic rings (e.g., benzene rings) and rings containing one or more heteroatoms (e.g., pyridine).

The term "heteroatom" or "hetero" as used herein alone or as part of another component refers to atoms other than carbon and hydrogen. The heteroatoms are independently selected from oxygen, nitrogen, sulfur, phosphorus, silicon, selenium and tin, but are not limited to these atoms. In embodiments where two or more heteroatoms are present, the two or more heteroatoms may be the same as each other, or some or all of the two or more heteroatoms may be different from each other.

The terms "fused" or "fused ring" as used herein, alone or in combination, refer to a cyclic structure in which two or more rings share one or more bonds.

The term "spiro" or "spirocyclic" as used herein, alone or in combination, refers to a cyclic structure in which two or more rings share one or more atoms.

The term "alkyl" as used herein alone or as part of another component (e.g., monoalkylamino) refers to an optionally substituted straight or optionally substituted branched chain monovalent saturated hydrocarbon having 1 to 12 carbon atoms, preferably 1 to 8 carbon atoms, more preferably 1 to 6 carbon atoms, attached to the rest of the molecule by a single bond, e.g., methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, n-hexyl, n-heptyl, 2-methylhexyl, 3 methylhexyl, n-octyl, n-nonyl, n-decyl, and the like.

The term "cycloalkyl" as used herein alone or as part of another ingredient refers to a stable monovalent non-aromatic monocyclic or polycyclic hydrocarbon group containing only carbon and hydrogen atoms, and may include fused, spiro or bridged ring systems containing from 3 to 15 ring-forming carbon atoms, preferably from 3 to 10 ring-forming carbon atoms, more preferably from 3 to 8 ring-forming carbon atoms, which may or may not be saturated, attached to the rest of the molecule by single bonds. Non-limiting examples of "cycloalkyl" include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and the like.

The terms "heterocyclyl", "heterocycloalkyl", "heterocycle", as used herein alone or as part of another ingredient, refer to a stable 3-18 membered monovalent non-aromatic ring comprising 2-12 carbon atoms, 1-6 heteroatoms selected from nitrogen, oxygen and sulfur. Unless otherwise specified, a heterocyclyl group may be a monocyclic, bicyclic, tricyclic or tetracyclic ring system, which may contain fused, spiro or bridged ring systems, the nitrogen, carbon or sulfur of the heterocyclyl group may optionally be oxidized, the nitrogen atom may optionally be quaternized, and the heterocyclyl group may be partially or fully saturated. The heterocyclic group may be attached to the rest of the molecule through a single bond via a carbon or heteroatom in the ring. The heterocyclic group containing fused rings may contain one or more aromatic or heteroaromatic rings, provided that the atoms on the non-aromatic ring are attached to the rest of the molecule. For purposes of this application, a heterocyclyl group is preferably a stable 4-11 membered monovalent non-aromatic monocyclic or bicyclic ring containing 1-3 heteroatoms selected from nitrogen, oxygen and sulfur, and more preferably a stable 4-8 membered monovalent non-aromatic monocyclic ring containing 1-3 heteroatoms selected from nitrogen, oxygen and sulfur. Non-limiting examples of heterocyclyl groups include azepanyl, azetidinyl, decahydroisoquinolinyl, dihydrofuranyl, indolinyl, dioxolanyl, 1-dioxo-thiomorpholinyl, imidazolidinyl, imidazolinyl, isothiazolidinyl, isoxazolidinyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, oxazinyl, piperazinyl, piperidinyl, 4-piperidinonyl, pyranyl, pyrazolidinyl, pyrrolidinyl, quinolizinyl, quinuclidinyl, tetrahydrofuranyl, tetrahydropyranyl and the like.

The term "polymorph" or "polymorph" as used herein means that the compounds of the present invention have multiple lattice morphologies. Some of the compounds of the present invention may have more than one crystal form, and the present invention encompasses all polymorphic forms or mixtures thereof.

Intermediate compounds of the present invention and polymorphs thereof are also within the scope of the present invention.

Unless otherwise specified, the compounds of the present invention contain olefinic double bonds including E and Z isomers.

It is understood that the compounds of the present invention may contain asymmetric centers. These asymmetric centers may independently be in the R or S configuration. It will be apparent to those skilled in the art that some of the compounds of the present invention may also exhibit cis-trans isomerism. It is to be understood that the compounds of the present invention include their individual geometric and stereoisomers as well as mixtures thereof, including racemic mixtures. These isomers may be separated from their mixtures by carrying out or modifying known methods such as chromatographic techniques and recrystallization techniques, or they may be prepared separately from the appropriate isomers of their intermediates.

The term "pharmaceutically acceptable salts" as used herein includes both acid and base salts.

"pharmaceutically acceptable acid addition salts" refers to those salts formed with inorganic acids such as, but not limited to, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like, or organic acids such as, but not limited to, acetic acid, 2-dichloroacetic acid, adipic acid, alginic acid, ascorbic acid, aspartic acid, benzenesulfonic acid, benzoic acid, capric acid, caproic acid, carbonic acid, cinnamic acid, citric acid, and the like, which retain the biological potency and properties of the free base of the compound, which are not biologically or otherwise undesirable. "pharmaceutically acceptable salt to be added to base" refers to those salts that retain the biological potency and properties of the free acid of the compound and are not biologically or otherwise undesirable. These salts are prepared by reacting the free acid with an inorganic or organic base. Salts formed by reaction with an inorganic base include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, and the like. Preferred inorganic salts are ammonium, sodium, potassium, calcium, and manganese salts.

Salt-forming organic bases include, but are not limited to, primary, secondary, tertiary, cyclic amines, and the like, such as ammonia, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, ethanolamine, dicyclohexylamine, ethylenediamine, purine, piperazine, piperidine, choline, caffeine, and the like. Particularly preferred organic bases are isopropylamine, diethylamine, ethanolamine, trimethylamine, dicyclohexylamine, choline, and caffeine.

Crystallization often produces solvates of the compounds of the present invention. The term "solvate" as used herein refers to a combination of one or more molecules of the compound of the present invention and one or more molecules of a solvent.

The solvent may be water, in which case the solvate is a hydrate. In addition, an organic solvent may be used. Thus, the compounds of the present invention may exist as hydrates, including monohydrate, dihydrate, hemihydrate, trihydrate, tetrahydrate and the like, as well as the corresponding solvated forms. The compounds of the present invention may be true solvates, but in other cases, the compounds of the present invention may also retain water only by chance or a mixture of water and some other solvent. The compounds of the invention may be reacted in a solvent or precipitated or crystallized in a solvent. Solvates of the compounds of the invention are also included within the scope of the invention.

The term "pharmaceutical composition" as used herein refers to a formulation mixed with a compound of the present invention and a vehicle generally accepted in the art for delivering biologically active compounds to a mammal, such as a human. Such media comprise all pharmaceutically acceptable carriers.

As used herein, the term "acceptable" in reference to a formulation, composition or ingredient means that there is no lasting deleterious effect on the overall health of the subject being treated.

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

"pharmaceutically acceptable carriers" include, but are not limited to, adjuvants, carriers, excipients, adjuvants, deodorants, diluents, preservatives, dyes/colorants, flavor enhancers, surfactants and wetting agents, dispersants, suspending agents, stabilizers, isotonic agents, solvents, or emulsifiers that have been approved by the relevant governmental authorities for use in humans and domestic animals.

The terms "subject," "patient," "subject" or "individual" as used herein refer to an individual having a disease, disorder or condition, and the like, including mammals and non-mammals. Examples of mammals include, but are not limited to, any member of the class mammalia: humans, non-human primates (e.g., chimpanzees and other apes and monkeys); livestock, such as cattle, horses, sheep, goats, pigs; domestic animals such as rabbits, dogs, and cats; laboratory animals, including rodents, such as rats, mice, and guinea pigs, and the like. Examples of non-human mammals include, but are not limited to, birds, fish, and the like. In one embodiment related to the methods and compositions provided herein, the mammal is a human.

