CN113583020B

CN113583020B - JAK2 inhibitor and application

Info

Publication number: CN113583020B
Application number: CN202010366980.2A
Authority: CN
Inventors: 不公告发明人
Original assignee: 百极弘烨（广东）医药科技有限公司; 百极弘烨(南通)医药科技有限公司
Current assignee: Baiji Hongye Guangdong Pharmaceutical Technology Co ltd; Baiji Hongye Nantong Pharmaceutical Technology Co Ltd
Priority date: 2020-04-30
Filing date: 2020-04-30
Publication date: 2022-04-22
Anticipated expiration: 2040-04-30
Also published as: CN113583020A; WO2021219140A1

Abstract

The invention provides a JAK2 inhibitor and application thereof, and particularly relates to a compound shown as a formula I, or a stereoisomer or an optical isomer, a pharmaceutically acceptable salt, a prodrug or a solvate thereof, a pharmaceutical composition of the compound, a JAK inhibitor prepared from the compound, and medical application of the compound in preparation of medicines for preventing and/or treating JAK, especially JAK2 related diseases.

Description

JAK2 inhibitor and application

Technical Field

The invention belongs to the field of medicinal chemistry, and particularly relates to a JAK2 inhibitor and application thereof.

Background

Protein Kinases (PKs) are a group of enzymes that regulate a variety of important biological processes, which constitute one of the largest families of enzymes in humans, including, inter alia, cellular kinases that catalyze the phosphorylation of proteins, lipids, sugars, nucleosides, and other cellular metabolites and play a key role in all aspects of eukaryotic cell physiology. Abnormal kinase activity has been implicated in a number of human diseases, including cancer, autoimmune and inflammatory diseases.

Janus kinases (JAKs) are cytoplasmic tyrosine kinases that transduce cytokine signals from membrane receptors to STAT transcription factors, which play an important role in cytokine signaling. The JAK family includes four members JAK1, JAK2, JAK3 and tyrosine kinase 2(TYK 2). JAKs are typically associated in pairs with cytokine receptors as homodimers or heterodimers. Cytokines bind to their receptors, cause dimerization of the receptor molecules, and JAKs coupled to the receptors approach each other and are activated by phosphorylation of interacting tyrosine residues. The JAK family delivers cytokine-mediated signals into cells via the JAK-STAT (signal transduction and activator of transcription) pathway.

Signal Transducers and Activators of Transcription (STATs) are a group of cytoplasmic proteins that bind to the DNA of the regulatory region of a target gene. As downstream substrates of JAKs, STATs can be activated by tyrosine phosphorylation under the stimulation of external signals and then transferred into the transcription of nuclear regulatory genes. When cytokines bind to their receptors, JAK family members are autophosphorylated and/or transphosphorylated to each other, followed by phosphorylation of STATs, which then migrate into the nucleus to regulate transcription.

Many abnormal immune responses, such as allergies, asthma, (allograft) rejection, rheumatoid arthritis, autoimmune diseases such as amyotrophic lateral sclerosis and multiple sclerosis, myeloproliferative disorders, hematologic malignancies such as leukemia and lymphoma, whose regulation is associated with the JAK/STAT signaling pathway.

It was found that point mutations in JAK2 pseudo kinase domain V617F were identified in greater than 90% of polycythemia vera patients and in about 50% of patients with primary myelofibrosis and primary thrombocythemia. This mutation was also found in Ph negative chronic myelogenous leukemia, chronic myelogenous monocytic leukemia, the pathogenesis of megakaryocytic acute myelogenous leukemia, and juvenile myelogenous monocytic leukemia (10-20%). Other mutations in the pseudo kinase domain of JAK2, including the point mutation of Arg683, have been detected in approximately 20% of down syndrome-associated acute lymphocytic and acute myeloid leukemias.

Currently, four JAK inhibitors are available on the market, but they all belong to pan JAK inhibitors and exhibit varying degrees of selectivity for JAK1, JAK2, JAK3 and TYK 2. Among JAK inhibitors, lack of selectivity for JAK2 can lead to several side effects, particularly an increased risk of infection. Therefore, there is a need in the art to develop small molecule inhibitors that target JAKs, particularly JAK2 kinase.

Disclosure of Invention

The invention provides a novel JAK-targeting inhibitor, which has a selective inhibition effect on JAK, particularly JAK2, and thus has clinical significance and application prospect.

In a first aspect of the invention, there is provided a compound of formula I or a stereoisomer or an optical isomer, a pharmaceutically acceptable salt, a prodrug or a solvate thereof,

in the formula (I), the compound is shown in the specification,

X₁、X₂、X₃and X₄Each independently selected from the group consisting of: n, C or C-R_d(ii) a And X₁、X₂、X₃And X₄0, 1,2 and 3 are N;

or, when X₁Is C-R_dWhen R is_dAnd X₂Fused to form a substituted or unsubstituted 5-6 membered cycloalkyl, heterocyclyl, aryl or heteroaryl group;

or, when X₂Is C-R_dWhen R is_dAnd X₁Fused to form a substituted or unsubstituted 5-6 membered cycloalkyl, heterocyclyl, aryl or heteroaryl group;

or, when X₃Is C-R_dWhen R is_dAnd X₄Fused to form a substituted or unsubstituted 5-6 membered cycloalkyl, heterocyclyl, aryl or heteroaryl group;

or, when X₄Is C-R_dWhen R is_dAnd X₃Fused to form a substituted or unsubstituted 5-6 membered cycloalkyl, heterocyclyl, aryl or heteroaryl group;

x is selected from the group consisting of: key, CR_bR_c、C(＝O)O-、O、N-R_b、S、SO、SO₂C3-C10 cycloalkylene, 3-10 membered heterocyclylene, 5-12 membered heteroarylene, C6-C12 arylene;

R₃、R₄、R₅、R₆、R₇、R₈、R₉、R_b、R_cand R_dEach independently selected from the group consisting of: H. d, halogen, amino, nitro, hydroxyl, cyano, carboxyl, sulfuryl, sulfoxide, amide, sulfonamide, ester, formyl, carboxamide, C1-C6 alkyl, C1-C6 alkoxy, C2-C6 alkenyl, C2-C6 alkynyl, 3-10 membered heterocyclic group, C3-C10 cycloalkyl, 5-12 membered heteroaryl, C6-C12 aryl;

or R₃And R₄Together with the C atom to which they are attached form a substituted or unsubstituted 5-6 membered cycloalkyl, heterocyclyl, aryl or heteroaryl group;

or R₅And R₆Together with the C atom to which they are attached form a substituted or unsubstituted 5-6 membered cycloalkyl, heterocyclyl, aryl or heteroaryl group;

or R_bAnd R_cTogether with the C atom to which they are attached form ═ O, or substituted or unsubstituted C3-C10 cycloalkylene, 3-10 membered heterocyclylene;

(CH₂) The H atom in n may optionally be substituted by one or more R_aSubstitution;

n is 1,2,3, 4, 5, 6, 7 or 8;

unless otherwise specified, the substitution refers to substitution by one or more groups selected from the group consisting of: D. halogen, amino, nitro, hydroxyl, cyano, carboxyl, sulfone, sulfoxide, amide, sulfonamide, ester, formyl, carboxamide, C1-C6 alkyl, C1-C6 alkoxy, C2-C6 alkenyl, C2-C6 alkynyl, 3-10 membered heterocyclic, C3-C10 cycloalkyl, 5-12 membered heteroaryl, C6-C12 aryl;

the above alkyl, alkoxy, alkenyl, alkynyl, heterocycloalkyl, cycloalkyl, heteroaryl, aryl groups may be further optionally substituted with 1 or more R, unless otherwise specified_aIn which each R is_aIndependently selected from the group consisting of: halogen, amino, nitro, hydroxyl, sulfydryl, cyano, carboxyl, sulfuryl, sulfoxide, amido, sulfamide, ester, formyl, formamido, C1-C6 alkyl, C1-C6 alkoxy, C2-C6 alkenyl, C2-C6 alkynyl, 3-10 membered heterocycloalkyl, C3-C10 cycloalkyl, 5-12 membered heteroaryl, C6-C12 aryl.

In another preferred embodiment, n is 1,2,3, 4 or 5.

In another preferred embodiment, X₁Is N.

In another preferred embodiment, X₂Is N.

In another preferred embodiment, X₃Is N.

In another preferred embodiment, X₄Is N.

