CN110724143B

CN110724143B - Preparation of target BTK protein degradation compound and application of target BTK protein degradation compound in treatment of autoimmune system diseases and tumors

Info

Publication number: CN110724143B
Application number: CN201910954295.9A
Authority: CN
Inventors: 饶燏; 刘万里; 杨子默; 孙永汇; 倪智豪; 朱璨; 杨冰; 李雨欣
Original assignee: Tsinghua University
Current assignee: Tsinghua University
Priority date: 2019-10-09
Filing date: 2019-10-09
Publication date: 2021-03-23
Anticipated expiration: 2039-10-09
Also published as: WO2021068380A1; CN110724143A

Abstract

The invention provides a compound, which is a compound shown in a formula I or a stereoisomer, a geometric isomer, a tautomer, a nitrogen oxide, a hydrate, a solvate, a metabolite, a pharmaceutically acceptable salt or a prodrug thereof, has a strong effect of degrading wild BTK, has no inhibition or degradation effect on other targets such as EGFR, ITK, TEC and the like, and has the effect of specifically targeting and degrading BTK protein. X-Y-Z is formula I.

Description

Preparation of target BTK protein degradation compound and application of target BTK protein degradation compound in treatment of autoimmune system diseases and tumors

Technical Field

The invention relates to the field of biological medicines, in particular to preparation of a target BTK protein degradation compound and application thereof in treating autoimmune system diseases and tumors.

Background

The protac (protein Targeting chiceras) technology, which is a protein Targeting degradation technology, is a chemical biological or drug discovery approach for inducing protein degradation by using the ubiquitin-proteasome system emerging in recent years. The function of the PROTAC small molecule is based on the process that E3 ubiquitin ligase performs ubiquitin ligation on a certain protein, one end of a PROTAC small molecule chimera can be specifically combined with a target protein, and the other end recruits E3 ubiquitin ligase to enable the PROTAC small molecule chimera to be close to the target protein, so that a subsequent degradation process occurs. The PROTAC chimera is synthesized on the basis of a selective small-molecule inhibitor, and the high homologous proteins can be distinguished through ingenious design, so that the PROTAC technology has certain advantages in the application of biological tools; in addition, the PROTAC molecules are non-covalently bound with the target protein, and can be dissociated after one molecule of protein is degraded to degrade another molecule of target protein, and the means for degrading the protein by the catalytic amount provides the possibility that the PROTAC technology is widely concerned and applied because of more directness and rapidness for the development of high-activity and low-toxicity medicine molecules.

BTK is a member of the non-receptor tyrosine kinase family, is a key kinase in the B cell antigen receptor (BCR) signaling pathway, can control the development and differentiation of B cells by activating cell cycle forward regulators and differentiation factors, and can also regulate the survival and proliferation of B cells by expression of pro-apoptotic and anti-apoptotic proteins. The sustained activation of BTK is a prerequisite for the development of many hematologic and autoimmune diseases, and thus modulation of BTK protein has promising prospects for the treatment of hematologic malignancies and autoimmune disorders.

Non-hodgkin's lymphoma (NHL) is a hematological cancer, a generic term for all lymphomas except hodgkin's lymphoma. In the united states, 2.1% of the population's lives are affected by it. In 2015, 430 million people had non-hodgkin's lymphoma, 231400 were deprived of life. The majority of non-Hodgkin's lymphomas in clinic are B cell types, accounting for 70% -85% of the total. Diffuse large B-cell lymphoma (DLBCL) is the most common non-hodgkin lymphoma (NHL), with 7-8 cases occurring in 10 million people per year in the united states and uk. Ibrutinib (Ibrutinib) is approved by the FDA for marketing for the treatment of Chronic Lymphocytic Leukemia (CLL), Mantle Cell Lymphoma (MCL), and fahrenheit macroglobulinemia indication (WM). Furthermore, ibrutinib has been reported to effectively inhibit the proliferation of some species of DLBCL. The mechanism of action of ibrutinib is that covalent cross-linking is carried out between the acrylamide structure of ibrutinib and the sulfhydryl of 481-bit cysteine of BTK protein in cells, so that BTK loses the function of phosphorylating downstream signal protein, and the ibrutinib plays a role in resisting cell proliferation.

Although ibrutinib appears to be very potent against BTK kinaseThe inhibitory activity of enzymes, but the drug itself is very strong due to off-target effects, causing many side effects. IC against EGFR₅₀A value of 5.6nM, capable of causing severe diarrhea and rash; IC for ITK₅₀The value is 10.7nM, which can cause the loss of natural killer cell function; IC for TEC₅₀The value is 78nM, which can cause coagulation defects, and PROTAC, as a technique capable of precisely and rapidly degrading target proteins, has great advantages in realizing high-efficiency anti-tumor and reducing off-target side effects.

The autoimmune disease refers to a disease caused by self-tissue damage due to the immune reaction of an organism to an autoantigen, and comprises organ-specific autoimmune diseases such as goodpasture's syndrome, pemphigus and the like, systemic autoimmune diseases such as arthritis, systemic lupus erythematosus and the like, the specific mechanism of the autoimmune disease is not completely clear, and one possible mechanism is that a large amount of BTK expression abnormally activates a BCR signal pathway, so that B cell dysfunction and immune tolerance state are changed, and the BTK is converted into autoreactive B cells, and a large amount of autoantibodies are secreted to induce the occurrence of the disease.

Based on the above mechanisms, BTK inhibitors have been increasingly studied for application to autoimmune diseases (including rheumatoid arthritis) in recent years. Although only ibrutinib is currently marketed as an autoimmune disease (chronic graft versus host disease (cGVHD)) with FDA approval in 2017, there are currently over a dozen small molecule inhibitors entering the clinical stage. BTK has become the most target of clinical drugs in the field of autoimmune diseases except TNF (tumor necrosis factor) and CD20, is expected to become a new target for treating autoimmune diseases in the future, and provides opportunities and challenges for a PROTAC technology capable of efficiently degrading BTK protein.

Therefore, BTK kinase degrading agents have promising prospects as drugs for antitumor or treatment of autoimmune diseases and need further development.

Disclosure of Invention

The present application is based on the discovery and recognition by the inventors of the following facts and problems:

ibbrutinib can inhibit BTK kinase in the prior art, but is reported (n.eng)l.J.Med.370,2352-2254(2014), the BTK inhibitory ability IC of Ibrutinib on C481S mutation was reported₅₀The drug has different degrees of effects on other targets, such as EGFR, ITK and TEC, and has obvious side effects. Based on the discovery of the above problems, the inventors propose a novel compound which adopts a dual-target molecular structure, the structure of which is shown in fig. 1, one end of the molecule is targeted to bind to E3 ligase, the other end of the molecule is targeted to bind to a target protein (BTK protein) to be degraded, and the structures at the two ends are connected through a chain (linker) to form a complete compound molecule. The compound has strong degradation effect on wild BTK, has no inhibition or degradation effect on other targets such as EGFR, ITK, TEC and the like, and has the efficacy of specifically targeting BTK protein.

To this end, in a first aspect of the present invention, the present invention provides a compound which is a compound represented by formula I or a stereoisomer, a geometric isomer, a tautomer, a nitrogen oxide, a hydrate, a solvate, a metabolite, a pharmaceutically acceptable salt or a prodrug thereof:

X-Y-Z

formula I

Wherein:

y is

And each E is independently an amide group, an ester group, a carbamate group, a urea group, a guanidine group, a heterocyclic group, a cycloalkyl group, a spiroheterobicyclic group, a fused heterobicyclic group, a bridged heterobicyclic group, or a C6-10 aryl group; each E is optionally substituted with 1,2,3 or 4 independent R_bSubstituted; each W is O, S, NH or Se, b is an integer of 1 to 30, d is an integer of 0 to 30, L₆、L₇Independently is a bond, -O-, -S (═ O)_t1-，-S-，-N(R_a)-，-C(＝O)O-，-N(R_a)-C(＝O)-，-C(＝O)-(CH₂)_t2-，-CH₂-，-C(＝O)-，-OC(＝O)-，-C(＝S)-，-C(＝O)-N(R_a)-，-C(＝S)-N(R_a)-，-(CH₂)_t2-C (═ O) -or triazolyl,

x is

Z is as follows:

or said Y is

d is an integer of 0 to 30, wherein,

d＝6，L₆、L₇independently is-N (R)_a) -, - (O) -, said X is

Z is as follows:

d≠6，L₆、L₇independently is a bond, -O-, -S (═ O)_t1-，-S-，-N(R_a)-，-C(＝O)O-，-N(R_a)-C(＝O)-，-CH₂-，-C(＝O)-，-OC(＝O)-，-C(＝S)-，-C(＝O)-N(R_a)-，-C(＝S)-N(R_a) -or triazolyl, said X is

Z is as follows:

or said Y is

And W is CH₂O, S, NH or Se, b is an integer of 0 to 30, d is an integer of 0 to 30,

x is

Z is as follows:

or said Y is

x is

When, Z is:

x is

When said Z is

Wherein each L₁、L₂、L₃Independently is a bond, -O-, -S (═ O)_t1-，-S-，-N(R_a)-，-C(＝O)O-，-N(R_a)-C(＝O)-，-C(＝O)-(CH₂)_t2-，-CH₂-，-C(＝O)-，-OC(＝O)-，-C(＝S)-，-C(＝O)-N(R_a)-，-C(＝S)-N(R_a)-，-(CH₂)_t2-C (═ O) -or triazolyl;

each R_aIndependently hydrogen, C1-4 alkyl, haloC 1-4 alkyl, C1-4 alkanoyl or hydroxy;

each t2 is independently 0, 1,2,3, or 4;

each t1 is independently 0, 1 or 2;

each X₁、X₂、X₃、X₄、X₅、X₆、X₇、X₈Q is independently CH, CH₂O, S, N, NH or Se;

each R₁、R₂、R₃、R₄、R₅Independently H, deuterium, amino, C1-4 amido, C1-4 alkyl, C1-4 heteroalkyl, C3-8 cycloalkyl, C2-10 heterocyclyl, C6-10 aryl, C1-9 heteroaryl, C6-10 aryl, C1-4 alkoxy, C1-4 alkenyl and C1-4 alkynyl, wherein the C1-4 alkyl, C1-4 heteroalkyl, C3-8 cycloalkyl, C2-10 heterocyclyl, C6-10 aryl, C1-9 heteroaryl, C6-10 aryl, C1-4 alkoxy, C1-4 alkenyl and C1-4 alkynyl may be optionally substituted with one or more groups selected from deuterium, hydroxy, amino, oxo, F, Cl, Br, I, cyano, C1-6 alkyl, C1-6 haloalkyl, C1-6 hydroxyalkyl, C1-6 aminoalkyl, C1-6 alkoxy C1-6 alkyl, C1-6 alkylamino C1-6 alkyl, C1-10 arylC 1-6 alkyl, C1-9 heteroaryl C1-6 alkyl, C2-10 heterocyclyl C1-6 alkyl, C3-10 cycloalkyl C1-6 alkyl, C1-6 alkoxy, C1-6 alkylamino, C6-10 aryl, C6-10 aryloxy, C6-10 arylamino, C1-9 heteroaryl, C1-9 heteroaryloxy, C2-6 alkenyl, C3-10 cycloalkyl, halogen-substituted C6-10 aryl, halogen-substituted C1-9 heteroaryl, halogen-substituted C2-10 heterocyclyl or C2-10 heterocyclyl;

each A is independently hydrogen, C1-4 alkyl, C1-4 haloalkyl, hydroxy, nitro, amino, cyano, halogen, carboxy, C1-4 alkoxy, C1-4 alkylamino, C1-4 alkylthio, C1-4 alkanoyl, C3-12 cycloalkyl, optionally substituted C3-9 heterocyclyl, optionally substituted C6-12 aryl, optionally substituted C1-9 heteroaryl;

each R_bIndependently hydrogen, C1-4 alkyl, C1-4 haloalkyl, hydroxy, nitro, amino, cyano, halogen, carboxy, C1-4 alkoxy, C1-4 alkylamino, C1-4 alkylthio, C1-4 alkanoyl, C3-12 cycloalkyl, C3-9 heterocyclyl, C6-12 aryl, C1-9 heteroaryl, amino C1-4 alkyl, hydroxy C1-4 alkyl, sulfo, aminosulfonyl or aminoacyl;

each of a, c, e, and f is independently an integer of 0 to 30.

According to an embodiment of the present invention, the above compound may further comprise at least one of the following additional technical features:

according to an embodiment of the present invention, each L₁Independently a bond, -O-, -S-, -NH-,

each L₂Independently a bond, - (C ═ O) CH ═ CH₂-，-O-，-S-，-S(＝O)-，-S(＝O)₂-，-NRa-，-C(＝O)-，-C(＝O)O-，-N(R_a)-C(＝O)-，-CH₂-，-OC(＝O)-，-C(＝S)-，-C(＝O)-N(R_a)-，-C(＝S)-N(R_a) Or a group selected from the group consisting of a triazole group,

each R_aIndependently hydrogen, C1-2 alkyl, haloC 1-2 alkyl, C1-2 alkanoyl or hydroxy,

each R₁、R₂、R₃、R₄Independently H, amino, C1-4 alkyl, C1-4 heteroalkyl, C5-7 cycloalkyl, C5-7 heterocyclyl, C6-7 aryl, C5-7 heteroaryl, wherein said C1-4 alkyl, C1-4 heteroalkyl, C5-7 cycloalkyl, C5-7 heterocyclyl, C6-7 aryl, C5-7 heteroaryl may be optionally substituted with one or more substituents selected from deuterium, hydroxy, amino, oxo, F, Cl, Br, I, cyano.

