CN113135943A - Boronic acid derivatives - Google Patents

Abstract

The present invention relates to boronic acid derivatives; the present invention provides compounds of formula (I) or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, pharmaceutical compositions comprising these compounds and the use of such compounds in the treatment of lmp7 related diseases.

Description

Boronic acid derivatives

Technical Field

The patent relates to a novel boric acid derivative shown as a formula (I) or a pharmaceutically acceptable salt thereof. Also provided herein are pharmaceutical compositions containing such compounds and methods of making the same. The compounds described herein are useful in the treatment or prevention of diseases associated with immunoproteasome.

Background

The ubiquitin-proteasome system (UPS) is present in all eukaryotic cells and is responsible for intracellular misfolding or degradation of redundant proteins. By regulating protein level homeostasis, nearly all important vital activities are regulated, such as signal transduction, transcriptional regulation, cell differentiation and apoptosis, etc. The 26S proteasome (the proteasome density gradient centrifugation has a sedimentation coefficient of 26S, and is also referred to as 26S proteasome) can be structurally divided into two parts, namely a 19S regulatory particle and a 20S core particle, wherein the 19S regulatory particle is responsible for identifying and unfolding proteins with ubiquitin markers, and finally conveying the unfolded proteins to the 20S core particle for degradation. The 20S proteasome has a barrel structure consisting of 4 rings. The outer two loops, each containing seven alpha subunits, act as a binding for the regulatory particle on the one hand and act as a "gate" on the other hand, preventing unregulated entry of the protein into the interior of the core particle. The inner two loops, each containing seven β subunits, contain protease active subunits β 1c, β 2c and β 5c for proteolytic reactions. In hematopoietic cells and cells stimulated by Interferon (IFN) - γ or Tumor Necrosis Factor (TNF) - α, these active subunits are replaced by β 1i (LMP2, low molecular weight polypeptide 2), β 2i (MECL-1, multicatalytic endopeptidase complex analog-1), and β 5i (LMP7) to form immunoproteasome [ Michael baseler et al, EMBO reports, 2018 ]. LMP7 is encoded by PSMB8 gene, 276 amino acids in total, and is a small molecule protein of about 30 kDa. LMP7 is the core catalytic subunit of immunoproteasome, has chymotrypsin activity and plays an important role in the process of immunoproteasome hydrolysis of proteins [ a. arkhjami et al, Immune and non-Immune functions of the immunoproteasome, Frontiers in Bioscience, 17 (1): 1904, 2012).

The function of the immunoproteasome in the course of immunization has been well studied, especially its antigen presentation function. The catalytic subunits of the immunoproteasome are hydrolyzed to produce polypeptides, which are presented on the cell surface by the histocompatibility complex (MHC) -1, causing cytotoxic T lymphocyte responses (CTLs). Compared to proteasomes, immunoproteases hydrolyze proteins and present antigens more efficiently, and the resulting antigens can elicit more potent CTLs. Several studies have demonstrated that immunoproteasome can regulate cytokine production. Similar phenomena were also observed in animal models of rheumatoid arthritis [ T.Muchamuel et al, A selective inhibitor of the immunological substrates and of the pathological precursors LMP7, cytokine production and expression of experimental arthritis, Nat Med, 15(7), 781-7, 2009 ] with selective inhibition of LMP7, IL-23 in monocytes and TNF-a and IL-6 in T cells by small molecule inhibitors. In addition, the function of immunoproteasome in T cell differentiation, proliferation and apoptosis has also been demonstrated in several studies [ c.m. caudil et al, T cells lacking immunoproteasome superbits MECL-1 and LMP7 superproproferase in response to polyclonal mitogens, j.immunol, 176(7), 4075-82, 2006 ]. In addition to immune function, immunoproteasome plays a role in maintaining protein homeostasis in the response to cytokine-induced oxidative stress. Oxidative stress releases free radicals, leading to the accumulation of a number of damaged proteins beyond the clearance of the normal proteasome, ultimately leading to cell death. Immunoproteasome can efficiently eliminate protein accumulation and maintain cellular homeostasis. In LMP7/β 5 i-and LMP2/β 1 i-deficient mice, the accumulation of oxidized and polyubiquinated proteins in the liver and brain was observed [ u.seifert et al, microprorotesomes previous protein in hormone-induced oxidative stress, Cell, 142 (4); 613-24, 2010 ].

Immunoproteasome is associated with a variety of diseases. Studies have shown that immunoproteasome is highly expressed in blood cancers and that selective inhibition of β 1i and LMP7 can effectively inhibit the growth of patient-derived cells and tumor models [ u.seifert et al, immunology precursors in tumor uptake on interferon-induced oxidative stress, Cell, 142(4), 613-24, 2010 ]. 668 Breast Cancer patients were studied and showed high Expression of LMP7 in 40% of patients' tumors [ M.Lee et al, Expression of Immunoproteasome Subunit LMP7 in Breast Cancer and Its associates with Immune-Related Markers, Cancer Research and Treatment, 51(1), 2018 ]. Immunoproteasome promotes the development and progression of colorectal cancer, and inhibitors of LMP7 can effectively inhibit the development of colorectal cancer in a mouse model [ j. koerner et al, Inhibition and specificity of the immunoproteasome outbuninbut LMP7 treatment and progression of clinical cancer in mice, Oncotarget, 8 (31): 50873 50888,2017. Recent evidence suggests that immunoproteasome is associated with autoimmune diseases and is expected to be a hot target for treating such diseases. Immunoproteasome is highly expressed in autoimmune diseases, such as rheumatoid Arthritis and inflammatory bowel disease [ T. Eger et al, Tissue-specific up-regulation of the protease subunit beta 5i (LMP7) in Sjogren's syndrome, Arthritis Rheum, 54(5), 1501-8, 2006 ]. In two mouse models of arthritis, selective inhibitors of LMP7 reduce the degree of inflammatory infiltration and cytokine levels, alleviating the symptoms of arthritis [ j. koerner et al, Inhibition and diagnosis of the immunological subset LMP7 suppression and progression of the immunological cancer in mice, Oncotarget, 8 (31): 50873 50888,2017. There are articles reporting that Immunoproteasome is associated with neurodegenerative diseases, with high expression of Immunoproteasome in the brain of alzheimer's patients [ m.d. i azHern ' ndez et al, neurological indication of the Immunoproteasome in Huntington's Disease, Journal of Neuroscience, 23 (37): 11653-11661, 2003 ].

The catalytic subunit LMP7 of immunoproteasome is used as a target for treating various diseases, is an innovative field and has wide development space. Compared with a broad-spectrum proteasome inhibitor, the LMP7 selective inhibitor has an absolute advantage in safety. Patents WO2019099582a1 and WO2019038250a1, among others, disclose certain LMP7 inhibitors and methods of using them to treat related diseases.

This patent describes a class of boronic acid derivatives which have good inhibitory activity against LMP7, while having good selectivity for other proteasomes. Moreover, such compounds have excellent oral bioavailability, plasma protein adsorption, pharmacokinetic profile, CYP-inhibition and stability.

Summary of The Invention

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

wherein,

R^aand R^bEach independently selected from H and C_1-6Alkyl, or R^aAnd R^bMay be joined together to form a 3-to 10-membered heterocyclic ring;

x is a bond, -O-, or-NR⁴-；

Y is a bond or- (CR)⁴R⁵)_m-；

R⁴And R⁵Each independently selected from H, C_1-6Alkyl and C_3-8A cycloalkyl group;

m is 1, 2, or 3;

R²selected from H and C_1-6An alkyl group;

R³is selected from C_6-10Aryl and C_5-10Heteroaryl, said aryl and heteroaryl being optionally substituted by halogen, -OH, -NH₂、-O-C_1-6Alkyl, -N (C)_1-6Alkyl) (C_1-6Alkyl), -CN, NO₂、C_1-6Alkyl radical, C_3-8Cycloalkyl, or C_3-8Heterocycloalkyl substitution;

R¹is selected from C_1-6Alkyl radical, C_3-8Cycloalkyl radical, C_3-8Heterocycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said alkyl, cycloalkyl, heterocycloalkyl, aryl and heteroaryl being substituted by

And may be optionally substituted by halogen, -OH, -NH₂、-(CH₂)_1-3-C_3-8Cycloalkyl, - (CH)₂)_0-6-CF₃、-O-C_1-6Alkyl, -NR⁹R⁸、-CN、NO₂、C_1-6Alkyl, - (CH)₂)_0-3-(CO)-R⁸、-(CH₂)_0-3-(CO)-NH-R⁸、-(CH₂)_0-3-NH-(CO)-R⁸Or R¹⁰Substitution;

R^6aand R^6bEach independently selected from C_1-6Alkyl radical, C_3-8Cycloalkyl radical, C_3-8Heterocycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said alkyl, cycloalkyl, heterocycloalkyl, aryl and heteroaryl being optionally substituted by halogen, C_1-6Alkyl radical, C_3-8Cycloalkyl, -OH, -MH₂、-O-C_1-6Alkyl, -NR⁹R⁸、-NO₂or-CN substitution, or

R^6aAnd R^6bMay be joined together to form a 3-8 membered heterocyclic ring;

R⁷selected from H, C_1-6Alkyl radical, C_3-8Cycloalkyl radical, C_3-8Heterocycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said alkyl, cycloalkyl, heterocycloalkyl, aryl and heteroaryl being optionally substituted by halogen, C_1-6Alkyl radical, C_3-8Cycloalkyl, -OH, -NH₂、-O-C_1-6Alkyl, -NR⁹R⁸、-NO₂or-CN substitution;