In another preferred embodiment, the compound or its stereoisomer or optical isomer, pharmaceutically acceptable salt, prodrug or solutionAn agent, wherein,

the moiety is a group selected from the group consisting of:

wherein the content of the first and second substances,

Y₁and Y₂Each independently selected from the group consisting of: CR_bR_c、N-R_b、O、S；

R_A、R_B、R_CAnd R_DEach independently selected from the group consisting of: H. deuterium, halogen, amino, nitro, hydroxyl, mercapto, cyano, carboxyl, sulfone, sulfoxide, amide, sulfonamide, ester, formyl, carboxamide, C1-C6 alkyl, C1-C6 alkoxy, C2-C6 alkenyl, C2-C6 alkynyl, 3-10 membered heterocycloalkyl, C3-C10 cycloalkyl, 5-12 membered heteroaryl, C6-C12 aryl;

p is 1,2,3, 4;

m is 1 or 2;

R_a、R_band R_cIs as defined above.

In another preferred embodiment, the compound is a stereoisomer or an optical isomer, a pharmaceutically acceptable salt, a prodrug or a solvate thereof, wherein X is selected from the group consisting of: key, CR_bR_cO, S; wherein R is_bAnd R_cIs as defined above.

In another preferred embodiment, the compound, or a stereoisomer or an optical isomer, a pharmaceutically acceptable salt, a prodrug or a solvate thereof, has the structure shown in formula II:

in the formula (I), the compound is shown in the specification,

R₁and R₂Each independently selected from the group consisting of: H. d, halogen, amino, nitro, hydroxyl, cyano, carboxyl, sulfuryl, sulfoxide, amide, sulfonamide, ester, formyl, carboxamide, C1-C6 alkyl, C1-C6 alkoxy, C2-C6 alkenyl, C2-C6 alkynyl, 3-10 membered heterocyclic group, C3-C10 cycloalkyl, 5-12 membered heteroaryl, C6-C12 aryl;

or R₁And R₂Fused together with the C atom to which they are attached to form a substituted or unsubstituted 5-6 membered cycloalkyl, heterocyclyl, aryl or heteroaryl group;

R_a、R₃、R₄、R₅、R₆、R₇、R₈、R₉x and n are as defined above;

In another preferred embodiment, the compound, or a stereoisomer or an optical isomer, a pharmaceutically acceptable salt, a prodrug or a solvate thereof, has the structure shown in formula a:

wherein the content of the first and second substances,

R₁、R₂、R_a、R₃、R₄、R₅、R₆x and n are as defined above.

In another preferred embodiment, X₁、X₂、X₃、X₄、R₃、R₄、R₅、R₆、R₇、R₈、R₉X and n are specific groups corresponding to the respective specific compounds in examples.

In another preferred embodiment, the compound or its stereoisomer or optical isomer, pharmaceutically acceptable salt, prodrug or solvate is selected from the group consisting of:

in another preferred embodiment, the compound is the compound shown in the examples.

In a second aspect of the present invention, there is provided a pharmaceutical composition comprising a compound of the first aspect, or a stereoisomer or an optical isomer thereof, or a pharmaceutically acceptable salt, prodrug or solvate thereof, and a pharmaceutically acceptable carrier or excipient.

In another preferred embodiment, the pharmaceutical composition further comprises a drug selected from the group consisting of:

PD-1 inhibitors (e.g., nivolumab, pembrolizumab, pidilizumab, cemipimab, JS-001, SHR-120, BGB-A317, IBI-308, GLS-010, GB-226, STW204, HX008, HLX10, BAT1306, AK105, LZM 009, or the like biologically similar to the above drugs), PD-L1 inhibitors (e.g., Duvalimab, alemtuzumab, Avelumab (avelumab), CS1001, KN035, HLX20, SHR-1316, BGB-A333, JS003, CS1003, KL-A167, F520, GR, MSB2311, or the like biologically similar to the above drugs), CD20 antibodies (e.g., rituximab, otuzumab, ofatumumab, ovuzumab, vetutumumab, tositumumab, 131I-tositumomab, ibritumomab, 90-Ititutentitiu 90, Iitumomab, In-90, and In-5956, or the like), and the like, CC-90002, TTI-621, TTI-622, OSE-172, SRF-231, ALX-148, NI-1701, SHR-1603, IBI188, IMM01, ALK inhibitors (e.g., Ceritinib, Aleptinib, Bugatinib, Laratinib, Ocatinib), PI3K inhibitors (e.g., Idiranib, Duvelisib, Dactlisib, Taselisib, Bimiralisib, Omipalisib, Bupalisib, etc.), BTK inhibitors (e.g., Ibrutinib, Tirabutinib, Acatinib, Zabritinib, Vebrutinib, etc.), EGFR inhibitors (e.g., Afatinib, Gefitinib, erlotinib, Lapatinib, Darkatinib, Icotinib, Netinib, Sapatinib, Napatinib, pyrroltinib, Hirtitinib, HDAC, erlotinib, Galatinib, e, Galatinib, e, vorinostat, fimepinastat, drosrinostat, entinostat, daciclast, Quisinostat, tacrine, etc.), CDK inhibitors (e.g., palbociclib, ribbociclib, Abemaciclib, micciib, Trilaciclib, Lerociclib, etc.), MEK inhibitors (e.g., semetinib (AZD6244), trametinib (GSK1120212), PD0325901, seru 0126, pimatiib (AS-703026), PD184352(CI-1040), etc.), mTOR inhibitors (e.g., vistuertib, etc.), SHP2 inhibitors (e.g., RMC-4630, JAB-3068, TNO155, etc.), or combinations thereof.

In another preferred embodiment, there is provided a method for preparing a pharmaceutical composition, comprising the steps of: mixing a pharmaceutically acceptable carrier with a compound according to the first aspect of the invention, or a stereoisomer or an optical isomer, a pharmaceutically acceptable salt, a prodrug or a solvate thereof, to form a pharmaceutical composition.

In another preferred embodiment, the compounds of the present invention can be prepared into powders, tablets, granules, capsules, solutions, emulsions, suspensions, and the like.

In a third aspect of the present invention, there is provided a use of a compound of the first aspect, or a stereoisomer or an optical isomer thereof, or a pharmaceutically acceptable salt, prodrug or solvate thereof, for the manufacture of a medicament for the prevention or treatment of a JAK 2-mediated disease; and/or for the preparation of a JAK2 inhibitor.

In another preferred embodiment, the JAK 2-mediated disease is myelodysplastic syndrome (MDS), eosinophilia, a tumor, an inflammatory disease, or an infection caused by a bacterium, virus, or fungus.

In another preferred embodiment, the tumor is selected from the group consisting of: myeloproliferative carcinoma (MPN), melanoma, lung cancer, kidney cancer, ovarian cancer, prostate cancer, breast cancer, colon cancer, bone cancer, pancreatic cancer, skin cancer, head and neck cancer, uterine cancer, rectal cancer, anal cancer, stomach cancer, testicular cancer, carcinoma of the fallopian tubes, carcinoma of the endometrium, carcinoma of the cervix, carcinoma of the vagina, carcinoma of the vulva, hodgkin's disease, non-hodgkin's lymphoma, carcinoma of the esophagus, carcinoma of the small intestine, carcinoma of the endocrine system, carcinoma of the thyroid gland, carcinoma of the parathyroid gland, carcinoma of the adrenal gland, sarcoma of soft tissue, carcinoma of the urethra, carcinoma of the penis, acute myeloid leukemia, chronic myeloid leukemia, acute lymphocytic leukemia, chronic lymphocytic leukemia, pediatric solid tumors, lymphocytic lymphoma, carcinoma of the bladder, carcinoma of the kidney or ureter, carcinoma of the renal pelvis, tumors of the Central Nervous System (CNS), primary CNS lymphoma, tumor angiogenesis, spinal axis tumor, brain stem glioma, Pituitary adenomas, kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, T-cell lymphoma.

In another preferred embodiment, the inflammatory disease is selected from the group consisting of: rheumatoid arthritis, ankylosing spondylitis, autoimmune hemolytic anemia, arthritis, myasthenia gravis, systemic lupus erythematosus, pernicious anemia, polymyositis.

In another preferred embodiment, the virus is selected from the group consisting of: hepatitis viruses (type a, b and c), herpesviruses, influenza viruses, adenoviruses, coronaviruses, measles viruses, dengue viruses, polio viruses, rabies viruses.

In another preferred embodiment, the bacteria are selected from the group consisting of: chlamydia, rickettsia, mycobacteria, staphylococci, pneumococcus, cholera, tetanus.

In another preferred embodiment, the fungus is selected from the group consisting of: candida, aspergillus, dermatitides.

In a fourth aspect, the present invention provides a JAK2 inhibitor comprising a compound of the first aspect, or a stereoisomer or an optical isomer thereof, or a pharmaceutically acceptable salt, prodrug or solvate thereof, or a pharmaceutical composition of the second aspect.

In a fifth aspect of the invention there is provided a method of preventing or treating a disease mediated by JAK2, the method comprising administering to a subject identified or diagnosed as having a disease associated with JAK2 a therapeutically effective amount of a compound of the first aspect or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition of the second aspect.