According to an embodiment of the invention, each X is independently a compound as shown below:

according to an embodiment of the invention, each A is independently hydrogen, optionally substituted C5-7 heterocyclyl, C5-7 cycloalkyl, optionally substituted C6-7 aryl, optionally substituted C5-7 heteroaryl,

each R₅Independently H, amino, C1-4 alkyl, C1-4 heteroalkyl, C5-7 cycloalkyl, C5-7 heterocyclyl, C6-7 aryl, C5-7 heteroaryl, wherein said C1-4 alkyl, C1-4 heteroalkyl, C5-7 cycloalkyl, C5-7 heterocyclyl, C6-7 aryl, C5-7 heteroaryl may be optionally substituted with one or more substituents selected from deuterium, hydroxy, amino, oxo, F, Cl, Br, I, cyano, C5-7 aryl, C5-7 heteroaryl, halogen substituted C5-7 aryl, halogen substituted C5-7 heteroaryl, halogen substituted C5-7 heterocyclyl;

each L₃Independently a bond, - (C ═ O) CH ═ CH₂-，-O-，-S-，-S(＝O)-，-S(＝O)₂-，-NRa-，-C(＝O)-，-C(＝O)O-，-N(R_a)-C(＝O)-，-CH₂-，-OC(＝O)-，-C(＝S)-，-C(＝O)-N(R_a)-，-C(＝S)-N(R_a) Or a group selected from the group consisting of a triazole group,

each R_aIndependently hydrogen, C1-2 alkyl, haloC 1-2 alkyl, C1-2 alkanoyl or hydroxy.

According to an embodiment of the invention, each A is independently

Each R_cIndependently hydrogen or C1-2 alkyl.

According to an embodiment of the invention, each Z is independently a compound as shown below:

according to an embodiment of the invention, each E is independently an amide group, an ester group, a carbamate group, a urea group, a guanidine group, a heterocyclic group, a cycloalkyl group or a C6-C8 aryl group; each E is optionally substituted with 1,2,3 or 4 independent R_bThe substitution is carried out on the raw materials,

each R_bIndependently hydrogen, C1-2 alkyl, C1-2 haloalkyl, hydroxy, nitro, amino, cyano, halogen, carboxy, C1-2 alkoxy, C1-2 alkylamino, C1-2 alkylthio, C1-2 alkanoyl, C5-7 cycloalkyl, C5-7 heterocyclyl, C6-7 aryl, C5-7 heteroaryl, amino C1-2 alkyl, hydroxy C1-2 alkyl, sulfo, aminosulfonyl or aminoacyl,

According to an embodiment of the invention, each Y is independently

According to an embodiment of the invention, X is

In some embodiments, Z is

In some embodiments, the Y is

Wherein m1 is an integer of 1-10, and n1 is an integer of 0-10.

According to an embodiment of the present invention, the compound includes a compound represented by any one of formulas (1) to (15) or a stereoisomer, a geometric isomer, a tautomer, a nitrogen oxide, a hydrate, a solvate, a metabolite, a pharmaceutically acceptable salt, or a prodrug thereof,

wherein m1 is an integer of 1-10, and n1 is an integer of 0-10.

In a second aspect of the invention, the invention features a compound. According to an embodiment of the present invention, the compound is a compound represented by any one of formulas 1 to 18, or a stereoisomer, a geometric isomer, a tautomer, a nitrogen oxide, a hydrate, a solvate, a metabolite, a pharmaceutically acceptable salt, or a prodrug thereof,

the compound provided by the embodiment of the invention has a strong effect on the degradation of wild BTK, has no inhibition or degradation effect on other targets such as EGFR, ITK, TEC and the like, and has the efficacy of specifically targeting and degrading BTK protein.

In a third aspect of the invention, the invention provides a pharmaceutical composition comprising a compound as described hereinbefore. The pharmaceutical composition provided by the embodiment of the invention has a strong effect on the degradation of wild BTK, has no inhibition or degradation effect on other targets such as EGFR, ITK, TEC and the like, and has the efficacy of specifically targeting and degrading BTK protein.

According to an embodiment of the present invention, the above pharmaceutical composition may further comprise at least one of the following additional technical features:

according to an embodiment of the present invention, the pharmaceutical composition further comprises a pharmaceutically acceptable carrier, excipient, diluent, adjuvant, vehicle, or combination thereof.

According to an embodiment of the present invention, the pharmaceutical composition further comprises other drugs for treating or preventing non-hodgkin's lymphoma or autoimmune diseases. The inventors have found that the combination is more effective in treating or preventing non-hodgkin's lymphoma or autoimmune disease.

According to an embodiment of the invention, the autoimmune disease is arthritis, pulmonary hemorrhage, systemic lupus erythematosus, pemphigus, chronic lymphocytic thyroiditis, hyperthyroidism, insulin dependent diabetes mellitus, myasthenia gravis, chronic ulcerative colitis, pernicious anemia with chronic atrophic gastritis, primary biliary cirrhosis, multiple sclerosis or acute idiopathic polyneuritis. In some embodiments, the autoimmune disease is arthritis or pulmonary hemorrhage.

According to an embodiment of the present invention, the other drug for treating or preventing non-hodgkin's lymphoma or autoimmune disease comprises ibrutinib.

In a fourth aspect of the invention, the invention proposes the use of a compound as described above or a pharmaceutical composition as described above for the manufacture of a medicament for degrading BTK or inhibiting BTK. The compound or the pharmaceutical composition provided by the embodiment of the invention has a strong effect of degrading wild BTK, has no inhibition or degradation effect on other targets such as EGFR, ITK, TEC and the like, and has the efficacy of specifically targeting BTK protein.

In a fifth aspect of the invention, the invention proposes the use of a compound as described above or a pharmaceutical composition as described above for the manufacture of a medicament for the treatment or prevention of a BTK-related disease. The compound or the pharmaceutical composition provided by the embodiment of the invention has a strong effect of degrading wild BTK, has no inhibition or degradation effect on other targets such as EGFR, ITK, TEC and the like, has the efficacy of specifically targeting BTK protein, and has a good treatment or prevention effect on BTK-related diseases.

According to an embodiment of the present invention, the above-mentioned use may further include at least one of the following additional technical features:

according to an embodiment of the invention, the BTK-related disease is non-hodgkin lymphoma or an autoimmune disease. The compound or the pharmaceutical composition according to the embodiment of the present invention has a better therapeutic or prophylactic effect on non-hodgkin's lymphoma or autoimmune diseases.

According to an embodiment of the invention, the autoimmune disease is arthritis, pulmonary hemorrhage, systemic lupus erythematosus, pemphigus, chronic lymphocytic thyroiditis, hyperthyroidism, insulin dependent diabetes mellitus, myasthenia gravis, chronic ulcerative colitis, pernicious anemia with chronic atrophic gastritis, primary biliary cirrhosis, multiple sclerosis or acute idiopathic polyneuritis.

According to an embodiment of the invention, the autoimmune disease is arthritis or pulmonary hemorrhage. The inventors have found that the compounds or pharmaceutical compositions described above can prevent and treat arthritis and pulmonary hemorrhage symptoms in mouse models, and that the prevention and improvement is significantly superior to ibrutinib.

In yet another aspect of the invention, the invention features general procedures for the synthesis of compounds of formula I. According to the embodiment of the invention, the compound shown in formula I can be connected through click reaction or amide condensation reaction between Pomalidomide or Lenanidomide or RG-7112 terminal derivatives and Ibrutinib terminal derivatives, as shown in FIG. 2, wherein the Pomalidomide terminal derivatives can be prepared by reference to Chemistry & Biology 22,755 763(2015), the Lenalidomide terminal derivatives can be prepared by reference to J.Med.chem (DOI:10.1021/ACS. jmedchem.6b01816), the RG-7112 terminal derivatives can be prepared by reference to bioorg.Med.chem.Lett.18,5904-5908(2008). ACS.Med.chem.Lett.4,466-469 (2013), the Ibrinib terminal derivatives can be synthesized by reference to Jaorg.Chem.29, PCT 3603, and PCT. The terminal alkyne required by Click chemistry is linked to the piperidine ring of the ibbrutinib parent nucleus by amide condensation, as described in j.chem.inf.model.50,446(2010). PCT int.appl.,2013170115,14Nov 2013.

Drawings

FIG. 1 is a basic technical scheme of PROTACs according to an embodiment of the present invention;

FIG. 2 is a schematic illustration of the construction of formula I by a click reaction and an amide condensation reaction according to an embodiment of the present invention;

FIG. 3 shows the results of inhibition of LPS-stimulated release of THP-1 cytokines by compounds according to examples of the present invention;

FIG. 4 is the degradation of BTK by compounds according to examples of the present invention.

Detailed Description

Reference will now be made in detail to embodiments of the present invention, examples of which are illustrated in the accompanying drawings. The embodiments described below with reference to the drawings are illustrative and intended to be illustrative of the invention and are not to be construed as limiting the invention.

As used herein, the term "administering to a patient a previously described compound, or a stereoisomer, geometric isomer, tautomer, nitrogen oxide, hydrate, solvate, metabolite, pharmaceutically acceptable salt or prodrug thereof, or a previously described pharmaceutical composition" refers to introducing a predetermined amount of a substance into the patient by some suitable means. The compound of formula I of the present invention, or a stereoisomer, geometric isomer, tautomer, nitrogen oxide, hydrate, solvate, metabolite, pharmaceutically acceptable salt or prodrug thereof, or pharmaceutical composition thereof, may be administered by any common route, as long as it reaches the intended tissue. Various modes of administration are contemplated, including peritoneal, intravenous, intramuscular, subcutaneous, cortical, oral, topical, nasal, pulmonary and rectal, but the invention is not limited to these exemplified modes of administration. However, because of oral administration, the active ingredients of orally administered compositions should be coated or formulated to prevent degradation in the stomach. Furthermore, the compounds of formula I according to the invention or the pharmaceutical compositions can be administered using specific devices for delivering the active ingredient to the target cells.

The administration frequency and dose of the pharmaceutical composition of the present invention can be determined by a number of relevant factors, including the type of disease to be treated, the administration route, the age, sex, body weight and severity of the disease of the patient and the type of drug as an active ingredient.

The term "therapeutically effective amount" refers to an amount of a compound sufficient to significantly ameliorate some of the symptoms associated with a disease or condition, i.e., to provide a therapeutic effect for a given condition and administration regimen. A therapeutically effective amount of a drug or compound need not cure a disease or condition, but will provide treatment for a disease or condition such that the onset of the disease or condition in an individual is delayed, prevented or prevented, or the symptoms of the disease or condition are alleviated, or the duration of the disease or condition is altered, or the disease or condition becomes less severe, or recovery is accelerated, for example.

The term "treatment" is used to refer to obtaining a desired pharmacological and/or physiological effect. The effect may be prophylactic in terms of complete or partial prevention of the disease or symptoms thereof, and/or may be therapeutic in terms of a partial or complete cure for the disease and/or adverse effects resulting from the disease. As used herein, "treatment" encompasses treatment of a disease in a mammal, particularly a human, including: (a) preventing the occurrence of a disease or disorder in an individual who is susceptible to the disease but has not yet been diagnosed with the disease; (b) inhibiting the disease; or (c) alleviating the disease, e.g., alleviating symptoms associated with the disease. As used herein, "treatment" encompasses any administration of a drug or compound to a subject to treat, cure, alleviate, ameliorate, reduce or inhibit a disease in the subject, including, but not limited to, administration of a composition comprising a compound of formula I or formula II as described herein to a subject in need thereof.

According to the embodiment of the invention, the auxiliary materials comprise medicinal excipients, lubricants, fillers, diluents, disintegrating agents, stabilizers, preservatives, emulsifiers, cosolvents, colorants and sweeteners which are well known in the field of preparation, and are prepared into different dosage forms such as tablets, pills, capsules, injections and the like.

Reference will now be made in detail to certain embodiments of the invention, examples of which are illustrated by the accompanying structural and chemical formulas. The invention is intended to cover alternatives, modifications and equivalents, which may be included within the scope of the invention as defined by the appended claims. Those skilled in the art will recognize that many methods and materials similar or equivalent to those described herein can be used in the practice of the present invention. The present invention is in no way limited to the methods and materials described herein. In the event that one or more of the incorporated documents, patents, and similar materials differ or contradict this application (including but not limited to defined terminology, application of terminology, described techniques, and the like), this application controls.

It will be further appreciated that certain features of the invention, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the invention which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable subcombination.

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 this invention belongs. All patents and publications referred to herein are incorporated by reference in their entirety.

The following definitions as used herein should be applied, unless otherwise indicated. For the purposes of the present invention, the chemical elements are in accordance with the CAS version of the periodic Table of the elements, and the handbook of chemistry and Physics, 75 th edition, 1994. In addition, general principles of Organic Chemistry can be referred to as described in "Organic Chemistry", Thomas Sorrell, University Science Books, Sausaltito: 1999, and "March's Advanced Organic Chemistry" by Michael B.Smith and Jerry March, John Wiley & Sons, New York:2007, the entire contents of which are incorporated herein by reference.

The articles "a," "an," and "the" as used herein are intended to include "at least one" or "one or more" unless otherwise indicated or clearly contradicted by context. Thus, as used herein, the articles refer to articles of one or more than one (i.e., at least one) object. For example, "a component" refers to one or more components, i.e., there may be more than one component contemplated for use or use in embodiments of the described embodiments.

The term "comprising" is open-ended, i.e. includes the elements indicated in the present invention, but does not exclude other elements.