The term "treatment" as used herein refers to the treatment of a disease or condition associated with a mammal, particularly a human, and includes

(i) Preventing the development of a disease or condition in a mammal, particularly a mammal that has previously been exposed to the disease or condition but has not been diagnosed as having the disease or condition;

(ii) inhibiting the disease or disorder, i.e., controlling its development;

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

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

The terms "disease" and "condition" as used herein may be used interchangeably and may have different meanings, as certain specific diseases or conditions have no known causative agent (and therefore the cause of the disease is not yet clear) and therefore are not considered as a disease but can be considered as an unwanted condition or syndrome, with more or less specific symptoms being confirmed by clinical researchers.

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

The terms "administering," "administration," "administering," and the like as used herein refer to a method capable of delivering a compound or composition to a desired site for biological action. These methods include, but are not limited to, oral routes, via the duodenal route, parenteral injection (including intravenous, subcutaneous, intraperitoneal, intramuscular, intraarterial injection or infusion), topical administration, and rectal administration. In preferred embodiments, the compounds and compositions discussed herein are administered orally.

Preparation of the Compounds of the invention

The following reaction scheme illustrates the process for preparing the compounds of the present invention.

It will be appreciated that in the following description, combinations of substituents and/or variables of the formula are permitted only in the context of forming stable compounds.

It will also be appreciated by those skilled in the art that in the schemes described below, the functional groups of the intermediate compounds may need to be protected by suitable protecting groups. These functional groups include hydroxyl, amino, mercapto and carboxyl groups. Suitable hydroxyl protecting groups include trialkylsilyl or diarylalkylsilyl groups (e.g., tert-butylmethylsilyl, tert-butyldiphenylsilyl or trimethylsilyl), tetrahydropyranyl, benzyl, and the like. Suitable amino, amidino and guanidine protecting groups include t-butyloxycarbonyl, benzyloxycarbonyl and the like. Suitable protecting groups for the thiol group include-C (O) -R '(R' represents an alkyl, aryl or arylalkyl group), p-methoxybenzyl, trityl, and the like. Suitable carboxyl protecting groups include alkyl, aryl or arylalkyl esters. Protecting groups may be added or removed by standard techniques known to those skilled in the art.

Examples

The following non-limiting examples are illustrative only and do not limit the invention in any way.

Unless otherwise indicated, temperatures are in degrees celsius. Reagents were purchased from commercial suppliers such as national drug group chemical reagents beijing ltd, Alfa Aesar (Alfa Aesar), or beijing carbofuran technologies ltd, and these reagents were used directly without further purification unless otherwise specified.

Unless otherwise stated, the following reactions are carried out at room temperature, in anhydrous solvents, under positive pressure of nitrogen or argon, or using a drying tube; the reaction bottle is provided with a rubber diaphragm so as to add the substrate and the reagent through an injector; glassware was dried and/or heat dried.

Unless otherwise stated, column chromatography purification was performed using 200-300 mesh silica gel from Qingdao oceanic plants; preparation of thin-layer chromatography silica gel precast slab (HSGF254) produced by Nicoti chemical industry research institute was used; MS is measured by a THERMOL LCQ FREet model (ESI) liquid chromatography-mass spectrometer; the optical rotation was measured by using an SGW-3 automatic polarimeter, Shanghai Spanish Meter, Ltd.

Nuclear magnetic data (¹H NMR) was run at 400MHz using a Varian instrument. The solvent used for nuclear magnetic data is CDCl₃、CD₃OD、D₂O, DMSO-d6, based on tetramethylsilane (0.00ppm) or based on residual solvent (CDCl)₃：7.26ppm；CD₃OD：3.31ppm；D₂O: 4.79 ppm; d 6-DMSO: 2.50 ppm). When indicating the diversity of the peak shapes, the following abbreviations represent the different peak shapes: s (singlet), d (doublet), t (triplet), q (quartet), m (multiplet), br (broad), dd (doublet of doublets), dt (doublet of triplets). If the coupling constant is given, it is given in Hertz (Hz).

Abbreviations:

Pd₂(dba)₃	tris (dibenzylideneacetone) dipalladium
		(±)BINAP	(±) -2, 2 '-bis (diphenylphosphino) -1, 1' -binaphthalene
Pd(dppf)Cl₂	1, 1' -bis (diphenylphosphino) ferrocene palladium dichloride
		TEA	Triethylamine
TFA	Trifluoroacetic acid
		NaBH(OAc)₃	Sodium triacetoxyborohydride
DMSO	Dimethyl sulfoxide
		KOAc	Potassium acetate
Toluene	Toluene
		K₂CO₃	Potassium carbonate
Pd(PPH₃)₄	Tetratriphenylphosphine palladium
		Cs₂CO₃	Cesium carbonate

Intermediate 21: synthesis of 4- (6- (3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) -6- (1-difluoromethyl-1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Step 1: 3- (5- (3-cyano-6- (trifluoromethanesulfonate) pyrazolo [1, 5-a ] pyridin-4-yl) pyridin-2-yl) -3, 6-diazabicyclo [3.1.1] heptane-6-carboxylic acid tert-butyl ester

Tert-butyl 3- (5- (3-cyano-6-hydroxypyrazolo [1, 5-a ] pyridin-4-yl) pyridin-2-yl) -3, 6-diazabicyclo [3.1.1] heptane-6-carboxylate (432mg) was dissolved in DMF (20mL), to which DIEA (260mg) was added, followed by N-phenylbis (trifluoromethanesulfonyl) imide (400 mg). After stirring at room temperature for 2 hours, the mixture was poured into water (100mL), and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure, and the residue was purified by column chromatography (eluent: petroleum ether: ethyl acetate ═ 2: 1 (V: V)) to give the objective compound. (450mg)

Step 2: 3- (5- (3-cyano-6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-yl) pyridin-2-yl) -3, 6-diazabicyclo [3.1.1] heptane-6-carboxylic acid tert-butyl ester

3- (5- (3-cyano-6- (trifluoromethanesulfonate) pyrazolo [1, 5-a ] pyridin-4-yl) pyridin-2-yl) -3, 6-diazabicyclo [3.1.1] heptane-6-carboxylic acid tert-butyl ester (282mg), 1-difluoromethylpyrazole-4-boronic acid (100mg), tetratriphenylphosphine palladium (55mg) and potassium carbonate (139mg) were successively added to a flask containing dioxane and water (50mL, volume ratio 5: 1), and the reaction flask was replaced with nitrogen three times. Heat to 80 ℃ and stir overnight. After cooling to room temperature, the reaction mixture was poured into water (150mL), followed by extraction with ethyl acetate. The organic phase was dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by column chromatography (eluent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the objective compound (200 mg).

And step 3: 4- (6- (3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) -6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Tert-butyl 3- (5- (3-cyano-6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-yl) pyridin-2-yl) -3, 6-diazabicyclo [3.1.1] heptane-6-carboxylate (200mg) was added at room temperature.

Dissolved in methylene chloride (20mL), trifluoroacetic acid (4mL) was added, the mixture was stirred for 2 hours, and the solvent was evaporated under reduced pressure in vacuo. Saturated aqueous sodium bicarbonate was added to the residue until the system was weakly basic, and the resulting solid of the objective compound (130mg) was filtered and dried.

Intermediate 22: synthesis of 4- (6- (3, 8-diazabicyclo [3.2.1] octan-3-yl) pyridin-3-yl) -6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Step 1: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4-hydroxypyrazolo [1, 5-a ] pyridine-3-carbonitrile

6-bromo-4-hydroxypyrazolo [1, 5-a ] pyridine-3-carbonitrile (238mg), 1-difluoromethylpyrazole-4-boronic acid (200mg), palladium tetratriphenylphosphine (110mg) and potassium carbonate (278mg) were successively added to a flask containing dioxane and water (50mL, volume ratio 5: 1), and the reaction flask was replaced with nitrogen gas three times. Heat to 80 ℃ and stir overnight. After cooling to room temperature, the reaction mixture was poured into water (150mL), followed by extraction with ethyl acetate. The organic phase was dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by column chromatography (eluent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the objective compound (250 mg).