In a sixth aspect of the invention, there is provided a method for inhibiting JAK2 kinase activity in a cell or a subject, the method comprising the step of contacting the cell or administering to the subject a compound of the first aspect or a pharmaceutical composition of the second aspect.

It is to be understood that within the scope of the present invention, the above-described features of the present invention and those specifically described below (e.g., in the examples) may be combined with each other to form new or preferred embodiments. Not to be reiterated herein, but to the extent of space.

Detailed Description

The inventors have conducted extensive and intensive studies and have unexpectedly found a series of macrocyclic compounds having novel structures. The compound can selectively inhibit JAK2 kinase, and the selectivity represented by the ratio of JAK1/JAK2 or the selectivity represented by the ratio of JAK3/JAK2 is averagely improved by tens of times or tens of times, so that the compound can become a small molecular lead compound for researching a JAK2 inhibitor, and further provides a brand-new material basis for the development of immune inflammation and anti-tumor drugs. The present invention has been completed based on this finding.

Term(s) for

In the present invention, unless otherwise specified, the terms used have the ordinary meanings well known to those skilled in the art.

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 term "about" when used in reference to a specifically recited value means that the value may vary by no more than 1% from the recited value. For example, as used herein, the expression "about 100" includes 99 and 101 and all values in between (e.g., 99.1, 99.2, 99.3, 99.4, etc.).

As used herein, the term "comprising" or "includes" can be open, semi-closed, and closed. In other words, the term also includes "consisting essentially of …," or "consisting of ….

As used herein, the term "alkyl" includes straight or branched chain alkyl groups. E.g. C₁-C₆Alkyl represents a straight or branched chain alkyl group having 1 to 6 carbon atoms, such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, and the like.

As used herein, the term "alkenyl" includes straight or branched chain alkenyl groups. E.g. C₂-C₆Alkenyl refers to straight or branched chain alkenyl groups having 2 to 6 carbon atoms, including but not limited to vinyl, allyl, 1-propenyl, isopropenyl, 1-butenyl, 2-butenyl, or the like.

As used herein, the term "alkynyl" includes straight or branched chain alkynyl groups. E.g. C₂-C₆Alkynyl refers to straight or branched chain alkynyl groups having 2 to 6 carbon atoms and includes, but is not limited to, ethynyl, propynyl, butynyl, or the like.

As used herein, the term "cycloalkyl" refers to cyclic alkyl groups containing the specified number of C atoms, such as "C3-C10 cycloalkyl" refers to cycloalkyl groups having 3-10 (preferably 3, 4, 5, 6, 7, or 8) carbon atoms. It may be a single ring, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, or the like. It may also be in the form of a double ring, for example a bridged or spiro ring. In the present invention, cycloalkyl is intended to include substituted cycloalkyl.

As used herein, the term "C1-C6 alkoxy" refers to a straight or branched chain alkoxy group having 1-6 carbon atoms; which has the formula C1-C6 alkyl-O-or-C1-C5 alkyl-O-C1-C5 alkyl (e.g., -CH₂-O-CH₂CH₃、-CH₂-O-(CH₂)₂CH₃、-CH₂CH₂-O-CH₂CH₃) Structures such as methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, tert-butoxy and the like.

As used herein, "heterocyclyl" refers to a saturated or partially saturated cyclic group having a heteroatom selected from N, S and O, and "3-10 membered heterocyclyl" refers to a saturated or partially saturated cyclic group having 3-10 atoms and wherein 1-3 atoms are heteroatoms selected from the group N, S and O. It may be monocyclic or may be in the form of a double ring, for example a bridged or spiro ring. The 3-to 10-membered heterocyclic group is preferably a 3-to 8-membered heterocyclic group, more preferably a 6-to 8-membered heterocyclic group. Specific examples may be oxetane, azetidine, tetrahydro-2H-pyranyl, piperidinyl, piperazinyl, tetrahydrofuranyl, morpholinyl, pyrrolidinyl, and the like.

As used herein, "aryl" refers to an aromatic ring group without heteroatoms in the ring, and "C6-C12 aryl" refers to an aromatic ring group having 6 to 12 carbon atoms without heteroatoms in the ring, which aryl group may be fused to a heteroaryl, heterocyclyl, or cycloalkyl ring, wherein the ring to which the parent structure is attached is an aryl ring. Such as phenyl (i.e., a six-membered aromatic ring), naphthyl, and the like, wherein a six-membered aryl is also intended to include a six-membered arylo 5-6-membered cycloalkyl and a six-membered arylo 5-6-membered heterocycloalkyl. The C6-C12 aryl group is preferably a C6-C10 aryl group. The aryl group may be optionally substituted or unsubstituted.

As used herein, "heteroaryl" refers to a cyclic aromatic group having 1 to 3 atoms as heteroatoms selected from the group consisting of N, S and O, and "5-12 membered heteroaryl" refers to a cyclic aromatic group having 5 to 12 atoms wherein 1 to 3 atoms are heteroatoms selected from the group consisting of N, S and O. It may be a single ring or a condensed ring form. Specific examples may be pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, pyrrolyl, pyrazolyl, imidazolyl, (1,2,3) -triazolyl and (1,2,4) -triazolyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl and the like. The heteroaryl ring may be fused to an aryl, heterocyclyl, or cycloalkyl ring, wherein the ring joined to the parent structure is a heteroaryl ring. Heteroaryl groups may be optionally substituted or unsubstituted. When substituted, the substituents are preferably one or more groups independently selected from alkyl, deuterated alkyl, haloalkyl, alkoxy, haloalkoxy, alkenyl, alkynyl, alkylthio, alkylamino, halogen, amino, nitro, hydroxy, mercapto, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, cycloalkylthio, oxo, amido, sulfonamido, formyl, carboxamido, carboxy, carboxylate, and the like.

When the substituent is a non-terminal substituent or the relevant group is deprived of an H atom, it is a subunit of the corresponding group, usually a divalent group, for example, an alkyl group deprived of an H atom is an alkylene group (e.g., methylene, ethylene, propylene, isopropylene)

) Butylene (e.g. butyl oxide)

) Pentylene (e.g. ethylene)

) Hexamethylene (e.g. hexamethylene)

) Heptylene (e.g. ethylene)

) Etc.), cycloalkyl corresponding cycloalkylene (e.g.:

etc.), heterocyclo-corresponding heterocyclylene (e.g.:

) Cycloalkyl corresponding heterocyclylene (e.g.:

etc.), alkoxy corresponding alkyleneoxy (-CH)₂O-、-CH₂CH₂-O-CH₂-、-CH₂OCH₂CH₂CH₂-) and the like.

As used herein, "halogen" or "halogen atom" refers to F, Cl, Br, and I. More preferably, the halogen or halogen atom is selected from F, Cl and Br.

In the present invention, the term "amido" refers to a group with the structure-CONRR ', wherein R and R' may independently represent hydrogen, alkyl, cycloalkyl, aryl, heterocyclyl, as defined above. R and R' may be the same or different in the dialkylamine fragment.

In the present invention, the term "sulfonamide group" means having the structure-SO₂NRR 'wherein R and R' may independently represent hydrogen, alkyl, cycloalkyl, aryl, heterocyclyl, as defined above. R and R' may be the same or different in the dialkylamine fragment.

In the present invention, the term "formyl" refers to a-CHO-containing group.

In the present invention, the term "carboxamide group" is meant to comprise

The carboxamide group is also intended to comprise a substituted carboxamide group of the formula

Wherein each R independently represents hydrogen, alkyl, cycloalkyl, cycloalkenyl, aryl, heteroaryl, heterocyclyl, as defined above. Each R may be the same or different.

In the present invention, "amino" means having the structure-N-RR ', R and R ' each independently represent hydrogen, alkyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, as defined above, and R ' may be the same or different.

In the present invention, "sulfoxide group" means having the formula-S (O) -R, R independently represents hydrogen, alkyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, as defined above.

In the present invention, "sulfone group" means a group having the formula-S (O)₂-R, R independently represent hydrogen, alkyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, as defined above.

In the present invention, "ester group" means having the structure-C (O) -O-R or R-C (O) -O-wherein R independently represents hydrogen, alkyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, as defined above.

In the present invention, the term "substituted" means that one or more hydrogen atoms on a specified group are replaced with a specified substituent. Particular substituents are those described correspondingly in the foregoing, or as appearing in the examples. Unless otherwise specified, a certain substituted group may have one substituent selected from a specific group at any substitutable site of the group, and the substituents may be the same or different at each position. It will be understood by those skilled in the art that the combinations of substituents contemplated by the present invention are those that are stable or chemically achievable.

