CN110734427A

CN110734427A - Alkenyl-containing pyrimidine formamide compound, composition and application thereof

Info

Publication number: CN110734427A
Application number: CN201811100041.2A
Authority: CN
Inventors: 张强; 刘彦生; 李兰涛; 李兴福; 胡晨明
Original assignee: Beijing Saite Mingqiang Medicine Technology Co Ltd
Current assignee: Beijing Saite Mingqiang Medicine Technology Co Ltd
Priority date: 2018-07-20
Filing date: 2018-09-20
Publication date: 2020-01-31
Anticipated expiration: 2038-09-20
Also published as: CN110734427B

Abstract

The invention relates to novel compounds serving as JAK inhibitors, a composition and application thereof, and particularly provides compounds with strong JAK inhibitory activity (shown as a formula (1)) or isomers, solvates, hydrates, pharmaceutically acceptable salts and prodrugs thereof, and a pharmaceutical composition containing the compounds.

Description

Alkenyl-containing pyrimidine formamide compound, composition and application thereof

Technical Field

The invention belongs to the field of medicinal chemistry, and particularly relates to pyrimidine carboxamides containing alkenyl, isomers, hydrates, solvates, pharmaceutically acceptable salts or prodrugs thereof, pharmaceutical compositions thereof, and applications thereof in preparation of medicines for treating autoimmune diseases and cancers related to tyrosine kinases JAK1, JAK2, JAK3 and TYK 2.

Background

JAK kinase (Janus kinase) and its downstream effector, signal transduction and transcriptional activator protein form an important cytokine signal transduction pathway, namely JAK-STAT pathway, researches show that the JAK-STAT pathway can be activated by various cytokines, growth factors and receptors and is involved in processes such as cell proliferation, differentiation, apoptosis, angiogenesis and immune regulation, the JAK kinase is a key kinase in the JAK-STAT signal pathway, and after more than two decades, JAK kinase inhibitors (tofacitinib) are approved for treating rheumatoid arthritis in 2012 [ Norman P, Selective JAKinhibitors in diabetes for rheumatoarthritis, Expert Opin investing drugs,2014,23: 1067-.

In a mammal, four members of JAK kinase family, JAK1, JAK2, JAK3 and TYK2, consist of more than 1100 amino acids, have a relative molecular mass of 120000-140000 and a homology of 40-70%, are divided into 7 homologous domains (JH) in sequence from C-terminal to N-terminal, JH1 is a kinase region highly conserved in the JAK family, JH2 is a kinase-like region or a "pseudo" kinase region, the pseudo kinase domain is a unique property of the JAK protein to distinguish other tyrosine proteins, the kinase region has no catalytic activity but has a regulatory effect on the activity of JH1, mutations in the domain often lead to enhancement or attenuation of the JAK kinase activity and further lead to certain diseases, JH3-JH4 is an SH2 domain (Src homology2 domain) containing about 100 amino acid residues, which can specifically recognize a ligand binding to the ligand of the JAK 3-JH 7, and the JAK 4642 is a conserved amino acid domain containing a tyrosine kinase domain which has a structural change with the same structure as a receptor kinase-7.

The JAK-STAT signaling pathway is intracellular signal transduction pathways important in the growth, activation, differentiation, apoptosis of various cells and their functions.STAT is a cytoplasmic protein of the class that binds to the regulatory region DNA of target genes, which is a downstream substrate of JAK.7 members of the STAT family include STAT1, STAT2, STAT3, STAT4, STAT5A, STAT5B, and STAT 6. the interaction between JAKs and STATs plays an important role in Cytokine receptor signaling pathways [ O' Sullivan LAet, Cytokine receptor stimulating hormone kinase, JAK-STAT-protein pathway, Mol stimulating hormone, 2007,44:2497, 2506] when Cytokine receptors on the cell surface bind to their respective Cytokine ligands, they cause receptor molecules to dimerize, so that kinases coupled to the receptors are close to each other and activate via the interactive tyrosine kinase, stimulating hormone receptor (IFN), macrophage stimulating hormone receptor (EGF), macrophage stimulating hormone receptor stimulating hormone, macrophage stimulating hormone, hormone stimulating hormone.

JAK1 and JAK2 are expressed in all tissue cells of a human body, JAK3 is mainly expressed in all hematopoietic tissue cells and mainly exists in bone marrow cells, thymocytes, NK cells, activated B lymphocytes and activated T lymphocytes, JAK1, JAK2 and the like can cause lethal injury of the human body, JAK3 can avoid toxic adverse reactions damaging other tissue cells [ Yamaoka K, et al, JAK3 newtivey regulated dendritic-cell cytokine expression and survival, Blood,2005,106:3227-3233], based on the functional characteristics and special tissue distribution of all subtypes in JAK kinase family, JAK3 has become a thermal target for treating autoimmune diseases, and more clinical researches focus on the treatment of rheumatoid arthritis on blocking JAK3 signal transduction, so that Tofacitinib, a selective JAK3 inhibitor, is approved clinically and used for treating rheumatoid arthritis for 2012.

Tofacitinib (CP690550) is pyrrolopyrimidine selective JAK3 kinase inhibitors developed by Pfizer, and has inhibitory activity (IC) on JAK3₅₀1nmol/L) is JAK2 (IC)₅₀20 times of 20nmol/L) and JAK1 (IC)₅₀112nmol/L) and the chiral structure of Tofacitinib determines that Tofacitinib can be specifically bound to JAK3 molecules, so that JAK3 phosphorylation is inhibited, and step leads to STAT phosphorylation inhibition, which results in inhibition of downstream inflammatory cytokine synthesisThe group showed significant statistical differences compared to the placebo group of equal volume, but Tofacitinib was found to be associated with increased risk of severe infection in clinical trial studies, and its long-term safety was left to studies.

The JAK-STAT signal pathway plays an important role in the cell differentiation and proliferation processes, and the change of the JAK activity also causes the change of the signal transmission of the pathway, thereby influencing the cell function. Based on the key role of JAK kinase in JAK-STAT signal transmission and the specific histiocyte distribution of JAK3 kinase, JAK3 becomes a good therapeutic target for diseases such as rheumatoid arthritis.

At present, JAK3 inhibitors are mainly used for treating patients with moderate-severe rheumatoid arthritis, the drugs show good treatment effect and better safety in treatment, but the long-term safety of the drugs is still required to be improved by .