Step 2: 3-cyano-6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-yl trifluoromethanesulfonate

6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4-hydroxypyrazolo [1, 5-a ] pyridine-3-carbonitrile (250mg) was dissolved in DMF (20mL), to which DIEA (300mg) was added, followed by N-phenylbis (trifluoromethanesulfonyl) imide (400 mg). After stirring at room temperature for 2 hours, the mixture was poured into water (50mL), and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure, and the residue was purified by column chromatography (eluent: petroleum ether: ethyl acetate 1: 1 (V: V)) to obtain the objective compound. (300mg)

And step 3: 3-cyano-6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-ylboronic acid

3-cyano-6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-yl trifluoromethanesulfonate (300mg), pinacol diboron (254mg), [1, 1' -bis (diphenylphosphino) ferrocene ] dichloropalladium (73mg) and potassium acetate (196mg) were added to dried dioxane (30mL) under nitrogen, heated to 90 ℃ and stirred overnight. Cooling to room temperature, filtering, washing a filter cake with dichloromethane, and evaporating a filtrate under reduced pressure to obtain a crude target compound which is directly used for the next reaction.

And 4, step 4: 3- (5- (3-cyano-6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-yl) pyridin-2-yl) -3, 6-diazabicyclo [3.2.1] octane-6-carboxylic acid tert-butyl ester

The boric acid (150mg) obtained above, tert-butyl 3- (5-bromopyridin-2-yl) -3, 8-diazabicyclo [3.2.1] octane-8-carboxylate (183mg), palladium tetratriphenylphosphine (50mg) and potassium carbonate (139mg) were sequentially added to a flask containing dioxane and water (50mL, volume ratio 5: 1), and the reaction flask was replaced with nitrogen gas three times. Heat to 80 ℃ and stir overnight. After cooling to room temperature, the reaction mixture was poured into water (150mL), followed by extraction with ethyl acetate. The organic phase was dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the objective compound (250 mg).

And 5: 4- (6- (3, 8-diazabicyclo [3.2.1] octan-3-yl) pyridin-3-yl) -6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Tert-butyl 3- (5- (3-cyano-6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-yl) pyridin-2-yl) -3, 6-diazabicyclo [3.2.1] octane-6-carboxylate (250mg) was added at room temperature

Dissolved in methylene chloride (20mL), trifluoroacetic acid (5mL) was added, the mixture was stirred for 1 hour, and the solvent was evaporated under reduced pressure in vacuo. To the residue was added a saturated aqueous sodium hydrogencarbonate solution until the system became weakly basic, and the resulting solid of the objective compound (180mg) was filtered and dried.

Intermediate 23: synthesis of 4- (5- (3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyrazin-2-yl) -6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Step 1: 3- (5- (3-cyano-6- (1-difluoromethyl-1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-yl) pyrazin-2-yl) -3, 6-diazabicyclo [3.1.1] heptane-6-carboxylic acid tert-butyl ester

Reacting 3-cyano-6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-ylboronic acid

(303mg) tert-butyl 3- (5-chloropyrazin-2-yl) -3, 6-diazabicyclo [3.1.1] heptane-6-carboxylate (250mg) from intermediate 4, step 2 was dissolved in a mixed solution of dioxane and water (5: 1) (50mL), and tetratriphenylphosphine palladium (110mg) and anhydrous potassium carbonate powder (278mg) were added in this order under nitrogen. Then, the reaction mixture was stirred at 85 ℃ for 2 hours, cooled to room temperature, poured into water (150mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether: ethyl acetate 2: 1 (V: V)) to give the title compound (400 mg).

Step 2: 4- (5- (3, 6-diaza-bicyclo [3.1.1] heptan-3-yl) pyrazin-2-yl) -6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Trifluoroacetic acid (5mL) was slowly added dropwise to a dichloromethane (20mL) solution of tert-butyl 3- (5- (3-cyano-6- (1-difluoromethyl-1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-yl) pyrazin-2-yl) -3, 6-diazabicyclo [3.1.1] heptane-6-carboxylate (400mg) obtained in step 1 at 0 ℃, and after completion of the addition, the mixture was warmed to room temperature and stirred for 1H. The reaction solution was concentrated, and then saturated aqueous sodium bicarbonate solution was added to adjust the pH to a weak alkaline, and the resulting solid was filtered and dried to obtain the title compound (240 mg).

Intermediate 24: synthesis of 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (piperazin-1-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Step 1: 4- (5- (3-cyano-6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-yl) pyridin-2-yl) piperazine-1-carboxylic acid tert-butyl ester

3-cyano-6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-ylboronic acid (303mg) and tert-butyl 4- (5-bromopyridin-2-yl) piperazine-1-carboxylate (342mg) were dissolved in a mixed solution of dioxane and water (5: 1) (50mL), and tetratriphenylphosphine palladium (110mg) and anhydrous potassium carbonate powder (278mg) were sequentially added under nitrogen protection. Then, the reaction mixture was stirred at 85 ℃ for 2 hours, cooled to room temperature, poured into water (150mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether: ethyl acetate 2: 1 (V: V)) to give the title compound (410 mg).

Step 2: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (piperazin-1-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Trifluoroacetic acid (5mL) was slowly added dropwise to a solution of tert-butyl 4- (5- (3-cyano-6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-yl) pyridin-2-yl) piperazine-1-carboxylate (410mg) obtained in step 1 in dichloromethane (20mL) at 0 ℃, and after completion of the addition, the mixture was warmed to room temperature and stirred for 1H. The reaction solution was concentrated, and then saturated aqueous sodium bicarbonate solution was added to adjust the pH to a weak alkaline, and the resulting solid was filtered and dried to obtain the title compound (220 mg).

Intermediate 25: synthesis of 4- (6- (3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) -6- (1-trifluoromethyl-1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Step 1: 3- (5- (3-cyano-6- (1- (trifluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-yl) pyridin-2-yl) -3, 6-diazabicyclo [3.1.1] heptane-6-carboxylic acid tert-butyl ester

3- (5- (3-cyano-6- (trifluoromethanesulfonate) pyrazolo [1, 5-a ] pyridin-4-yl) pyridin-2-yl) -3, 6-diazabicyclo [3.1.1] heptane-6-carboxylic acid tert-butyl ester (282mg), 1-trifluoromethylpyrazole-4-boronic acid (110mg), tetratriphenylphosphine palladium (50mg) and potassium carbonate (139mg) were sequentially added to a flask containing dioxane and water (40mL, volume ratio 5: 1), and the reaction flask was replaced with nitrogen three times. Heat to 80 ℃ and stir overnight. After cooling to room temperature, the reaction mixture was poured into water (150mL), followed by extraction with ethyl acetate. The organic phase was dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the objective compound (170 mg).

Step 2: synthesis of 4- (6- (3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) -6- (1-trifluoromethyl-1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Tert-butyl 3- (5- (3-cyano-6- (1- (trifluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridin-4-yl) pyridin-2-yl) -3, 6-diazabicyclo [3.1.1] heptane-6-carboxylate (170mg) was added at room temperature

Dissolved in methylene chloride (20mL), trifluoroacetic acid (4mL) was added, the mixture was stirred for 2 hours, and the solvent was evaporated under reduced pressure in vacuo. Saturated aqueous sodium bicarbonate was added to the residue until the system was weakly basic, and the resulting solid of the objective compound (120mg) was filtered and dried.

Example 59: 6- (1-methyl-1H-pyrazol-4-yl) -4- (6- (6- (4- (methylthio) benzyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 2(40mg) was dissolved in THF (10mL) at 0 deg.C, and 4-methylthiobenzaldehyde (31mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extracts were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (30 mg).