"stereoisomers" refers to compounds having the same chemical structure but differing in the arrangement of atoms or groups in space. Stereoisomers include enantiomers, diastereomers, conformers (rotamers), geometric isomers (cis/trans), atropisomers, and the like.

"chiral" is a molecule having the property of not overlapping its mirror image; and "achiral" refers to a molecule that can overlap with its mirror image.

"enantiomer" refers to two isomers of a compound that are not overlapping but are in mirror image relationship to each other.

"diastereomer" refers to a stereoisomer having two or more chiral centers and whose molecules are not mirror images of each other. Diastereomers have different physical properties, such as melting points, boiling points, spectral properties, and reactivities. Mixtures of diastereomers may be separated by high resolution analytical procedures such as electrophoresis and chromatography, e.g., HPLC.

The stereochemical definitions and rules used in the present invention generally follow the general definitions of S.P. Parker, Ed., McGraw-Hill Dictionary of Chemical Terms (1984) McGraw-Hill Book Company, New York; and Eliel, E.and Wilen, S., "Stereochemistry of Organic Compounds", John Wiley & Sons, Inc., New York, 1994.

Many organic compounds exist in an optically active form, i.e., they have the ability to rotate the plane of plane polarized light. In describing optically active compounds, the prefixes D and L or R and S are used to denote the absolute configuration of a molecule with respect to one or more of its chiral centers. The prefixes d and l or (+) and (-) are the symbols used to specify the rotation of plane polarized light by the compound, where (-) or l indicates that the compound is left-handed. Compounds prefixed with (+) or d are dextrorotatory. A particular stereoisomer is an enantiomer and a mixture of such isomers is referred to as an enantiomeric mixture. A50: 50 mixture of enantiomers is referred to as a racemic mixture or racemate, which may occur when there is no stereoselectivity or stereospecificity in the chemical reaction or process.

Any asymmetric atom (e.g., carbon, etc.) of a compound disclosed herein can exist in racemic or enantiomerically enriched forms, such as the (R) -, (S) -or (R, S) -configuration. In certain embodiments, each asymmetric atom has at least 50% enantiomeric excess, at least 60% enantiomeric excess, at least 70% enantiomeric excess, at least 80% enantiomeric excess, at least 90% enantiomeric excess, at least 95% enantiomeric excess, or at least 99% enantiomeric excess in the (R) -or (S) -configuration.

Depending on the choice of starting materials and methods, the compounds of the invention may exist as one of the possible isomers or as mixtures thereof, for example as racemates and diastereomeric mixtures (depending on the number of asymmetric carbon atoms). Optically active (R) -or (S) -isomers can be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques. If the compound contains a double bond, the substituents may be in the E or Z configuration; if the compound contains a disubstituted cycloalkyl group, the substituents of the cycloalkyl group may have cis or trans configuration.

Any resulting mixture of stereoisomers may be separated into pure or substantially pure geometric isomers, enantiomers, diastereomers, depending on differences in the physicochemical properties of the components, for example, by chromatography and/or fractional crystallization.

The racemates of any of the resulting end products or intermediates can be resolved into the optical enantiomers by known methods using methods familiar to those skilled in the art, e.g., by separation of the diastereomeric salts obtained. The racemic product can also be separated by chiral chromatography, e.g., High Performance Liquid Chromatography (HPLC) using a chiral adsorbent. In particular, Enantiomers can be prepared by asymmetric synthesis, for example, see Jacques, et al, Enantiomers, racemes and solutions (Wiley Interscience, New York, 1981); principles of Asymmetric Synthesis (2)^nd Ed.Robert E.Gawley,Jeffrey Aubé,Elsevier,Oxford,UK,2012)；Eliel,E.L.Stereochemistry of Carbon Compounds(McGraw-Hill,NY,1962)；Wilen,S.H.Tables of Resolving Agents and Optical Resolutions p.268(E.L.Eliel,Ed.,Univ.of Notre Dame Press,Notre Dame,IN 1972)；Chiral Separation Techniques:A Practical Approach(Subramanian,G.Ed.,Wiley-VCH Verlag GmbH&Co.KGaA,Weinheim,Germany,2007)。

The term "tautomer" or "tautomeric form" refers to structural isomers having different energies that can interconvert by a low energy barrier (low energy barrier). If tautomerism is possible (e.g., in solution), then the chemical equilibrium of the tautomer can be reached. For example, proton tautomers (also known as proton transfer tautomers) include interconversions by proton migration, such as keto-enol isomerization and imine-enamine isomerization. Valence tautomers (valenctautomers) include interconversion by recombination of some of the bonding electrons. A specific example of keto-enol tautomerism is the tautomerism of the pentan-2, 4-dione and 4-hydroxypent-3-en-2-one tautomers. Another example of tautomerism is phenol-ketone tautomerism. One specific example of phenol-ketone tautomerism is the tautomerism of pyridin-4-ol and pyridin-4 (1H) -one tautomers. Unless otherwise indicated, all tautomeric forms of the compounds of the invention are within the scope of the invention.

The compounds of the invention may be optionally substituted with one or more substituents, as described herein, in compounds of the general formula above, or as specifically exemplified, sub-classes, and classes of compounds encompassed by the invention. It is understood that the term "optionally substituted" may be used interchangeably with the term "substituted or unsubstituted". In general, the term "substituted" means that one or more hydrogen atoms in a given structure are replaced with a particular substituent. Unless otherwise indicated, an optional substituent group may be substituted at each substitutable position of the group. When more than one position in a given formula can be substituted with one or more substituents selected from a particular group, the substituents may be substituted at each position, identically or differently.

In addition, unless otherwise explicitly indicated, the descriptions of the terms "… independently" and "… independently" and "… independently" used in the present invention are interchangeable and should be understood in a broad sense to mean that the specific items expressed between the same symbols do not affect each other in different groups or that the specific items expressed between the same symbols in the same groups do not affect each other.

In the various parts of this specification, substituents of the disclosed compounds are disclosed in terms of group type or range. It is specifically intended that the invention includes each and every independent subcombination of the various members of these groups and ranges. For example, the term "C1-6 alkyl" refers specifically to independently disclosed methyl, ethyl, C3 alkyl, C4 alkyl, C5 alkyl, and C6 alkyl.

Each of a, b, c, d, e, and f is independently an integer of 0 to 30. In one embodiment, each of a, b, c, d, e, f is independently an integer between 0 and 25. In another embodiment, each of a, b, c, d, e, f is independently an integer between 0 and 20. In another embodiment, each of a, b, c, d, e, f is independently an integer between 0 and 15. In another embodiment, each of a, b, c, d, e, f is independently an integer between 0 and 10. In another embodiment, each of a, b, c, d, e, f is independently an integer between 0 and 5.

In each of the parts of the invention, linking substituents are described. Where the structure clearly requires a linking group, the markush variables listed for that group are understood to be linking groups. For example, if the structure requires a linking group and the markush group definition for the variable recites "alkyl" or "aryl," it is understood that the "alkyl" or "aryl" represents an attached alkylene group or arylene group, respectively.

The term "alkyl" or "alkyl group" as used herein, denotes a saturated, straight or branched chain monovalent hydrocarbon radical containing from 1 to 20 carbon atoms, wherein the alkyl group may be optionally substituted with one or more substituents as described herein. Unless otherwise specified, alkyl groups contain 1-20 carbon atoms. In one embodiment, the alkyl group contains 1 to 12 carbon atoms; in another embodiment, the alkyl group contains 1 to 6 carbon atoms; in yet another embodiment, the alkyl group contains 1 to 4 carbon atoms; in yet another embodiment, the alkyl group contains 1 to 3 carbon atoms.

Examples of alkyl groups include, but are not limited to, methyl (Me, -CH3), ethyl (Et, -CH2CH3), n-propyl (n-Pr, -CH2CH2CH3), isopropyl (i-Pr, -CH (CH3)2), n-butyl (n-Bu, -CH2CH2CH2CH3), isobutyl (i-Bu, -CH2CH (CH3)2), sec-butyl (s-Bu, -CH (CH3) CH2CH3), tert-butyl (t-Bu, -C (CH3)3), n-pentyl (-CH2CH2CH2CH 3), 2-pentyl (-CH (CH3) CH2CH2CH3), 3-pentyl (-CH (CH2CH3)2), 2-methyl-2-butyl (-C (CH3)2CH2CH3), 3-methyl-2-butyl (-CH (CH3) CH (CH3), 3-methyl-2-butyl (-CH2CH 3-3 CH3)2CH2CH 3-2 CH), 2-methyl-1-butyl (-CH2CH 3), n-hexyl (-CH2CH2CH2CH2CH3), 2-hexyl (-CH (CH3) CH2CH2CH3), 3-hexyl (-CH (CH2CH3) (CH2CH2CH3)), 2-methyl-2-pentyl (-C (CH3)2CH2CH2CH3), 3-methyl-2-pentyl (-CH (CH3) CH (CH3) CH2CH3), 4-methyl-2-pentyl (-CH (CH3) CH2CH (CH3)2), 3-methyl-3-pentyl (-C (CH3) (CH2CH3)2), 2-methyl-3-pentyl (-CH (CH2CH3) CH (CH3)2), 2, 3-dimethyl-2-butyl (-C (CH 2) 2 (-CH 69556), 2-dimethyl-3-butyl (-CH 8653) 2CH 8653), n-heptyl, n-octyl, and the like.

The term "alkylene" refers to a saturated divalent hydrocarbon radical resulting from the removal of two hydrogen atoms from a saturated straight or branched chain hydrocarbon radical. Unless otherwise specified, the alkylene group contains 1 to 12 carbon atoms. In one embodiment, the alkylene group contains 1 to 6 carbon atoms; in another embodiment, the alkylene group contains 1 to 4 carbon atoms; in yet another embodiment, the alkylene group contains 1 to 3 carbon atoms; in yet another embodiment, the alkylene group contains 1 to 2 carbon atoms. Examples of such include methylene (-CH2-), ethylene (-CH2CH2-), isopropylidene (-CH (CH3) CH2-), and the like.

The term "alkenyl" denotes a straight or branched chain monovalent hydrocarbon radical containing 2 to 12 carbon atoms, wherein there is at least one site of unsaturation, i.e., one carbon-carbon sp2 double bond, wherein the alkenyl radical may be optionally substituted with one or more substituents described herein, including the positioning of "cis" and "tans", or the positioning of "E" and "Z". In one embodiment, the alkenyl group contains 2 to 8 carbon atoms; in another embodiment, the alkenyl group contains 2 to 6 carbon atoms; in yet another embodiment, the alkenyl group contains 2 to 4 carbon atoms. Examples of alkenyl groups include, but are not limited to, vinyl (-CH ═ CH2), allyl (-CH2CH ═ CH2), and the like.

The term "alkynyl" denotes a straight or branched chain monovalent hydrocarbon radical containing 2 to 12 carbon atoms, wherein there is at least one site of unsaturation, i.e. a carbon-carbon sp triple bond, wherein said alkynyl radical may optionally be substituted with one or more substituents as described herein. In one embodiment, alkynyl groups contain 2-8 carbon atoms; in another embodiment, alkynyl groups contain 2-6 carbon atoms; in yet another embodiment, alkynyl groups contain 2-4 carbon atoms. Examples of alkynyl groups include, but are not limited to, ethynyl (-C.ident.CH), propargyl (-CH2℃ ident.CH), 1-propynyl (-C.ident.C-CH 3), and the like.

The term "heteroalkyl" denotes an alkyl chain interrupted by one or more heteroatoms, wherein the alkyl group and the heteroatoms have the meaning as described herein. Unless otherwise specified, the heteroalkyl group has from 2 to 10 carbon atoms, in other embodiments the heteroalkyl group has from 2 to 8 carbon atoms, in other embodiments the heteroalkyl group has from 2 to 6 carbon atoms, in other embodiments the heteroalkyl group has from 2 to 4 carbon atoms, and in other embodiments the heteroalkyl group has from 2 to 3 carbon atoms. Examples include, but are not limited to, CH3OCH2-, CH3CH2OCH2-, CH3SCH2-, (CH3)2NCH2-, (CH3)2CH2OCH2-, CH3OCH2CH2-, CH3CH2OCH2CH2-, and the like.

The term "alkenylene" denotes an alkenyl group derived from a straight or branched chain alkene by the removal of two hydrogen atoms. And the alkenylene group may be substituted or unsubstituted, wherein the substituent may be, but is not limited to, deuterium, hydroxyl, amino, halogen, cyano, aryl, heteroaryl, alkoxy, alkyl, alkenyl, alkynyl, heterocyclic, mercapto, nitro, or aryloxy. Examples include, but are not limited to, ethenylene (-CH ═ CH-), isopropenylene (-C (CH3) ═ CH-), 3-methoxypropene-1, 1-diyl, 2-methylbutene-1, 1-diyl, and the like.

The term "carbocyclylene" ("cycloalkylene") denotes a saturated divalent hydrocarbon ring obtained by removing two hydrogen atoms from a monocyclic ring having 3 to 12 carbon atoms or a bicyclic ring having 7 to 12 carbon atoms, wherein carbocyclyl or cycloalkyl have the meaning as defined in the present invention, and examples thereof include, but are not limited to, cyclopropylene, cyclobutylene, cyclopentylene, 1-cyclopent-1-enylene, 1-cyclopent-2-enylene and the like.