R⁸is selected from C_1-6Alkyl radical, C_3-8Cycloalkyl radical, C_3-8Heterocycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said alkyl, cycloalkyl, heterocycloalkyl, aryl and heteroaryl being optionally substituted by halogen, C_1-6Alkyl, -O-C_1-6Alkyl radical, C_6-10Aryl, or C_5-10Heteroaryl substitution;

R⁹selected from H and C_1-6An alkyl group;

R¹⁰is selected from C_3-8Cycloalkyl radical, C_3-8Heterocycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said cycloalkyl, heterocycloalkyl, aryl and heteroaryl being optionally substituted by halogen, -OH, -NH₂、-O-C_1-6Alkyl, -N (C)_1-6Alkyl) (C_1-6Alkyl), -CN, NO₂Or C_1-6Alkyl substitution;

in some embodiments, R^aAnd R^bIs H;

in some embodiments, X is a bond, -O-, or-NR⁴-，R⁴Is H;

in some embodiments, Y is a bond or- (CR)⁴R⁵)_m-，R⁴And R⁵Each independently selected from H and C_1-6Alkyl, m is 1, 2, or 3;

in some embodiments, Y is a bond or- (CR)⁴R⁵)_m-，R⁴And R⁵Is H, m is 1, 2, or 3;

in some embodiments, Y is a bond or- (CR)⁴R⁵)_m-，R⁴And R⁵Each independently selected from H and C_1-6Alkyl, m is 1 or 2;

in some embodiments, Y is a bond or- (CR)⁴R⁵)_m-，R⁴And R⁵Is H, m is 1 or 2;

in some embodiments, Y is a bond or- (CR)⁴R⁵)_m-，R⁴And R⁵Each independently selected from H and C_1-6Alkyl, m is 1;

in some embodiments, Y is a bond or- (CR)⁴R⁵)_m-，R⁴And R⁵Is H, m is 1;

in some embodiments, R²Is H;

in some embodiments, R³Is selected from C_5-10Heteroaryl, said heteroaryl being optionally substituted by halogen, NO₂、C_1-6Alkyl radical, C_3-8Cycloalkyl, or C_3-8Heterocycloalkyl substitution;

in some embodiments, R³Is composed of

The above-mentioned

Optionally substituted by halogen, NO₂、C_1-6Alkyl radical, C_3-8Cycloalkyl, or C_3-8Heterocycloalkyl substitution;

in some embodiments, R¹Is selected from C_3-8Cycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said cycloalkyl, aryl and heteroaryl being substituted by

And may optionally be substituted by halogen, -CF₃、-O-C_1-6Alkyl radical, NO₂、C_1-6Alkyl, - (CO) -R⁸、-(CO)-NH-R⁸Or R¹⁰Substitution;

R^6aand R^6bEach independently selected from C_1-6Alkyl and C_3-8Cycloalkyl, or R^6aAnd R^6bMay be joined together to form a 3-8 membered heterocyclic ring;

R⁷selected from H, C_1-6Alkyl and C_3-8A cycloalkyl group;

R⁸is selected from C_1-6Alkyl and C_3-8Cycloalkyl, said cycloalkyl being optionally substituted by C_1-6Alkyl substitution;

R¹⁰is selected from C_3-8Cycloalkyl and C_3-8Heterocycloalkyl, said cycloalkyl and heterocycloalkyl being optionally substituted by C_1-6Alkyl substitution;

in some embodiments, the present invention provides the following compounds or pharmaceutically acceptable salts, solvates, polymorphs, or isomers thereof

The compounds of formula (I) of the present invention may be useful in the treatment of diseases associated with lmp 7; in some embodiments, the disease associated with lmp7 activity is a hematological malignancy, solid tumor, or immunoregulatory abnormality, more preferably multiple myeloma, acute myelogenous leukemia, myelocytic leukemia, mantle cell lymphoma, chronic lymphocytic leukemia, acute lymphocytic leukemia, diffuse large B-cell lymphoma, plasmacytoma, follicular lymphoma, immune cell tumor, breast cancer, liver cancer, colorectal cancer, ovarian cancer, esophageal cancer, lung cancer, head and neck cancer, pancreatic cancer, kidney cancer, stomach cancer, thyroid cancer, prostate cancer, bladder cancer, rheumatoid arthritis, systemic lupus erythematosus, inflammatory bowel disease, multiple sclerosis, scleroderma, ankylosing spondylitis, atherosclerosis, behcet disease, crohn's disease, inflammatory bowel disease, ulcerative colitis, autoimmune hepatitis, sjogren's syndrome, lupus nephritis, multiple sclerosis, autoimmune dysgenopathy, autoimmune diseases, and other diseases, Asthma, Amyotrophic Lateral Sclerosis (ALS), psoriasis, type a immunoglobulin nephropathy, allergic purpura, Alzheimer's Disease (AD);

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

in another aspect, the present invention provides a method of treating a disease associated with lmp7 activity, the method comprising administering to a subject an effective amount of a compound of the present invention, or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, or a composition thereof; in some embodiments, the disease associated with lmp7 activity is multiple myeloma, acute myelogenous leukemia, myelocytic leukemia, mantle cell lymphoma, chronic lymphocytic leukemia, acute lymphocytic leukemia, diffuse large B-cell lymphoma, plasmacytoma, follicular lymphoma, immune cell tumor, breast cancer, liver cancer, colorectal cancer, ovarian cancer, esophageal cancer, lung cancer, head and neck cancer, pancreatic cancer, kidney cancer, stomach cancer, thyroid cancer, prostate cancer, bladder cancer, rheumatoid arthritis, systemic lupus erythematosus, inflammatory bowel disease, multiple sclerosis, scleroderma, ankylosing spondylitis, atherosclerosis, behcet's disease, crohn's disease, inflammatory bowel disease, ulcerative colitis, autoimmune hepatitis, sjogren's syndrome, lupus nephritis, asthma, Amyotrophic Lateral Sclerosis (ALS), psoriasis, immunoglobulin a-type a nephropathy, autoimmune hepatitis, rheumatoid arthritis, systemic lupus nephritis, rheumatoid arthritis, psoriasis, rheumatoid arthritis, and other autoimmune arthritis, rheumatoid arthritis, Allergic purpura, Alzheimer's Disease (AD);

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

in another aspect, the present invention provides the use of a compound of the invention, or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, in the manufacture of a medicament for the treatment of a disease associated with lmp7 activity; in some embodiments, the disease associated with lmp7 activity is multiple myeloma, acute myelogenous leukemia, myelocytic leukemia, mantle cell lymphoma, chronic lymphocytic leukemia, acute lymphocytic leukemia, diffuse large B-cell lymphoma, plasmacytoma, follicular lymphoma, immune cell tumor, breast cancer, liver cancer, colorectal cancer, ovarian cancer, esophageal cancer, lung cancer, head and neck cancer, pancreatic cancer, kidney cancer, stomach cancer, thyroid cancer, prostate cancer, bladder cancer, rheumatoid arthritis, systemic lupus erythematosus, inflammatory bowel disease, multiple sclerosis, scleroderma, ankylosing spondylitis, atherosclerosis, behcet's disease, crohn's disease, inflammatory bowel disease, ulcerative colitis, autoimmune hepatitis, sjogren's syndrome, lupus nephritis, asthma, Amyotrophic Lateral Sclerosis (ALS), psoriasis, immunoglobulin a-type a nephropathy, autoimmune hepatitis, rheumatoid arthritis, systemic lupus nephritis, rheumatoid arthritis, psoriasis, rheumatoid arthritis, rheumatoid, Allergic purpura, Alzheimer's Disease (AD).

Detailed Description

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

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

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

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

Certain chemical terms

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

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

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

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

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

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

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

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

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

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

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

The term "alkenyl" as used herein, alone or in combination, refers to an optionally substituted straight or optionally substituted branched chain monovalent hydrocarbon radical having one or more C ═ C double bonds and having from 2 to about 10 carbon atoms, more preferably from 2 to about 6 carbon atoms. The double bond in these groups may be in either the cis or trans conformation and should be understood to encompass both isomers. Examples include, but are not limited to, ethenyl (CH ═ CH)₂) 1-propenyl (CH)₂CH＝CH₂) Isopropenyl (C (CH3) ═ CH₂) Butenyl, 1, 3-butadienyl and the like.

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

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

The term "aryl" refers to an all-carbon monocyclic or fused ring having a fully conjugated pi-electron system, having 6 to 14 carbon atoms, preferably 6 to 12 carbon atoms, most preferably 6 carbon atoms. Aryl groups may be unsubstituted or substituted with one or more substituents, examples of which include, but are not limited to, alkyl, alkyloxy, aryl, aralkyl, amino, halo, hydroxy, sulfonyl, sulfinyl, phosphoryl, and heteroalicyclic. Non-limiting examples of unsubstituted aryl groups include, but are not limited to, phenyl, naphthyl, and anthracenyl.

The term "heteroaryl" refers to a monocyclic or fused ring of 5 to 12 ring atoms, having 5, 6, 7, 8, 9, 10, 11 or 12 ring atoms, containing 1, 2, 3 or 4 ring atoms selected from N, O, S, the remaining ring atoms being C, and having a fully conjugated pi-electron system. Heteroaryl groups may be unsubstituted or substituted, and the substituents include, but are not limited to, alkyl, alkyloxy, aryl, aralkyl, amino, halo, hydroxy, cyano, nitro, carbonyl, and heteroalicyclic. Non-limiting examples of unsubstituted heteroaryl groups include, but are not limited to, pyrrolyl, furanyl, thienyl, imidazolyl, oxazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyrazinyl, quinolinyl, isoquinolinyl, tetrazolyl, triazinyl.