Unless specifically stated to be "substituted or unsubstituted", the groups of the present invention may be substituted with a substituent selected from the group consisting of: deuterium, halogen, cyano, nitro, hydroxyl, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, 3-10 membered heterocycloalkyl, C3-C10 cycloalkyl, 5-12 membered heteroaryl, C6-C12 aryl.

In the present invention, the term "plurality" refers independently to 2,3, 4, 5.

As used herein, the term "tautomer" means that structural isomers having different energies may exceed the low energy barrier, thereby converting with each other. For example, proton tautomers (i.e., proton transmutations) include interconversion by proton shift, such as 1H-indazoles and 2H-indazoles. Valence tautomers include interconversion by recombination of some of the bonding electrons.

As used herein, the term "solvate" refers to a complex of a compound of the present invention coordinated to solvent molecules in a specific ratio.

The term "substituted" as used herein means that one or more hydrogen atoms on a particular group are replaced with a particular substituent. The specific substituents may be those described above in correspondence with the description, or may be specific substituents appearing in each example or substituents conventional in the art. Therefore, in the present invention, the substituents in the general formula may also each independently be the corresponding group in the specific compounds in the examples; that is, the present invention includes both combinations of the respective substituents in the above general formulae and combinations of partial substituents shown in the general formulae with other specific substituents appearing in the examples. Preparing compounds having such combinations of substituents and testing the resulting compounds for activity is readily accomplished by those skilled in the art based on routine skill in the art.

Based on the teachings of the present invention and the general knowledge in the art, one skilled in the art will appreciate that various groups in the compounds of the present invention can be further substituted to provide derivatives that have the same or similar activity as the specifically disclosed compounds of the present invention. Each group in the compounds of the present invention may be substituted with various substituents which are conventional in the art, as long as such substitution does not violate the rules of chemical synthesis or the rules of valency.

Active ingredient

As used herein, the terms "compound of the invention" or "active ingredient of the invention" are used interchangeably to refer to formula (I) or a stereoisomer or optical isomer, pharmaceutically acceptable salt, prodrug or solvate thereof. The term also includes racemates, optical isomers,

in the formula I, X₁、X₂、X₃、X₄、R₃、R₄、R₅、R₆、R₇、R₈、R₉N has the definitions as described above.

Preferably, the compounds of the present invention have the structure shown in formula II:

wherein R is₁、R₂、R₃、R₄、R₅、R₆、R₇、R₈、R₉X and n are as defined above.

Preferably, the compounds of the present invention have the structure shown in formula a:

wherein the content of the first and second substances,

R₁、R₂、R₃、R₄、R₅、R₆、R₇、R₈、R₉x and n are as defined above.

The salts which the compounds of the invention may form are also within the scope of the invention. Unless otherwise indicated, the compounds of the present invention are understood to include salts thereof. The term "salt" as used herein refers to a salt formed from an inorganic or organic acid and a base in either an acidic or basic form. Furthermore, when a compound of the present invention contains a basic moiety, including but not limited to pyridine or imidazole, and an acidic moiety, including but not limited to carboxylic acid, zwitterions ("inner salts") that may form are included within the scope of the term "salt(s)". Pharmaceutically acceptable (i.e., non-toxic, physiologically acceptable) salts are preferred, although other salts are also useful, e.g., in isolation or purification steps during manufacture. The compounds of the invention may form salts, for example, by reacting compound I with an amount of acid or base, e.g. an equivalent amount, and salting out in a medium, or lyophilizing in an aqueous solution.

The compounds of the invention may contain basic moieties, including but not limited to amine or pyridine or imidazole rings, which may form salts with organic or inorganic acids. Typical acids which may form salts include acetates (e.g. with acetic acid or trihaloacetic acid such as trifluoroacetic acid), adipates, alginates, ascorbates, aspartates, benzoates, benzenesulfonates, bisulfates, borates, butyrates, citrates, camphorates, camphorsulfonates, cyclopentanepropionates, diglycolates, dodecylsulfates, ethanesulfonates, fumarates, glucoheptonates, glycerophosphates, hemisulfates, heptanoates, hexanoates, hydrochlorides, hydrobromides, hydroiodides, hydroxyethanesulfonates (e.g. 2-hydroxyethanesulfonates), lactates, maleates, methanesulfonates, naphthalenesulfonates (e.g. 2-naphthalenesulfonates), nicotinates, nitrates, oxalates, pectinates, persulfates, phenylpropionates (e.g. 3-phenylpropionates), phosphates, propionates, citrates, and the like, Picrates, pivalates, propionates, salicylates, succinates, sulfates (e.g., with sulfuric acid), sulfonates, tartrates, thiocyanates, tosylates, e.g., p-toluenesulfonate, dodecanoate, and the like

Acidic moieties that certain compounds of the present invention may contain, including but not limited to carboxylic acids, may form salts with various organic or inorganic bases. Typical salts with bases include ammonium salts, alkali metal salts such as sodium, lithium, potassium salts, alkaline earth metal salts such as calcium, magnesium salts, and salts with organic bases (e.g., organic amines) such as benzathine, dicyclohexylamine, hydrabamine (salt with N, N-bis (dehydroabietyl) ethylenediamine), N-methyl-D-glucamine, N-methyl-D-glucamide, t-butylamine, and salts with amino acids such as arginine, lysine, and the like. The basic nitrogen-containing groups may be combined with halide quaternary ammonium salts, such as small molecule alkyl halides (e.g., methyl, ethyl, propyl, and butyl chlorides, bromides, and iodides), dialkyl sulfates (e.g., dimethyl, diethyl, dibutyl, and diamyl sulfates), long chain halides (e.g., decyl, dodecyl, tetradecyl, and tetradecyl chlorides, bromides, and iodides), aralkyl halides (e.g., benzyl and phenyl bromides), and the like.

Prodrugs and solvates of the compounds of the invention are also contemplated. The term "prodrug" as used herein refers to a compound that undergoes chemical conversion by metabolic or chemical processes to yield a compound, salt, or solvate of the present invention when used in the treatment of a related disease. The compounds of the present invention include solvates, such as hydrates.

The compounds, salts or solvates of the invention may exist in tautomeric forms (e.g. amides and imino ethers). All of these tautomers are part of the present invention.

All stereoisomers of the compounds (e.g., those asymmetric carbon atoms that may exist due to various substitutions), including enantiomeric and diastereomeric forms thereof, are contemplated within the invention. The individual stereoisomers of the compounds of the invention may not be present in combination with the other isomers (e.g. as a pure or substantially pure optical isomer having a particular activity), or may be present as a mixture, e.g. as a racemate, or as a mixture with all or a portion of the other stereoisomers. The chiral center of the invention has two S or R configurations, and is defined by the International Union of theory and applied chemistry (IUPAC) proposed in 1974. The racemic forms can be resolved by physical methods such as fractional crystallization, or by separation of the crystals by derivatization into diastereomers, or by chiral column chromatography. The individual optical isomers can be obtained from the racemates by any suitable method, including, but not limited to, conventional methods such as salt formation with an optically active acid followed by crystallization.

The compounds of the present invention, obtained by preparing, isolating and purifying the compound in sequence, have a weight content of 90% or more, for example, 95% or more, 99% or more ("very pure" compounds), as set forth in the text. Such "very pure" compounds of the invention are also part of the invention herein.

All configurational isomers of the compounds of the invention are within the scope of the invention, whether in mixture, pure or very pure form. The definition of compounds in the present invention encompasses both cis (Z) and trans (E) olefin isomers, as well as cis and trans isomers of carbocyclic and heterocyclic rings.

Throughout the specification, groups and substituents may be selected to provide stable fragments and compounds.

Specific functional groups and definitions of chemical terms are detailed below. For purposes of the present invention, the chemical Elements are compatible with the Periodic Table of the Elements, CAS version, Handbook of Chemistry and Physics,75^thD. as defined in. The definition of a particular functional group is also described herein. In addition, the basic principles of Organic Chemistry, as well as specific functional groups and reactivities are also described in "Organic Chemistry", Thomas Sorrell, University Science Books, Sausaltito: 1999, which is incorporated by reference in its entirety.

Certain compounds of the present invention may exist in specific geometric or stereoisomeric forms. The present invention encompasses all compounds, including cis and trans isomers, R and S enantiomers, diastereomers, (D) isomer, (L) isomer, racemic mixtures and other mixtures thereof. Further the asymmetric carbon atom may represent a substituent such as an alkyl group. All isomers, as well as mixtures thereof, are encompassed by the present invention.