¹H NMR(400MHz，CDCl₃)δ8.66(1H，d，J＝1.2Hz)，8.45(1H，d，J＝2.4Hz)，8.27(1H，s)，7.85(1H，dd，J＝8.4Hz，2.4Hz)，7.80(1H，s)，7.70(1H，s)，7.50(2H，d，J＝8.0Hz)，7.43(1H，d，J＝1.2Hz)，7.24(2H，d，J＝8.0Hz)，6.73(1H，d，J＝8.4Hz)，4.18-4.32(2H，m)，3.80-4.09(9H，m)，3.16-3.30(1H，m)，2.47(3H，s)，1.82-1.89(1H，m)。

Example 60: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (6- ((6- (methylthio) pyridin-3-yl) methyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 21(43mg) was dissolved in THF (10mL) at 0 deg.C, and 6- (methylthio) nicotinaldehyde (31mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (27 mg).

¹H NMR(400MHz，CDCl₃)δ8.72(1H，d，J＝1.6Hz)，8.44(1H，d，J＝2.4Hz)，8.41(1H，d，J＝2.0Hz)，8.31(1H，s)，8.14(1H，s)，7.97(1H，s)，7.82(1H，dd，J＝8.8Hz，2.4Hz)，7.64-7.70(1H，m)，7.43(1H，d，J＝1.6Hz)，7.25(1H，t，J＝60.4Hz)，7.16(1H，d，J＝8.4Hz)，6.71(1H，d，J＝8.8Hz)，3.84-3.94(4H，m)，3.62-3.73(4H，m)，2.78-2.86(1H，m)，2.55(3H，s)，1.71(1H，d，J＝8.4Hz)。

Example 61: (E) -4- (6- (6- (4-methoxybut-2-enoyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) -6- (1-methyl-1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 2(40mg) was dissolved in THF (10mL) at 0 deg.C, and (E) -4-methoxybut-2-enoic acid (12mg) and triethylamine (30mg) were added, followed by warming to room temperature and stirring for 1 hour. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (37 mg).

¹H NMR(400MHz，CDCl3)δ8.63(1H，d，J＝1.2Hz)，8.37(1H，d，J＝2.4Hz)，8.25(1H，s)，7.78(1H，s)，7.76(1H，dd，J＝8.8Hz，2.4Hz)，7.68(1H，s)，7.38(1H，d，J＝1.2Hz)，6.88(1H，dt，J＝15.6Hz，4.0Hz)，6.65(1H，d，J＝8.8Hz)，6.16(1H，dt，J＝15.6Hz，2.0Hz)，4.64-4.72(2H，m)，4.17(1H，d，J＝12.0Hz)，4.06-4.09(2H，m)，3.98(3H，s)，3.88-3.91(2H，m)，3.64-3.71(1H，m)，3.39(3H，s)，2.78-2.84(1H，m)，1.73(1H，d，J＝8.8Hz)。

Example 62: (E) -6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (6- (4-methoxybut-2-enoyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 21(43mg) was dissolved in THF (10mL) at 0 deg.C, and (E) -4-methoxybut-2-enoic acid (12mg) and triethylamine (30mg) were added, followed by warming to room temperature and stirring for 1 hour. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (39 mg).

¹H NMR(400MHz，CDCl₃)δ8.70(1H，d，J＝1.2Hz)，8.38(1H，d，J＝2.4Hz)，8.29(1H，s)，8.13(1H，s)，7.96(1H，s)，7.76(1H，dd，J＝8.8Hz，2.4Hz)，7.39(1H，d，J＝1.6Hz)，7.25(1H，t，J＝60.4Hz)，6.89(1H，dt，J＝15.2Hz，4.0Hz)，6.64(1H，d，J＝8.8Hz)，6.16(1H，dt，J＝15.2Hz，2.0Hz)，4.64-4.72(2H，m)，4.18(1H，d，J＝10.8Hz)，4.06-4.09(2H，m)，3.88-3.92(2H，m)，3.67(1H，d，J＝10.8Hz)，3.39(3H，s)，2.78-2.84(1H，m)，1.74(1H，d，J＝8.8Hz)。

Example 63: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (4- ((6- (methylsulfonyl) pyridin-3-yl) methyl) piperazin-1-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 24(42mg) was dissolved in THF (10mL) at 0 deg.C, and 6- (methylsulfonyl) nicotinaldehyde (37mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (25 mg).

¹H NMR(400MHz，CDCl3)δ8.72(1H，s)，8.70(1H，d，J＝1.6Hz)，8.36(1H，d，J＝2.4Hz)，8.29(1H，s)，8.13(1H，s)，8.07(1H，s)，8.02(1H，d，J＝7.2Hz)，7.96(1H，s)，7.75(1H，dd，J＝9.2Hz，2.4Hz)，7.40(1H，d，J＝1.6Hz)，7.25(1H，t，J＝60.4Hz)，6.78(1H，d，J＝8.8Hz)，3.64-3.74(6H，m)，3.25(3H，s)，2.58-2.64(4H，m)。

Example 64: 6- (3, 3-Difluoroazetidin-1-yl) -4- (6- (6- (3- (methylsulfonyl) benzyl) -3, 6-diazabicyclo [3.1.1] Heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 9(64mg), 3-difluorotrimethylene imine hydrochloride (26mg), tris (dibenzylideneacetone) dipalladium (9mg), (±) -2, 2 '-bis (diphenylphosphino) -1, 1' -binaphthyl (12mg), and cesium carbonate (98mg) were successively added to a flask containing toluene (20mL), and the reaction flask was replaced with nitrogen three times. Heat to 110 ℃ and stir overnight. Cooled to room temperature, filtered, the filter cake was washed with dichloromethane, the filtrates were combined and concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (29 mg).

¹H NMR(400MHz，CDCl₃)δ8.38(1H，d，J＝2.4Hz)，8.19(1H，s)，7.98(1H，s)，7.82(1H，d，J＝8.0Hz)，7.77-7.80(2H，m)，7.69(1H，d，J＝6.8Hz)，7.52(1H，t，J＝8.0Hz)，6.75(1H，d，J＝2.0Hz)，6.69(1H，d，J＝9.2Hz)，4.31(4H，t，J＝11.6Hz)，3.78-3.88(4H，m)，3.72(2H，s)，3.58-3.68(2H，m)，3.06(3H，s)，2.73-2.82(1H，m)，1.69(1H，d，J＝8.4Hz)。

Example 65: 4- (6- (6- (3- (methylsulfonyl) benzyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) -6- (3-hydroxy-3-methylazepin-1-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 9(64mg), 3-methyl-3-acridinium alkoxide (27mg), tris (dibenzylideneacetone) dipalladium (9mg), (±) -2, 2 '-bis (diphenylphosphino) -1, 1' -binaphthyl (12mg), and cesium carbonate (98mg) were successively added to a flask containing toluene (20mL), and the reaction flask was replaced with nitrogen three times. Heat to 110 ℃ and stir overnight. Cooled to room temperature, filtered, the filter cake was washed with dichloromethane, the filtrates were combined and concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (21 mg).

¹H NMR(400MHz，CDCl₃)δ8.37(1H，d，J＝2.4Hz)，8.15(1H，s)，7.98(1H，s)，7.81(1H，d，J＝7.6Hz)，7.78(1H，dd，J＝8.8Hz，2.4Hz)，7.71(1H，d，J＝2.0Hz)，7.67(1H，d，J＝8.0Hz)，7.51(1H，t，J＝8.0Hz)，6.74(1H，d，J＝2.0Hz)，6.68(1H，d，J＝8.8Hz)，3.91(2H，d，J＝8.0Hz)，3.77-3.87(6H，m)，3.73(2H，s)，3.58-3.66(2H，m)，3.06(3H，s)，2.72-2.79(1H，m)，1.67(1H，d，J＝9.2Hz)，1.66(3H，s)。

Example 66: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (6- ((5- (methylthio) pyridin-2-yl) methyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 21(43mg) was dissolved in THF (10mL) at 0 deg.C, and 5- (methylthio) pyridine-2-carbaldehyde (31mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (31 mg).