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

Preparation of the Compounds of the invention

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

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

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

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

Nuclear magnetic data (¹H NMR) was run at 400MHz using a Varian instrument. The solvent used for nuclear magnetic data is CDCl₃、CD₃OD、D₂O、DMSO-d₆Etc., based on tetramethylsilane (0.00ppm) or based on residual solvent (CDCl)₃：7.26ppm；CD3OD：3.31ppm；D2O：4.79ppm；d₆-DMSO: 2.50 ppm). When indicating the diversity of the peak shapes, the following abbreviations represent the different peak shapes: s (singlet), d (doublet), t (triplet), q (quartet), m (multiplet), br (broad), dd (doublet of doublets), dt (doublet of triplets). If the coupling constant is given, it is given in Hertz (Hz).

Example 1

((1R) -2- (benzofuran-3-yl) -1- (((((((2R) -1- (N- (tert-butyl) phenylsulfonylimino) pyrrolidin-2-yl) methoxy) carbonyl) amino) ethyl) boronic acid

Step A: benzofuran-3-yl carbinols

N₂Under protection, a methanol (200mL) solution containing benzofuran-3-yl-formaldehyde (15.0g) was cooled to 0 ℃, sodium borohydride (5.9g) was added to the system in portions, and after naturally returning to room temperature, the mixture was stirred for 2 hours, after completion of the reaction was monitored, the solvent was removed by rotary evaporation, the residue was separated into ethyl acetate and a 1mol/L hydrochloric acid aqueous solution, the organic phase was dried over anhydrous sodium sulfate, and then the solvent was filtered and evaporated to dryness to obtain a product (14.8 g).

¹H NMR(400MHz，CDCl₃)δ7.64-7.66(m，2H)，7.58(s，1H)，7.46-7.48(m，1H)，7.23-7.32(m，2H)，4.81(s，2H)。

And B: 3-chloromethyl benzofurans

A dichloromethane (100mL) solution containing benzofuran-3-yl methanol (10.0g) is cooled to 0 ℃, phosphorus pentachloride (18.2g) is added into the system in batches, the temperature naturally returns to room temperature after the addition is finished, water is added after stirring for 1 hour to quench the reaction, an organic phase is washed by saturated sodium bicarbonate water solution and saturated salt water in sequence, the organic phase is dried by anhydrous sodium sulfate, and the solvent is evaporated after filtering to obtain a product (12.1 g).

¹H NMR(400MHz，CDCl₃)δ7.65-7.70(m，2H)，7.49(d，J＝8.4Hz，1H)，7.30-7.35(m，2H)，4.75(s，2H)。

And C: benzofuran-3-ylmethylboronic acid pinacol ester

Cooling a suspension of N, N-dimethylformamide (80mL) containing 3-chloromethylbenzofuran (12.0g), cuprous iodide (1.3g), pinacol ester of diboronic acid (18.9) and triphenylphosphine (1.78g) to 0 ℃, adding lithium tert-butoxide (8.68g) into the system in batches, naturally returning the reaction solution to room temperature after the addition is finished, and stirring for 2 hours; after quenching the reaction with water, the reaction solution was extracted with dichloromethane and washed with saturated brine, the organic phase was dried over anhydrous sodium sulfate, filtered and the solvent was evaporated, and the residue was purified by silica gel column chromatography (elution with 10-20% ethyl acetate mixed solution) to give the product (14.1 g).

¹H NMR(400MHz，CDCl₃)δ7.50-7.55(m，2H)，7.43(d，J＝8.4Hz，1H)，7.20-7.25(m，2H)，2.21(s，2H)，1.26(s，12H)。

Step D: benzofuran-3-ylmethylboronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester

Benzofuran-3-ylmethylboronic acid pinacol ester (14.0g) and (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol (18.4g) were added to anhydrous ether (200mL), the suspension was stirred overnight at room temperature, after completion of the detection reaction, washed three times with water, the organic phase was dried over anhydrous sodium sulfate, filtered and the solvent was evaporated to dryness, and the residue was purified by silica gel column chromatography (50% dichloromethane/petroleum ether) to give a product (10.5 g).

¹H NMR(400MHz，CDCl₃)δ7.44-7.56(m，2H)，7.43(d，J＝8.4Hz，1H)，7.20-7.26(m，2H)，4.31(dd，J＝8.4Hz，1.6Hz，1H)，2.30-2.36(m，1H)，2.26(s，2H)，2.17-2.21(m，1H)，2.06(t，J＝6.0Hz，1H)，1.86-1.91(m，2H)，1.40(s，3H)，1.27(s，3H)，1.11(d，J＝11.2Hz，1H)，0.87(s，3H)。

Step E: 2- (benzofuran-3-yl) -1- (S) -chloroethylboronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester

Dissolving dried dichloromethane (4.11g) in anhydrous tetrahydrofuran (30mL), fully replacing nitrogen, cooling the solution to-100 ℃, slowly dripping 2.5 mol/L butyl lithium n-hexane solution (9.6mL) into the system along the inner wall of a reaction bottle for not less than 10 minutes to obtain milky white suspension, continuously stirring at-100 ℃ for half an hour, slowly adding anhydrous tetrahydrofuran (30mL) solution of benzofuran-3-yl methyl boronic acid (+) -pinanediol ester (5.0g) into the system along the inner wall of the reaction bottle, after 10 minutes, dripping 1mol/L zinc chloride tetrahydrofuran solution (8.8mL) into the reaction system, gradually returning to room temperature, and stirring overnight. The reaction was quenched with water, the reaction was partitioned between ethyl acetate (100mL) and saturated aqueous ammonium chloride (50mL), the organic phase was dried over anhydrous sodium sulfate, filtered and evaporated to dryness to give the crude product which was used directly in the next reaction. (6.0 g).

¹H NMR(400MHz，CDCl₃)δ7.58-7.60(m，1H)，7.48-7.55(m，2H)，7.25-7.30(m，2H)，4.30-4.33(m，1H)，3.72-3.76(m，1H)，3.29(dd，J＝15.2Hz，8.0Hz，1H)，3.18(dd，J＝15.2Hz，8.0Hz，1H)，2.28-2.35(m，1H)，2.06-2.17(m，2H)，1.86-1.91(m，2H)，1.27(s，3H)，1.22(s，3H)，1.12-1.17(m，1H)，0.84(s，3H)。

Step F: 2- ((benzofuran-3-yl) -1- (R) -aminoethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester hydrochloride

2- ((S) -2- (benzofuran-3-yl) -1-chloroethyl) boric acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester (6.0g) is dissolved in anhydrous n-hexane, nitrogen is sufficiently replaced, the solution is cooled to-78 ℃, 1mol/L lithium bistrimethylsilyl amide tetrahydrofuran solution (16mL) is slowly dripped into the system, the temperature is slowly returned to the room temperature, and the mixture is stirred overnight. The resulting suspension was filtered through celite and rinsed with n-hexane, the resulting mother liquor was cooled to 0 ℃, a 4 mol/l solution of 1, 4-dioxane (16mL) of hydrogen chloride was slowly added dropwise to the solution, and after the addition was complete, the solution was allowed to return to room temperature and stirred for 2 hours. And (3) removing the solvent from the reaction solution by rotary evaporation, adding n-hexane, fully stirring, filtering, and leaching the obtained solid with n-hexane to obtain a product (2.91 g).

¹H NMR(400MHz，CDCl₃)δ8.32(brs，3H)，7.81(s，1H)，7.67(d，J＝7.6Hz，1H)，7.43(d，J＝8.0Hz，1H)，7.22-7.28(m，2H)，4.27(d，J＝8.0Hz，1H)，3.30-3.36(m，3H)，2.19-2.22(m，1H)，2.08-2.15(m，1H)，1.95-2.05(m，1H)，1.80-1.88(m，2H)，1.27(s，3H)，1.25(s，3H)，1.03-1.06(m，1H)，0.70(s，3H)。

Step G: (R) -2- (((Chlorocarbonyl) oxy) methyl) pyrrolidine-1-carboxylic acid tert-butyl ester

Tert-butyl (R) -2- (hydroxymethyl) pyrrolidine-1-carboxylate (314mg) was placed in a three-necked flask, dichloromethane (10mL) and diisopropylethylamine (402mg) were added, the mixture was cooled to 0 ℃ under nitrogen, a solution of bis (trichloromethyl) carbonate (232mg) in dichloromethane was added dropwise to the reaction mixture, and after completion, the mixture was stirred at 0 ℃ for 2 hours and used directly in the next step.

Step H: ((1R) -2- (benzofuran-3-yl) -1- (((((((2R) -N-tert-butoxycarbonyl-pyrrolidin-2-yl) methoxy) carbonyl) amino) ethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester

2- (benzofuran-3-yl) -1- (R) -aminoethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester hydrochloride (470mg) and diisopropylethylamine (273mg) were added to dried dichloromethane, under the protection of nitrogen, cooling to 0 ℃, dropwise adding a dichloromethane solution of (R) -2- (((chlorocarbonyl) oxy) methyl) pyrrolidine-1-tert-butyl formate into the reaction solution, returning to room temperature after the completion of the addition, stirring for 2 hours, quenching the reaction by water, extracting with dichloromethane, washing organic phase with saturated salt water, drying organic phase with anhydrous sodium sulfate, the residue obtained by filtration to dryness of the solvent was purified by silica gel column chromatography (1: 1 ethyl acetate/petroleum ether) to give the product (350 mg).