According to the present invention, the mixture of isomers may contain a variety of isomer ratios. For example, in a mixture of only two isomers, the following combinations are possible: 50:50, 60:40, 70:30, 80:20, 90:10, 95:5, 96:4, 97:3, 98:2, 99:1, or 100:0, all ratios of isomers are within the scope of the invention. Similar ratios, as well as ratios that are mixtures of more complex isomers, are also within the scope of the invention, as would be readily understood by one of ordinary skill in the art.

The invention also includes isotopically-labelled compounds, equivalent to the original compounds disclosed hereinAnd opening. In practice, however, it will often occur that one or more atoms are replaced by an atom having a different atomic weight or mass number. Examples of isotopes that can be listed as compounds of the invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine and chlorine, respectively²H、³H、¹³C、¹¹C、¹⁴C、¹⁵N、¹⁸O、¹⁷O、³¹P、³²P、³⁵S、¹⁸F and³⁶and (4) Cl. The compounds of the present invention, or enantiomers, diastereomers, isomers, or pharmaceutically acceptable salts or solvates thereof, wherein isotopes or other isotopic atoms comprising such compounds are within the scope of the present invention. Certain isotopically-labelled compounds of the invention, e.g.³H and¹⁴among these, the radioactive isotope of C is useful in tissue distribution experiments of drugs and substrates. Tritium, i.e.³H and carbon-14, i.e.¹⁴C, their preparation and detection are relatively easy. Is the first choice among isotopes. In addition, heavier isotopes such as deuterium, i.e.²H, due to its good metabolic stability, may be advantageous in certain therapies, such as increased half-life in vivo or reduced dose, and therefore, may be preferred in certain circumstances. Isotopically labeled compounds can be prepared by conventional methods by substituting a readily available isotopically labeled reagent for a non-isotopically labeled reagent using the protocols disclosed in the examples.

If it is desired to design the synthesis of a particular enantiomer of a compound of the invention, it may be prepared by asymmetric synthesis or by derivatization with chiral auxiliary agents, separation of the resulting diastereomeric mixture and removal of the chiral auxiliary agent to give the pure enantiomer. Alternatively, if the molecule contains a basic functional group, such as an amino acid, or an acidic functional group, such as a carboxyl group, diastereomeric salts can be formed therewith with an appropriate optically active acid or base, and the isolated enantiomers can be obtained in pure form by conventional means such as fractional crystallization or chromatography.

Metabolites of the compounds and pharmaceutically acceptable salts thereof to which this application relates, and prodrugs that can be converted in vivo to the structures of the compounds and pharmaceutically acceptable salts thereof to which this application relates, are also included in the claims of this application.

Preparation method

The compounds of the present invention may be prepared according to conventional routes or methods, or may be obtained according to the methods or routes described herein.

The following more specifically describes the preparation of the compounds of the present invention, but these specific methods do not set any limit to the present invention. The compounds of the present invention may also be conveniently prepared by optionally combining various synthetic methods described in the present specification or known in the art, and such combinations may be readily carried out by those skilled in the art to which the present invention pertains.

In general, in the preparative schemes, each reaction is usually carried out in an inert solvent at a temperature ranging from room temperature to reflux temperature (e.g., from 0 ℃ to 150 ℃, preferably from 10 ℃ to 100 ℃). The reaction time is usually 0.1 to 60 hours, preferably 0.5 to 48 hours.

Preferably, the invention provides a synthetic method of formula I, the synthetic route is as follows:

A-I can be obtained by conventional methods either commercially or as reported in the literature;

in the formula, X₁、X₂、X₃、X₄、R₃、R₄、R₅、R₆、R₇、R₈、R₉X and n are as defined above;

(i) in an inert solvent (such as DMF), A-I is mixed with halide in base (such as NaH, LiAlH)₄Etc.) to obtain A-II;

(ii) in an inert solvent (such as methanol, ethanol, etc.), a reducing agent (such as palladium/carbon, SnCl₂Etc.) in the presence of the acid, the A-II is subjected to reduction reaction to obtain A-III;

(iii) in an inert solvent in the presence of a base (such as sodium carbonate, potassium carbonate, cesium carbonate, etc.) and a palladium catalyst (such as tris (bis) phosphonium hydroxide)Benzalacetone) dipalladium, bis (triphenylphosphine) palladium dichloride, etc.), A-III and

carrying out coupling reaction to obtain A-IV;

(iv) hydrolyzing A-IV in an inert solvent (such as tetrahydrofuran and/or water) under basic (such as lithium hydroxide) conditions to obtain A-V;

(v) in an inert solvent (such as dioxane) and under an acidic (such as HCl) condition, removing Boc protecting groups from A-V to obtain amino acids A-VI;

(vi) under the common condensation conditions (such as inert solvent (such as dichloromethane) and alkaline (such as N, N-diisopropylethylamine)), A-VI is subjected to condensation reaction to obtain the target products A-VII (namely the compound of the formula I).

In the above-mentioned synthetic steps, the starting materials and reagents used are either commercially available or synthesized by literature-reported methods.

Pharmaceutical compositions and methods of administration

Since the compound of the present invention has excellent inhibitory activity against JAK kinases, particularly JAK2 kinase, the compound of the present invention and various crystal forms, pharmaceutically acceptable inorganic or organic salts, hydrates or solvates thereof, and a pharmaceutical composition containing the compound of the present invention as a main active ingredient are useful for preventing and/or treating (stabilizing, alleviating or curing) JAK kinase-associated diseases (e.g., myelodysplastic syndrome (MDS), hypereosinophilia, tumors, inflammatory diseases or infections caused by bacteria, viruses or fungi, etc.).

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

"pharmaceutically acceptable carrier" refers to: a orA wide variety of compatible solid or liquid fillers or gel materials which are suitable for human use and must be of sufficient purity and sufficiently low toxicity. By "compatible" is meant herein that the components of the composition are capable of intermixing with and with the compounds of the present invention without significantly diminishing the efficacy of the compounds. Examples of pharmaceutically acceptable carrier moieties are cellulose and its derivatives (e.g., sodium carboxymethylcellulose, sodium ethylcellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (e.g., stearic acid, magnesium stearate), calcium sulfate, vegetable oils (e.g., soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (e.g., propylene glycol, glycerin, mannitol, sorbitol, etc.), emulsifiers (e.g., propylene glycol, glycerin, mannitol, sorbitol, etc.), and the like

) Wetting agents (e.g., sodium lauryl sulfate), coloring agents, flavoring agents, stabilizers, antioxidants, preservatives, pyrogen-free water, and the like.

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

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

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

Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups or tinctures. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly employed in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, propylene glycol, 1, 3-butylene glycol, dimethylformamide and oils, in particular, cottonseed, groundnut, corn germ, olive, castor and sesame oils or mixtures of such materials and the like.

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

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

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

The compounds of the present invention may be administered alone or in combination with other pharmaceutically acceptable compounds (e.g., anti-HBV agents).

When administered in combination, the pharmaceutical composition further comprises one or more (2, 3, 4, or more) other pharmaceutically acceptable compounds. One or more (2, 3, 4, or more) of the other pharmaceutically acceptable compounds may be used simultaneously, separately or sequentially with the compounds of the invention for the prevention and/or treatment of JAK, in particular JAK 2-associated diseases.

When the pharmaceutical composition is used, a safe and effective amount of the compound of the present invention is suitable for mammals (such as human beings) to be treated, wherein the administration dose is a pharmaceutically-considered effective administration dose, and for a human body with a weight of 60kg, the daily administration dose is usually 1 to 2000mg, preferably 20 to 500 mg. Of course, the particular dosage will depend upon such factors as the route of administration, the health of the patient, and the like, and is within the skill of the skilled practitioner.

The main advantages of the invention are:

1. the compound has a novel structure and excellent JAK kinase, especially JAK2 kinase inhibitor effect;

2. the compounds of the invention are more selective for JAK 2.

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

The test materials and reagents used in the following examples are commercially available without specific reference.

General materials and test methods:

methods of synthesis of the compounds of the invention are shown in the schemes, methods and examples below. The starting materials are commercially available or can be prepared according to known methods in the art or described herein. The compounds of the present invention are illustrated by the specific examples shown below. However, these specific embodiments should not be construed as the only variety of the present invention. These examples further illustrate the preparation of the compounds of the present invention. Those skilled in the art will readily appreciate that known variations of conditions and processes may be used to prepare these compounds. All temperatures are in degrees celsius unless otherwise indicated.

Thin Layer Chromatography (PTLC) was performed on 20X 20cm plates (500 μm thick silica gel). Silica gel chromatography was performed using a Biotage flash chromatography system.