¹H NMR(400MHz，CDCl₃)δ8.74(1H，s)，8.45(1H，d，J＝2.4Hz)，8.43(1H，s)，8.32(1H，s)，8.15(1H，s)，7.98(1H，s)，7.86(1H，dd，J＝8.8Hz，2.4Hz)，7.76(1H，d，J＝8.0Hz)，7.43(1H，s)，7.35(1H，d，J＝8.0Hz)，7.26(1H，t，J＝60.4Hz)，6.75(1H，d，J＝8.8Hz)，4.38(2H，d，J＝5.6Hz)，3.99(2H，s)，3.89(4H，s)，3.24-3.32(1H，m)，2.56(3H，s)，1.86(1H，d，J＝10.4Hz)。

Example 67: 6- (3, 3-Difluoroazetidin-1-yl) -4- (6- (6- (2- (methylsulfonyl) benzyl) -3, 6-diazabicyclo [3.1.1] Heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 20(64mg), 3-difluorotrimethylene imine hydrochloride (26mg), tris (dibenzylideneacetone) dipalladium (9mg), (±) -2, 2 '-bis (diphenylphosphino) -1, 1' -binaphthyl (12mg), and cesium carbonate (98mg) were successively added to a flask containing toluene (20mL), and the reaction flask was replaced with nitrogen three times. Heat to 110 ℃ and stir overnight. Cooled to room temperature, filtered, the filter cake was washed with dichloromethane, the filtrates were combined and concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (19 mg).

¹H NMR(400MHz，CDCl3)δ8.39(1H，d，J＝2.0Hz)，8.18(1H，s)，8.09(1H，d，J＝8.0Hz)，7.76-7.79(2H，m)，7.43-7.60(3H，m)，6.75(1H，d，J＝2.0Hz)，6.72(1H，d，J＝8.8Hz)，4.31(4H，t，J＝11.6Hz)，4.06-4.19(2H，m)，3.90-3.99(2H，m)，3.76-3.88(2H，m)，3.59-3.71(2H，m)，3.34(3H，s)，2.52-2.72(1H，m)，1.71(1H，d，J＝9.2Hz)。

Example 68: 4- (6- (6- (4- (methylsulfinyl) benzyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) -6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 21(43mg) was dissolved in THF (10mL) at 0 deg.C, and 4-methylsulfinylbenzaldehyde (34mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (32 mg).

¹H NMR(400MHz，CDCl₃)δ8.72(1H，s)，8.44(1H，d，J＝2.4Hz)，8.31(1H，s)，8.14(1H，s)，7.97(1H，s)，7.82(1H，dd，J＝8.8Hz，2.4Hz)，7.58-7.64(4H，m)，7.43(1H，d，J＝1.2Hz)，7.25(1H，t，J＝60.4Hz)，6.72(1H，d，J＝8.4Hz)，3.83-3.98(4H，m)，3.79(2H，s)，3.63-3.73(2H，m)，2.82-2.92(1H，m)，2.72(3H，s)，1.73(1H，d，J＝9.2Hz)。

Example 69: 6- (1-difluoromethyl-1H-pyrazol-4-yl) -4- (6- (8- (4- (methylsulfonyl) benzyl) -3, 8-diazabicyclo [3.2.1] oct-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 22(45mg) was dissolved in THF (15mL) at 0 deg.C, 4-methylsulfonylbenzaldehyde (37mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extracts were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 15: 1 (V: V)) to give the title compound (30 mg).

¹H NMR(400MHz，CDCl₃)δ8.70(1H，d，J＝1.2Hz)，8.36(1H，d，J＝2.8Hz)，8.29(1H，s)，8.12(1H，s)，7.96(1H，s)，7.92(2H，d，J＝8.0Hz)，7.74(1H，dd，J＝8.8Hz，2.8Hz)，7.67(2H，d，J＝8.0Hz)，7.40(1H，s)，7.25(1H，t，J＝60.4Hz)，6.68(1H，d，J＝8.8Hz)，3.91(2H，d，J＝11.2Hz)，3.74(2H，s)，3.35(2H，s)，3.26(2H，d，J＝11.2Hz)，3.07(3H，s)，2.04-2.10(2H，m)，1.78-1.85(2H，m)。

Example 70: 4- (5- (6- (4- (methylsulfonyl) benzyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyrazin-2-yl) -6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 23(43mg) was dissolved in THF (15mL) at 0 deg.C, 4-methylsulfonylbenzaldehyde (37mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 15: 1 (V: V)) to give the title compound (17 mg).

¹H NMR(400MHz，CDCl₃)δ8.74(1H，d，J＝1.6Hz)，8.57(1H，d，J＝1.6Hz)，8.35(1H，s)，8.28(1H，d，J＝1.6Hz)，8.16(1H，s)，7.99(1H，s)，7.90(2H，d，J＝8.8Hz)，7.67(1H，d，J＝1.2Hz)，7.60(2H，d，J＝8.0Hz)，7.26(1H，t，J＝60.4Hz)，3.82-3.90(4H，m)，3.76(2H，s)，3.68(2H，d，J＝12.8Hz)，3.05(3H，s)，2.77-2.84(1H，m)，1.71(1H，d，J＝9.2Hz)。

Example 71: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (6- ((6- (methylthio) pyridin-3-yl) methyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 21(43mg) was dissolved in THF (10mL) at 0 deg.C, and 6- (methylthio) pyridine-3-carbaldehyde (31mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (32 mg).

¹H NMR(400MHz，CDCl₃)δ8.68(1H，d，J＝1.6Hz)，8.31(1H，d，J＝2.8Hz)，8.28(1H，s)，8.11(1H，s)，7.94(1H，s)，7.89(1H，d，J＝2.8Hz)，7.71(1H，dd，J＝8.8Hz，2.8Hz)，7.37(1H，s)，7.23(1H，t，J＝60.4Hz)，7.03(1H，d，J＝8.4Hz)，6.81(1H，dd，J＝8.8Hz，3.2Hz)，6.56(1H，d，J＝8.8Hz)，4.49(2H，d，J＝6.0Hz)，4.06(2H，d，J＝12.0Hz)，3.55-3.63(2H，m)，2.88-2.94(1H，m)，2.49(3H，s)，1.82(1H，d，J＝8.8Hz)。

Example 72: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (6- ((2- (methylthio) pyridin-4-yl) methyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 21(43mg) was dissolved in THF (10mL) at 0 deg.C, and 2- (methylthio) pyridine-4-carbaldehyde (31mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (33 mg).

¹H NMR(400MHz，CDCl₃)δ8.71(1H，d，J＝1.6Hz)，8.43(1H，d，J＝2.8Hz)，8.35(1H，d，J＝4.8Hz)，8.31(1H，s)，8.13(1H，s)，7.97(1H，s)，7.81(1H，dd，J＝8.8Hz，2.4Hz)，7.43(1H，d，J＝1.6Hz)，7.25(1H，t，J＝60.4Hz)，7.23(1H，s)，7.01(1H，d，J＝4.0Hz)，6.70(1H，d，J＝9.2Hz)，3.77-3.84(4H，m)，3.56-3.66(4H，m)，2.73-2.82(1H，m)，2.56(3H，s)，1.68(1H，d，J＝8.8Hz)。

Example 73: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (8- ((6- (methylthio) pyridin-3-yl) methyl) -3, 8-diazabicyclo [3.2.1] oct-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 22(45mg) was dissolved in THF (15mL) at 0 deg.C, and 6- (methylthio) pyridine-3-carbaldehyde (31mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 15: 1 (V: V)) to give the title compound (14 mg).