¹H NMR(400MHz，CDCl₃)δ7.56(d，J＝7.2Hz，1H)，7.42-7.45(m，2H)，7.27(t，J＝8.0Hz，1H)，7.21(t，J＝7.6Hz，1H)，4.93(s，1H)，4.28(d，J＝8.0Hz，1H)，3.82-4.17(m，3H)，3.48-3.55(m，1H)，3.24-3.40(m，2H)，3.11(dd，J＝14.8Hz，6.0Hz，1H)，2.96(dd，J＝14.8Hz，7.2Hz，1H)，2.28-2.33(m，1H)，2.07-2.14(m，1H)，1.97(t，J＝5.6Hz，1H)，1.76-1.92(m，6H)，1.45(s，9H)，1.25(s，3H)，1.20(s，3H)，0.85(d，J＝6.8Hz，1H)，0.80(s，3H)。

Step J: ((1R) -2- (benzofuran-3-yl) -1- (((((((2R) -pyrrolidin-2-yl) methoxy) carbonyl) amino) ethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester trifluoroacetate

((1R) -2- (benzofuran-3-yl) -1- (((((((2R) -N-tert-butoxycarbonyl-pyrrolidin-2-yl) methoxy) carbonyl) amino) ethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester (191mg) was added to dichloromethane (5mL), followed by trifluoroacetic acid (1mL), stirred at room temperature for 3 hours, and the solvent was evaporated to dryness under reduced pressure to give a product (190 mg).

The intermediate was confirmed by mass spectrometry, [ M + H ]⁺]＝467。

Step K: n- (tert-butyl) benzenesulfinamide

Tert-butylamine (414mg) was added to dried dichloromethane, nitrogen blanketed and cooled to 0 ℃ and triethylamine (1.14g) was added to the reaction mixture followed by dropwise addition of phenylsulfonyl chloride (63mg), followed by dropwise slow addition of triphenylphosphine (1.48g) in dichloromethane to the above reaction mixture at 0 ℃ after completion of the addition, and the addition was completed over 1 hour. The mixture was allowed to warm to room temperature and stirred overnight, and the residue obtained by evaporation of the solvent was purified by silica gel column chromatography (1: 4 ethyl acetate/petroleum ether) to give the product (280 mg).

¹H NMR(400MHz，CDCl₃)，7.66-7.69(m，2H)，7.43-7.48(m，3H)，3.86(s，1H)，1.39(s，9H)。

Step L: n- (tert-butyl) anilinosulphenyl chloride

Adding N- (tert-butyl) benzene sulfinamide (280mg) into dried dichloromethane, cooling to 0 ℃ under the protection of nitrogen, dropwise adding tert-butyl hypochlorite (185mg) into the reaction solution, returning to room temperature after the reaction is finished, stirring for 1 hour, quenching the reaction with water, extracting with dichloromethane, washing an organic phase with saturated salt water, drying the organic phase with anhydrous sodium sulfate, filtering and evaporating the solvent to dryness to obtain a product (140 mg).

Step M: ((1R) -2- (benzofuran-3-yl) -1- (((((((2R) -1- (N- (tert-butyl) phenylsulfonylimino) pyrrolidin-2-yl) methoxy) carbonyl) amino) ethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester

((1R) -2- (benzofuran-3-yl) -1- (((((((2R) -pyrrolidin-2-yl) methoxy) carbonyl) amino) ethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester trifluoroacetate (190mg) and diisopropylethylamine (132mg) were added to dry dichloromethane, under the protection of nitrogen, cooling to 0 ℃, dropwise adding a dichloromethane solution of N- (tert-butyl) anilinosulfino chloride (106mg) into the reaction solution, returning to room temperature after the completion of the addition, stirring for 2 hours, quenching the reaction with water, extracting with dichloromethane, washing organic phase with saturated salt water, drying organic phase with anhydrous sodium sulfate, the residue obtained by filtration to dryness of the solvent was purified on a silica gel preparation plate (1: 4 ethyl acetate/petroleum ether) to give the product (90 mg).

¹H NMR(400MHz，CDCl₃)δ7.87-7.97(m，2H)，7.41-7.58(m，6H)，7.26(t，J＝7.6Hz，1H)，7.20(t，J＝8.0Hz，1H)，4.86-4.96(m，1H)，3.78-4.29(m，4H)，2.92-3.70(m，5H)，2.27-2.48(m，1H)，1.56-2.21(m，8H)，1.37(s，3H)，1.33(s，3H)，1.24(s，3H)，1.22(s，3H)，1.19(s，3H)，1.12(d，J＝7.2Hz，1H)，0.92(s，0.5Hz)，0.79(s，2.5H).

And step N: ((1R) -2- (benzofuran-3-yl) -1- (((((((2R) -1- (N- (tert-butyl) phenylsulfonylimino) pyrrolidin-2-yl) methoxy) carbonyl) amino) ethyl) boronic acid

((1R) -2- (benzofuran-3-yl) -1- (((((2R) -1- (N- (tert-butyl) phenylsulfonylimino) pyrrolidin-2-yl) methoxy) carbonyl) amino) ethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester (90mg) was dissolved in methanol, isobutylboronic acid (45mg), 1mol/l hydrochloric acid (3mL) and N-hexane were added to the solution, stirred at room temperature overnight, the upper N-hexane was removed by liquid separation, the methanol phase was washed three times with N-hexane and concentrated to dryness at 30 ℃, saturated saline (15mL) was added, after extraction three times with dichloromethane, the organic phase was dried over anhydrous sodium sulfate, the residue obtained by filtration to dryness of the solvent was purified by a silica gel preparation plate (1: 40 methanol/dichloromethane) to obtain a diradical group The isomers at the sulfur atom were isomer 1(30mg) and isomer 2(20mg), respectively.

Isomer 1

¹H NMR(400MHz，CD₃OD)δ7.89(d，J＝7.2Hz，2H)，7.49-7.60(m，5H)，7.40(d，J＝8.0Hz，1H)，7.17-7.26(m，2H)，4.20(dd，J＝10.8Hz，4.0Hz，1H)，4.11(dd，J＝10.0Hz，6.8Hz，1H)，3.80-3.86(m，1H)，3.28-3.32(m，1H)，3.12-3.26(m，2H)，2.93(dd，J＝14.4Hz，6.8Hz，1H)，2.85(dd，J＝14.4Hz，8.0Hz，1H)，1.50-1.70(m，3H)，1.36(s，9H)，1.08-1.16(m，1H)。

Isomer 2

¹H NMR(400MHz，CD₃OD)δ7.82-7.90(m，2H)，7.44-7.58(m，5H)，7.41(d，J＝8.0Hz，1H)，7.25(t，J＝7.6Hz，1H)，7.20(t，J＝7.6Hz，1H)，4.20(dd，J＝10.4Hz，2.8Hz，1H)，4.04(dd，J＝10.4Hz，7.2Hz，1H)，3.91-3.98(m，1H)，3.22-3.30(m，2H)，3.10-3.15(m，1H)，2.91(dd，J＝14.4Hz，7.2Hz，1H)，2.83(dd，J＝14.4Hz，7.6Hz，1H)，1.61-1.86(m，4H)，1.32(s，9H)。

Example 2

((1R) -2- (benzofuran-3-yl) -1- (((((((2R) -1- (N, S-dimethylsulfonylimino) pyrrolidinyl-2-yl) methoxy) carbonyl) amino) ethyl) boronic acid

Step A: n-sulfinyl triphenylmethylamine

Triphenylmethylamine (10g) and triethylamine (7.8g) were added to dry ether, the mixture was cooled to 0 ℃ under nitrogen protection, thionyl chloride (4.6g) was added dropwise to the reaction solution, and after completion of stirring at 0 ℃ for 2 hours, the mixture was filtered through celite, rinsed with ether, and the solvent was evaporated under reduced pressure at room temperature to give a white solid product (11 g).

And B: (±) -N-trityl methane sulfinamide

Dissolving N-sulfinyl tritylamine (6g) in anhydrous tetrahydrofuran, cooling to 0 ℃ under the protection of nitrogen, dropwise adding 3 mol/L methylmagnesium bromide tetrahydrofuran solution (5.2mL) into the reaction solution, returning to room temperature after the reaction is finished, stirring for 1 hour, quenching the reaction with saturated ammonium chloride, extracting with ethyl acetate, washing an organic phase with saturated salt water, drying the organic phase with anhydrous sodium sulfate, filtering to evaporate the solvent to obtain a residue, pulping the residue with petroleum ether, and performing suction filtration to obtain a solid product (4.5 g).

¹H NMR(400MHz，CDCl₃)δ7.24-7.33(m，15H)，4.83(s，1H)，2.57(s，3H)。

And C: (R) -2- (((tert-butylmethylsilyl) oxy) methyl) pyrrolidine

Adding D-prolinol (1g) and imidazole (1.3g) into dichloromethane, cooling to 0 ℃, dropwise adding tert-butyldiphenylchlorosilane (5.7g) into the reaction solution, returning to room temperature after the reaction is finished, stirring for 1 hour, performing suction filtration to obtain a filtrate, adding a saturated ammonium chloride aqueous solution into the filtrate, extracting with dichloromethane, drying an organic phase by anhydrous sodium sulfate, filtering and evaporating a solvent to obtain a residue, and purifying the residue by silica gel column chromatography (1: 7 methanol/dichloromethane) to obtain a product (3.1 g).