1H NMR with a Bruker Ascend TM400 spectrometer, 400MHz, 298 ℃ K, and chemical shifts (ppm) of residual protons in the deuterated reagent are referenced:

CHCl3 δ 7.26ppm, CH3OH or CH3OD δ 3.30ppm, DMSO-d6 δ 2.50ppm

LCMS chromatography uses Agilent technologies 1200 series or 6120 quadrupole spectrometers. For LC mobile phase acetonitrile (a) and water (B) and 0.01% formic acid, eluent gradient: 5-95% A at 6.0 min, 60-95% A at 5.0 min, 80-100% A at 5.0 min and 85-100% A at 10 min, using a capillary column SBC1850 mm by 4.6mm by 2.7 μm.

Mass Spectra (MS) were determined by electrospray ion mass spectrometry (ESI).

HPLC mass spectrometry conditions:

LC 1: column: SB-C1850 mm X4.6 mm X2.7 μm

Temperature: 50 deg.C

Eluent: 5:95 to 95:5 v/v acetonitrile/water + 0.01% formic acid, 6 minutes.

Flow rate: 1.51.5mL/min, 5 uL injection

And (3) detection: PDA, 200 ion 600nm

MS: mass range 150-; positive ion electrospray ionization

LC 2: column: SB-C1850 mm X4.6 mm X2.7 μm

Temperature: 50 deg.C

Eluent: 5:95 to 95:5 v/v acetonitrile/water + 0.05% TFA gradient over 3.00 minutes.

Flow rate: 1.51.5mL/min, 5 uL injection

And (3) detection: PDA, 200 ion 600nm

MS: mass range 150-; positive ion electrospray ionization

LC3 column: SB-C1850 mm X4.6 mm X2.7 μm

Temperature: 50 deg.C

Eluent: 10:90 to 98:2 v/v acetonitrile/water + 0.05% TFA gradient over 3.75 minutes.

Flow rate: 1.0mL/min, 10. mu.L injection

And (3) detection: PDA, 200 ion 600nm

MS: mass range 150-; positive ion electrospray ionization

Abbreviation list:

AcOH ═ acetic acid

Alk is alkyl

AR is aryl

Boc ═ tert-butoxycarbonyl

bs ═ broad peak

CH₂Cl₂Methylene chloride ═

d ═ double peak

ddd-doublet

DBU ═ 1, 8-diazabicyclo [5.4.0] undec-7-ene

DCM ═ dichloromethane

DEAD ═ azodicarboxylic acid diethyl ester

DMF ═ N, N-dimethylformamide

DMSO ═ dimethyl sulfoxide

EA is ethyl acetate

ESI-electrospray ionization

Et is ethyl

EtOAc ═ ethyl acetate

EtOH ═ ethanol

h is hour

HOAc ═ acetic acid

LiOH lithium hydroxide

m is multiple

Me is methyl

MeCN ═ acetonitrile

MeOH ═ methanol

MgSO4 magnesium sulfate

min is minutes

MS mass spectrum

NaCl sodium chloride

NaOH (sodium hydroxide)

Na2SO4 ═ sodium sulfate

NMR spectrum

PE-Petroleum Ether

PG ═ protecting group

Ph ═ phenyl

rt-room temperature

s ═ singlet

t is triplet

TFA ═ trifluoroacetic acid

THF ═ tetrahydrofuran

TS ═ p-toluenesulfonyl (toluenesulfonyl)

EXAMPLE 1 Synthesis of Compound A1

The first step is as follows: 5- (2-morpholine-5-nitrobenzyloxy) pentanoic acid methyl ester (A1-2)

Under the protection of nitrogen, 2-morpholine-5-nitrobenzol (A1-1, 500mg, 2.1mmol) is added into anhydrous N, N-dimethylformamide (5ml), the mixture is cooled to 0 ℃ in an ice bath, sodium hydride (60%, 168mg, 4.2mmol) is added, the ice bath is continuously stirred for 30min, then a solution of methyl 5-bromovalerate (614mg, 3.15mmol) in N, N-dimethylformamide (6ml) is slowly added, the mixture is naturally warmed to room temperature, and the stirring is continuously carried out for 16 h. After confirming that the reaction was substantially complete by TLC, the reaction was quenched with saturated ammonium chloride solution under ice bath, extracted with EA, washed three times with deionized water, dried over anhydrous sodium sulfate, concentrated in EA solution, and the residue was purified by column chromatography to give methyl 5- (2-morpholine-5-nitrobenzyloxy) valerate (A1-2, 505mg, 68% yield).¹H NMR(400MHz,CDCl3)δ8.36(d,J＝2.6Hz,1H),8.15(dd,J＝8.9,2.7Hz,1H),7.08(d,J＝8.9Hz,1H),4.54(s,2H),3.93–3.85(m,4H),3.69(s,3H),3.57(t,J＝6.0Hz,2H),3.13–3.02(m,4H),2.37(t,J＝7.1Hz,2H),1.31(s,2H),1.28(s,2H)。

The second step is that: 5- (2-morpholine-5-aminobenzyloxy) pentanoic acid methyl ester (A1-3)

Methyl 5- (2-morpholine-5-nitrobenzyloxy) valerate (A1-2, 505mg, 1.43mmol) was added to methanol (5ml) and palladium on carbon (25mg, 10% on carbon (wet with ca. 55% Water) was added, stirring was carried out at room temperature for 6H, after completion of the reaction was confirmed by TLC, suction filtration was carried out, the filtrate was concentrated, and the residue was purified by column chromatography to give methyl 5- (2-morpholine-5-aminobenzyloxy) valerate (A1-3,285mg, yield 61.7%), MS (ESI) M/z as a yellow oil, theoretical value (calcd)323.20(M + H), found value (found) 323.42.

The third step: (4- (2-Chloropyrimidine-4-) phenyl) carbamic acid tert-butyl ester (A1-4)

Under nitrogen, 2, 4-dichloropyrimidine (1.50g,10.1mmol) and 4- (N-Boc-amino) phenylboronic acid (2.37g,10mmol) were added to a mixed solvent of N, N-dimethylformamide: 1, 4-dioxane: 12ml of water: 7.5 ml: to 3ml was added sodium carbonate (2.12g,20mmol) followed by palladium bis (triphenylphosphine) dichloride (73mg,0.1mmol), and the mixture was heated to 80 ℃ and stirred for 16 h. TLC confirmed the reaction was complete, 1, 4-dioxane was concentrated, EA extracted, the organic phase was washed three times with deionized water, dried over anhydrous sodium sulfate, EA concentrated, and the residue was purified by column chromatography to give a white solid a1-4(2.50g, 82% yield). MS (ESI) M/z theoretical 306.10(M + H), found 306.32;¹H NMR(400MHz,CDCl3):δ8.73(d,J＝5.2Hz,1H),8.24–8.16(m,4H),7.73(d,J＝5.2Hz,1H),3.99(s,3H)。

the fourth step: methyl 5- (5- ((4- (4- (tert-butoxycarbonylamino) phenyl) pyrimidin-2-) amino) -2-morpholinobenzyloxy) pentanoate (A1-5)

Under nitrogen protection, methyl 5- (2-morpholine-5-aminobenzyloxy) valerate (A1-3, 142.5mg, 0.44mmol), (4- (2-chloropyrimidin-4-) phenyl) carbamic acid tert-butyl ester (A1-4, 162mg, 0.53mmol) and potassium carbonate (122mg, 0.88mmol) were added to 1, 4-dioxane (10ml), stirred for 5min, tris (dibenzylideneacetone) dipalladium (40.5mg, 0.04mmol) and 2-dicyclohexylphosphonium-2 ',4',6' -triisopropylbiphenyl (42mg, 0.09mmol) were added in one portion, the temperature was raised to 100 ℃, and stirred for 16 h. After confirming completion of the reaction by TLC, 1, 4-dioxane was concentrated, EA and deionized water were added, liquid was extracted, the organic layer was dried over anhydrous sodium sulfate, EA was concentrated, and the residue was purified by column chromatography to give a1-5(175mg, yield 67%).

The fifth step: 5- (5- ((4- (4- (tert-butoxycarbonylamino) phenyl) pyrimidin-2-) amino) -2-morpholinobenzyloxy) pentanoic acid (A1-6)

Methyl 5- (5- ((4- (4- (tert-butoxycarbonylamino) phenyl) pyrimidin-2-) amino) -2-morpholinobenzyloxy) pentanoate (A1-5, 175mg, 0.30mmol) was added to tetrahydrofuran (3ml) and water (1ml), followed by addition of lithium hydroxide monohydrate (62mg, 1.50mmol), and stirring at room temperature for 16 h. After confirming the reaction by TLC, 1mol/L HCl was added dropwise to adjust pH to 5 to 6, tetrahydrofuran was concentrated, EA and deionized water were added, the mixture was extracted and separated into layers, dried over anhydrous sodium sulfate, EA was concentrated, and the residue was purified by column chromatography to obtain 5- (5- ((4- (4- (tert-butoxycarbonylamino) phenyl) pyrimidin-2-) amino) -2-morpholinobenzyloxy) pentanoic acid (a1-6, 110mg, yield 64.4%).