¹H NMR(400MHz，CDCl₃)δ8.69(1H，d，J＝1.6Hz)，8.41(1H，d，J＝2.0Hz)，8.35(1H，d，J＝2.4Hz)，8.29(1H，s)，8.12(1H，s)，7.95(1H，s)，7.72(1H，dd，J＝8.8Hz，2.4Hz)，7.64(1H，dd，J＝8.4Hz，2.0Hz)，7.39(1H，d，J＝1.6Hz)，7.25(1H，t，J＝60.4Hz)，7.18(1H，d，J＝8.4Hz)，6.66(1H，d，J＝9.2Hz)，3.88(2H，d，J＝10.4Hz)，3.55(2H，s)，3.28-3.34(2H，m)，3.18(2H，d，J＝11.2Hz)，2.58(3H，s)，1.98-2.08(2H，m)，1.75-1.81(2H，m)。

Example 74: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (6- ((6- (methylthio) pyridin-2-yl) methyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 21(43mg) was dissolved in THF (10mL) at 0 deg.C, and 6- (methylthio) pyridine-2-carbaldehyde (31mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (20 mg).

¹H NMR(400MHz，CDCl₃)δ8.71(1H，d，J＝1.6Hz)，8.44(1H，d，J＝2.4Hz)，8.31(1H，s)，8.14(1H，s)，7.97(1H，s)，7.82(1H，dd，J＝8.8Hz，2.4Hz)，7.47(1H，t，J＝8.0Hz)，7.42(1H，d，J＝1.6Hz)，7.25(1H，t，J＝60.4Hz)，7.12-7.17(1H，m)，7.06(1H，d，J＝7.2Hz)，6.73(1H，d，J＝8.4Hz)，3.53-4.03(8H，m)，2.73-2.93(1H，m)，2.54(3H，s)，1.73(1H，d，J＝8.8Hz)。

Example 75: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (4- ((6- (methylthio) pyridin-3-yl) methyl) piperazin-1-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 24(42mg) was dissolved in THF (10mL) at 0 deg.C, and 6- (methylthio) pyridine-3-carbaldehyde (31mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (36 mg).

¹H NMR(400MHz，CDCl₃)δ8.69(1H，d，J＝1.6Hz)，8.39(1H，d，J＝1.6Hz)，8.36(1H，d，J＝2.8Hz)，8.29(1H，s)，8.12(1H，s)，7.96(1H，s)，7.73(1H，dd，J＝8.8Hz，2.8Hz)，7.54(1H，dd，J＝8.4Hz，2.0Hz)，7.39(1H，d，J＝1.6Hz)，7.25(1H，t，J＝60.4Hz)，7.17(1H，d，J＝8.4Hz)，6.76(1H，d，J＝8.8Hz)，3.64-3.66(4H，m)，3.51(2H，s)，2.54-2.58(7H，m)。

Example 76: 6- (1- (methyl) -1H-pyrazol-4-yl) -4- (6- (6- ((6- (methylsulfonyl) pyridin-3-yl) methyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 2(42mg) was dissolved in THF (10mL) at 0 deg.C, and 6- (methylsulfonyl) nicotinaldehyde (37mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (21 mg).

¹H NMR(400MHz，CDCl₃)δ8.73(1H，s)，8.65(1H，d，J＝1.6Hz)，8.44(1H，d，J＝2.4Hz)，8.28(1H，s)，8.01-8.09(2H，m)，7.82(1H，dd，J＝8.8Hz，2.4Hz)，7.79(1H，s)，7.69(1H，s)，7.42(1H，d，J＝1.6Hz)，6.71(1H，d，J＝8.8Hz)，3.99(3H，s)，3.77-3.84(6H，m)，3.62-3.70(2H，m)，3.23(3H，s)，2.73-2.82(1H，m)，1.71(1H，d，J＝8.4Hz)。

Example 77: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (6- ((6- (methylsulfonyl) pyridin-3-yl) methyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 21(43mg) was dissolved in THF (10mL) at 0 deg.C, and 6- (methylsulfonyl) nicotinaldehyde (37mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (23 mg).

¹H NMR(400MHz，CDCl₃)δ8.73(1H，s)，8.72(1H，d，J＝1.6Hz)，8.44(1H，d，J＝2.4Hz)，8.31(1H，s)，8.14(1H，s)，8.04(1H，d，J＝7.6Hz)，8.01(1H，dd，J＝7.6Hz，1.6Hz)，7.97(1H，s)，7.82(1H，dd，J＝9.2Hz，2.4Hz)，7.43(1H，d，J＝1.6Hz)，7.26(1H，t，J＝60.4Hz)，6.71(1H，d，J＝8.4Hz)，3.78-3.82(4H，m)，3.76(2H，s)，3.60-3.69(2H，m)，3.23(3H，s)，2.71-2.79(1H，m)，1.70(1H，d，J＝8.8Hz)。

Example 78: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (6- ((5- (methylsulfonyl) pyridin-2-yl) methyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 21(43mg) was dissolved in THF (10mL) at 0 deg.C, and 5- (methylsulfonyl) pyridine-2-carbaldehyde (37mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (26 mg).

¹H NMR(400MHz，CDCl₃)δ9.05(1H，d，J＝2.0Hz)，8.71(1H，d，J＝1.6Hz)，8.43(1H，d，J＝2.4Hz)，8.30(1H，s)，8.18(1H，dd，J＝8.0Hz，2.4Hz)，8.14(1H，s)，7.97(1H，s)，7.80(1H，dd，J＝8.8Hz，2.4Hz)，7.68(1H，d，J＝8.0Hz)，7.42(1H，d，J＝1.6Hz)，7.26(1H，t，J＝60.4Hz)，6.72(1H，d，J＝8.8Hz)，3.86-3.94(6H，m)，3.61-3.71(2H，m)，3.10(3H，s)，2.76-2.84(1H，m)，1.72(1H，d，J＝8.4Hz)。

Example 79: 6- (1- (methyl) -1H-pyrazol-4-yl) -4- (6- (6- ((5- (methylsulfonyl) pyridin-2-yl) methyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 2(42mg) was dissolved in THF (10mL) at 0 deg.C, and 5- (methylsulfonyl) pyridine-2-carbaldehyde (37mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (28 mg).

¹H NMR(400MHz，DMSO-d6)δ9.22(1H，s)，8.94(1H，s)，8.64(1H，s)，8.43(1H，s)，8.38(1H，s)，8.28(1H，d，J＝7.6Hz)，8.33(1H，s)，7.87(1H，d，J＝8.0Hz)，7.78(1H，s)，7.75(1H，d，J＝8.4Hz)，6.80(1H，d，J＝8.4Hz)，3.86(3H，s)，3.72-3.84(6H，m)，3.48-3.64(2H，m)，3.29(3H，s)，2.42-2.52(1H，m)，1.60-1.70(1H，m)。

Example 80: 6- (1- (difluoromethyl) -1H-pyrazol-4-yl) -4- (6- (4- ((5- (methylsulfonyl) pyridin-2-yl) methyl) piperazin-1-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 24(42mg) was dissolved in THF (10mL) at 0 deg.C, and 5- (methylsulfonyl) pyridine-2-carbaldehyde (37mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (25 mg).

¹H NMR(400MHz，CDCl₃)δ9.01(1H，d，J＝2.4Hz)，8.70(1H，d，J＝1.6Hz)，8.37(1H，d，J＝2.8Hz)，8.29(1H，s)，8.22(1H，dd，J＝8.4Hz，2.0Hz)，8.13(1H，s)，7.96(1H，s)，7.73-7.76(2H，m)，7.40(1H，d，J＝1.6Hz)，7.25(1H，t，J＝60.4Hz)，6.79(1H，d，J＝9.2Hz)，3.84(2H，s)，3.68-3.75(4H，m)，3.13(3H，s)，2.64-2.70(4H，m)。

Example 81: 4- (6- (4- (4- (methylthio) benzyl) piperazin-1-yl) pyridin-3-yl) -6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 24(42mg) was dissolved in THF (10mL) at 0 deg.C, and 4-methylthiobenzaldehyde (31mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (35 mg).