¹H NMR(400MHz，CDCl₃)δ7.64-7.69(m，4H)，7.35-7.44(m，6H)，3.85(dd，J＝11.2Hz，4.8Hz，1H)，3.74(dd，J＝11.2Hz，5.6Hz，1H)，3.58-3.65(m，1H)，3.23(t，J＝7.2Hz，2H)，1.85-1.99(m，3H)，1.67-1.75(m，1H)，1.06(s，9H)。

Step D: (2R) -2- (((tert-butyldiphenylsilyl) oxy) methyl) -1- (S-methylsulfonylimino) pyrrolidine

(±) -N-tritylmethanesulfinamide (2g) was added to dry dichloromethane, under the protection of nitrogen, cooling to 0 ℃, dropwise adding tert-butyl hypochlorite (676mg) into the reaction solution, stirring for 1 hour at 0 ℃, the reaction solution was then added dropwise to a solution of (R) -2- (((tert-butylmethylsilyl) oxy) methyl) pyrrolidine (1.4g) and triethylamine (1.2g) in methylene chloride, stirring overnight at room temperature under nitrogen protection, adding p-toluenesulfonic acid monohydrate (10.6g), stirring at room temperature for 30 minutes, adding 1mol/L sodium hydroxide aqueous solution to adjust pH to alkalinity, extracting with dichloromethane, washing organic phase with saturated salt water, drying organic phase with anhydrous sodium sulfate, the residue obtained by filtration and evaporation of the solvent was purified by silica gel column chromatography (pure ethyl acetate) to give the product (1.6 g).

¹H NMR(400MHz，CDCl₃)δ7.62-7.68(m，4H)，7.34-7.43(m，6H)，3.82(dd，J＝10.4Hz，4.0Hz，0.5H)，3.69-3.79(m，1H)，3.64(dd，J＝10.4Hz，4.0Hz，0.5H)，3.51-3.59(m，1H)，3.22-3.42(m，2H)，2.78(s，3H)，2.17-2.21(m，1H)，1.78-2.13(m，4H)，1.05(s，9H)。

Step E: (2R) -2- (((tert-butyldiphenylsilyl) oxy) methyl) -1- (N, S-dimethylsulfonylimino) pyrrolidine

Adding (2R) -2- (((tert-butyldiphenylsilyl) oxy) methyl) -1- (S-methylsulfonimidoyl) pyrrolidine (1.2g) into dried tetrahydrofuran, cooling to 0 ℃, adding sodium hydride (230mg) to the reaction solution in batches, stirring at 0 ℃ for 30 minutes, dropwise adding methyl iodide (809mg), returning to room temperature after the completion of stirring overnight, quenching the reaction with water, extracting with ethyl acetate, washing the organic phase with saturated saline, drying the organic phase with anhydrous sodium sulfate, filtering and evaporating the solvent to dryness to obtain a product (1.2 g).

¹H NMR(400MHz，CDCl₃)δ7.62-7.67(m，4H)，7.35-7.44(m，6H)，3.77-3.82(m，1H)，3.67-3.72(m，1H)，3.53-3.62(m，1H)，3.16-3.30(m，2H，，2.76(s，1.7H)，2.75(s，1.3s)，2.64(s，1.3s)，2.51(s，1.7H)，2.04-2.13(m，1H)，1.78-1.98(m，3H)，1.05(s，9H)。

Step F: ((2R) -1- (N, S-dimethylsulfonylimino) pyrrolidin-2-yl) methanol

(2R) -2- (((tert-butyldiphenylsilyl) oxy) methyl) -1- (N, S-dimethylsulfonylimino) pyrrolidine (1.2g) was dissolved in tetrahydrofuran, tetrabutylammonium fluoride monohydrate (1.2g) was added to the solution, the mixture was stirred at 40 ℃ for 2 hours, and the residue obtained by evaporating the solvent was purified by silica gel column chromatography (methanol: ethyl acetate 3/40) to give a product (400 mg).

¹H NMR(400MHz，CDCl₃)δ3.57-3.70(m，2H)，3.51-3.54(m，2H)，3.24-3.31(m，2H)，2.85(s，0.6H)，2.80(s，2.4H)，2.67(s，0.6H)，2.65(s，2.4H)，1.97-2.06(m，1H)，1.66-1.95(m，3H)。

Step G: ((1R) -2- (benzofuran-3-yl) -1- (((((((2R) -1- (N, S-dimethylsulfonylimino) pyrrolidinyl-2-yl) methoxy) carbonyl) amino) ethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester

2- (benzofuran-3-yl) -1- (R) -aminoethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester hydrochloride (150mg) and triethylamine (162mg) were added to dried dichloromethane, nitrogen gas was used for protection and cooling to-60 ℃, a dichloromethane solution of bis (trichloromethyl) carbonate (47mg) was added dropwise to the reaction solution, after completion of stirring at-60 ℃ for 1 hour, -a dichloromethane solution of ((2R) -1- (N, S-dimethylsulfonylimino) pyrrolidin-2-yl) methanol (93mg) was added dropwise to the above solution at-60 ℃, stirred at-60 ℃ for 1 hour, quenched with water and extracted with dichloromethane, the organic phase was dried over anhydrous sodium sulfate, filtered and the solvent evaporated to dryness and the residue was purified on silica gel preparation plates (1: 30 methanol/dichloromethane) to give the product (25 mg).

¹H NMR(400MHz，CDCl₃)δ7.56(d，J＝6.8Hz，1H)，7.45(s，1H)，7.43(d，J＝8.0Hz，1H)，7.26(t，J＝7.2Hz，1H)，7.21(t，J＝7.2Hz，1H)，4.96(d，J＝5.6Hz，1H)，4.27-4.30(m，1H)，3.96-4.14(m，2H)，3.87-3.94(m，0.6H)，3.72-3.80(m，0.4H)，3.49-3.55(m，1H)，3.18-3.28(m，2H)，3.10(dd，J＝14.8Hz，5.2Hz，1H)，2.96(dd，J＝14.8Hz，7.2Hz，1H)，2.83(s，1.8H)，2.80(s，1.2H)，2.67(s，1.8H)，2.65(s，1.2H)，2.06-2.48(m，3H)，1.59-2.01(m，6H)，1.24(s，3H)，1.20(s，3H)，1.05(d，J＝10.8Hz，1H)，0.79(s，3H)。

Step H: ((1R) -2- (benzofuran-3-yl) -1- (((((((2R) -1- (N, S-dimethylsulfonylimino) pyrrolidinyl-2-yl) methoxy) carbonyl) amino) ethyl) boronic acid

((1R) -2- (benzofuran-3-yl) -1- ((((((2R) -1- (N, S-dimethylsulfonylimino) pyrrolidinyl-2-yl) methoxy) carbonyl) amino) ethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester (25mg) was dissolved in methanol, isobutylboronic acid (14mg), 1 mole/liter hydrochloric acid (0.1mL) and N-hexane were added to this solution, stirred overnight at room temperature, the upper N-hexane was removed by liquid separation, and the methanol phase was washed three times with N-hexane, concentrated to dryness and diluted with dichloromethane and the product was washed to the aqueous phase with 2 mole/liter aqueous sodium hydroxide solution (5mL), the aqueous phase was washed three times with dichloromethane and acidified to pH acidity with 3 mole/liter hydrochloric acid, after three extractions with dichloromethane, the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated at 30 ℃ to remove the solvent to give the product (10 mg).

¹H NMR(400MHz，CD₃OD)δ7.59(d，J＝8.0Hz，1H)，7.54(s，1H)，7.42(d，J＝7.6Hz，1H)，7.26(t，J＝7.2Hz，1H)，7.21(t，J＝7.2Hz，1H)，4.17-4.30(m，2H)，3.96-4.07(m，1H)，3.62(s，1H)，3.58(s，2H)，3.33-3.50(m，3H)，2.83-2.97(m，2H)，2.79(s，2H)，2.76(s，1H)，1.94-2.20(m，4H)。

Example 3

(R) - (2- (benzofuran-3-yl) -1- (2- (3-dimethylsulfinimidophenyl) acetamido) ethyl) boronic acid

Step A: 2- (3-dimethylsulfinylphenyl) acetic acid

Dissolving 3-bromobenzeneacetic acid (431mg) in 1, 4-dioxane, sequentially adding tris (dibenzylideneacetone) dipalladium (184mg), 4, 5-bis (diphenylphosphino) -9, 9-dimethylxanthene (230mg) and cesium carbonate (1.96g), fully replacing nitrogen in the system, heating to 100 ℃ for overnight reaction, monitoring complete conversion of the raw materials, adding water for dilution, extracting with dichloromethane, combining organic phases, sequentially washing with water and saturated common salt water, drying the organic phase with anhydrous sodium sulfate, filtering, evaporating the solvent to dryness, and purifying the residue with a silica gel preparation plate (1: 10 methanol/dichloromethane) to obtain a product (67 mg).

¹H NMR(400MHz，CDCl₃)δ7.15(t，J＝8.0Hz，1H)，6.96-6.98(m，2H)，6.89(d，J＝7.6Hz，1H)，3.54(s，2H)，3.13(s，6H)。

And B: (R) -2- (benzofuran-3-yl) -1- (2- (3-dimethylsulfinimidophenyl) acetamido) ethylboronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester

Adding 2- (3-dimethylsulfinylphenyl) acetic acid (67mg), 2- (benzofuran-3-yl) -1- (R) -aminoethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester hydrochloride (111mg), 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate (146mg), N, N-diisopropylethylamine (114mg) and 4-dimethylaminopyridine (36mg) into dried dichloromethane, stirring at room temperature overnight after completion, quenching the reaction with water, extracting with dichloromethane, washing the organic phase with saturated saline, drying the organic phase with anhydrous sodium sulfate, filtering and evaporating the solvent to dryness to obtain a residue, and purifying the residue with a silica gel preparation plate (20: 1 methanol/dichloromethane) Product (30 mg).