And a sixth step: 5- ((5- ((4- (4-aminophenyl) pyrimidine-2-) amino) -2-morpholinobenzyloxy) pentanoic acid (A1-7)

5- (5- ((4- (4- (tert-butoxycarbonylamino) phenyl) pyrimidin-2-) amino) -2-morpholinobenzyloxy) pentanoic acid (A1-6, 110mg, 0.19mmol) was added to 1, 4-Dioxane (2ml), and a 4mol/L solution of HCl in Dioxane (2ml) was added dropwise and stirred at room temperature for 16 h. After completion of the reaction was confirmed by LCMS, 1, 4-dioxane was concentrated, saturated aqueous sodium bicarbonate was added to adjust pH to neutral, EA was extracted, dried, EA was concentrated, and the residue was purified by reverse phase column and lyophilized to give 5- ((5- ((4- (4-aminophenyl) pyrimidin-2-) amino) -2-morpholinobenzyloxy) pentanoic acid (a1-7, 30mg, yield 33%).

The seventh step: 4⁴Morpholine-6-oxa-3, 12-diaza-2 (4,2) -pyrimidinehetero-1 (1,4),4(1,3) -diphenylheterocyclodododecan-11-one (A1)

2- (7-Benzotolyltriazole) -N, N, N ', N' -tetramethyluronium hexafluorophosphate (47.8mg, 0.13mmol) and N, N-diisopropylethylamine (16.3mg, 0.13mmol) were added to dichloromethane (2ml) and stirred for 5 min. Slowly adding a mixed solution of DCM (8ml) and DMF (1ml) of 5- ((5- ((4- (4-aminophenyl) pyrimidine-2-) amino) -2-morpholinobenzyloxy) pentanoic acid (A1-7, 30mg, 0.06mmol) dropwise at room temperature, stirring at room temperature for 5h after dropwise adding to confirm that the reaction is complete, adding deionized water to quench the reaction, extracting and separating the liquid, drying an organic layer by anhydrous sodium sulfate, concentrating DCM, purifying the residue by a reverse phase column, and freeze-drying to obtain a white solid 4⁴Morpholine-6-oxa-3, 12-diaza-2 (4,2) -pyrimidinehetero-1 (1,4),4(1,3) -diphenylheterocyclodecan-11-one (A1, 8.3mg, yield 28.7%). MS (ESI) M/z theoretical 460.23(M + H), found 460.41;¹H NMR(400MHz,DMSO)δ9.55(d,J＝6.7Hz,2H),8.52(d,J＝5.0Hz,1H),8.48(s,1H),7.88(d,J＝8.3Hz,2H),7.36(d,J＝8.2Hz,2H),7.22(d,J＝5.1Hz,1H),7.10–7.06(m,1H),7.03(s,1H),4.47(s,2H),3.75–3.68(m,4H),3.30–3.29(m,2H),2.85–2.80(m,4H),2.28–2.22(m,2H),1.33–1.28(m,4H)。

example 2 Synthesis of Compound A2

First step diethyl 2-morpholine-5-nitrobenzylphosphonate (A2-1)

4- (2-bromomethyl-4-nitrophenyl) morpholine (800mg,2.66mmol) and triethyl phosphite (0.55mL,3.20mmol) were added to a sealed tube and stirred at 140 ℃ for 3h, after completion of the reaction by LCMS, the reaction mixture was directly purified on silica gel column to give diethyl 2-morpholine-5-nitrobenzylphosphate (A2-1, 950mg, 99% yield). MS (ESI) M/z theoretical 359.13(M + H), found 359.49.

Second step ethyl 6-oxohexanoate (A2-2)

Ethyl 6-hydroxycaproate (640mg,4.00mmol) was dissolved in EA (30mL), 2-iodoxybenzoic acid (1.68g,6.00mmol) was added, stirring was carried out at 80 ℃ for 3h, TLC showed that all the starting material was converted to product A2-2, the solid was filtered off, and the filtrate was concentrated to give ethyl 6-oxohexanoate as a colorless oil (A2-2,630mg, yield 99%).

Third step 7- (2-morpholine-5-nitro) phenyl-6-heptenoic acid ethyl ester (A2-3)

Under the protection of nitrogen, A2-1(238mg,0.66mmol) was dissolved in anhydrous THF (30mL), cooled to 0 ℃ in an ice bath, NaH (40mg,0.99mmol, 60%) was slowly added, stirring was continued for 30min in an ice bath, then A2-2(125mg,0.80mmol) in THF (3mL) was slowly added, the ice bath was removed, and stirring was continued for 3h after naturally rising to room temperature. TLC confirmed the reaction was complete, quenched with saturated ammonium chloride solution in an ice bath, concentrated under reduced pressure to remove THF, EA extracted, washed three times with deionized water, dried over anhydrous sodium sulfate, concentrated EA solution, and the residue purified by column chromatography to afford A2-3(128mg, 53.5% yield). MS (ESI) M/z theoretical 363.18(M + H), found 363.07.

The fourth step, ethyl 7- (2-morpholin-5-amino) phenyl-heptanoate (A2-4)

A2-3(128mg,0.35mmol) was added to absolute ethanol (30ml), raney nickel (20mg) was added, catalytic hydrogenation was carried out under a hydrogen pressure of 1atm, and stirring was carried out at room temperature for 5 hours. After completion of the reaction was confirmed by LCMS, the catalyst was filtered off, and the filtrate was concentrated to give A2-4(118mg, yield 99%) as a colorless oil. MS (ESI) M/z theoretical 335.23(M + H), found 335.30;

the fifth step Ethyl 7- (5- ((4- (4- (tert-butoxycarbonylamino) phenyl) pyrimidin-2-) amino) -2-morpholinophenyl) heptanoate (A2-5)

Under nitrogen, ethyl 7- (2-morpholin-5-amino) phenyl-heptanoate (A2-4, 118mg, 0.35mmol), (4- (2-chloropyrimidin-4-yl) phenyl) carbamate (A1-4, 128.4mg, 0.42mmol) and potassium carbonate (97mg, 0.70mmol) were added to 1, 4-dioxane (20ml), stirred for 5min, tris (dibenzylideneacetone) dipalladium (64mg, 0.07mmol) and 2-dicyclohexylphosphonium-2 ',4',6' -triisopropylbiphenyl (67mg, 0.14mmol) were added in one portion, the temperature was raised to 100 ℃ and stirred for 16 h. After confirming completion of the reaction by TLC, 1, 4-dioxane was concentrated, EA and deionized water were added, liquid was extracted, the organic layer was dried over anhydrous sodium sulfate, EA was concentrated, and the residue was purified by column chromatography to give ethyl 7- (5- ((4- (4- (tert-butoxycarbonylamino) phenyl) pyrimidin-2-) amino) -2-morpholinophenyl) heptanoate (a2-5, 132mg, yield 63%). MS (ESI) M/z theoretical 604.34(M + H), found 604.48.

The sixth step 7- (5- ((4- (4- (tert-butoxycarbonylamino) phenyl) pyrimidin-2-) amino) -2-morpholinophenyl) heptanoic acid (A2-6)

Ethyl (4- (tert-butoxycarbonylamino) phenyl) pyrimidin-2-) amino) -2-morpholinophenyl) heptanoate (A2-5, 132mg, 0.22mmol) THF (3ml) and water (1ml) were added followed by lithium hydroxide monohydrate (92mg, 2.20mmol) and stirring at 70 ℃ for 5 h. After confirming the completion of the reaction by TLC, 1mol/L HCl was added dropwise to adjust the pH to 5 to 6, and the reaction mixture was purified by reverse phase column and lyophilized to obtain 7- (5- ((4- (4- (tert-butoxycarbonylamino) phenyl) pyrimidin-2-) amino) -2-morpholinophenyl) heptanoic acid (a2-6, 120mg, yield 94.5%). MS (ESI) M/z theoretical 576.31(M + H), found 576.51.

Seventh step 7- ((5- ((4- (4-aminophenyl) pyrimidin-2-) amino) -2-morpholinophenyl) heptanoic acid (A2-7)

7- (5- ((4- (4- (tert-Butoxycarbonylamino) phenyl) pyrimidin-2-) amino) -2-morpholinophenyl) heptanoic acid (A2-6, 120mg, 0.21mmol) was added to EA (3ml), 3mol/L HCl in EA (3ml) was added dropwise, and the mixture was stirred at room temperature for 5 hours. After completion of the reaction was confirmed by LCMS, the reaction was concentrated and the crude product was purified by reverse phase column chromatography and lyophilized to give 7- ((5- ((4- (4-aminophenyl) pyrimidin-2-) amino) -2-morpholinophenyl) heptanoic acid (A2-7, 83mg, yield 84%). MS (ESI) M/z: theoretical 476.26(M + H), found 476.50.