The JAK inhibitors of series which are disclosed at present have improved space in the aspects of curative effect, selectivity and safety, and still need to be developed with better drug effect and better safety.

Disclosure of Invention

In view of the above discussion, the present invention aims to provide pyrimidine carboxamides containing alkenyl group, isomers, hydrates, solvates, pharmaceutically acceptable salts or prodrugs thereof, and pharmaceutical compositions thereof, and their use in the preparation of medicaments for treating autoimmune diseases and cancers associated with tyrosine kinases JAK1, JAK2, JAK3 and TYK 2.

According to aspects of the invention there are provided alkenyl-containing pyrimidinecarboxamides having the structural formula (I):

wherein X is N;

R¹is composed of

n₁Is an integer of 0 to 8, n₂Is an integer of 0 to 1, n₃Is an integer of 0 to 8, and n₁、n₂、n₃The sum is 10 or less;

L₁is composed of

R⁴、R⁵Each independently is H, C₁-C₃Alkyl radical, R⁴、R⁵The same or different, or a combination thereof,

R³is unsubstituted or C₁-C₃Alkyl substituted C₂-C₈Alkenyl or unsubstituted or C₁-C₃Alkyl substituted C₄-C₈Cycloalkenyl group of (a);

R²is composed of

n₄Is an integer of 0 to 8, n₅Is an integer of 0 to 1, n₆Is an integer of 0 to 8, and n₄、n₅、n₆The sum is 10 or less;

L₂is composed of

R⁷、R⁸Each independently is H, C₁-C₃Alkyl radical, R⁷、R⁸The same or different, or a combination thereof,

R⁶is-H, hydroxyalkyl, C₁-C₅Alkyl radical, C₃-C₈Cycloalkyl radical, C₁-C₅Alkoxyalkyl group, C₁-C₅Alkylthioalkyl, 5-6 membered heterocyclyl, or-NR 'R ", wherein R', R" are each independently H, C₁-C₃Alkyl group of (1).

Preferably, in the compound of formula (I), R³Is composed of

R⁹、R¹⁰、R¹¹、R¹²、R¹³Each independently is H, C₁-C₃An alkyl group, m is an integer of 0 to 2,

m₁is an integer of 0 to 5, m₂Is an integer of 0 to 5, and m₁,m₂The sum is less than or equal to 5.

Preferably, in the compound of formula (I), n₁Is an integer of 0 to 2, n₂Is an integer of 0 to 1, n₃Is an integer of 0 to 3, R⁴、R⁵Each independently is H, methyl, R⁴、R⁵The same or different;

R⁹、R¹⁰、R¹¹、R¹²、R¹³each independently is H, methyl or ethyl, and m is 0 or 1. More preferably, n₁、n₂、n₃The sum of the amounts is 5 or less.

More preferablyIn the compound of the formula (I), the R is³Is composed of

R⁹、R¹¹、R¹²、R¹³Each independently is H, methyl or ethyl.

Preferably, R³Is composed of

R⁹、R¹⁰Each independently is H, C₁-C₃Alkyl radical, R¹¹、R¹²、R¹³Zhongren is C₄-C₆Alkyl, the remainder are each independently H, C₁-C₃Alkyl, m is an integer of 0 to 2.

Preferably, in the compound of formula (I), n₄Is an integer of 0 to 3, n₅Is an integer of 0 to 1, n₆Is an integer of 0 to 5, R⁷、R⁸Each independently is H, methyl, R⁷、R⁸The same or different;

R⁶is-H, hydroxyethyl, hydroxypropyl, C₁-C₅Alkyl radical, C₃-C₈Cycloalkyl radical, C₁-C₅Alkoxyethyl radical, C₁-C₅Alkoxypropyl group, C₁-C₅An alkylthioethyl group, a 5-6 membered heterocyclyl group, or-NR 'R ", wherein R', R" are each independently H, C₁-C₃Alkyl group of (1). More preferably, n₄、n₅、n₆The sum of the amounts is 5 or less.

More preferably, in the compound of formula (I), R⁶Is H, hydroxyethyl, hydroxypropyl, methyl, ethyl, propyl, isopropyl, tert-butyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, methoxyethyl, ethoxyethyl, propoxyethyl, isopropoxyethyl, methoxypropyl, ethoxypropyl, propoxypropyl, isopropoxypropyl, methylthioethyl, ethylthioethyl, propylthioethyl, isopropylthioEthyl, 5-6 membered heterocyclyl, or-NR 'R ", wherein R', R" are each independently H, methyl, ethyl;

the 5-6 membered heterocyclic group is unsubstituted or substituted by 1-2 groups selected from hydroxy, C₁-C₃Alkyl radical, C₁-C₃Acyl substituents substituted heterocyclyl containing 1-2 heteroatoms selected from N, O, S.

More specifically, the heterocyclic group containing 1 to 2 heteroatoms selected from N, O, S is any selected from:

wherein R is¹⁴H, methyl, ethyl, propyl and isopropyl.

According to embodiments of the present application, the pharmaceutically acceptable salt of the alkenyl-containing pyrimidine carboxamide compound is or more selected from the group consisting of hydrochloride, hydrobromide, hydroiodide, perchlorate, sulfate, nitrate, phosphate, formate, acetate, propionate, glycolate, lactate, succinate, maleate, tartrate, malate, citrate, fumarate, gluconate, benzoate, mandelate, methanesulfonate, isethionate, benzenesulfonate, oxalate, palmitate, 2-naphthalenesulfonate, p-toluenesulfonate, cyclamate, salicylate, gluconate, trifluoroacetate, aluminum salt, calcium salt, chloroprocaine salt, choline salt, diethanolamine salt, ethylenediamine salt, lithium salt, magnesium salt, potassium salt, sodium salt and zinc salt of the compound.

Another aspect of the present invention relates to the use of the alkenyl-containing pyrimidinecarboxamides, their isomers, hydrates, solvates, pharmaceutically acceptable salts or prodrugs thereof in the manufacture of a medicament for the treatment of autoimmune diseases and cancers associated with tyrosine kinases JAK1, JAK2, JAK3, TYK2, wherein said autoimmune diseases and cancers associated with tyrosine kinases JAK1, JAK2, JAK3, TYK2 include ocular fundus disease, dry eye, psoriasis, vitiligo, dermatitis, alopecia areata, rheumatoid arthritis, colitis, multiple sclerosis, systemic lupus erythematosus, crohn's disease, atheroma, pulmonary fibrosis, hepatic fibrosis, myelofibrosis, non-small cell lung cancer, breast cancer, pancreatic cancer, glioma, glioblastoma, ovarian cancer, cervical cancer, colorectal cancer, melanoma, endometrial cancer, prostate cancer, bladder cancer, leukemia, gastric cancer, liver cancer, gastrointestinal stromal tumors, thyroid cancer, acute myelogenous leukemia, non-hodgkin's lymphoma, chronic lymphoma, brain cell lymphoma, multiple myeloma, cholangiocarcinoma, multiple myeloma, and cholangiocarcinoma.