¹H NMR(400MHz，CDCl₃)δ7.44(d，J＝8.0Hz，1H)，7.38(d，J＝8.0Hz，1H)，7.22-7.26(m，2H)，7.17(d，J＝7.2Hz，1H)，7.13(t，J＝8.0Hz，1H)，6.97(d，J＝8.0Hz，1H)，6.83(s，1H)，6.72(d，J＝7.2Hz，1H)，6.21(s，1H)，4.24(d，J＝8.0Hz，1H)，3.61(d，J＝16.8Hz，1H)，3.54(d，J＝16.8Hz，1H)，3.08(s，6H)，2.92-2.99(m，2H)，2.73-2.80(m，1H)，2.30-2.35(m，1H)，2.11-2.16(m，1H)，1.99(t，J＝5.6Hz，1H)，1.80-1.89(m，2H)，1.40(d，J＝10.4Hz，1H)，1.37(s，3H)，1.26(s，3H)，0.85(s，3H)。

And C: (R) - (2- (benzofuran-3-yl) -1- (2- (3-dimethylsulfinimidophenyl) acetamido) ethyl) boronic acid

2(R) -2- (benzofuran-3-yl) -1- (2- (3-dimethylsulfinimidophenyl) acetamido) ethylboronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester (30mg) was dissolved in methanol (1mL), isobutylboronic acid (28mg), 1mol/l hydrochloric acid (0.2mL) and n-hexane (1mL) were added to the solution, stirred at room temperature overnight, liquid-separated to remove the upper n-hexane, and the methanol phase was washed three times with n-hexane and then concentrated to dryness at 30 ℃ to give a crude product, ether was added and slurried, the solid was collected by filtration, and the solid was dried to give a product (15 mg).

¹H NMR(400MHz，CD₃OD)δ7.57(s，1H)，7.56(d，J＝8.4Hz，1H)，7.42-7.47(m，2H)，7.24-7.33(m，4H)，7.21(t，J＝7.6Hz，1H)，3.74-3.77(m，8H)，3.01(dd，J＝9.6Hz，5.2Hz，1H)，2.89(dd，J＝14.8Hz，4.8Hz，1H)，2.71(dd，J＝14.8Hz，9.6Hz，1H)。

Example 4

(R) - (1- (3- (6- ((dimethyl)Radical (oxo) -lambda⁶-Thioalkylene) amino) pyridin-2-yl) ureido) -2-phenylethyl) boronic acid

Step A: ((6-bromopyridin-2-yl) imino) dimethyl-lambda⁶-sulfenimide

N₂A suspension of 1, 4-dioxane (100mL) containing 2, 6-dibromopyridine (6.00g), dimethylsulfinimide (1.63g), 4, 5-bis (diphenylphosphino) -9, 9-dimethylxanthene (1.50g), tris (dibenzylideneacetone) dipalladium (0.793g) and cesium carbonate (8.59g) was heated to reflux overnight under protection. After cooling to room temperature, filtration and rinsing with dichloromethane, the filtrate was freed of solvent and purified by column chromatography on silica gel (100% AcOEt) to give the product (4.0 g).

¹H NMR(400MHz，CDCl₃)δ7.31-7.35(m，1H)，6.93(dd，J＝7.6Hz，0.8Hz，1H)，6.69(dd，J＝7.6Hz，0.8Hz，1H)，3.36(s，6H)。

And B: ((6-aminopyridin-2-yl) imino) dimethyl-lambda⁶-sulfenimide

Will contain ((6-bromopyridin-2-yl) imino) dimethyl-lambda⁶A mixture of-sulfinimide (4.0g), cuprous oxide (0.230g), N, N' -dimethylethylenediamine (0.142g), anhydrous potassium carbonate (4.43g), 25% aqueous ammonia (20mL) and ethylene glycol (20mL) was heated to 60 ℃ and stirred overnight. After cooling to room temperature, filtration through celite and rinsing with dichloromethane, the aqueous phase after separation was extracted with dichloromethane, the organic phase was dried over anhydrous sodium sulphate and filtered, and the solvent was removed by evaporation to dryness to give the product (1.4 g).

¹H NMR(400MHz，CDCl₃)δ7.27-7.31(m，1H)，6.19(d，J＝7.6Hz，1H)，6.04(d，J＝7.6Hz，1H)，4.26(brs，2H)，3.35(s，6H)。

And C: benzylboronic acid pinacol ester

Referring to the procedure of step C of example 1, benzyl chloride, cuprous iodide, pinacol ester of diboronic acid, triphenylphosphine and lithium tert-butoxide were reacted to give the product (2.5 g).

¹H NMR(400MHz，CDCl₃)δ7.23-7.27(m，2H)，7.17-7.19(m，2H)，7.13(t，J＝7.6Hz，1H)，2.29(s，2H)，1.25(s，12H)。

Step D: benzylboronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester

Referring to the procedure of step D of example 1, benzylboronic acid pinacol ester and (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol were reacted as main raw materials to obtain a product (2.0 g).

¹H NMR(400MHz，CDCl₃)δ7.22-7.27(m，2H)，7.18-7.20(m，2H)，7.13(t，J＝7.6Hz，1H)，4.27(dd，J＝8.4Hz，2.0Hz，1H)，2.32(s，2H)，2.17-2.30(m，2H)，2.04(t，J＝6.0Hz，1H)，1.80-1.90(m，2H)，1.37(s，3H)，1.25(s，3H)，1.06(d，J＝10.4Hz，1H)，0.87(s，3H)。

Step E: 1- (S) -chloro-2-phenylethylboronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester

Referring to step E of example 1, benzylboronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester, dichloromethane, butyllithium and zinc chloride tetrahydrofuran solution were reacted as main raw materials to obtain a crude product (2.3g) as a pale yellow oil to be directly used for the next reaction.

¹H NMR(400MHz，CDCl₃)δ7.22-7.31(m，3H)，7.12-7.16(m，1H)，4.77(d，J＝8.8Hz，1H)，3.59-3.63(m，1H)，3.18-3.23(m，1H)，3.08-3.12(m，1H)，2.32-2.37(m，1H)，2.15-2.22(m，2H)，2.05(t，J＝5.6Hz，1H)，1.82-1.91(m，2H)，1.35(s，3H)，1.27(s，3H)，1.06(d，J＝10.8Hz，1H)，0.85(s，3H)。

Step F: 2-phenyl-1- (R) -aminoethylboronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester hydrochloride

Referring to the procedure of step F in example 1, 1- (S) -chloro-2-phenylethylboronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester, lithium bistrimethylsilylamide tetrahydrofuran solution and 1, 4-dioxane solution of hydrogen chloride were reacted as main raw materials to obtain a product (0.70 g).

¹H NMR(400MHz，CDCl₃)δ7.97-8.16(brs，3H)，7.34-7.43(m，3H)，7.17-7.26(m，2H)，4.35(d，J＝8.4Hz，1H)，2.80-3.06(m，3H)，2.17-2.20(m，1H)，1.95-2.13(m，2H)，1.80-1.88(m，2H)，1.27(s，3H)，1.20(s，3H)，1.03-1.06(m，1H)，0.76(s，3H)。

Step G: (R) - (1- (3- (6- ((dimethyl (oxo) -lambda)⁶-Thioalkylene) amino) pyridin-2-yl) ureido) -2-phenylethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester

2-phenyl-1- (R) -aminoethylboronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester hydrochloride (150mg) and triethylamine (162mg) were added to the dry solutionUnder nitrogen, cooling to-60 deg.C, adding dropwise bis (trichloromethyl) carbonate (48mg) in dichloromethane to the reaction solution, stirring at-60 deg.C for 1 hr, and reacting at-60 deg.C⁶A solution of sulfinimide (81mg) in dichloromethane was added dropwise to the above solution, stirred at-60 ℃ for 1 hour, quenched with water, extracted with dichloromethane, washed with saturated brine, the organic phase was dried over anhydrous sodium sulfate, filtered and the solvent evaporated to dryness to give a residue which was purified on silica gel preparation plates (1: 30 methanol/dichloromethane) to give a product (20 mg).

¹H NMR(400MHz，CDCl₃)δ8.97(s，1H)，7.48(s，1H)，7.43(t，J＝8.0Hz，1H)，7.28(d，J＝7.2Hz，2H)，7.21(t，J＝7.6Hz，2H)，7.11(t，J＝7.2Hz，1H)，6.36(d，J＝8.4Hz，1H)，6.28(d，J＝8.0Hz，1H)，4.18(d，J＝7.2Hz，1H)，3.14-3.21(m，1H)，3.12(s，3H)，2.94-3.05(m，5H)，2.18-2.22(m，1H)，1.85-1.94(m，2H)，1.71-1.81(m，2H)，1.31(s，3H)，1.20(s，3H)，1.04(d，J＝9.2Hz，1H)，0.80(s，3H)。

Step H: (R) - (1- (3- (6- ((dimethyl (oxo) -lambda)⁶-Thioalkylene) amino) pyridin-2-yl) ureido) -2-phenylethyl) boronic acid

Mixing (R) - (1- (3- (6- ((dimethyl (oxo) -lambda)⁶-Thioalkylene) amino) pyridin-2-yl) ureido) -2-phenylethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester (20mg) was dissolved in methanol (1mL), to the solution were added isobutylboronic acid (12mg), 1mol/l hydrochloric acid (0.1mL) and n-hexane (1mL), stirred at room temperature overnight, and the upper n-hexane layer was removed by liquid separation, and the methanol phase was washed three times with n-hexane, concentrated to dryness at 30 ℃ diluted with dichloromethane and the product washed to the aqueous phase with 2 mol/l aqueous sodium hydroxide (5mL), the aqueous phase washed three times with dichloromethane and acidified to acidic pH with 3 mol/l hydrochloric acid, extracted three times with dichloromethane, the organic phase dried over anhydrous sodium sulfate, filtered and concentrated.Concentration at 30 ℃ removed the solvent to give the product (10 mg).