The term "heterocyclylene" denotes a monocyclic, bicyclic or tricyclic ring system wherein one or more atoms in the ring are independently selected from heteroatoms and may be fully saturated or contain one or more unsaturations, but not belonging to the aromatic class, having two points of attachment to the rest of the molecule, wherein the heterocyclyl group has the meaning as described herein. Examples include, but are not limited to, piperidine-1, 4-diyl, piperazine-1, 4-diyl, tetrahydrofuran-2, 4-diyl, tetrahydrofuran-3, 4-diyl, azetidine-1, 3-diyl, pyrrolidine-1, 3-diyl, and the like.

The term "alkoxy" means an alkyl group attached to the rest of the molecule through an oxygen atom, wherein the alkyl group has the meaning as described herein. Unless otherwise specified, the alkoxy group contains 1 to 12 carbon atoms. In one embodiment, the alkoxy group contains 1 to 6 carbon atoms; in another embodiment, the alkoxy group contains 1 to 4 carbon atoms; in yet another embodiment, the alkoxy group contains 1 to 3 carbon atoms. The alkoxy group may be optionally substituted with one or more substituents described herein.

Examples of alkoxy groups include, but are not limited to, methoxy (MeO, -OCH3), ethoxy (EtO, -OCH2CH3), 1-propoxy (n-PrO, n-propoxy, -OCH2CH2CH3), 2-propoxy (i-PrO, i-propoxy, -OCH (CH3)2), 1-butoxy (n-BuO, n-butoxy, -OCH2CH2CH2CH3), 2-methyl-l-propoxy (i-BuO, i-butoxy, -OCH2CH (CH3)2), 2-butoxy (s-BuO, s-butoxy, -OCH (CH3) CH2CH3), 2-methyl-2-propoxy (t-BuO, t-butoxy, -OC (CH3)3), 1-pentoxy (n-pentoxy, n-pentoxy, -OCH2CH3), 2-pentyloxy (-OCH (CH3) CH2CH3), 3-pentyloxy (-OCH (CH2CH3)2), 2-methyl-2-butoxy (-OC (CH3)2CH 3), 3-methyl-2-butoxy (-OCH (CH3) CH (CH3)2), 3-methyl-l-butoxy (-OCH2CH2CH (CH3)2), 2-methyl-l-butoxy (-OCH2CH (CH3) CH2CH3), and the like.

The terms "haloalkyl", "haloalkenyl" or "haloalkoxy" denote alkyl, alkenyl or alkoxy groups substituted with one or more halogen atoms, examples of which include, but are not limited to, trifluoromethyl, trifluoromethoxy and the like.

The term "hydroxyalkyl" or "hydroxy-substituted alkyl" means that the alkyl group is substituted with one or more hydroxy groups, wherein the alkyl group has the meaning described herein. Examples include, but are not limited to, hydroxymethyl, hydroxyethyl, 1, 2-dihydroxyethyl, and the like.

The term "carbocyclyl" or "carbocycle" denotes a monovalent or multivalent, non-aromatic, saturated or partially unsaturated monocyclic, bicyclic or tricyclic ring system containing 3 to 12 carbon atoms. Carbobicyclic groups include spirocarbocyclic and fused carbocyclic groups, and suitable carbocyclic groups include, but are not limited to, cycloalkyl, cycloalkenyl and cycloalkynyl groups. Examples of carbocyclyl groups further include cyclopropyl, cyclobutyl, cyclopentyl, 1-cyclopentyl-1-alkenyl, 1-cyclopentyl-2-alkenyl, 1-cyclopentyl-3-alkenyl, cyclohexyl, 1-cyclohexyl-1-alkenyl, 1-cyclohexyl-2-alkenyl, 1-cyclohexyl-3-alkenyl, cyclohexadienyl, cycloheptyl, cyclooctyl, cyclononyl, cyclodecyl, cycloundecyl, cyclododecyl, and the like.

The term "cycloalkyl" denotes a monovalent or polyvalent saturated monocyclic, bicyclic or tricyclic ring system containing from 3 to 12 carbon atoms. In one embodiment, the cycloalkyl group contains 3 to 12 carbon atoms; in another embodiment, cycloalkyl contains 3 to 8 carbon atoms; in yet another embodiment, the cycloalkyl group contains 3 to 6 carbon atoms. The cycloalkyl groups may be independently unsubstituted or substituted with one or more substituents described herein.

The terms "heterocyclyl" and "heterocycle" are used interchangeably herein and refer to a saturated or partially unsaturated monocyclic, bicyclic or tricyclic ring containing 3 to 12 ring atoms, wherein at least one ring atom is selected from the group consisting of nitrogen, sulfur and oxygen atoms. Unless otherwise indicated, heterocyclyl groups may be carbon-or nitrogen-based, and the-CH 2-group may optionally be replaced by-c (o) -. The sulfur atom of the ring may optionally be oxidized to the S-oxide. The nitrogen atom of the ring may optionally be oxidized to an N-oxygen compound. Examples of heterocyclyl groups include, but are not limited to: oxiranyl, azetidinyl, oxetanyl, thietanyl, pyrrolidinyl, 2-pyrrolinyl, 3-pyrrolinyl, pyrazolinyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, tetrahydrofuryl, dihydrofuranyl, tetrahydrothienyl, dihydrothienyl, 1, 3-dioxolanyl, dithiocyclopentyl, tetrahydropyranyl, dihydropyranyl, 2H-pyranyl, 4H-pyranyl, tetrahydrothiopyranyl, piperidinyl, morpholinyl, thiomorpholinyl, piperazinyl, dioxanyl, dithianyl, thiaxanyl, homopiperazinyl, homopiperidinyl, oxepanyl, thiepanyl, oxazepanyl, thiazepinyl, thiazepanyl, homopiperazinyl, homopiperidinyl, oxazepanyl, and the like

Radical, diaza

Radical, S-N-aza

Radicals, indolinyl, 1,2,3, 4-tetrahydroisoquinolinyl, 1, 3-benzodioxolyl, 2-oxa-5-azabicyclo [2.2.1]Hept-5-yl. Examples of substitutions of the-CH 2-group by-C (O) -in the heterocyclic group include, but are not limited to, 2-oxopyrrolidinyl, oxo-1, 3-thiazolidinyl, 2-piperidinonyl, 3, 5-dioxopiperidinyl and pyrimidinedione. Examples of the sulfur atom in the heterocyclic group being oxidized include, but are not limited to, sulfolane group, 1-dioxothiomorpholinyl group. The heterocyclyl group may be optionally substituted with one or more substituents as described herein.

In one embodiment, heterocyclyl is a 4-7 atom heterocyclyl and refers to a saturated or partially unsaturated monocyclic ring containing 4-7 ring atoms in which at least one ring atom is selected from the group consisting of nitrogen, sulfur, and oxygen atoms. Unless otherwise indicated, a heterocyclic group of 4-7 atoms may be carbon-or nitrogen-based, and the-CH 2-group may optionally be replaced by-c (o) -. The sulfur atom of the ring may optionally be oxidized to the S-oxide. The nitrogen atom of the ring may optionally be oxidized to an N-oxygen compound. Examples of heterocyclic groups consisting of 4 to 7 atoms include, but are not limited to: azetidinyl, oxetanyl, thietanyl, pyrrolidinyl, 2-pyrrolinyl, 3-pyrrolinyl, pyrazolinyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, tetrahydrofuranyl, dihydrofuranyl, tetrahydrothienyl, dihydrothienyl, 1, 3-dioxolanyl, dithiocyclopentyl, tetrahydropyranyl, dihydropyranyl, 2H-pyranyl, 4H-pyranyl, tetrahydrothiopyranyl, piperidinyl, morpholinyl, thiomorpholinyl, piperazinyl, dioxanyl, dithianyl, thioxanyl, homopiperazinyl, homopiperidinyl, oxepanyl, thiepanyl, oxacycloheptanyl, oxazepanyl, thiazepanyl, thiazepan

Radical, diaza

Radical, S-N-aza

And (4) a base. Examples of substitutions of the-CH 2-group by-C (O) -in the heterocyclic group include, but are not limited to, 2-oxopyrrolidinyl, oxo-1, 3-thiazolidinyl, 2-piperidinonyl, 3, 5-dioxopiperidinyl and pyrimidinedione. Examples of the sulfur atom in the heterocyclic group being oxidized include, but are not limited to, sulfolane group, 1-dioxothiomorpholinyl group. Said heterocyclyl group of 4 to 7 atoms may be optionally substituted by one or more substituents as described herein.

In another embodiment, heterocyclyl is a 4-atom heterocyclyl and refers to a saturated or partially unsaturated monocyclic ring containing 4 ring atoms in which at least one ring atom is substituted by a member selected from the group consisting of nitrogen, sulfur, and oxygen atoms. Unless otherwise indicated, a heterocyclic group consisting of 4 atoms may be carbon-or nitrogen-based, and the-CH 2-group may optionally be replaced by-c (o) -. The sulfur atom of the ring may optionally be oxidized to the S-oxide. The nitrogen atom of the ring may optionally be oxidized to an N-oxygen compound. Examples of heterocyclic groups consisting of 4 atoms include, but are not limited to: azetidinyl, oxetanyl, thietanyl. The 4-atom heterocyclyl group may be optionally substituted with one or more substituents described herein.

In another embodiment, heterocyclyl is a 5 atom heterocyclyl and refers to a saturated or partially unsaturated monocyclic ring containing 5 ring atoms, wherein at least one ring atom is selected from the group consisting of nitrogen, sulfur, and oxygen atoms. Unless otherwise indicated, a heterocyclic group consisting of 5 atoms may be carbon-or nitrogen-based, and the-CH 2-group may optionally be replaced by-c (o) -. The sulfur atom of the ring may optionally be oxidized to the S-oxide. The nitrogen atom of the ring may optionally be oxidized to an N-oxygen compound. Examples of 5-atom heterocyclic groups include, but are not limited to: pyrrolidinyl, 2-pyrrolinyl, 3-pyrrolinyl, pyrazolinyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, tetrahydrofuranyl, dihydrofuranyl, tetrahydrothienyl, dihydrothienyl, 1, 3-dioxolanyl, dithiocyclopentyl. Examples of the-CH 2-group substituted by-C (O) -in the heterocyclic group include, but are not limited to, 2-oxopyrrolidinyl, oxo-1, 3-thiazolidinyl. Examples of the sulfur atom in the heterocyclic group being oxidized include, but are not limited to, sulfolane group. The 5-atom heterocyclyl group may be optionally substituted with one or more substituents described herein.

In another embodiment, heterocyclyl is a 6 atom heterocyclyl and refers to a saturated or partially unsaturated monocyclic ring containing 6 ring atoms, wherein at least one ring atom is selected from the group consisting of nitrogen, sulfur, and oxygen atoms. Unless otherwise indicated, a heterocyclic group of 6 atoms may be carbon-or nitrogen-based, and the-CH 2-group may optionally be replaced by-c (o) -. The sulfur atom of the ring may optionally be oxidized to the S-oxide. The nitrogen atom of the ring may optionally be oxidized to an N-oxygen compound. Examples of heterocyclic groups consisting of 6 atoms include, but are not limited to: tetrahydropyranyl, dihydropyranyl, 2H-pyranyl, 4H-pyranyl, tetrahydrothiopyranyl, piperidinyl, morpholinyl, thiomorpholinyl, piperazinyl, dioxanyl, dithianyl, thioxanyl. Examples of substitutions of the-CH 2-group in the heterocyclyl by-C (O) -include, but are not limited to, 2-piperidinonyl, 3, 5-dioxopiperidinyl and pyrimidinedione. Examples of the sulfur atom in the heterocyclic group being oxidized include, but are not limited to, 1, 1-dioxothiomorpholinyl. The 6-atom heterocyclyl group may be optionally substituted with one or more substituents described herein.

In yet another embodiment, heterocyclyl is a 7-12 atom heterocyclyl and refers to a saturated or partially unsaturated spiro-or fused-bicyclic ring containing 7-12 ring atoms in which at least one ring atom is selected from the group consisting of nitrogen, sulfur and oxygen atoms. Unless otherwise indicated, a heterocyclic group of 7-12 atoms may be carbon-or nitrogen-based, and the-CH 2-group may optionally be replaced by-c (o) -. The sulfur atom of the ring may optionally be oxidized to the S-oxide. The nitrogen atom of the ring may optionally be oxidized to an N-oxygen compound. Examples of heterocyclic groups consisting of 7 to 12 atoms include, but are not limited to: indolinyl, 1,2,3, 4-tetrahydroisoquinolinyl, 1, 3-benzodioxolyl, 2-oxa-5-azabicyclo [2.2.1] hept-5-yl. Said heterocyclyl group of 7 to 12 atoms may be optionally substituted by one or more substituents as described herein.

The terms "fused bicyclic ring," "fused bicyclic group," and "fused ring group" are used interchangeably herein and all refer to a monovalent or multivalent saturated or partially unsaturated bridged ring system, which refers to a non-aromatic bicyclic ring system. Such systems may contain independent or conjugated unsaturated systems, but the core structure does not contain aromatic or heteroaromatic rings (although aromatic groups may be substituted thereon).

The terms "spirocyclic", "spiro", "spirobicyclic" or "spirobicyclic" are used interchangeably herein to refer to a monovalent or multivalent saturated or partially unsaturated ring system in which one ring is derived from a specific ring carbon atom on another ring. For example, as described below, one saturated bridged ring system (rings B and B') is referred to as "fused bicyclic ring", while ring a and ring B share one carbon atom in two saturated ring systems, referred to as "spiro" or "spirobicyclic ring". Each ring in the fused bicyclic and spirobicyclic groups may be a carbocyclic or heterocyclic group, and each ring is optionally substituted with one or more substituents described herein.

The term "heterocycloalkyl" refers to a monovalent or polyvalent saturated monocyclic, bicyclic, or tricyclic ring system containing 3 to 12 ring atoms, wherein at least one ring atom is selected from nitrogen, sulfur, or oxygen atoms.