In a further aspect, the invention provides pharmaceutical compositions comprising the alkenyl-containing pyrimidinecarboxamides of the present application, their isomers, hydrates, solvates, pharmaceutically acceptable salts, or prodrugs thereof, and or more pharmaceutically acceptable carriers or excipients.

According to embodiments of the present application, the pharmaceutical composition may also include or more additional therapeutic agents.

Advantageous effects

The alkenyl-containing pyrimidine carboxamide compound shows strong inhibitory activity of tyrosine kinases JAK1, JAK2, JAK3 and TYK2, and the alkenyl-containing pyrimidine carboxamide compound, isomers, hydrates, solvates, pharmaceutically acceptable salts or prodrugs thereof and pharmaceutical compositions thereof are expected to be used for preparing medicines for treating autoimmune diseases and cancers related to tyrosine kinases JAK1, JAK2, JAK3 and TYK2, and are particularly suitable for treating autoimmune diseases (such as psoriasis) and the like through transdermal administration.

Detailed Description

Unless otherwise indicated, the following terms used in the present application (including the specification and claims) have the definitions given below. In this application, the use of "or" and "means" and/or "unless stated otherwise. Furthermore, the use of the terms "including" and other forms, such as "including", "comprising", and "having", are not limiting. The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.

"alkyl" refers to an aliphatic hydrocarbon group, "alkyl" moiety, whether saturated or unsaturated, and may be branched or straight chain, "alkyl" may have 1 to 5 carbon atoms, preferably 1 to 3 carbon atoms in aspect the alkyl group is selected from the group consisting of methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, sec-butyl, and tert-butyl.

The term "cycloalkyl" refers to a monocyclic or polycyclic aliphatic nonaromatic group in which each atom (i.e., a backbone atom) making up the ring is a carbon atom.cycloalkyl can be saturated or partially unsaturated.cycloalkyl can be fused to an aromatic ring and the point of attachment is on a carbon other than an aromatic ring carbon atom.cycloalkyl includes groups having 3 to 10 ring atoms.in embodiments the cycloalkyl is selected from cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloheptyl, and cyclooctyl the cycloalkyl can be substituted or unsubstituted. depending on the structure, cycloalkyl can be a monovalent group or a divalent group (i.e., cycloalkylene such as, but not limited to, cyclopropyl-1, 1-diyl, cyclobutyl-1, 1-diyl, cyclopentyl-1, 1-diyl, cyclohexyl-1, 4-diyl, cyclohept-1, 1-diyl, etc.. in aspects, cycloalkyl is C₃-C₆A cycloalkyl group.

"alkenyl" or "cycloalkenyl" refers to a straight or cyclic hydrocarbon chain radical consisting solely of carbon and hydrogen atoms, containing at least double bonds.

"alkoxyalkyl" refers to a (alkyl) O (alkyl) -group, and "alkylthio" refers to a (alkyl) S (alkyl) -group, wherein alkyl is as defined herein. Preferably, the alkoxyalkyl group is C₁-C₅Alkoxyalkyl group, more preferably C₁-C₅Alkoxy radical C₁-C₃Alkyl, more preferably C₁-C₅Alkoxyethyl or C₁-C₅An alkoxypropyl group. Preferably, said alkylthioalkyl is C₁-C₅Alkylthioalkyl, more preferably C₁-C₅Alkylthio group C₁-C₃Alkyl, more preferably C₁-C₅An alkylthio ethyl group.

The term "heterocyclyl" in the term "5-6 membered heterocyclyl" refers to both aromatic heterocycles (also known as heteroaryls) and heterocycloalkyl rings (also known as heteroalicyclic groups) containing or more heteroatoms in the ring, wherein each heteroatom in the ring is selected from O, S and N, wherein each heterocyclic group contains 5 to 6 atoms in its ring system, and the 5-6 membered heterocyclyl may be unsubstituted or substituted with 1 to 2 heteroatoms selected from hydroxy, C₁-C₃Alkyl radical, C₁-C₃Acyl substituents substituted heterocyclyl containing 1-2 heteroatoms selected from N, O, S.

wherein R is¹⁴H, methyl, ethyl, propyl and isopropyl.

Certain compounds described herein contain or more asymmetric centers and thus can produce enantiomers, diastereomers, and other stereoisomeric forms that can be defined as (R) -or (S) -in terms of absolute stereochemistry.

Specifically, for example, for compounds of formula (I) herein, when n is₂Or n₅Is not 0, and R⁴、R⁵Is different from R⁷、R⁸At different times, L₁

Contains a chiral carbon atom in the structure, in which case L₁

The structure of (A) is meant to include all possible isomers, including racemic mixtures, optically active (R) -and (S) -isomers in optically pure form, and the like, i.e., R⁴、R⁵At different times, L₁Is composed of

Comprises that

And racemates; r⁷、R⁸At different times, L₂Is composed of

Comprises that

And racemates.

When a compound described herein contains an olefinic double bond, it is meant that the compound includes various cis-trans isomers, unless otherwise specified.

"tautomers" are structurally different isomers that can interconvert by tautomerization, "tautomerization" is the form of isomerization and includes proton shift or proton shift tautomerization, which can be considered a subset of acid-base chemistry, "proton shift tautomerization" or "proton shift tautomerization" involves proton shift with a shift in bond order, often an exchange of a single bond with an adjacent double bond.

In the present application, the compounds of structural formula (I), isomers, crystals or prodrugs and pharmaceutically acceptable salts thereof may exist in solvated as well as unsolvated forms. For example, the solvated form may be water soluble. The present invention includes all such solvated and unsolvated forms.