¹H NMR(400MHz，CDCl₃)δ8.69(1H，d，J＝1.6Hz)，8.36(1H，d，J＝2.8Hz)，8.29(1H，s)，8.12(1H，s)，7.96(1H，s)，7.73(1H，dd，J＝8.8Hz，2.8Hz)，7.39(1H，d，J＝1.2Hz)，7.27-7.31(2H，m)，7.24(1H，t，J＝60.4Hz)，7.23(2H，d，J＝8.4Hz)，6.76(1H，d，J＝8.8Hz)，3.62-3.70(4H，m)，3.53(2H，s)，2.54-2.60(4H，m)，2.49(3H，s)。

Example 82: 6- (1-difluoromethyl-1H-pyrazol-4-yl) -4- (6- (6- (4- (methylthio) benzyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 21(43mg) was dissolved in THF (20mL) at 0 deg.C, and 4-methylthiobenzaldehyde (31mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (33 mg).

¹H NMR(400MHz，CDCl₃)δ8.73(1H，s)，8.46(1H，s)，8.31(1H，s)，8.15(1H，s)，7.98(1H，s)，7.85(1H，d，J＝7.2Hz)，7.47-7.55(2H，m)，7.44(1H，d，J＝1.2Hz)，7.25-7.26(2H，m)，7.24(1H，t，J＝60.4Hz)，6.73(1H，d，J＝9.2Hz)，4.13-4.37(2H，m)，3.78-4.10(6H，m)，3.10-3.38(1H，m)，2.47(3H，s)，1.80-1.90(1H，m)

Example 83: 4- (6- (6- (4-nitrobenzyl) -3, 6-diaza-bicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) -6- (1- (difluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 21(43mg) was dissolved in THF (20mL) at 0 deg.C, and 4-nitrobenzaldehyde (30mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by stirring at room temperature for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (34 mg).

¹H NMR(400MHz，CDCl₃)δ8.72(1H，d，J＝1.6Hz)，8.44(1H，d，J＝2.8Hz)，8.31(1H，s)，8.18(2H，d，J＝8.8Hz)，8.13(1H，s)，7.97(1H，s)，7.82(1H，dd，J＝8.8Hz，2.8Hz)，7.58(2H，d，J＝7.6Hz)，7.43(1H，d，J＝1.2Hz)，7.26(1H，t，J＝60.4Hz)，6.71(1H，d，J＝8.8Hz)，3.73-3.88(6H，m)，3.59-3.68(2H，m)，2.72-2.82(1H，m)，1.70(1H，d，J＝9.2Hz).

Example 84: 4- (6- (6- (4- (methylsulfonyl) benzyl) -3, 6-diazabicyclo [3.1.1] heptan-3-yl) pyridin-3-yl) -6- (1- (trifluoromethyl) -1H-pyrazol-4-yl) pyrazolo [1, 5-a ] pyridine-3-carbonitrile

Intermediate 25(45mg) was dissolved in THF (20mL) at 0 deg.C, 4-methylsulfonylbenzaldehyde (37mg) and sodium triacetoxyborohydride (100mg) were added in this order, followed by warming to room temperature and stirring for 12 hours. The reaction mixture was poured into saturated aqueous sodium bicarbonate (50mL), extracted with dichloromethane, the extract was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by thin layer chromatography (developing solvent: dichloromethane: methanol ═ 20: 1 (V: V)) to give the title compound (33 mg).

¹H NMR(400MHz，CDCl3)δ8.72(1H，d，J＝1.6Hz)，8.44(1H，d，J＝2.4Hz)，8.32(1H，s)，8.13(1H，s)，8.07(1H，s)，7.89(2H，d，J＝8.4Hz)，7.81(1H，dd，J＝8.8Hz，2.4Hz)，7.60(2H，d，J＝8.4Hz)，7.41(1H，d，J＝1.2Hz)，6.71(1H，d，J＝8.4Hz)，3.78-3.84(4H，m)，3.74(2H，s)，3.58-3.68(2H，m)，3.05(3H，s)，2.72-2.78(1H，m)，1.69(1H，d，J＝8.4Hz)。

Biological Activity assay

RET compound bioactivity test method

1. Compound pair RET^WTIn vitro enzymatic Activity assay

The compound pair RET in this patent^WTInhibition of enzymatic Activity IC of₅₀The value determination was carried out by the homogeneous time-resolved fluorescence (HTRF) method. Compounds were diluted in 100% DMSO in 5-fold gradients (7 concentrations) starting at 0.2mM, and 2. mu.L of each compound was added to 48. mu.L of reaction buffer (50mM HEPES pH7.5, 0.1mM Na)₃VO₄，5mM MgCl₂1mM DTT, 0.001% Tween 20 and 100. mu.g/ml BSA) were diluted and mixed well. 2.5. mu.L of GST-RET was added to 384 well plates (OptiPlate-384, purchased from Perkinelmer) followed by 5. mu.L of GST-RET^WT(658-1114aa, final concentration 0.5nM), mixing by centrifugation, and starting the reaction by adding 2.5. mu.L of ATP (final concentration 25. mu.M) and TK Peptide Substrate mixture (final concentration 1. mu.M, purchased from Cisbio) in a total reaction volume of 10. mu.L. The 384 well plates were placed in an incubator at 23 ℃ for 2 hours, and then 5. mu.L of TKAntibody (purchased from Cisbio) and 5. mu.L of Streptavidin-XL665 (purchased from Cisbio) were added to stop the reaction. After a further 1 hour incubation in the incubator, fluorescence values (320nm excitation, detecting 665nm and 620nm emission as the enzyme activity signal) were read on Envision (purchased from PerkinElmer). RET was determined at 7 concentrations for each compound^WTThe data used for the calculation of IC of the compound using GraphPad Prism software₅₀The value is obtained.

2. Compound pair RET^V804MIn vitro enzymatic Activity assay

The compound pair RET in this patent^V804MInhibition of enzymatic Activity IC of₅₀The value determination was carried out by the homogeneous time-resolved fluorescence (HTRF) method. Compounds were diluted in 100% DMSO in 5-fold gradients (7 concentrations) starting at 0.2mM, and 2. mu.L of each compound was added to 48. mu.L of reaction buffer (50mM HEPES pH7.5, 0.1mM Na)₃VO₄，5mM MgCl₂1mM DTT, 0.001% Tween 20, 20nM SEB (purchased from Cisbio) and 100. mu.g/ml BSA). 2.5. mu.L of GST-RET was added to 384 well plates (OptiPlate-384, purchased from Perkinelmer) followed by 5. mu.L of GST-RET^V804M(658-1114aa, final concentration 0.02nM), centrifuged and mixed, and then 2.5. mu.L ATP (final concentration 1.5. mu.M) and TK Peptide Substrate mixture (final concentration 1. mu.M, purchased from Cisbio) were added to start the reaction, the total reaction volume was 10. mu.L. The 384 well plates were placed in an incubator at 23 ℃ for 1 hour, and then 5. mu.L of TK Antibody (purchased from Cisbio) and 5. mu.L of Streptavidin-XL665 (purchased from Cisbio) were added to stop the reaction. After a further 1 hour incubation in the incubator, fluorescence values (320nm excitation, detecting 665nm and 620nm emission as the enzyme activity signal) were read on Envision (purchased from PerkinElmer). Each of themCompounds were tested for RET at 7 concentrations each^V804MThe data used for the calculation of IC of the compound using GraphPad Prism software₅₀The value is obtained.