The term "n-atomic" where n is an integer typically describes the number of ring-forming atoms in a molecule in which the number of ring-forming atoms is n. For example, piperidinyl is a heterocycloalkyl group of 6 atoms, while 1,2,3, 4-tetrahydronaphthalene is a cycloalkyl group of 10 atoms.

The term "unsaturated" as used herein means that the group contains one or more unsaturations.

The term "heteroatom" refers to O, S, N, P and Si, including N, S and any oxidation state form of P; primary, secondary, tertiary amines and quaternary ammonium salt forms; or a form in which a hydrogen on a nitrogen atom in the heterocycle is substituted, for example, N (like N in 3, 4-dihydro-2H-pyrrolyl), NH (like NH in pyrrolidinyl) or NR (like NR in N-substituted pyrrolidinyl).

The term "halogen" refers to fluorine (F), chlorine (Cl), bromine (Br) or iodine (I).

The term "aryl" denotes monocyclic, bicyclic and tricyclic carbon ring systems containing 6 to 14 ring atoms, or 6 to 12 ring atoms, or 6 to 10 ring atoms, wherein at least one ring system is aromatic, wherein each ring system comprises a ring of 3 to 7 atoms with one or more attachment points to the rest of the molecule. The term "aryl" may be used interchangeably with the term "aromatic ring". Examples of the aryl group may include phenyl, naphthyl, and anthracene. The aryl group may independently be optionally substituted with one or more substituents described herein.

The term "heteroaryl" denotes monocyclic, bicyclic and tricyclic ring systems containing 5 to 12 ring atoms, or 5 to 10 ring atoms, or 5 to 6 ring atoms, wherein at least one ring system is aromatic and at least one ring system contains one or more heteroatoms, wherein each ring system contains a ring of 5 to 7 atoms with one or more attachment points to the rest of the molecule. The term "heteroaryl" may be used interchangeably with the terms "heteroaromatic ring" or "heteroaromatic compound". The heteroaryl group is optionally substituted with one or more substituents described herein. In one embodiment, a heteroaryl group of 5-10 atoms contains 1,2,3, or 4 heteroatoms independently selected from O, S, and N.

Examples of heteroaryl groups include, but are not limited to, 2-furyl, 3-furyl, N-imidazolyl, 2-imidazolyl, 4-imidazolyl, 5-imidazolyl, 3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl, 2-oxazolyl, 4-oxazolyl, 5-oxazolyl, N-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidinyl, 4-pyrimidinyl, 5-pyrimidinyl, pyridazinyl (e.g., 3-pyridazinyl), 2-thiazolyl, 4-thiazolyl, 5-thiazolyl, tetrazolyl (e.g., 5-tetrazolyl), triazolyl (e.g., 2-triazolyl and 5-triazolyl), and the like, 2-thienyl, 3-thienyl, pyrazolyl (e.g., 2-pyrazolyl), isothiazolyl, 1,2, 3-oxadiazolyl, 1,2, 5-oxadiazolyl, 1,2, 4-oxadiazolyl, 1,2, 3-triazolyl, 1,2, 3-thiadiazolyl, 1,3, 4-thiadiazolyl, 1,2, 5-thiadiazolyl, pyrazinyl, 1,3, 5-triazinyl; the following bicyclic rings are also included, but are in no way limited to these: benzimidazolyl, benzofuranyl, benzothienyl, indolyl (e.g., 2-indolyl), purinyl, quinolyl (e.g., 2-quinolyl, 3-quinolyl, 4-quinolyl), isoquinolyl (e.g., 1-isoquinolyl, 3-isoquinolyl, or 4-isoquinolyl), imidazo [1,2-a ] pyridyl, pyrazolo [1,5-a ] pyrimidinyl, imidazo [1,2-b ] pyridazinyl, [1,2,4] triazolo [4,3-b ] pyridazinyl, [1,2,4] triazolo [1,5-a ] pyrimidinyl, [1,2,4] triazolo [1,5-a ] pyridyl, and the like.

The term "carboxy", whether used alone or in combination with other terms, such as "carboxyalkyl", denotes-CO₂H; the term "carbonyl", whether used alone or in combination with other terms, such as "aminocarbonyl" or "acyloxy", denotes- (C ═ O) -.

The term "alkylamino" includes "N-alkylamino" and "N, N-dialkylamino" in which the amino groups are each independently substituted with one or two alkyl groups. In some of these embodiments, the alkylamino group is one or two C_1-6Lower alkylamino groups in which the alkyl group is attached to the nitrogen atom. In other embodiments, the alkylamino group is C_1-3Lower alkylamino groups of (a). Suitable alkylamino groups can be monoalkylamino or dialkylamino, and such examples include, but are not limited to, N-methylamino, N-ethylamino, N-dimethylamino, N-diethylamino, and the like.

The term "arylamino" denotes an amino group substituted with one or two aryl groups, examples of which include, but are not limited to, N-phenylamino. In some embodiments, the aromatic ring on the arylamino group may be further substituted.

The term "aminoalkyl" includes C substituted with one or more amino groups_1-10A straight or branched alkyl group. In some of these embodiments, aminoalkyl is C substituted with one or more amino groups_1-6Examples of "lower aminoalkyl" radicals include, but are not limited to, aminomethyl, aminoethyl, aminopropyl, aminobutyl, and aminohexyl.

The term "prodrug", as used herein, represents a compound that is converted in vivo to a compound of formula (I). Such conversion is effected by hydrolysis of the prodrug in the blood or by enzymatic conversion to the parent structure in the blood or tissue. The prodrug compound of the invention can be ester, and in the prior invention, the ester can be used as the prodrug and comprises phenyl ester and aliphatic (C)_1-24) Esters, acyloxymethyl esters, carbonates, carbamates and amino acid esters. For example, a compound of the present invention contains a hydroxy group, i.e., it can be acylated to provide the compound in prodrug form. Other prodrug forms include phosphate esters, such as those obtained by phosphorylation of a hydroxyl group on the parent. For a complete discussion of prodrugs, reference may be made to the following: T.Higuchi and V.Stella, Pro-drugs as Novel Delivery Systems, Vol.14of the A.C.S.Symphosis Series, Edward B.Roche, ed., Bioreversible Carriers in Drug designs, American Pharmaceutical Association and Pergamon Press,1987, J.Rautio et al, Prodrugs in Design and Clinical Applications, Nature Review Delivery, 2008,7,255 and 270, S.J.Herer et al, Prodrugs of pharmaceuticals and pharmaceuticals, Journal of chemical Chemistry,2008,51,2328 and 5.

"metabolite" refers to the product of a particular compound or salt thereof obtained by metabolism in vivo. Metabolites of a compound can be identified by techniques well known in the art, and its activity can be characterized by assay methods as described herein. Such products may be obtained by administering the compound by oxidation, reduction, hydrolysis, amidation, deamidation, esterification, defatting, enzymatic cleavage, and the like. Accordingly, the present invention includes metabolites of compounds, including metabolites produced by contacting a compound of the present invention with a mammal for a sufficient period of time.

As used herein, "pharmaceutically acceptable salts" refer to organic and inorganic salts of the compounds of the present invention. Pharmaceutically acceptable salts are well known in the art, as are: berge et al, description of the scientific acceptable salts in detail in J. pharmaceutical Sciences,1977,66:1-19. Pharmaceutically acceptable non-toxic acid salts include, but are not limited to, inorganic acid salts formed by reaction with amino groups such as hydrochloride, hydrobromide, phosphate, sulfate, perchlorate, and organic acid salts such as acetate, oxalate, maleate, tartrate, citrate,succinate, malonate, or by other methods described in the literature, such as ion exchange. Other pharmaceutically acceptable salts include adipates, alginates, ascorbates, aspartates, benzenesulfonates, benzoates, bisulfates, borates, butyrates, camphorates, camphorsulfonates, cyclopentylpropionates, digluconates, dodecylsulfates, ethanesulfonates, formates, fumarates, glucoheptonates, glycerophosphates, gluconates, hemisulfates, heptanoates, hexanoates, hydroiodides, 2-hydroxy-ethanesulfonates, lactobionates, lactates, laurates, malates, malonates, methanesulfonates, 2-naphthalenesulfonates, nicotinates, nitrates, oleates, palmitates, pamoates, pectinates, persulfates, 3-phenylpropionates, picrates, pivalates, propionates, stearates, thiocyanate, p-toluenesulfonate, undecanoate, valerate, and the like. Salts obtained with appropriate bases include alkali metals, alkaline earth metals, ammonium and N⁺(C_1-4Alkyl radical)₄A salt. The present invention also contemplates quaternary ammonium salts formed from compounds containing groups of N. Water-soluble or oil-soluble or dispersion products can be obtained by quaternization. Alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like. Pharmaceutically acceptable salts further include suitable, non-toxic ammonium, quaternary ammonium salts and amine cations resistant to formation of counterions, such as halides, hydroxides, carboxylates, sulfates, phosphates, nitrates, C_1-8Sulfonates and aromatic sulfonates.

"solvate" of the present invention refers to an association of one or more solvent molecules with a compound of the present invention. Solvents that form solvates include, but are not limited to, water, isopropanol, ethanol, methanol, dimethyl sulfoxide, ethyl acetate, acetic acid, and aminoethanol. The term "hydrate" refers to an association of solvent molecules that is water.

The term "treating" or "treatment" as used herein refers, in some embodiments, to ameliorating a disease or disorder (i.e., slowing or arresting or reducing the development of the disease or at least one clinical symptom thereof). In other embodiments, "treating" or "treatment" refers to moderating or improving at least one physical parameter, including physical parameters that may not be perceived by the patient. In other embodiments, "treating" or "treatment" refers to modulating the disease or disorder, either physically (e.g., stabilizing a perceptible symptom) or physiologically (e.g., stabilizing a parameter of the body), or both. In other embodiments, "treating" or "treatment" refers to preventing or delaying the onset, occurrence, or worsening of a disease or disorder.

Pharmaceutically acceptable acid addition salts may be formed with inorganic and organic acids, for example, acetate, aspartate, benzoate, benzenesulfonate, bromide/hydrobromide, bicarbonate/carbonate, bisulfate/sulfate, camphorsulfonate, chloride/hydrochloride, chlorotheophylline, citrate, edisylate, fumarate, glucoheptonate, gluconate, glucuronate, hippurate, hydroiodide, isethionate, lactate, lactobionate, lauryl sulfate, malate, maleate, malonate, mandelate, methanesulfonate, methylsulfate, naphthoate, naphthalenesulfonate, nicotinate, nitrate, octadecanoate, oleate, oxalate, palmitate, pamoate, phosphate/biphosphate/dihydrogen phosphate, dihydrogenphosphate, Polysilonolactates, propionates, stearates, succinates, sulfosalicylates, tartrates, tosylates and trifluoroacetates.

Inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like.

Organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, sulfosalicylic acid, and the like.

Pharmaceutically acceptable base addition salts can be formed with inorganic and organic bases.

Inorganic bases from which salts can be derived include, for example, ammonium salts and metals of groups I to XII of the periodic table. In certain embodiments, the salts are derived from sodium, potassium, ammonium, calcium, magnesium, iron, silver, zinc, and copper; particularly suitable salts include ammonium, potassium, sodium, calcium and magnesium salts.

Organic bases from which salts can be derived include primary, secondary and tertiary amines, and substituted amines include naturally occurring substituted amines, cyclic amines, basic ion exchange resins, and the like. Some organic amines include, for example, isopropylamine, benzathine (benzathine), choline salts (cholinate), diethanolamine, diethylamine, lysine, meglumine (meglumine), piperazine, and tromethamine.

The pharmaceutically acceptable salts of the present invention can be synthesized from the parent compound, basic or acidic moiety, by conventional chemical methods. In general, such salts can be prepared by reacting the free acid forms of these compounds with a stoichiometric amount of the appropriate base (e.g., Na, Ca, Mg, or K hydroxide, carbonate, bicarbonate, etc.), or by reacting the free base forms of these compounds with a stoichiometric amount of the appropriate acid. Such reactions are usually carried out in water or an organic solvent or a mixture of both. Generally, where appropriate, it is desirable to use a non-aqueous medium such as diethyl ether, ethyl acetate, ethanol, isopropanol or acetonitrile. In, for example, "Remington's Pharmaceutical Sciences", 20 th edition, Mack Publishing Company, Easton, Pa., (1985); and "handbook of pharmaceutically acceptable salts: properties, Selection and application (Handbook of Pharmaceutical Salts: Properties, Selection, and Use) ", Stahl and Wermuth (Wiley-VCH, Weinheim, Germany, 2002) may find some additional lists of suitable Salts.

In addition, the compounds disclosed herein, including their salts, may also be obtained in the form of their hydrates or in the form of solvents containing them (e.g., ethanol, DMSO, etc.), for their crystallization. The compounds disclosed herein may form solvates with pharmaceutically acceptable solvents (including water), either inherently or by design; thus, the present invention is intended to include both solvated and unsolvated forms.