Yet another object of the present invention is to provide methods for treating autoimmune diseases and cancer comprising administering to a subject a therapeutically effective amount of a composition comprising a compound of the present invention autoimmune diseases, cancer that can be so treated are noted elsewhere herein, including autoimmune diseases, cancer, etc. that have resistance to treatment with Tofacitinib, peicitinib, Roxolitinib, decentotinib, or other kinase inhibitors.

or more other therapies may also be used in combination in the treatment of cancer, including surgery, radiation therapy (e.g., gamma-rays, neutron beam radiation therapy, electron beam radiation therapy, proton therapy, brachytherapy, and systemic radioisotopes, etc.), endocrine therapy, biological response modifiers (e.g., interferons, interleukins, and Tumor Necrosis Factor (TNF)), hyperthermia, cryotherapy, attenuating any adverse effects (e.g., antiemetics), and other therapeutic drugs.

The invention also includes the use of a compound of the invention, or a pharmaceutically acceptable derivative thereof, for the manufacture of a medicament for the treatment of diseases such as ocular fundus disease, dry eye, psoriasis, rheumatoid arthritis, skin rash, eczema, alopecia areata, atheroma, pulmonary fibrosis, liver fibrosis, myelofibrosis, autoimmune diseases such as enteritis sexually, and tumors, including those that confer resistance to or more other therapeutic agents as noted elsewhere herein.

The invention also provides a method for preparing the corresponding compound, which can be prepared by the following method. The synthetic route for the series (I) of compounds is shown below:

synthetic route to the compounds of formula (I)

Wherein R is¹And R²The specific reaction conditions in steps 1 and 2 are not particularly limited as long as the reaction can be carried out, and reaction conditions are 1) DIEA, THF,25 ℃ and 2-16 h, and 2) TFA and s-BuOH, 60-100 ℃ and 2-16 h.

The present invention will be described in further detail with reference to specific examples for the purpose of making clear the objects, aspects and advantages of the invention, it being understood that the specific examples described herein are intended to be illustrative only and are not intended to be limiting of the invention.

For the synthesis of some of the intermediates used in this application, see chinese patent application No. 2017108562180.

Example 14 preparation of- (butyl-3-en-1-ylamino) -2- ((1- (tert-butyl) -1H-pyrazol-4-yl) amino ] pyrimidine-5-carboxamide:

step 1): preparation of 2-chloro-4- (butyl-3-en-1-ylamino) pyrimidine-5-carboxamide:

2, 4-dichloropyrimidine-5-carboxamide (400mg, 2.08mmol) and triethylamine (633mg, 6.25mmol) were dissolved in tetrahydrofuran (10mL), and butyl-3-en-1-amine hydrochloride (225mg, 2.1mmol) was added and reacted at 25 ℃ for 3 hours. Adding saturated saline solution (200mL), stirring for 15 min, filtering, washing the filter cake with petroleum ether to obtain 380mg of white solid, MS:227[ M + H ]]⁺。

Step 2): preparation of 4- (butyl-3-en-1-ylamino) -2- ((1- (tert-butyl) -1H-pyrazol-4-yl) amino ] pyrimidine-5-carboxamide:

2-chloro-4- (butyl-3-en-1-ylamino) pyrimidine-5-carboxamide (70mg, 0.29mmol) was dissolved in sec-butanol (3mL), 1-tert-butyl-1H-pyrazol-4-amine (49mg, 0.35mmol) and trifluoroacetic acid (0.1mL) were added, and the reaction was blocked at 100 ℃ for 2 hours. The reaction solution was concentrated, filtered, and the solid was washed with acetonitrile to obtain 50mg of a white solid.¹H NMR(400MHz,DMSO-d₆) Delta 10.52(s,1H),10.04(s,1H),8.53(s,1H), 8.30-8.10 (m,1H),8.02(s,1H), 7.71-7.52 (m,2H), 5.92-5.72 (m,1H), 5.20-4.97 (m,2H), 3.68-3.53 (m,2H), 2.43-2.28 (m,2H),1.52(s,9H)₁₆H₂₃N₇O，MS:330(M+H)⁺.

Example 22 preparation of- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((2-methylallyl) amino) pyrimidine-5-carboxamide:

the procedure is as in example 1, replacing butyl-3-en-1-amine hydrochloride in step 1) with 2-methylpropyl-2-en-1-amine to give a white solid,¹H NMR(400MHz,DMSO-d₆) δ 10.36(s,1H),10.12(s,1H),8.55(s,1H), 8.29-8.09 (m,1H),7.98(s,1H), 7.70-7.49 (m,2H), 4.90-4.80 (m,2H),4.10(d, J ═ 5.7Hz,2H),1.76(s,3H),1.51(s,9H). formula C₁₆H₂₃N₇O，MS:330(M+H)⁺.

Example preparation of 32- ((1- (tert-butyl) -1H-pyrazol-4-yl) amino) -4- ((3-methylbutyl-2-en-1-yl) amino) pyrimidine-5-carboxamide:

the procedure is as in example 1, 3-methylbutyl-2-en-1-amine is used instead of butyl-3-en-1-amine hydrochloride in step 1) to give a white solid,¹H NMR(400MHz,DMSO-d₆) Delta 10.48(s,1H),9.94(s,1H),8.51(s,1H), 8.36-8.10 (m,1H),8.06(s,1H), 7.81-7.40 (m,2H), 5.41-5.22 (m,1H), 4.24-3.92 (m,2H),1.71(s,3H),1.67(s,3H),1.50(s,9H). chemical formula C₁₇H₂₅N₇O，MS:344(M+H)⁺.

Example 42 preparation of- ((1- (tert-butyl) -1H-pyrazol-4-yl) amino) -4- (pentyl-4-en-1-ylamino) pyrimidine-5-carboxamide:

the procedure is as in example 1, pentyl-4-en-1-amine is used instead of butyl-3-en-1-amine hydrochloride in step 1) to give a white solid,¹H NMR(400MHz,DMSO-d₆) Δ 10.39(s,1H),10.00(s,1H),8.49(s,1H),8.18(s,1H),8.02(s,1H), 7.71-7.50 (m,2H), 5.91-5.73 (m,1H), 5.10-4.91 (m,2H), 3.60-3.45 (m,2H), 2.14-2.02 (m,2H), 1.78-1.63 (m,2H),1.52(s,9H)₁₇H₂₅N₇O，MS:344(M+H)⁺.