Eighth step 4⁴Morpholine-3, 12-diaza-2 (4,2) -pyrimidinehetero-1 (1,4),4(1,3) -diphenylheterocyclodecanone-11-one (A2)

2- (7-Benzotolyltriazole) -N, N, N ', N' -tetramethyluronium hexafluorophosphate (132.3mg, 0.35mmol) and N, N-diisopropylethylamine (225mg, 1.74mmol) were added to dichloromethane (20ml) and stirred for 5 min. Slowly adding a mixed solution of DCM (8ml) and DMF (1ml) of 7- ((5- ((4- (4-aminophenyl) pyrimidine-2-) amino) -2-morpholinophenyl) heptanoic acid (A2-7, 83mg, 0.17mmol) dropwise at room temperature, stirring at room temperature for 24H after dropwise adding to confirm that the reaction is complete, adding deionized water to quench the reaction, extracting and separating liquid, drying an organic layer anhydrous sodium sulfate, concentrating DCM, purifying a residue by a reverse phase column, and freeze-drying to obtain a light yellow solid A2(20mg, yield 25.2%). MS (ESI) M/z: theoretical value 458.26(M + H), measured value 458.58;¹H NMR(400MHz,DMSO-d6)δ9.63(s,1H),9.52(s,1H),8.51(d,J＝5.0Hz,1H),8.37(s,1H),7.93(d,J＝8.0Hz,2H),7.37(d,J＝8.0Hz,2H),7.22(d,J＝5.0Hz,1H),7.04-6.94(m,2H),3.73-3.70(m,4H),2.81-2.73(m,4H),2.80-2.51(m,2H),2.38-2.35(m,2H),1.37-1.33(m,4H),1.27-1.25(m,4H).

with reference to the experimental procedure of examples 1 and 2, with different starting materials, examples 3-5 were obtained, as shown in table 1 below,

TABLE 1

Example 6: biological test method

The JAK kinase activity measuring method is homogeneous phase time resolution fluorescence technology. The reaction of this method was carried out in 384-well shallow plates, and the total volume of the reaction was 10. mu.L. The mixture of kinase protein, compound, ATP and substrate was incubated at 50mM Hepes (pH7.0), NaN₃0.02％，BSA 0.01％，0.1mM Orthocanadate，5mM MgCl₂1mM DTT, and after 1 hour of reaction, an antibody capable of recognizing phosphorylation of the substrate and a dye XL-615 and a detection buffer containing EDTA (Cisbio) were added to the system. The reaction signal of the kinase was detected by a multi-well plate detector of PE. The parameter settings were excitation light 320nm, emission light 615nm and 665 nm. JAK viability is reflected indirectly by the ratio of signals at 665nm and 615 nm. A background well without enzyme and a full-enzyme active well without compound are set in the reaction.

Compound inhibiting protein IC₅₀Is given by the formula: y100/(1 +10^ ((LogIC50-X) × HillSlope)). In the JAK1 reaction system, the ATP concentration was 2. mu.M, and the JAK1 protein concentration was 0.2 ng/. mu.L.

In the JAK2 reaction system, the ATP concentration was 2. mu.M, and the JAK2 protein concentration was 0.01 ng/. mu.L.

In the JAK3 reaction system, the ATP concentration was 2. mu.M, and the JAK3 protein concentration was 0.04 ng/. mu.L.

In the TYK2 reaction system, the concentration of ATP was 2. mu.M, and the concentration of TYK2 protein was 0.2 ng/. mu.L.

The test data is divided into the following: a is IC₅₀<10nM；B:IC₅₀11-100nM；C:IC₅₀101-1000nM；D:IC₅₀1001-10000nM；E:IC₅₀>10000nM。

The test results are shown in table 2.

TABLE 2 JAK inhibitory Activity Table

The ratio of JAK1/JAK2, JAK3/JAK2 and TYK2/JAK2 are calculated by C1/C2, wherein C1 is the IC50 value of a compound of the invention against JAK1, JAK3 and TYK 2; c2 is the IC50 value of the same compound against JAK 2;

+ represents 1 to 5 in the ratios JAK1/JAK2, JAK1/JAK3, TYK2/JAK 2; + represents 6-20; , + ++ represents 21-50; , + +++ denotes > 50.

Discussion:

the above experimental results suggest:

(1) the compounds of formula I of the present invention exhibit very excellent JAK inhibitory activity, especially against JAK 2. The IC50 values of the compounds of the invention against JAK2 may be as low as in the order of 10nM or less, so that for subjects weighing about 70kg (e.g., patients, especially rheumatoid arthritis or psoriasis patients), a daily dose of usually 10mg to 30mg is extremely effective in inhibiting JAK 2.

(2) The compound of formula I of the invention shows very excellent JAK selectivity, namely the ratio of IC50 of JAK1/JAK2 and/or the ratio of IC50 of JAK3/JAK2 and/or the ratio of IC50 of TYK2/JAK2 is superior to or equal to the existing compound.

All documents referred to herein are incorporated by reference into this application as if each were individually incorporated by reference. Furthermore, it should be understood that various changes and modifications of the present invention can be made by those skilled in the art after reading the above teachings of the present invention, and these equivalents also fall within the scope of the present invention as defined by the appended claims.

Claims

1. A compound shown in formula I or a stereoisomer or an optical isomer and a pharmaceutically acceptable salt thereof,

in the formula (I), the compound is shown in the specification,

is partially made of

Wherein R is_A、R_B、R_CAnd R_DEach independently selected from the group consisting of: H. deuterium, halogen, amino, nitro, hydroxy, mercapto, cyano, carboxyl, C1-C6 alkyl, C1-C6 alkoxy;

x is selected from the group consisting of: key, CR_bR_c、O、N-R_b、S；

R₃、R₄、R₅、R₆、R₇、R₈、R₉、R_bAnd R_cEach independently selected from the group consisting of: H. d, halogen, amino, nitro, hydroxyl, cyano, carboxyl, C1-C6 alkyl, C1-C6 alkoxy, C2-C6 alkenyl, C2-C6 alkynyl, 3-10 membered heterocyclyl, C3-C10 cycloalkyl, 5-12 membered heteroaryl, C6-C12 aryl;

n is 1,2,3, 4, 5, 6, 7 or 8;

the above alkyl, alkoxy, alkenyl, alkynyl, heterocyclyl, cycloalkyl, heteroaryl, aryl may be further optionally substituted with 1 or more R_aIn which each R is_aIndependently selected from the group consisting of: halogen, amino, hydroxyl, C1-C6 alkyl, C1-C6 alkoxy.

2. The compound of claim 1, or a stereoisomer or optical isomer, pharmaceutically acceptable salt thereof

R_A、R_B、R_CAnd R_DEach independently selected from the group consisting of: H. deuterium, halogen, C1-C6 alkyl, C1-C6 alkoxy.

3. The compound of claim 1, or a stereoisomer or optical isomer, pharmaceutically acceptable salt thereof, wherein X is selected from the group consisting of: CR_bR_cO; wherein R is_bAnd R_cIs as defined in claim 1.

4. The compound of claim 1, or a stereoisomer or optical isomer, pharmaceutically acceptable salt thereof, having the structure of formula II:

in the formula (I), the compound is shown in the specification,

R₁and R₂Each independently selected from the group consisting of: H. d, halogen, C1-C6 alkyl;

R_a、R₃、R₄、R₅、R₆、R₇、R₈、R₉x and n are as defined in claim 1.

5. The compound of claim 1, or a stereoisomer or optical isomer, pharmaceutically acceptable salt thereof, having the structure of formula a:

wherein the content of the first and second substances,

R₁and R₂As defined in claim 4;

R_a、R₃、R₄、R₅、R₆x and n are as defined in claim 1.

6. The compound of claim 1, or a stereoisomer or optical isomer, pharmaceutically acceptable salt thereof, selected from the group consisting of:

7. a pharmaceutical composition comprising a compound of any one of claims 1-6, or a stereoisomer or an optical isomer thereof, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier or excipient.

8. Use of a compound of any one of claims 1-6, or a stereoisomer or optical isomer thereof, or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the prevention or treatment of a JAK 2-mediated disease; and/or for the preparation of a JAK2 inhibitor.

9. The use according to claim 8, wherein the JAK 2-mediated disease is myelodysplastic syndrome, eosinophilia, tumor, inflammatory disease, or infection by bacteria, viruses, or fungi.

10. A JAK2 inhibitor comprising the compound of any one of claims 1 to 6, or a stereoisomer or an optical isomer thereof, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of claim 7.