Any formulae given herein are also intended to represent the non-isotopically enriched forms of these compounds as well as isotopically enriched formsIn the form of (1). Isotopically enriched compounds have the structure depicted by the formulae given herein, except that one or more atoms are replaced by an atom having a selected atomic mass or mass number. Exemplary isotopes that can be incorporated into compounds of the invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine and chlorine, such as²H，³H，¹¹C，¹³C，¹⁴C，¹⁵N，¹⁷O，¹⁸O，¹⁸F，³¹P，³²P，³⁵S，³⁶Cl and¹²⁵I。

in another aspect, the compounds of the invention include isotopically enriched compounds as defined herein, e.g. wherein a radioisotope, e.g. is present³H，¹⁴C and¹⁸those compounds of F, or in which a non-radioactive isotope is present, e.g.²H and¹³C. the isotopically enriched compounds can be used for metabolic studies (use)¹⁴C) Reaction kinetics study (using, for example²H or³H) Detection or imaging techniques such as Positron Emission Tomography (PET) or Single Photon Emission Computed Tomography (SPECT) including drug or substrate tissue distribution determination, or may be used in radiotherapy of a patient.¹⁸F-enriched compounds are particularly desirable for PET or SPECT studies. Isotopically enriched compounds of formula I or formula II can be prepared by conventional techniques known to those skilled in the art or by the procedures and examples described in the present specification using a suitable isotopically labelled reagent in place of the original used unlabelled reagent.

In addition, heavier isotopes are, in particular, deuterium (i.e.,²substitution of H or D) may provide certain therapeutic advantages resulting from greater metabolic stability. For example, increased in vivo half-life or decreased dosage requirements or improved therapeutic index. The concentration of such heavier isotopes, particularly deuterium, can be defined by isotopic enrichment factors. If a substituent of a compound of the invention is designated as deuterium, the compound has at least 3500 (52.5% deuterium incorporation at each designated deuterium atom), at least 4000 (60% deuterium incorporation) for each designated deuterium atom) An isotopic enrichment factor of at least 4500 (67.5% deuterium incorporation), at least 5000 (75% deuterium incorporation), at least 5500 (82.5% deuterium incorporation), at least 6000 (90% deuterium incorporation), at least 6333.3 (95% deuterium incorporation), at least 6466.7 (97% deuterium incorporation), at least 6600 (99% deuterium incorporation), or at least 6633.3 (99.5% deuterium incorporation). Pharmaceutically acceptable solvates of the invention include those in which the crystallization solvent may be isotopically substituted, e.g. D₂O, acetone-d₆、DMSO-d₆Those solvates of (a).

In another aspect, the invention relates to intermediates for the preparation of compounds encompassed by formula I or formula II.

In another aspect, the invention relates to a process for the preparation, isolation and purification of a compound encompassed by formula I or formula II.

In another aspect, the present invention provides a pharmaceutical composition comprising a compound of the present invention, a pharmaceutically acceptable carrier, excipient, diluent, adjuvant, vehicle, or combination thereof. In some embodiments, the pharmaceutical composition may be in a liquid, solid, semi-solid, gel, or spray dosage form.

"combination" means a fixed combination or a kit of parts for combined administration in the form of a single dosage unit, wherein a compound disclosed in the invention and a combination partner may be administered separately at the same time or may be administered separately within certain time intervals, in particular such that the combination partners show a cooperative, e.g. synergistic, effect. Such as the terms "co-administration" or "co-administration" and the like are intended to encompass administration of the selected combination partners to a single individual in need thereof (e.g., a patient), and are intended to encompass treatment regimens in which the substances are not necessarily administered by the same route of administration or simultaneously. The term "pharmaceutical combination" as used herein denotes a product resulting from mixing or combining more than one active ingredient and includes both fixed and non-fixed combinations of active ingredients. The term "fixed combination" means that the active ingredients, such as the compounds disclosed herein and the combination partners, are administered to the patient simultaneously in the form of a single entity or dosage. The term "non-fixed combination" means that the active ingredients, such as the disclosed compounds and combination partners, are both administered to a patient as separate entities simultaneously, jointly or sequentially with no specific time limits, wherein the administration provides therapeutically effective levels of both compounds in the patient.

The experimental procedures used in the following examples are all conventional procedures unless otherwise specified.

Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.

The Pomalidomide terminal derivatives used in the following examples were prepared according to the methods disclosed in the literature Chemistry & Biology 22, 755-. Lenalidomide terminal derivatives were prepared according to the method disclosed in document j.med.chem (DOI:10.1021/acs.jmedchem.6b01816) and RG-7112 terminal carboxylic acid derivatives were prepared according to the methods disclosed in documents bioorg.med.chem.lett.18,5904-5908(2008) and ACS med.chem.lett.4,466-469(2013).

The Ibrutinib terminal derivatives used in the following examples were prepared as follows: the terminal alkynes required for Click chemistry were attached to Ibrutinib intermediate (cas: 1022150-12-4) by amide condensation. The preparation process comprises the following steps:

(1) preparation of intermediate 1a

A25 mL round bottom flask was charged with 193mg Ibrutinib intermediate (cas: 1022150-12-4), 3mL anhydrous chloroform and 31. mu.L propiolic acid. 103mg of DCC was dissolved in 2mL of anhydrous chloroform, and the reaction solution was stirred at 0 ℃ while the anhydrous chloroform solution of DCC was slowly added to the reaction solution, and stirred at 0 ℃ for 1 hour. Cooling the reaction solution to-20 ℃, filtering, keeping the filtrate, repeating the filtering twice, spin-drying the filtrate, and separating and purifying by using a 200-mesh and 300-mesh silica gel chromatographic column, wherein the mobile phase is dichloromethane: methanol 60: 1, obtaining the intermediate 1a with the yield of 72 percent.

Preparation of intermediate 1b

A25 mL round bottom flask was charged with 193mg Ibrutinib intermediate (cas: 1022150-12-4), 3mL anhydrous chloroform and 42mg propiolic acid. 103mg of DCC was dissolved in 2mL of anhydrous chloroform, and the reaction solution was stirred at 0 ℃ while the anhydrous chloroform solution of DCC was slowly added to the reaction solution, and stirred at 0 ℃ for 1 hour. Cooling the reaction solution to-20 ℃, filtering, keeping the filtrate, repeating the filtering twice, spin-drying the filtrate, and separating and purifying by using a 200-mesh and 300-mesh silica gel chromatographic column, wherein the mobile phase is dichloromethane: methanol 60: 1, obtaining an intermediate 1b with the yield of 62 percent.

Preparation of intermediate 1c

A25 mL round bottom flask was charged with 193mg Ibrutinib intermediate (cas: 1022150-12-4), 3mL anhydrous DMF, 75mg HOBT, 106mg EDCI, 49mg pentynoic acid, 77. mu.L triethylamine and 10mg DMAP. The reaction solution was stirred at room temperature for 24 hours. The reaction was quenched by addition of 15mL of saturated aqueous sodium chloride solution, the mixture was extracted three times with 15mL of × 3 ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, the solvent was dried, and the separation and purification were carried out using a 200-mesh 300-mesh silica gel column, the mobile phase was dichloromethane: 40 parts of methanol: 1, obtaining an intermediate 1c with the yield of 61%.

Preparation of intermediate 1d

A25 mL round bottom flask was charged with 193mg Ibrutinib intermediate (cas: 1022150-12-4), 3mL anhydrous DMF, 75mg HOBT, 106mg EDCI, 56mg hexynoic acid, 77. mu.L triethylamine and 10mg DMAP. The reaction solution was stirred at room temperature for 24 hours. The reaction was quenched by addition of 15mL of saturated aqueous sodium chloride solution, the mixture was extracted three times with 15mL of × 3 ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, the solvent was dried, and the separation and purification were carried out using a 200-mesh 300-mesh silica gel column, the mobile phase was dichloromethane: 40 parts of methanol: 1, obtaining an intermediate 1d with the yield of 71 percent.

(2) Preparation of intermediate 2a

In a 100mL round bottom flask were placed 2.67mL of 3-aminopropan-1-ol and 50mL of anhydrous dichloromethane, the reaction was cooled to 0 ℃ and 6.8mL of triethylamine and 9.7mL of Boc were added₂And O. The reaction solution was allowed to spontaneously return to room temperature with stirring, and stirring was continued for 12 hours. The reaction was quenched by addition of 100mL of saturated aqueous sodium bicarbonate, the mixture was extracted three times with 50mL × 3 dichloromethane, the organic phases were combined, dried over anhydrous sodium sulfate, the solvent was dried, and the separation and purification were performed using a 200-: methanol 20: 1, obtaining the intermediate 2a with the yield of 72 percent.

Preparation of intermediate 2b

In a 100mL round bottom flask was added 3.22mL of 4-aminobutan-1-ol and 50mL of anhydrous dichloromethane, the reaction was cooled to 0 ℃ and 6.8mL of triethylamine and 9.7mL of Boc were added₂And O. The reaction solution was allowed to spontaneously return to room temperature with stirring, and stirring was continued for 12 hours. The reaction was quenched by the addition of 100mL of saturated aqueous sodium bicarbonate solution, the mixture was extracted three times with 50 mL. times.3 dichloromethane, the organic phases were combined, dried over anhydrous sodium sulfate,spin-drying the solvent, and separating and purifying by using a 200-mesh 300-mesh silica gel chromatographic column, wherein the mobile phase is dichloromethane: methanol 20: 1, obtaining the intermediate 2b with the yield of 81 percent.

Preparation of intermediate 2c

A100 mL round bottom flask was charged with 1.6g of sodium azide and 25mL of water, and 4.9g of 2- (2-hydroxyethoxy) ethyl-4-methylbenzenesulfonate was added. The reaction was stirred at 90 ℃ for 24 hours. The reaction was quenched by the addition of 50mL of saturated aqueous sodium bicarbonate, the mixture was extracted three times with 40mL × 3 dichloromethane, the organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was spin dried to give the intermediate in 46% yield. A50 mL round-bottom flask was charged with 787mg of the obtained product, 8mL of anhydrous dichloromethane and 1.67mL of triethylamine, and a solution of 1.72g of TsCl in anhydrous dichloromethane was slowly added to the above reaction solution with stirring, followed by stirring at room temperature for 24 hours. Quenching the reaction by adding 100mL of saturated aqueous sodium bicarbonate solution, extracting the mixture with 50mL × 3 dichloromethane for three times, combining the organic phases, drying with anhydrous sodium sulfate, spin-drying the solvent, and separating and purifying by using a 200-mesh 300-mesh silica gel chromatographic column, wherein the mobile phase is petroleum ether: ethyl acetate ═ 5: 1, obtaining the intermediate 2c with the yield of 67%.

Preparation of intermediate 2d

526mg of intermediate 2a and 15mL of anhydrous THF were added to a 50mL round-bottom flask, 150mg of NaH (60% dispersed in mineral oil) was slowly added with stirring at 0 deg.C, the reaction solution was stirred at 0 deg.C for 1 hour, 285mg of intermediate 2c in anhydrous THF was slowly added to the reaction solution, the reaction solution was naturally warmed to room temperature with stirring, and stirring was continued for 15 hours. Dropwise adding methanol into the reaction solution for quenching reaction, spin-drying the solvent, and separating and purifying by using a 200-plus-300-mesh silica gel chromatographic column, wherein the mobile phase is petroleum ether: ethyl acetate ═ 6:1, obtaining the intermediate with the yield of 26 percent. In a 25mL round-bottom flask, 230mg of the obtained product, 5mL of dichloromethane and 0.5mL of trifluoroacetic acid were added, and the mixture was stirred at room temperature for 6 hours. Adding 15mL of toluene, spin-drying the solvent, and separating and purifying by using a 200-mesh 300-mesh silica gel chromatographic column, wherein the mobile phase is dichloromethane: methanol 10: 1, obtaining the intermediate 2d with the yield of 62 percent.

Preparation of intermediate 2e

A100 mL round bottom flask was charged with 2.8g of intermediate 2b and 50mL of anhydrous THF, 800mg of NaH (60% dispersed in mineral oil) was added slowly with stirring at 0 deg.C, the reaction was stirred at 0 deg.C for 1 hour, 1.43g of intermediate 2c in anhydrous THF was added slowly to the reaction, the reaction was allowed to spontaneously warm to room temperature with stirring, and stirring was continued for 15 hours. Dropwise adding methanol into the reaction solution for quenching reaction, spin-drying the solvent, and separating and purifying by using a 200-plus-300-mesh silica gel chromatographic column, wherein the mobile phase is petroleum ether: ethyl acetate ═ 6:1, obtaining the intermediate with the yield of 22 percent. In a 25mL round bottom flask, 800mg of the resulting product, 15mL of dichloromethane and 1.5mL of trifluoroacetic acid were added and stirred at room temperature for 2 hours. Adding 25mL of toluene, spin-drying the solvent, and separating and purifying by using a 200-mesh 300-mesh silica gel chromatographic column, wherein the mobile phase is dichloromethane: methanol 10: 1, obtaining the intermediate 2e with the yield of 66 percent.

(3) Preparation of intermediate 4a

A5 mL round bottom flask was charged with 24mg of 8-aminoctanoic acid, 70. mu.L of DIEA, 0.3mL of DMF, and 35mg of 2- (2, 6-dioxyperidin-3-yl) -4-fluorooisolinone-1, 3-dione. The reaction solution was stirred for 3 hours at 90 ℃. The reaction was quenched by addition of 5mL of saturated aqueous sodium chloride, the mixture was extracted three times with 5mL of × 3 ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, the solvent was dried, and the separation and purification were carried out using a 200-: 40 parts of methanol: 1, obtaining the intermediate 4a with the yield of 22 percent.

Preparation of intermediate 4b

A5 mL round bottom flask was charged with 26mg of 9-aminoctanoic acid, 70. mu.L of DIEA, 0.3mL of DMF, and 35mg of 2- (2, 6-diozoperidin-3-yl) -4-fluorooisolinone-1, 3-dione. The reaction solution was stirred for 3 hours at 90 ℃. The reaction was quenched by addition of 5mL of saturated aqueous sodium chloride, the mixture was extracted three times with 5mL of × 3 ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, the solvent was dried, and the separation and purification were carried out using a 200-: 40 parts of methanol: 1, obtaining the intermediate 4b with the yield of 31 percent.