Example preparation of- ((2-methylallyl) amino) -2- ((1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-yl) amino) pyrimidine-5-carboxamide:

the procedure is as in example 1, replacing butyl-3-en-1-amine hydrochloride in step 1) with 2-methylpropyl-2-en-1-amine and 1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-amine with 1-tert-butyl-1H-pyrazol-4-amine in step 2) to give a white solid,¹H NMR(400MHz,DMSO-d₆)δ10.55(s,1H),10.16(s,1H),8.59(s,1H), 8.37-8.16 (m,1H),7.95(s,1H), 7.72-7.53 (m,2H), 4.92-4.76 (m,2H), 4.45-4.31 (m,1H),4.09(d, J ═ 5.8Hz,2H), 4.01-3.88 (m,2H), 3.48-3.41 (m,2H), 2.02-1.82 (m,4H),1.76(s,3H)₁₇H₂₃N₇O₂，MS:358(M+H)⁺.

Example preparation of 64- ((3-methylbutyl-2-en-1-yl) amino) -2- ((1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-yl) amino) pyrimidine-5-carboxamide:

the same procedure as in example 1 was followed, except that 3-methylbutyl-2-en-1-amine hydrochloride in step 1) was used in place of butyl-3-en-1-amine hydrochloride and 1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-amine was used in place of 1-tert-butyl-1H-pyrazol-4-amine in step 2), to give a white solid,¹H NMR(400MHz,DMSO-d₆) Delta 10.47(s,1H),9.93(s,1H),8.50(s,1H), 8.29-8.07 (m,1H),8.00(s,1H), 7.71-7.50 (m,2H), 5.36-5.24 (m,1H), 4.46-4.30 (m,1H), 4.20-4.06 (m,2H), 4.00-3.87 (m,2H), 3.54-3.38 (m,2H), 2.02-1.80 (m,4H),1.71(s,3H),1.68(s,3H)₁₈H₂₅N₇O₂，MS:372(M+H)⁺.

Example preparation of 74- (butyl-3-en-1-ylamino) -2- ((1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-yl) amino) pyrimidine-5-carboxamide:

the procedure of example 1 was followed, replacing 1-tert-butyl-1H-pyrazol-4-amine in step 2) with 1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-amine to give a white solid,¹H NMR(400MHz,DMSO-d₆) Delta 10.63(s,1H),10.06(s,1H),8.55(s,1H), 8.38-8.12 (m,1H),7.99(s,1H), 7.76-7.45 (m,2H), 5.95-5.73 (m,1H), 5.21-5.02 (m,2H), 4.48-4.29 (m,1H), 4.04-3.87 (m,2H), 3.69-3.53 (m,2H), 3.52-3.34 (m,2H), 2.43-2.28 (m,2H), 2.01-1.81 (m,4H) of the formula C₁₇H₂₃N₇O₂，MS:358(M+H)⁺.

Example preparation of 82- ((1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-yl) amino) -4- (pentyl-4-en-1-ylamino) pyrimidine-5-carboxamide:

the procedure is as in example 1, pentyl-4-en-1-amine is used instead of butyl-3-en-1-amine hydrochloride in step 1) and 1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-amine is used instead of 1-tert-butyl-1H-pyrazol-4-amine in step 2) to give a white solid,¹H NMR(400MHz,DMSO-d₆) Delta 10.41(s,1H),10.00(s,1H),8.48(s,1H), 8.28-8.07 (m,1H),7.97(s,1H), 7.70-7.52 (m,2H), 5.91-5.75 (m,1H), 5.10-4.92 (m,2H), 4.46-4.33 (m,1H), 4.01-3.91 (m,2H), 3.57-3.41 (m,4H), 2.15-2.05 (m,2H), 2.02-1.86 (m,4H), 1.77-1.64 (m,2H) formula C₁₈H₂₅N₇O₂，MS:372(M+H)⁺.

Example preparation of 92- ((1- (tert-butyl) -1H-pyrazol-4-yl) amino) -4- ((cyclopentyl-3-en-1-ylmethyl) amino) pyrimidine-5-carboxamide:

step 1): preparation of 2-chloro-4- ((cyclopentyl-3-en-1-ylmethyl) amino) pyrimidine-5-carboxamide:

2, 4-dichloropyrimidine-5-carboxamide (400mg, 2.08mmol) and triethylamine (633mg, 6.25mmol) were dissolved in tetrahydrofuran (10mL), and cyclopentyl-3-en-1-ylmethylamine hydrochloride (281mg, 2.1mmol) was added and reacted at 25 ℃ for 3 hours. Adding saturated saline solution (200mL), stirring for 15 min, filtering, washing the filter cake with petroleum ether to obtain 380mg of white solid, MS:253[ M + H ]]⁺。

Step 2): preparation of 2- ((1- (tert-butyl) -1H-pyrazol-4-yl) amino) -4- ((cyclopentyl-3-en-1-ylmethyl) amino) pyrimidine-5-carboxamide:

2-chloro-4- ((cyclopentyl-3-en-1-ylmethyl) amino) pyrimidine-5-carboxamide (127mg, 0.5mmol) was dissolved in sec-butanol (3mL), 1-tert-butyl-1H-pyrazol-4-amine (84mg, 0.6mmol) and trifluoroacetic acid (0.1mL) were added, and the reaction was blocked at 100 ℃ for 2 hours. The reaction solution was concentrated and purified by column chromatography to obtain 50mg of a white solid.¹H NMR(400MHz,DMSO-d₆)δ10.58(s,1H),10.16(s,1H),8.55(s,1H),8.37–8.14(m,1H),8.02(s,1H),7.71–7.52(m,2H),5.75–5.63(m,2H),3.57–3.44(m,2H),2.72–2.56(m,1H),2.49–2.39(m,2H),2.15–2.01(m,2H),1.52(s,9H).MS:356[M+H]⁺。

Example preparation of 104- ((cyclopentyl-3-en-1-ylmethyl) amino) -2- ((1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-yl) amino) pyrimidine-5-carboxamide:

step 1: same as example 9) step 1;

step 2) preparation of 4- ((cyclopentyl-3-en-1-ylmethyl) amino) -2- ((1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-yl) amino) pyrimidine-5-carboxamide:

the procedure is as in example 9, step 2) using 1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-amine instead of 1-tert-butyl-1H-pyrazol-4-amine to give a white solid.¹H NMR(400MHz,DMSO-d₆)δ10.62(s,1H),10.16(s,1H),8.56(s,1H),8.26(s,1H),7.98(s,1H),7.71–7.53(m,2H),5.75–5.65(m,2H),4.46–4.31(m,1H),4.01–3.90(m,2H),3.57–3.39(m,4H),2.70–2.55(m,1H),2.48–2.36(m,2H),2.17–2.04(m,2H),2.03–1.82(m,4H).MS:384[M+H]⁺。