3. In vitro enzymatic Activity assay for VEGFR2 with Compounds

The compounds of this patent inhibit the enzymatic Activity of VEGFR2 IC₅₀The value measurement was carried out by a method of homogeneous time-resolved fluorescence technique (HTRF). Compounds were diluted in 100% DMSO in 5-fold gradients starting at 1mM (7 concentrations in total), and 2. mu.L of compound was added to 48. mu.L of reaction buffer (50mM HEPES pH7.5, 0.1mM Na)₃VO₄，5mM MgCl₂，1mM MnCl₂1mM DTT, 0.001% Tween 20 and 100. mu.g/ml BSA). mu.L of the mixture was added to a 384 well plate (OptiPlate-384, purchased from Perkinelmer), 5. mu.L of GST-VEGFR2 (final concentration 0.1nM) was added, the mixture was centrifuged and mixed, and 2.5. mu.L of ATP (final concentration 2.5. mu.M) and TK Peptide Substrate mixture (final concentration 1. mu.M, purchased from Cisbio) were added to initiate the reaction in a total reaction volume of 10. mu.L. The 384 well plates were placed in an incubator at 23 ℃ for 1.5 hours, and then 5. mu.L of TK Antibody (purchased from Cisbio) and 5. mu.L of Streptavidin-XL665 (purchased from Cisbio) were added to stop the reaction. After a further 1 hour incubation in the incubator, fluorescence values (320nm excitation, detecting 665nm and 620nm emission as the enzyme activity signal) were read on Envision (purchased from PerkinElmer). The signal for the enzymatic activity of VEGFR2 was measured at 7 concentrations for each compound, and the data was calculated using GraphPad Prism software to obtain the IC of the compound₅₀The value is obtained.

4. Determination of cell proliferation Activity of Compounds in TT

Human thyroid carcinoma cell line TT cells were cultured using Ham' S F-12K medium plus 15% fetal bovine serum (FBS, available from Biological Industries, BI) and 1% penicillin/streptomycin diabody (P/S, available from Life Techology) at 37 deg.C, 5% CO₂. The day before compound detection, TT cells were plated in 96-well plates (#3917, purchased from Corning) at a concentration of 5000 cells/195 μ L/well. After 24 hours the compounds were dosed starting at 10mM in 100% DMSOLine 3 times gradient dilution (total 10 concentrations), then each concentration to take 2L compound added to 48L Ham's F-12K medium dilution. mu.L of each diluted compound was added to the plated Cell suspension, the compound and cells were co-incubated in a Cell incubator for 120h (5 days), the medium was aspirated off, and 25. mu.L of Cell-Titer Glo (G7570, purchased from Promega) reagent was added and incubated for a further 5-10 minutes. Fluorescence values were then read on Envision and the data were calculated using GraphPad Prism software to obtain the IC of the compound for inhibition of cell proliferation₅₀The value is obtained.

5. Compound proliferation activity assay in KIF5B-RET/HEK293T cells

RET fusion-type cell line KIF5B-RET/HEK293T cells constructed by lentivirus infection were cultured using DMEM medium plus 10% fetal bovine serum (FBS, purchased from Biological Industries, BI) and 1% penicillin/streptomycin diabody (P/S, purchased from Life technologies) at 37 ℃ with 5% CO₂. The day before compound detection, KIF5B-RET/HEK293T cells were plated in 96-well plates (#3917, purchased from Corning) at a concentration of 2000 cells/195. mu.L/well. After 24 hours, compounds were diluted in 100% DMSO in 3-fold gradient starting at 10mM (10 concentrations total), and then 2 μ L of compound was added to 48 μ L of DMEM medium for dilution at each concentration. mu.L of each diluted compound was added to the plated Cell suspension, the compound and cells were incubated together for 72h (3 days) in a Cell incubator, and 25. mu.L of Cell-Titer Glo (G7570, purchased from Promega) reagent was added after the medium was aspirated off for another 5-10 min incubation. Fluorescence values were then read on Envision and the data were calculated using GraphPad Prism software to obtain the IC of the compound for inhibition of cell proliferation₅₀The value is obtained.

Protein Activity and cytology data

"-" indicates no test

4. Pharmacokinetic data for compounds:

male SD rats are from Beijing Wittingle laboratory animal technology, Inc., the rats are grouped into 3 groups, and suspension of samples to be tested (5mg/kg, suspension MC 0.5%) is orally administrated. Animals were fasted overnight prior to the experiment, with the fasting time ranging from 10 hours prior to dosing to 4 hours post-dosing. Blood was collected at 0.25, 0.5, 1, 2, 4, 6, 8, and 24 hours post-dose, respectively. After isoflurane anesthesia by using a small animal anesthesia machine, 0.3mL of whole blood is collected through an eyeground venous plexus, the whole blood is placed in a heparin anticoagulation tube, a sample is centrifuged at 4000rpm for 5min at 4 ℃, and the plasma is transferred to a centrifugal tube and stored at-80 ℃ until analysis. Samples from plasma were extracted using protein precipitation and the extracts were analyzed by LC/MS.

Claims

1. A compound of formula (I), or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof:

wherein,

x and Y are each independently selected from CH and N;

a is selected from 5-6 membered heteroaryl, 3-12 membered heterocyclyl, -O-C_1-6Alkyl, -S-C_1-6Alkyl and-SO₂-C_1-6Alkyl, said heteroaryl and heterocyclyl being each optionally substituted by C_1-6Alkyl, -CN, -OH, -NR⁵R⁶Halogen, -O-C_1-6Alkyl, -S-C_1-6Alkyl, or-SO₂-C_1-6Alkyl substitution, said C_1-6Alkyl may optionally be substituted by-OH, halogen, or- (CO) N (CH)₃)₂Is substituted, and R⁵And R⁶Each independently selected from H and C_1-6An alkyl group;

R¹、R¹’、R²and R²Each independently is H or C_1-6Alkyl, or R¹And R¹' one of the compounds with R²And R²One of' may be taken together to form a bond or- (CH)₂)_m-；

Z is selected from- (CH)₂)_m-S-C_1-6Alkyl, - (CS) -N (R)⁸)₂、

And- (CH)₂)_n-Ar；

each m is independently selected from 1 and 2;

n is selected from 0, 1 and 2.

2. A compound according to claim 1 or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, wherein

A is selected from the group consisting of 5-6 membered heteroaryl and 3-12 membered heterocyclyl,

the heteroaryl and heterocyclyl groups may each be optionally substituted by C_1-6Alkyl, -CN, -OH, -NR⁵R⁶Halogen, -O-C_1-6Alkyl, -S-C_1-6Alkyl, or-SO₂-C_1-6Alkyl substitution, said C_1-6Alkyl may optionally be substituted by-OH, halogen, or- (CO) N (CH)₃)₂Is substituted and

R⁵and R⁶Each independently selected from H and C_1-6An alkyl group.

3. A compound according to claim 1 or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, wherein

A is selected from 5-6 membered heteroaryl and 4-9 membered heterocyclyl, each of which may optionally be substituted with C_1-6Alkyl, -CN, -OH, -NR⁵R⁶Halogen, -O-C_1-6Alkyl, -S-C_1-6Alkyl, or-SO₂-C_1-6Alkyl substitution, said C_1-6Alkyl may optionally be substituted by-OH, halogen, or- (CO) N (CH)₃)₂Is substituted and

R⁵and R⁶Each independently selected from H and C_1-6An alkyl group.

4. A compound selected from:

or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof.

5. A pharmaceutical composition comprising a compound according to any one of claims 1-4, or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, and a pharmaceutically acceptable carrier.

6. Use of a compound according to any one of claims 1-4, or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, for the manufacture of a medicament for the treatment of a RET-associated disease.

7. The use of claim 6, wherein the RET-associated disease is lung cancer, papillary thyroid carcinoma, medullary thyroid carcinoma, differentiated thyroid carcinoma, recurrent thyroid carcinoma, refractory differentiated thyroid carcinoma, multiple endocrine tumors of type 2A or 2B, pheochromocytoma, parathyroid hyperplasia, breast cancer, colorectal cancer, papillary renal cell carcinoma, gastrointestinal mucosal ganglion cell tumor, or cervical cancer.

8. The use according to claim 6, wherein the RET associated disease is RET fusion lung cancer or medullary thyroid cancer.

9. The use of claim 6, wherein the RET-associated disease is small cell lung cancer, non-small cell lung cancer, or bronchiolar lung cancer or lung adenocarcinoma.