(4) Preparation of intermediate 5a

A100 mL round bottom flask was charged with 3g octane-1,8-diol, 20mL anhydrous THF, and 1.3g potassium tert-butoxide. The reaction mixture was stirred for 15 minutes under argon atmosphere at room temperature, and 1.2g of 3-bromopropyne was added dropwise. The reaction solution was stirred for 12 hours while maintaining the room temperature under argon atmosphere. Spin-drying the solvent, adding 30mL of saturated aqueous sodium chloride solution to quench the reaction, adjusting the pH to 3-5 with 1M HCl, extracting the mixture with 30mL × 3 dichloromethane three times, combining the organic phases, drying with anhydrous sodium sulfate, spin-drying the solvent, and separating and purifying with a 200-mesh 300-mesh silica gel chromatography column, wherein the mobile phase is petroleum ether: ethyl acetate ═ 3: 1, obtaining the intermediate 5a with the yield of 55 percent.

Preparation of intermediate 5b

A100 mL round bottom flask was charged with 12.6g of 2, 2' - (ethane-1,2-diylbis (oxy)) bis (ethane-1-ol), 50mL of anhydrous THF, and 5g of potassium tert-butoxide. The reaction mixture was stirred for 15 minutes under argon atmosphere at room temperature, and 3.5mL of 3-bromopropyne was added dropwise. The reaction solution was stirred for 12 hours while maintaining the room temperature under argon atmosphere. Spin-drying the solvent, adding 30mL of saturated aqueous sodium chloride solution to quench the reaction, adjusting the pH to 3-5 with 1M HCl, extracting the mixture with 30mL × 3 dichloromethane three times, combining the organic phases, drying with anhydrous sodium sulfate, spin-drying the solvent, and separating and purifying with a 200-mesh 300-mesh silica gel chromatography column, wherein the mobile phase is petroleum ether: ethyl acetate ═ 3: 1, obtaining the intermediate 5b with the yield of 77%.

Preparation of intermediate 5c

A100 mL round-bottom flask was charged with 4.4mL pentane-1,5-diol, 30mL anhydrous THF, and 2.5g potassium tert-butoxide. The reaction mixture was stirred for 15 minutes under argon atmosphere at room temperature, and 1.73mL of 3-bromopropyne was added dropwise. The reaction solution was stirred for 12 hours while maintaining the room temperature under argon atmosphere. Spin-drying the solvent, adding 30mL of saturated aqueous sodium chloride solution to quench the reaction, adjusting the pH to 3-5 with 1M HCl, extracting the mixture with 30mL × 3 dichloromethane three times, combining the organic phases, drying with anhydrous sodium sulfate, spin-drying the solvent, and separating and purifying with a 200-mesh 300-mesh silica gel chromatography column, wherein the mobile phase is petroleum ether: ethyl acetate ═ 3: 1, obtaining the intermediate 5c with the yield of 74 percent.

(4) Preparation of intermediate 6a

276mg of acetylene lithium ethylenediamine complex is added into a 25mL round-bottom flask, dissolved in 8mLDMSO, placed into an ice bath to be cooled to 0 ℃, 6-bromohexanoic acid is dissolved in 8mLDMSO, slowly dripped into the reaction solution, stirred in the ice bath for 10 minutes continuously, slowly heated to room temperature and stirred for 2 hours. Pouring the reaction solution into ice water, acidifying with 1M hydrochloric acid, extracting the mixture for three times with 30mL multiplied by 3 dichloromethane, combining the organic phases, drying by using anhydrous sodium sulfate, spin-drying the solvent, and separating and purifying by using a 200-plus-300-mesh silica gel chromatographic column, wherein the mobile phase is petroleum ether: ethyl acetate 10: 1, obtaining the intermediate 6a with the yield of 35%.

Preparation of intermediate 6b

In a 250mL round bottom flask, 7.3g of methyl 3-bromoo-2-methylbenezoate, 6.5g of NBS and 529mg of AIBN were added, dissolved in 100mL of dry chloroform, and then refluxed at 90 ℃ for 5 hours. After the solvent was spin-dried, 60mL of saturated aqueous sodium chloride solution was added to quench the reaction, the mixture was extracted three times with 60mL of × 3 dichloromethane, the organic phases were combined, dried over anhydrous sodium sulfate, and the solvent was spin-dried to obtain an intermediate. 7.6g of the intermediate thus obtained was mixed with 5.3g of 2, 6-dioxolane-3-ammonium chloride, 4.6mL of Et₃N, 90mL of dry acetonitrile was added, and the reaction mixture was stirred at 80 ℃ for 10 hours. Spin-drying the solvent, and separating and purifying by using a 200-mesh 300-mesh silica gel chromatographic column, wherein the mobile phase is dichloromethane: methanol 20: 1-5: 1, obtaining the intermediate 6b with the yield of 60 percent.

Preparation of intermediate 6c

A25 mL round bottom flask was charged with 35mg of intermediate 6b, 30mg of intermediate 6a, 5mg of cuprous iodide, 15mg of Pd (dppf)₂Cl₂6mL DMF and 3mL Et₃And N, reacting for 3 hours at 80 ℃ under the protection of nitrogen. Spin-drying the solvent, and separating and purifying by using a 200-mesh 300-mesh silica gel chromatographic column, wherein the mobile phase is dichloromethane: 25 parts of methanol: 1, obtaining the intermediate 6c with the yield of 82%.

Preparation of intermediate 6d

A25 mL round bottom flask was charged with 35mg of intermediate 6b, 30mg of intermediate 6a, 5mg of cuprous iodide, 15mg of Pd (dppf)₂Cl₂6mL DMF and 3mL Et₃And N, reacting for 3 hours at 80 ℃ under the protection of nitrogen. Spin-drying the solvent, and separating and purifying by using a 200-mesh 300-mesh silica gel chromatographic column, wherein the mobile phase is dichloromethane: 25 parts of methanol: 1, obtaining the intermediate 6d with the yield of 81 percent.

Example 1 preparation of Compounds represented by formula 1 to formula 18

A10 mL round bottom flask was charged with 50mg of Ibrutinib intermediate (cas: 1022150-12-4), 56mg of intermediate 4b, 21mg of HOBT, 30mg of EDCI, 20. mu.l of Et₃N, 2mg DMAP and 2ml DMF were dissolved and stirred at room temperature for 12 hours. 10mL of saturated aqueous sodium chloride solution was added to the reaction solution, the mixture was extracted with 10mL of X3 ethyl acetate three times, the organic phases were combined, dried over anhydrous sodium sulfate, the solvent was dried by spinning, and the separation and purification were carried out using a 200-mesh 300-mesh silica gel column, and the mobile phase was dichloromethane: 40 parts of methanol: 1, obtaining the formula 10 with the yield of 85%.

A10 mL round bottom flask was charged with 50mg of Ibrutinib intermediate (cas: 1022150-12-4), 54mg of intermediate 4a, 21mg of HOBT, 30mg of EDCI, 20. mu.l of Et₃N, 2mg DMAP and 2ml DMF were dissolved and stirred at room temperature for 12 hours. 10mL of saturated aqueous sodium chloride solution was added to the reaction solution, the mixture was extracted with 10mL of X3 ethyl acetate three times, the organic phases were combined, dried over anhydrous sodium sulfate, the solvent was dried by spinning, and the separation and purification were carried out using a 200-mesh 300-mesh silica gel column, and the mobile phase was dichloromethane: 40 parts of methanol: 1, obtaining the formula 12 with the yield of 86%.

A10 mL round bottom flask was charged with 30mg of Ibrutinib intermediate (cas: 1022150-12-4), 30mg of intermediate 6d, 13mg of HOBT, 18mg of EDCI, 15. mu.l of Et₃N, 6mg DMAP and 2ml DMF were dissolved and stirred at room temperature for 12 hours. 10mL of saturated aqueous sodium chloride solution was added to the reaction solution, the mixture was extracted with 10mL of X3 ethyl acetate three times, the organic phases were combined, dried over anhydrous sodium sulfate, the solvent was dried by spinning, and the separation and purification were carried out using a 200-mesh 300-mesh silica gel column, and the mobile phase was dichloromethane: methanol 30: 1, obtaining an intermediate, dissolving the intermediate in 4mL of methanol and 1mL of DMF, adding 20mg of palladium carbon, and reacting for 8 hours at 38 ℃ in hydrogen. Filtering the reaction solution by using kieselguhr, spin-drying the solvent, and separating and purifying by using a 200-mesh and 300-mesh silica gel chromatographic column, wherein the mobile phase is dichloromethane: methanol 30: 1, obtaining the formula 16 with the yield of 72 percent.

A10 mL round bottom flask was charged with 30mg of Ibrutinib intermediate (cas: 1022150-12-4), 30mg of intermediate 6d, 13mg of HOBT, 18mg of EDCI, 15. mu.l of Et₃N, 6mg DMAP and 2ml DMF were dissolved and stirred at room temperature for 12 hours. 10mL of saturated aqueous sodium chloride solution was added to the reaction solution, the mixture was extracted with 10mL of X3 ethyl acetate three times, the organic phases were combined, dried over anhydrous sodium sulfate, the solvent was dried by spinning, and the separation and purification were carried out using a 200-mesh 300-mesh silica gel column, and the mobile phase was dichloromethane: methanol 30: 1, obtaining an intermediate, dissolving the intermediate in 4mL of methanol and 1mL of DMF, adding 20mg of palladium carbon, and reacting for 8 hours at 38 ℃ in hydrogen. Filtering the reaction solution by using kieselguhr, spin-drying the solvent, and separating and purifying by using a 200-mesh and 300-mesh silica gel chromatographic column, wherein the mobile phase is dichloromethane: methanol 30: 1, obtaining the formula 18 with the yield of 69%.

Example 2 biological Activity testing of Compounds at the level of Western Blot

Cell treatment (wild type cells):

ramos or HBL-1 or Mino or IgE MM or Jurkat or HeLa cells in the logarithmic growth phase of Set up are placed in a 6-well plate and treated by adding a compound; the cells were harvested after 48 h.

Cell treatment (C481S or C481A mutant BTK plasmid transiently transfected cells):

293T or HeLa cells in the logarithmic growth phase are placed in a 6-well plate, after 18 hours, the culture solution is replaced after the cells are attached to the wall, after 1.5 hours, 500 microliters of newly prepared Opti-MEM culture medium containing PEI and plasmids is added into each well (the preparation method of the Opti-MEM culture medium containing PEI and plasmids is shown as the following steps), the mixture is uniformly mixed, after 8 hours, the normal culture solution is replaced, a compound is added, and the treatment is carried out for 48 hours; the medium was changed and the compound was added, and the cells were harvested after an additional 48 hours of treatment.

The preparation method of the Opti-MEM culture medium containing PEI and the plasmid comprises the following steps:

adding 0.2 microgram or 0.5 microgram of plasmid into 250 microliter of preheated Opti-MEM culture medium, shaking gently and mixing uniformly, and standing for 5 minutes; adding 0.6 microgram or 1.5 microgram PEI to 250 microliter preheated Opti-MEM culture medium, shaking gently and mixing uniformly, and standing for 5 minutes; slowly adding the PEI-containing Opti-MEM culture medium into the plasmid-containing Opti-MEM culture medium, gently shaking and uniformly mixing while adding, and standing for 30 minutes to obtain the PEI-containing Opti-MEM culture medium.

Extracting cell whole protein:

collecting cells: the treated cells were scraped off in a culture medium, suspended thoroughly, centrifuged at 300g for 5 minutes and collected, washed once with PBS, and the PBS was discarded.

Cell lysis: 150 mul of 2 XLoading Buffer is added into each sample, the mixture is fully shaken and mixed evenly, denatured for 15 minutes at 95 ℃, and the mixture is stored at minus 20 ℃ after being mixed evenly or is directly used for Western Blot detection.

The formula of 5 × Loading Buffer is as follows: 250mM Tris-HCl (pH6.8), 10% (W/V) SDS, 0.5% (W/V) bromophenol blue, 50% (V/V) glycerol, 5% (W/V) beta-mercaptoethanol (2-ME). The 2 XLoading Buffer is prepared by adding 1.5 times volume of dd water into 5 Xloading Buffer.

The Western Blot detection method comprises the following specific steps:

1) SDS-PAGE gels were prepared at appropriate concentrations. A separation gel was prepared at a suitable concentration according to the molecular cloning laboratory Manual (scientific Press, second edition) at page 883, table 18.3, and a 4% concentration concentrated gel was prepared according to page 883, table 18.4.

2) Samples were prepared. Protein samples were prepared according to experimental requirements, denatured at 95 ℃ for 15 minutes, centrifuged, mixed and loaded onto SDS-PAGE gel wells. The loading volume was adjusted appropriately according to the protein quantification results, typically 4. mu.l per well.

3) And (4) electrophoresis. The power supply is switched on, the voltage of the protein sample in the concentrated gel is 83 volts, and when the protein sample enters the separation gel, the voltage is adjusted to 110 volts to continue electrophoresis. The electrophoresis was stopped when bromophenol blue almost completely ran off the PAGE gel.

4) And (5) transferring the film. And (3) taking down the gel after electrophoresis is finished, and installing a film transfer device according to the following sequence: (negative electrode), filter paper, gel, activated PVDF membrane, filter paper, (positive electrode). It is absolutely impossible to bubble between the gel and the PVDF membrane. Then the clamping and transferring device is placed in a film transferring buffer solution, and finally the ice box is placed in a refrigerator with the temperature of 4 ℃ and the constant voltage power-on is carried out for 1.5 hours at 100V.