Example 114 preparation of (allylamino) -2- ((1- (tert-butyl) -1H-pyrazol-4-yl) amino) pyrimidine-5-carboxamide:

the procedure is as in example 9, allylamine hydrochloride being used instead of cyclopentyl-3-en-1-ylmethylamine hydrochloride to give a white-like solid.¹HNMR(400MHz,DMSO-d₆)δ10.71(s,1H),10.16(s,1H),8.62(s,1H),8.31(s,1H),8.00(s,1H),7.77–7.54(m,2H),6.07–5.89(m,1H),5.27–5.11(m,2H),4.25–4.12(m,2H),1.51(s,9H).MS:316[M+H]⁺。

EXAMPLE 12 preparation of (Z) -2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((pentan-2-en-1-yl) amino) pyrimidine-5-carboxamide

The procedure is as in example 9, substituting (Z) -pentan-2-en-1-amine for cyclopentyl-3-en-1-ylmethylamine hydrochloride to give an off-white solid.¹H NMR(400MHz,DMSO-d₆)δ9.44(s,1H),9.18(d,J＝28.1Hz,1H),8.46(s,1H),8.02(d,J＝9.3Hz,1H),7.83–7.56(m,1H),7.48(s,1H),7.23–6.97(m,1H),5.72–5.52(m,2H),4.14–4.03(m,2H),2.14–1.98(m,2H),1.49(s,9H),0.99–0.91(m,3H).MS:344[M+H]⁺。

Example 13 preparation of (E) -2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((4-methylpentan-2-en-1-yl) amino) pyrimidine-5-carboxamide

The procedure is as in example 9, substituting (E) -4-methylpentane-2-en-1-amine for cyclopentyl-3-en-1-ylmethylamine hydrochloride to give an off-white solid.¹H NMR(400MHz,DMSO-d₆)δ9.45(s,1H),9.24(s,1H),8.46(s,1H),8.01(s,1H),7.87–7.51(m,1H),7.48(s,1H),7.39–6.79(m,1H),5.75–5.39(m,2H),4.25–3.86(m,2H),2.40–2.10(m,1H),1.49(s,9H),0.94(d,J＝6.7Hz,6H).MS:358[M+H]⁺。

Example 14 preparation of (E) -2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((pentan-3-en-1-yl) amino) pyrimidine-5-carboxamide

The procedure is as in example 9, substituting (E) -pentan-3-en-1-amine for cyclopentyl-3-en-1-ylmethylamine hydrochloride to give an off-white solid.¹H NMR(400MHz,DMSO-d₆)δ9.41(s,1H),9.19(s,1H),8.44(s,1H),8.00(s,1H),7.67–7.53(m,1H),7.50(s,1H),7.21–6.93(m,1H),5.58–5.36(m,2H),3.52–3.47(m,2H),2.37–2.20(m,2H),1.63(d,J＝5.9Hz,3H),1.49(s,9H).MS:344[M+H]⁺。

Example preparation of 152- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((3-methylbutyl-3-en-1-yl) amino) pyrimidine-5-carboxamide

The procedure is as in example 9, 3-methylbutyl-3-en-1-ylamine is used instead of cyclopentyl-3-en-1-ylmethylamine hydrochloride to give an off-white solid.¹H NMR(400MHz,DMSO-d₆)δ9.40(s,1H),9.17(s,1H),8.44(s,1H),7.97(s,1H),7.82–7.59(m,1H),7.52(s,1H),7.18–6.87(m,1H),4.84–4.77(m,1H),4.77–4.72(m,1H),3.67–3.50(m,2H),2.39–2.22(m,2H),1.73(s,3H),1.49(s,9H).MS:344[M+H]⁺。

Example 162 preparation of- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((4-methylpentan-3-en-1-yl) amino) pyrimidine-5-carboxamide

The procedure is as in example 9, 4-methylpentane-3-en-1-amine is used instead of cyclopentyl-3-en-1-ylmethylamine hydrochloride to give an off-white solid.¹H NMR(400MHz,DMSO-d₆)δ9.41(s,1H),9.16(s,1H),8.45(s,1H),8.01(s,1H),7.82–7.55(m,1H),7.50(s,1H),7.24–6.87(m,1H),5.21–5.11(m,1H),3.54–3.43(m,2H),2.36–2.22(m,2H),1.67(s,3H),1.57(s,3H),1.49(s,9H).MS:358[M+H]⁺。

Example preparation of 172- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((4-methylpentan-4-en-1-yl) amino) pyrimidine-5-carboxamide

The procedure is as in example 9, 4-methylpentane-4-en-1-amine is used instead of cyclopentyl-3-en-1-ylmethylamine hydrochloride to give an off-white solid.¹H NMR(400MHz,DMSO-d₆)δ9.46(s,1H),9.27(s,1H),8.45(s,1H),8.00(s,1H),7.86–7.59(m,1H),7.51(s,1H),7.31–6.76(m,1H),4.78–4.63(m,2H),3.52–3.44(m,2H),2.06(t,J＝7.7Hz,2H),1.80–1.70(m,2H),1.69(s,3H),1.50(s,9H).MS:358[M+H]⁺。

Example 182 preparation of- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((5-methylhexan-4-en-1-yl) amino) pyrimidine-5-carboxamide

The procedure is as in example 9, 5-methylhexan-4-en-1-amine is used instead of cyclopentyl-3-en-1-ylmethylamine hydrochloride to give an off-white solid.¹H NMR(400MHz,DMSO-d₆)δ9.40(s,1H),9.21(s,1H),8.44(s,1H),7.98(s,1H),7.82–7.54(m,1H),7.52(s,1H),7.26–6.84(m,1H),5.21–5.01(m,1H),2.07–1.99(m,2H),1.64(s,6H),1.57–1.53(m,4H),1.49(s,9H).MS:372[M+H]⁺。

Example 19 preparation of (E) -2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((5-methylhexan-2-en-1-yl) amino) pyrimidine-5-carboxamide