¹H NMR(400MHz，CD₃OD)δ7.81(t，J＝8.0Hz，1H)，7.15-7.27(m，5H)，6.76(d，J＝7.6Hz，1H)，6.46(d，J＝8.4Hz，1H)，3.44(s，3H)，3.43(s，3H)，3.28-3.31(m，1H)，2.84-2.94(m，2H)。

Example 5

((1R) -1- (((((((2R) -1- (N, S-dimethylsulfonylimino) pyrrolidin-2-yl) methoxy) carbonyl) amino) -2-phenylethyl) boronic acid

Step A: ((1R) -1- (((((((2R) -1- (N, S-dimethylsulfonylimino) pyrrolidin-2-yl) methoxy) carbonyl) amino) -2-phenylethyl) boronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester

Adding 2-phenyl-1- (R) -aminoethylboronic acid- (1S, 2S, 3R, 5S) - (+) -pinane-2, 3-diol ester hydrochloride (150mg) and triethylamine (162mg) to dried dichloromethane, protecting with nitrogen and cooling to-60 ℃, dropwise adding a dichloromethane solution of bis (trichloromethyl) carbonate (48mg) to the reaction solution, stirring at-60 ℃ for 1 hour after completion, dropwise adding a dichloromethane solution of ((2R) -1- (N, S-dimethylsulfonylimino) pyrrolidin-2-yl) methanol (93mg) to the above solution at-60 ℃, stirring at-60 ℃ for 1 hour, quenching with water, extracting with dichloromethane, washing with saturated brine, drying the organic phase over anhydrous sodium sulfate, the residue obtained by filtration to dryness of the solvent was purified on a silica gel preparation plate (1: 35 methanol/dichloromethane) to give the product (105 mg).

¹H NMR(400MHz，CDCl₃)δ7.24-7.28(m，2H)，7.16-7.21(m，3H)，4.81(d，J＝5.2Hz，1H)，4.32(d，J＝8.8Hz，1H)，4.04-4.14(m，2H)，3.86-3.94(m，0.4H)，3.73-3.80(m，0.6H)，3.38-3.44(m，1H)，3.14-3.32(m，2H)，3.01(dd，J＝14.0Hz，4.8Hz，1H)，2.77-2.88(m，4H)，2.67(s，1.2H)，2.66(s，1.8H)，2.29-2.34(m，1H)，2.13-2.18(m，1H)，1.81-2.01(m，7H)，1.31(s，3H)，1.26(s，3H)，1.11(d，J＝11.2Hz，1H)，0.82(s，3H)。

And B: ((1R) -1- (((((((2R) -1- (N, S-dimethylsulfonylimino) pyrrolidin-2-yl) methoxy) carbonyl) amino) -2-phenylethyl) boronic acid

((1R) -1- ((((((2R) -1- (N, S-dimethylsulfonylimino) pyrrolidin-2-yl) methoxy) carbonyl) amino) -2-phenylethyl) boronic acid- (1S, 2S, 3R5S) - (+) -pinane-2, 3-diol ester (105mg) was dissolved in methanol, isobutylboronic acid (62mg), 1 mole/liter hydrochloric acid (0.1mL) and N-hexane were added to the solution, stirred at room temperature overnight, the upper N-hexane was removed by liquid separation, and the methanol phase was washed three times with N-hexane and then concentrated at 30 ℃ to dryness and diluted with dichloromethane and the product was washed with 2M aqueous sodium hydroxide solution (5mL), the aqueous phase was washed three times with dichloromethane and then acidified to pH acidity with 3 mole/liter hydrochloric acid, after extraction with dichloromethane three times, the organic phase was dried over anhydrous sodium sulfate, filtered and concentrated at 30 ℃ to remove the solvent to give the product (40 mg).

¹H NMR(400MHz，CD₃OD)δ7.16-7.26(m，5H)，3.94-4.22(m，3H)，3.33-3.43(m，5H)，3.20-3.25(m，1H)，2.73-2.90(m，5H)，1.94-2.18(m，4H)。

Biological assay

1. Determination of the enzymatic inhibitory activity of the compounds on LMP 7:

LMP7 is the catalytic subunit of immunoproteasome, and this experiment utilizes its hydrolase activity to establish the enzymology detection method platform and is used for the activity detection of compound. The amount of fluorophore AMC (7-Amino-4-methylcoumarin ) released after hydrolysis was enzymatically active using Ac-ANW-AMC (Bonston biochem, Cat # S-320) as a substrate for LMP 7. MOLT-4 cells are human acute lymphoblastic leukocytesA leukemia cell identified as a cell highly expressing LMP 7. We used MOLT-4 cell lysate as enzyme source of LMP7 to establish the enzymatic detection method of LMP7 by the compound and carry out the compound inhibition activity (half inhibition concentration, IC)₅₀) Detection of (3).

MOLT-4 cells were cultured in RPMI-1640(Biological Industries) medium containing 10% fetal bovine serum (Biological Industries) and 1% Pen Strep (Gibco) at 75cm²Culturing was carried out in a cell culture flask (Corning) (37 ℃, 95% air and 5% CO)₂) And passage 2-3 times a week. Collection of 1X10⁷MOLT-4 cells were resuspended in 1ml PBS (Solarbio), 3000rpm and centrifuged for 5 minutes. The supernatant was aspirated off. The cells were resuspended in 500. mu.l lysis buffer (20mM Tris, pH 8.0, 5mM EDTA, Protease Inhibitor (1: 1000) and Phosphatase Inhibitor (1: 100) in time) and placed on ice for 30 minutes. Cells were sonicated, 0.5S on, 0.5S off, sonication time 2.5S. Centrifuge at 12000rpm, 4 ℃ for 10 minutes. The supernatant was a cell lysate, and the protein was quantified by BCA method (Thermo, # 23225).

Compounds were diluted in 5-fold gradients with 100% DMSO at 9 concentrations, and 2 μ l of each was added to 48 μ l of reaction buffer (20mM Tris, pH 8.0, 0.5mM EDTA) and mixed well as 4 × compound (final concentration 2000, 400, 80, 16, 3.2, 0.64, 0.128, 0.0256, 0nM) for use. 4 MOLT-4 cell lysates were prepared in reaction buffer at 20 ng/. mu.l, 2 Ac-ANW-AMC at 100. mu.M. Mu.l of 4X compound was added to a 384 well plate (OptiPlate-384, purchased from Perkinelmer), 5. mu.l of 4X cell lysate was added, centrifuged, and reacted in an incubator at 23 ℃ for 1 hour. 10. mu.l of 2X Ac-ANW-AMC was added, and the reaction was started by centrifugation and then carried out for 2 hours at 23 ℃ in the absence of light. After the reaction is finished, CLARIO star^PlusThe signal values (excitation wavelength 345 nm/emission wavelength 445nm) were read (purchased from BMG LRBTECH). The enzyme activity was measured at 9 concentrations for each compound, and the data were processed using GraphPad Prism software to calculate the half inhibitory concentration of the compound against LMP7, i.e., IC₅₀The value is obtained.

2. Assay of compounds for β 5 enzymatic inhibitory activity:

beta 5 is a catalytic subunit of the proteasomeThe experiment utilizes the hydrolase activity to establish an enzymology detection method platform and is used for detecting the activity of the compound. The amount of fluorophore AMC (7-Amino-4-methylcoumazin, 7-Amino-4-methylcoumarin) released after hydrolysis can be reacted enzymatically using Ac-WLA-AMC (Bonston biochem, Cat # S-330) as substrate for β 5. HEK-293 is a human embryonic kidney cell, constitutively expresses proteasomes, and does not express immunoproteasome. We established an enzymatic detection method of beta 5 by using HEK-293 cell lysate as an enzyme source of beta 5 and carried out compound inhibitory activity (semi-inhibitory concentration, IC)₅₀) Detection of (3). The enzymatic inhibitory activity of the compound on beta 5 is used as an index for detecting the selectivity of the compound.

HEK-293 cells were cultured in DMEM (Biological Industries) medium containing 10% fetal bovine serum (Biological Industries) and 1% Pen Strep (Gibco) at 75cm²Culturing was carried out in a cell culture flask (Corning) (37 ℃, 95% air and 5% CO)₂) And passage 2-3 times a week. Collection of 1X10⁷HEK-293 cells were resuspended in 1ml PBS (Solarbio), 3000rpm and centrifuged for 5 min. The supernatant was aspirated off. The cells were resuspended in 500. mu.l lysis buffer (20mM Tris, pH 8.0, 5mM EDTA, Protease Inhibitor (1: 1000) and Phosphatase Inhibitor (1: 100) in time) and placed on ice for 30 minutes. Cells were sonicated, 0.5S on, 0.5S off, sonication time 2.5S. Centrifuge at 12000rpm, 4 ℃ for 10 minutes. The supernatant was a cell lysate, and the protein was quantified by BCA method (Thermo, # 23225).