5) And (5) sealing. After the completion of the membrane transfer, the PVDF membrane was removed, immersed in TBST buffer containing 5% skimmed milk powder, and shaken on a shaker at room temperature for 1 hour.

6) Primary antibody incubation. After blocking, the cells were washed 3 times with TBST buffer, and then primary antibody was added at a moderate dilution rate overnight at 4 ℃. The primary antibody was recovered and the PVDF membrane was washed 3 times with TBST buffer with 10 minutes shaking.

7) And (5) incubating a secondary antibody. The TBST buffer is discarded, secondary antibodies (mouse or rabbit antibodies, determined as primary antibodies) are added at a dilution ratio (usually 1: 3000-1: 5000), and the mixture is shaken in a shaker at room temperature for 1 hour. The secondary antibody was discarded and the PVDF membrane was washed 3 times with TBST buffer with shaking for 10 minutes. Finally, the mixture is washed with TBST buffer for 10 minutes.

8) Color development and tabletting. And (3) uniformly covering the ECL chromogenic substrate on the PVDF membrane, and developing for 0.5-15 minutes at room temperature.

The degradation activity of the compounds according to the examples of the present invention on BTK is as follows:

the compound provided by the embodiment of the invention has strong effect of degrading wild-type BTK, can be obviously degraded under 10-50nM, has no inhibition or degradation effect on other targets such as EGFR, ITK, TEC and the like, has the efficacy of specifically targeting and degrading BTK protein, and partial results are shown in FIG. 4, wherein L18I represents the compound

Example 3 inhibition of BTK, ITK, EGFR by Compounds

1. 1 Xkinase matrix buffer and stop buffer were prepared.

1)1 Xkinase matrix buffer

50mM HEPES,pH 7.5

0.01％Brij-35

2) Stop buffer

100mM HEPES,pH 7.5

0.015％Brij-35

0.2％Coating Reagent#3

50mM EDTA

2. A compound is prepared.

1) Compounds were diluted with 100% DMSO to reach 50X of the highest concentration of inhibitor ultimately desired in the reaction. If compounds were tested at 1. mu.M, a 50. mu.M DMSO solution of compound was prepared in this step.

2) Compounds were serially diluted in 3-fold gradients to give a total of 10 concentrations.

3) 100 μ l of 100% DMSO was added to 2 empty wells of a 96-well plate as a no-compound control and a no-enzyme control.

4) A middle plate is prepared.

10 microliters of compound was transferred from the source plate to a new 96-well plate as an intermediate plate.

90 microliters of 1x kinase buffer was added to each well of the intermediate plate.

Mix the compound in the middle plate on a shaker for 10 minutes

3. An enzyme-linked plate was prepared.

1) Transfer 5 microliters per well of the 96-well intermediate plate to 384-well plates in duplicate. For example, 96-well plate a1 was transferred to 384-well plates a1 and a2, 96-well plate a2 was transferred to 384-well plates A3 and a4, and so on.

4. And (3) kinase reaction.

1) Preparation of 2.5 Xenzyme solution

The enzyme was added to 1x kinase matrix buffer.

2) Preparation of 2.5 Xpeptide solution

FAM-labeled peptide and ATP were added to 1x kinase matrix buffer.

3) The 2.5x enzyme solution was transferred to an enzyme conjugate plate.

4) The enzyme-linked plate already contains 5 microliters of compound in 10% DMSO.

5) 10 microliters of 2.5x enzyme solution was added to each well of the 384-well enzyme-linked plate.

6) Incubate for 10 minutes at room temperature.

7) The 2.5x peptide solution was transferred to an enzyme conjugate plate.

10 microliters of 2.5x peptide solution was added to each well of the 384-well enzyme-linked plate.

8) Kinase reaction and cessation

Incubate at 28 degrees celsius for the indicated time.

The reaction was stopped by adding 25. mu.l of stop buffer.

9) Data was collected on the Caliper.

5. Fitting of curves

1) The translation data is copied from the Caliper program.

2) The converted value is converted into a suppressed value.

The inhibition ratio is (max-conversion value)/(max-min) × 100.

"max" represents DMSO control; "min" represents low control.

3) IC was obtained by fitting the data to XLFit excel add-in version 5.4.0.8₅₀The value is obtained.

Using the equation of

Y＝Bottom+(Top-Bottom)/(1+(LogIC₅₀/X)*HillSlope))

Inhibitory Activity IC of Ibrutinib on BTK₅₀1.4 nM; ITK inhibitory Activity IC of Ibrutinib₅₀34nM, inhibitory Activity IC on EGFR₅₀Was 1.3 nM. Inhibitory Activity of Compounds according to embodiments of the invention on BTK kinase IC₅₀IC for ITK, EGFR of 10-100nM₅₀The values are all higher than 1000nM, which shows that the compounds according to the examples of the invention have no significant inhibitory effect on the target of the side effects of Ibrutinib.

Example 4 inhibition of cell proliferation by Compounds of formula 1-18

MTT test reagent:

reagent: RPIM 1640 medium; DMEM medium; 100 × non-essential amino acids (NEAA); 100 times streptomycin mixed liquor; 50mM beta mercaptoethanol; calf serum (FBS, previously inactivated).

Medium a (500 ml): RPIM 1640medium (450ml) +100 XNEAA (5ml) +100 Xstreptomycin mixed liquor (5ml) + calf serum (50ml) +50mM beta mercaptoethanol (0.5 ml).

B medium (500 ml): DMEM medium (450ml) +100 XNEAA (5ml) +100 Xstreptomycin mixture (5ml) + calf serum (50ml) +50mM beta mercaptoethanol (0.5 ml). .

CCK-8 Kit (Cell Counting Kit-8)

MTT Experimental flow (Ramos cell and HBL-1 cell):

1) cells were collected in log phase and cell suspension concentration was adjusted to 6.6X 10 using A medium⁴/ml。

2) The small molecule concentration was diluted from 100nM to 2nM with a 2-fold gradient of a medium. Preparing into small molecule solution.

3) 45 μ L of cell suspension was added to a 96-well plate (marginal wells filled with sterile PBS, 3000 cells/well). Negative controls (45. mu.L of cell suspension and 45. mu.L of A medium) were set for each plate, and 3 wells were set for each group.

4) Standing at 37 deg.C for 5% CO₂After 1 hour of incubation, 45 μ L of the corresponding small molecule solution was added to each well of the 96-well plate. Then at 37 ℃ with 5% CO₂Incubate for 72-96 hours. In the system, the concentration of the small molecules is diluted in a 2-fold gradient from 50nM to 1 nM.

Add 10. mu.l cck-8 solution per well and incubate for 4 h. The absorbance of each well was measured by direct enzyme-linked immunosorbent assay OD490 nm.

The results of inhibition of cell proliferation by the compounds according to the examples of the present invention measured by the above-described methods are shown in table 1. Table 1: MTT assay (GI50 value) for inhibitory potency of compounds of formula 1-formula 18 on HBL-1cell line and Ramos cell line:

wherein n.d represents no detectable inhibitory activity.

As can be seen from Table 1 above, the compounds according to the examples of the present invention have an inhibitory ability against HBL-1cell proliferation comparable to or even superior to Ibrutinib.

EXAMPLE 5 inhibition of THP-1 Release of inflammatory factors upon LPS stimulation by Compounds

1. LPS was prepared.

Adding RPMI1640 medium without FBS into LPS solid powder to a final concentration of 1mg/ml, and subpackaging and storing.

2. Compounds (see table 2 for compound structures and numbers) were prepared.

1) Compounds were diluted with 100% DMSO to reach 100X of the highest concentration of inhibitor ultimately desired in the reaction. If compounds were tested at 100nM, a 10. mu.M DMSO solution of compound was prepared in this step.

2) The compounds were diluted 100-fold in the medium.

3. The cells are treated with a drug.

1) THP-1 cells were diluted to the required concentration and six well plates were seeded with 1X10^6 cells per well supplemented to 2ml medium.

2) Diluted drugs and DMSO controls were dispensed into corresponding wells at 500ul, while blank controls without drug and DMSO were placed and incubated in an incubator at 37 ℃ for 24 h. Each group was repeated 3 times.

LPS stimulation.

The corresponding volume of 1mg/ml LPS solution was taken, the medium was diluted and added to a six-well plate with a constant concentration of 0.5ug/ml, and incubated for 24 hours.

RT-PCR detection of inflammatory factor levels

1) The treated THP-1 cells were collected, centrifuged at 100g for 5 min, washed 2 times with PBS, and total RNA was extracted from the cells using Trizol method, followed by cDNA synthesis using CWBIO easy quick RT MasterMix as a template for qPCR.

2) PCR primers for detecting IL-1 beta, TNF alpha, IL-6, IL-10 and internal reference GAPDH were designed based on the gene sequence of human proinflammatory cytokine, and fluorescent quantitative PCR was performed using the designed primers (see Table 3 for specific conditions).

The results of the inhibition of the release of the inflammatory cytokines by the compounds of the present invention measured by the above methods are shown in fig. 3 and table 4, and the exemplified compounds down-regulate the RNA levels of IL-1 β, TNF α, IL-6, IL-10 in the cells to different degrees, wherein the inhibition of the release of the inflammatory cytokines by the compounds of formula 10 is most significant, which indicates that the compounds of the present invention can inhibit the BTK pathway and thus the release of the inflammatory cytokines.

Table 2: compound structure and numbering

Table 3: RT-PCR

Wherein, target gene refers to target gene, species, primer name refers to primer name, sequence guide sequence.

Table 4: modulation of inflammatory factor mRNA levels by compounds

	P9I	L8I	mL8I
				IL-1β	down	down	down
TNFα	down	down	down
				IL-6	down	down	down
IL-10	down	down	down

Wherein down indicates down regulation.

Example 6 Effect of Compounds in mouse arthritis, pulmonary hemorrhage model

1. Arthritis (arthritis)

The experimental scheme is as follows: bovine type II Collagen (CII) was thoroughly emulsified with equal volume of Freund's complete adjuvant (CFA) to make an emulsion containing CII 1 mg/mL. Under sterile conditions, 100 mu L of mice are injected into the tail root of each group, after successful model induction, the mice are randomly divided into 4 groups, namely a control group, an ibrutinib group, an exemplary compound 50mg/kg group and an exemplary compound 100mg/kg group, and corresponding medicaments are respectively injected. Each group of mice was scored daily for Arthritis (AI): score 0, normal; 1 point, erythema and slight swelling of the ankle; erythema and slight swelling of the ankle to the metatarsal or metacarpal joints in score 2; 3 points, erythema and moderate swelling of the ankle joint to the metatarsophalangeal joint or the metacarpal joint; erythema and severe swelling of the ankle to toe joints in 4 points; maximum score 16 per mouse.

The experimental results are as follows: ibrutinib significantly improved the symptoms of swelling of the mouse feet, and the exemplary compounds improved the symptoms of arthritis compared to the blank group.

2. Pulmonary hemorrhage

Experimental protocol: b6 mice were divided into healthy group (3), blank group (7), ibrutinib group (7) and exemplified compound group (7), except for healthy group, 1ml of norphytane was intraperitoneally injected uniformly, blank group, ibrutinib group and exemplified compound were injected with vehicle, ibrutinib and exemplified compound respectively starting one week before norphytane was given, intraperitoneally injected, 100mg/kg daily, mice were sacrificed after continuous injection for 18 days after norphytane injection, lungs and spleens were removed, weighed and observed for bleeding in lungs.

The experimental results are as follows: in a mouse pulmonary hemorrhage model, taking ibrutinib as a control, the exemplified compound can effectively improve the symptoms of pulmonary hemorrhage, and the lung weight and the spleen weight are also obviously lower than those of a control combined ibrutinib group.

In the description herein, references to the description of the term "one embodiment," "some embodiments," "an example," "a specific example," or "some examples," etc., mean that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the invention. In this specification, the schematic representations of the terms used above are not necessarily intended to refer to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples. Furthermore, various embodiments or examples and features of different embodiments or examples described in this specification can be combined and combined by one skilled in the art without contradiction.

Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention, and that variations, modifications, substitutions and alterations can be made to the above embodiments by those of ordinary skill in the art within the scope of the present invention.

Claims

1. A compound of any one of formulas 10, 16, and 18, or a pharmaceutically acceptable salt thereof,

2. a pharmaceutical composition comprising a compound of claim 1.

3. The pharmaceutical composition of claim 2, further comprising a pharmaceutically acceptable carrier, excipient, diluent, adjuvant, vehicle, or combination thereof.

4. Use of a compound of claim 1 or a pharmaceutical composition of claim 2 or 3 in the manufacture of a medicament for degrading BTK or inhibiting BTK.

5. Use of a compound of claim 1 or a pharmaceutical composition of claim 2 or 3 in the manufacture of a medicament for treating or preventing a BTK-related disease.

6. The use according to claim 5, wherein the BTK-associated disease is non-Hodgkin's lymphoma or an autoimmune disease.

7. Use according to claim 6, wherein the autoimmune disease is arthritis, pulmonary hemorrhage, systemic lupus erythematosus, pemphigus, chronic lymphocytic thyroiditis, hyperthyroidism, insulin dependent diabetes mellitus, myasthenia gravis, chronic ulcerative colitis, pernicious anemia with chronic atrophic gastritis, primary biliary cirrhosis, multiple sclerosis or acute idiopathic polyneuritis.

8. Use according to claim 6, wherein the autoimmune disease is arthritis or pulmonary hemorrhage.