The procedure is as in example 9, substituting (E) -5-methylhexan-2-en-1-amine for cyclopentyl-3-en-1-ylmethylamine hydrochloride to give an off-white solid.¹H NMR(400MHz,DMSO-d₆)δ9.40(s,1H),9.18(s,1H),8.44(s,1H),8.01(s,1H),7.77–7.54(m,1H),7.50(s,1H),7.22–6.80(m,1H),5.56–5.44(m,1H),5.44–5.31(m,1H),3.55–3.42(m,2H),2.35–2.14(m,3H),1.50(s,9H),0.93(d,J＝6.7Hz,6H).MS:372[M+H]⁺。

Example 202 preparation of- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- (((2E,4E) -hexane-2, 4-dien-1-yl) amino) pyrimidine-5-carboxamide

The procedure is as in example 9, replacing cyclopentyl-3-en-1-ylmethylamine hydrochloride with (2E,4E) -hexane-2, 4-dien-1-amine to give an off-white solid.¹H NMR(400MHz,DMSO-d₆)δ9.48(s,1H),9.29(s,1H),8.47(s,1H),7.98(s,1H),7.86–7.58(m,1H),7.49(s,1H),7.34–6.92(m,1H),6.22–5.99(m,2H),5.82–5.70(m,1H),5.70–5.55(m,1H),4.19–4.02(m,2H),1.69(d,J＝6.7Hz,3H),1.48(s,9H).MS:356[M+H]⁺。

Example 212 preparation of- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- (((2E,4E) -heptane-2, 4-dien-1-yl) amino) pyrimidine-5-carboxamide

The procedure is as in example 9, replacing cyclopentyl-3-en-1-ylmethylamine hydrochloride with (2E,4E) -heptane-2, 4-dien-1-amine to give an off-white solid.¹H NMR(400MHz,DMSO-d₆)δ9.44(s,1H),9.25(s,1H),8.47(s,1H),7.98(s,1H),7.88–7.58(m,1H),7.48(s,1H),7.32–6.86(m,1H),6.16(dd,J＝15.1,10.4Hz,1H),6.04(dd,J＝15.1,10.5Hz,1H),5.86–5.60(m,2H),4.21–4.05(m,2H),2.12–1.96(m,2H),1.48(s,9H),0.94(t,J＝7.5Hz,3H).MS:370[M+H]⁺。

Experimental example 1 test for inhibition of JAK1, JAK2, JAK3, TYK2 kinase Activity by Small molecule Compounds

In the in vitro assembled enzymatic reaction, compounds with different concentrations are added to detect the inhibition of the specific enzymatic reaction by the compounds, and the specific test method is as follows:

instruments, materials and reagents

Second, testing method

Specific experimental conditions for JAK1, JAK2, JAK3 and TYK2, exemplified by JAK3, are described below.

1. Preparing a reagent:

preparation of EDTA (0.5M pH8.0) solution by accurately weighing 14.612g of EDTA powder, adding ultrapure water, and adjusting the volume to 100mL (if insoluble, heating to 37 deg.C, adjusting pH to 8.0 with 1N NaOH solution)

1 × Kinase Assay Buffer: into a reagent bottle were added 25mL of HEPES solution (1M), 190.175mgEGTA, 5mL of MgCl₂The solution (1M), 1mL DTT, 50. mu.L Tween-20, and ultrapure water were added to make a volume of 500mL (pH adjusted to 7.5).

1 × Detection Buffer 1mL of 10 × Detection Buffer was added to 9mL of water and mixed.

4 × stop solution: 0.8mL of the above EDTA (0.5M, pH8.0) solution, 1mL of 10 XDetection Buffer, and 8.2mL of ultrapure water were mixed together.

4 XJAK 3 Kinase solution A Kinase stock solution was diluted with 1 XKinase Assay Buffer to a concentration of 0.36nM, mixed well and stored on ice.

4 × substrate solution: substrate ULight was diluted with 1 × Kinase Assay Buffer^TM-Labeled JAK-1(Tyr1023) Peptide stock solution to 200nM and mixed well.

4 × ATP solution: the ATP stock was diluted with 1 XKinase Assay Buffer to a concentration of 40. mu.M and mixed well.

4 × detection solution: the Detection antibody Europium-anti-phosphorus-tyrosine antibody (PT66) was diluted with 1 XDeprotection Buffer to a concentration of 8nM and mixed well.

2 × substrate/ATP mixture: the 4 Xsubstrate solution and 600. mu.l of 4 XATP solution were mixed in equal amounts (prepared before use).

2. Experimental procedure

1) The dilution of the compound is carried out,

in a 96-well plate, compounds were diluted with DMSO solutions at a ratio of 3 times to form 11 gradients, and pure DMSO solutions were used as positive controls, and new 96-well plates were prepared, and the solutions were diluted 25 times with ultrapure water (DMSO concentration: 4%)

2) Rotating Compounds to 384 well plates

The compound solution diluted with ultrapure water in the above 96-well plate was transferred to the corresponding well of a 384-well plate according to a standard 2-well carousel.

3) Add 4 × kinase solution: mu.l of the 4 Xkinase solution was added to the corresponding reaction well of the 384-well plate using a line gun, and pre-reacted at room temperature for 5 minutes.

4) Add 2 Xsubstrate/ATP mix 5. mu.l of the above 2 Xsubstrate/ATP mix to the corresponding reaction well of a 384 well plate using a discharge gun.

5) Negative control: a negative control well was set in a 384-well plate, and 2.5. mu.l/well of 4 Xsubstrate, 2.5. mu.l of 4 Xenzyme solution, 2.5. mu.l of 1 XKinase Assay Buffer, and 2.5. mu.l of ultrapure water containing 4% DMSO were added to the well.

6) And (4) centrifuging, mixing uniformly, and reacting for 60min at room temperature in the dark.

7) Termination of the enzymatic reaction:

5. mu.l of the 4 Xstop solution was pipetted into the corresponding well of 384-well plate, centrifuged and mixed, and reacted at room temperature for 5 minutes.

8) And (3) color development reaction:

5. mu.l of the 4 × detection solution was pipetted into the mesopores of 384-well plate, centrifuged and mixed, and reacted at room temperature for 60 min.

9) The 384 well plate is placed in a plate reader, and a corresponding program detection signal is called.

10) Inhibition rate and IC₅₀And (3) calculating:

pore reading 10000 × EU 665/EU 615 values

Inhibition rate [1- (experimental well reading-negative control well reading)/(positive control well reading-negative control well reading) ]. 100%

Inputting the drug concentration and the corresponding inhibition rate into GraphPad Prism5 for processing and calculating the corresponding IC₅₀The value is obtained.