Compounds were diluted in 5-fold gradients with 100% DMSO at 9 concentrations, and 2 μ l of each was added to 48 μ l of reaction buffer (20mM Tris, pH 8.0, 0.5mM EDTA) and mixed well as 4 × compound (final concentration 100000, 20000, 4000, 800, 160, 32, 6.4, 1.28, 0nM) for use. 4 HEK-293 cell lysates were prepared in reaction buffer at 25 ng/. mu.l, 2 Ac-WLA-AMC at 20. mu.M. Mu.l of 4 compounds were added to a 384 well plate (OptiPlate-384, purchased from Perkinelmer), 5. mu.l of 4 HEK-293 cell lysate was added, centrifuged, and reacted in an incubator at 23 ℃ for 1 hour. 10. mu.l of 2. mu.Ac-WLA-AMC was added, and the reaction was started by centrifugation and then carried out at 23 ℃ for 2 hours in the absence of light. After the reaction is finished, CLARIO star^PlusThe signal values (excitation wavelength 345 nm/emission wavelength 445nm) were read (purchased from BMG LRBTECH). The activity of the enzyme was measured at 9 concentrations for each compound, and the data were processed using GraphPad Prism software to calculate the half inhibitory concentration of the compound against β 5, i.e., IC₅₀The value is obtained.

The foregoing "+" means multiplication, indicating a multiple.

The results of the tests on the above partial compounds are shown in table 1.

Table 1: results of in vitro Activity test

3. Animal pharmacokinetic studies of compounds:

animal drug experiments 3 healthy adult male rats from experimental animal technology ltd, viton, beijing were used. The compound is suspended in 20 percent solution (W/W/V) of sulfobutyl ether-beta-cyclodextrin, the concentration of the solution is 1mg/mL, the administration volume is 5mL/kg, and the dosage is 5mg/kg through single intragastric administration. Animals were fasted overnight prior to the experiment, with the fasting time ranging from 10 hours prior to dosing to 4 hours post-dosing. Blood was collected at 0.25, 0.5, 1, 2, 4, 6, 8 and 24 hours post-dose. Animals were lightly anesthetized with isoflurane, approximately 0.4mL of whole blood was collected from the orbital venous plexus using a glass blood collection tube, placed in a heparin anticoagulation tube, the sample centrifuged at 4 ℃, 4200rpm for 5min, and the plasma transferred to a centrifuge tube and stored at-80 ℃ until analysis. Plasma sample analysis test compounds and internal standards (warfarin or propranolol) were extracted from rat plasma using acetonitrile protein precipitation and the extracts were analyzed by LC/MS/MS. The measured plasma concentration-time data of individual animals were analyzed using a non-compartmental model of the software WinNonlin (version 5.2.1; Pharsight corporation) to obtain the following pharmacokinetic parameters: maximum (peak) plasma drug concentration C_max(ii) a Time to peak T_max(ii) a Half life T_1/2And the area under the plasma concentration-time curve AUC extrapolated to infinite time_0-inf。

TABLE 2 pharmacokinetic test results

Claims (10)

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

wherein,

R^aand R^bEach independently selected from H and C_1-6Alkyl, or R^aAnd R^bMay be joined together to form a 3-to 10-membered heterocyclic ring;

x is a bond, -O-, or-NR⁴-；

Y is a bond or- (CR)⁴R⁵)_m-；

R⁴And R⁵Each independently selected from H, C_1-6Alkyl and C_3-8A cycloalkyl group;

m is 1, 2, or 3;

R²selected from H and C_1-6An alkyl group;

R³is selected from C_6-10Aryl and C_5-10Heteroaryl, said aryl and heteroaryl being optionally substituted by halogen, -OH, -NH₂、-O-C_1-6Alkyl, -N (C)_1-6Alkyl) (C_1-6Alkyl), -CN, NO₂、C_1-6Alkyl radical, C_3-8Cycloalkyl, or C_3-8Heterocycloalkyl substitution;

R¹is selected from C_1-6Alkyl radical, C_3-8Cycloalkyl radical, C_3-8Heterocycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said alkyl, cycloalkyl, heterocycloalkyl, aryl and heteroaryl being substituted by

And can be substituted by one ofOptionally substituted by halogen, -OH, -NH₂、-(CH₂)_1-3-C_3-8Cycloalkyl, - (CH)₂)_0-6-CF₃、-O-C_1-6Alkyl, -NR⁹R⁸、-CN、NO₂、C_1-6Alkyl, - (CH)₂)_0-3-(CO)-R⁸、-(CH₂)_0-3-(CO)-NH-R⁸、-(CH₂)_0-3-NH-(CO)-R⁸Or R¹⁰Substitution;

R^6aand R^6bEach independently selected from C_1-6Alkyl radical, C_3-8Cycloalkyl radical, C_3-8Heterocycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said alkyl, cycloalkyl, heterocycloalkyl, aryl and heteroaryl being optionally substituted by halogen, C_1-6Alkyl radical, C_3-8Cycloalkyl, -OH, -NH₂、-O-C_1-6Alkyl, -NR⁹R⁸、-NO₂or-CN substitution, or

R^6aAnd R^6bMay be joined together to form a 3-8 membered heterocyclic ring;

R⁷selected from H, C_1-6Alkyl radical, C_3-8Cycloalkyl radical, C_3-8Heterocycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said alkyl, cycloalkyl, heterocycloalkyl, aryl and heteroaryl being optionally substituted by halogen, C_1-6Alkyl radical, C_3-8Cycloalkyl, -OH, -NH₂、-O-C_1-6Alkyl, -NR⁹R⁸、-NO₂or-CN substitution;

R⁸is selected from C_1-6Alkyl radical, C_3-8Cycloalkyl radical, C_3-8Heterocycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said alkyl, cycloalkyl, heterocycloalkyl, aryl and heteroaryl being optionally substituted by halogen, C_1-6Alkyl, -O-C_1-6Alkyl radical, C_6-10Aryl, or C_5-10Heteroaryl substitution;

R⁹selected from H and C_1-6An alkyl group;

R¹⁰is selected from C_3-8Cycloalkyl radical, C_3-8Heterocycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said cycloalkyl, heterocycloalkyl, aryl and heteroaryl being optionally substituted by halogen, -OH, -NH₂、-O-C_1-6Alkyl, -N (C)_1-6Alkyl) (C_1-6Alkyl), -CN, NO₂Or C_1-6Alkyl substitution.

2. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, wherein X is a bond, -O-, or-NR⁴-，R⁴Is H.

3. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, wherein Y is a bond or- (CR)⁴R⁵)_m-，R⁴And R⁵Each independently selected from H and C_1-6Alkyl, m is 1 or 2.

4. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, wherein R²Is H.

5. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, wherein R³Is selected from C_5-10Heteroaryl, said heteroaryl being optionally substituted by halogen, NO₂、C_1-6Alkyl radical, C_3-8Cycloalkyl, or C_3-8Heterocycloalkyl substituted.

6. The compound of claim 1, or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, wherein R¹Is selected from C_3-8Cycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said cycloalkyl, aryl and heteroaryl being substituted by

And may optionally be substituted by halogen, -CF₃、-O-C_1-6Alkyl radical, NO₂、C_1-6Alkyl, - (CO) -R⁸、-(CO)-NH-R⁸Or R¹⁰Substitution;

R^6aand R^6bEach independently selected from C_1-6Alkyl radical, C_3-8Cycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said alkyl, cycloalkyl, aryl and heteroaryl optionally being substituted by halogen, C_1-6Alkyl, -O-C_1-6Alkyl radical, C_3-8Cycloalkyl radical, C_3-8Heterocycloalkyl, or NR⁸R⁹Substituted, or R^6aAnd R^6bMay be joined together to form a 3-8 membered heterocyclic ring;

R⁷selected from H, C_1-6Alkyl radical, C_3-8Cycloalkyl radical, C_6-10Aryl and C_5-10Heteroaryl, said alkyl, cycloalkyl, aryl and heteroaryl optionally being substituted by halogen, C_1-6Alkyl, -O-C_1-6Alkyl radical, C_3-8Cycloalkyl radical, C_3-8Heterocycloalkyl, or NR⁸R⁹Substitution;

R⁸is selected from C_1-6Alkyl and C_3-8Cycloalkyl, said cycloalkyl being optionally substituted by C_1-6Alkyl substitution;

R⁹selected from H and C_1-6Alkyl radical

R¹⁰Is selected from C_3-8Cycloalkyl and C_3-8Heterocycloalkyl, said cycloalkyl and heterocycloalkyl being optionally substituted by C_1-6Alkyl substitution.

7. The following compounds

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

8. A pharmaceutical composition comprising a compound according to any one of claims 1-7, or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof, and a pharmaceutically acceptable carrier.

9. Use of a compound according to any one of claims 1 to 7 or a pharmaceutically acceptable salt, solvate, polymorph or isomer thereof or a composition according to claim 8 in the manufacture of a medicament for the treatment of a disease associated with lmp7 activity.

10. The use of claim 9, wherein the disease associated with lmp7 activity is multiple myeloma, acute myelogenous leukemia, myeloid leukemia, mantle cell lymphoma, chronic lymphocytic leukemia, acute lymphocytic leukemia, diffuse large B-cell lymphoma, plasmacytoma, follicular lymphoma, immune cell tumor, breast cancer, liver cancer, colorectal cancer, ovarian cancer, esophageal cancer, lung cancer, head and neck cancer, pancreatic cancer, kidney cancer, stomach cancer, thyroid cancer, prostate cancer, bladder cancer, rheumatoid arthritis, systemic lupus erythematosus, inflammatory bowel disease, multiple sclerosis, scleroderma, ankylosing spondylitis, atherosclerosis, behcet's disease, crohn's disease, inflammatory bowel disease, ulcerative colitis, autoimmune hepatitis, sjogren's syndrome, lupus nephritis, asthma, Amyotrophic Lateral Sclerosis (ALS), Psoriasis, immunoglobulin A nephropathy, allergic purpura, Alzheimer's Disease (AD).