Test conditions:

JAK1 kinase activity assay:

JAK1 (final concentration 10 nM); ATP (final concentration 10 μ M); ULight^TM-labeled JAK-1(Tyr1023) Peptide (final concentration 100 nM); the enzymatic reaction time was 2 hours. The maximum final concentration of the compound was 2.5. mu.M, and after 3-fold gradient dilution, the minimum final concentration was 0.042 nM. The final concentration of DMSO is 1%.

JAK2 kinase activity assay:

JAK2 (final concentration 0.25 nM); ATP (final concentration 5 μ M); ULight^TM-labeled JAK-1(Tyr1023) Peptide (final concentration 50 nM); the enzymatic reaction time was 1 hour. The maximum final concentration of the compound was 2.5. mu.M, and after 3-fold gradient dilution, the minimum final concentration was 0.042 nM. The final concentration of DMSO is 1%.

JAK3 kinase activity assay:

JAK3 (final concentration 0.36 nM); ATP (final concentration 10 μ M); ULight^TM-labeled JAK-1(Tyr1023) Peptide (final concentration 50 nM); the enzymatic reaction time was 1 hour. The maximum final concentration of the compound was 2.5. mu.M, and after 3-fold gradient dilution, the minimum final concentration was 0.042 nM. The final concentration of DMSO is 1%.

TYK2 kinase activity assay:

TYK2 (final concentration 8 nM); ATP (final concentration 20 μ M); ULight^TM-labeled JAK-1(Tyr1023) Peptide (final concentration 100 nM); the enzymatic reaction time was 2 hours. The maximum final concentration of the compound was 2.5. mu.M, and after 3-fold gradient dilution, the minimum final concentration was 0.042 nM. The final concentration of DMSO is 1%.

Table 1 shows the results of measurement of inhibitory activity of some of the compounds of the present invention against tyrosine kinases JAK1, JAK2, JAK3, TYK 2. IC in the following Table₅₀The values represent the concentration of compound at which the enzyme inhibits maximally 50%, and NT means that the corresponding enzyme was not tested.

TABLE 1 results of measurement of inhibitory Activity of partial compounds of the present invention against JAK1, JAK2, JAK3 and TYK2 tyrosine kinases

Experimental example 2 test of inhibitory Activity of Compound of the present invention on mouse spleen cell proliferation

The specific experimental steps are as follows:

1) compound dilution: a total of 9 concentrations were obtained starting from the highest concentration of 5000nM after 3-fold gradient dilution (5000 nM for the maximum final concentration of drug and 0.76nM for the minimum final concentration in this experiment).

2) Placing a cell filter sieve with apertures of 70 mu m in a culture dish with diameters of 6 cm, and adding 2mL of HBSS solution into the sieve to infiltrate the bottom of the culture dish;

3) euthanasia adult Balb/c mice with carbon dioxide, soaking in 75% alcohol for 1 minute, placing into a safety cabinet, cutting a small opening in the middle of the left ventral side of the mice, exposing the abdominal wall, and finding out the spleen;

4) taking spleen, removing surrounding adipose tissues, putting the spleen on a cell filtering sieve in a culture dish, and properly shearing;

5) gently grinding the spleen with the flat part at the top end of the syringe piston to obtain a cell suspension;

6) collecting cell suspension from a culture dish, and slowly adding the cell suspension into a 15ml centrifuge tube containing 5ml of Ficoll-Paque PLUS;

7) centrifugation was carried out at 400g for 30 minutes at room temperature;

8) after the centrifugation is finished, the upper layer is slowly removed by a suction pipe, and then the middle layer, namely spleen cells, is slowly sucked out;

9) placing the collected spleen cell suspension into another 15ml centrifuge tube, adding 10ml RPMI1640 complete culture medium, centrifuging at 300g and 4 ℃ for 4 minutes;

10) discarding the supernatant, adding the complete culture medium to resuspend the cells, and then counting the cells, and washing the cell suspension times according to the ninth step;

11) transferring the cells to a culture dish (containing 2.5 mu g/mL of concanavalin A) at a cell density of 200-500 ten thousand/mL for overnight culture;

12) the next day, the cells were transferred to a 15mL centrifuge tube and centrifuged at 300g for 5 min;

13) discarding the supernatant, adding 5mL of RPMI1640 complete culture solution, blowing and beating uniformly, mixing 10 μ L of cell suspension and 10 μ L of trypan blue uniformly, counting by using a cell counter, and recording the number of cells and the survival rate.

14) Inoculating the cell suspension into a 96-well plate, and inoculating 80 mu l of cell suspension into each well, wherein the density of the cell suspension is 100000 cells/well;

15) adding 20 μ L of the 5 × compound solution diluted with the culture solution into each well, and mixing well;

16) after 72 hours of culture, 10. mu.L of CCK-8 reagent is added into each well, and the mixture is incubated for 2 hours (the reaction time can be adjusted according to the color depth);

17) the OD value was read at 450nm on a multifunction plate reader.

18) Data processing:

cell survival (%) - (As-Ab)/(Ac-Ab) ]. 100%

As: OD value of the experimental well (cell-containing medium, CCK-8, compound),

ac: OD value of control well (cell-containing medium, CCK-8),

ab: OD of blank wells (medium without cells and compounds, CCK-8),

the values were then introduced into Graphpad Prism5 software for curve fitting and IC50 was calculated.

Table 2 shows the results of the test of the inhibitory activity of some compounds of the present invention on mouse splenocyte proliferation.

TABLE 2 test results of the inhibitory Activity of some of the Compounds of the invention on mouse splenocyte proliferation

Use, formulation, administration

Medical use and indications

The compounds of the invention are also useful for the treatment of autoimmune diseases, including but not limited to psoriasis, vitiligo, dermatitis, alopecia areata, rheumatoid arthritis, colitis, multiple sclerosis, systemic lupus erythematosus, crohn's disease, including primary and metastatic cancers, such cancers, including solid tumors, such cancers include but not limited to non-small cell lung cancer, breast cancer, pancreatic cancer, glioma, glioblastoma, ovarian cancer, cervical cancer, colorectal cancer, melanoma, endometrial cancer, prostate cancer, leukemia, gastric cancer, gastrointestinal stromal tumor, thyroid cancer, chronic myelogenous leukemia, chronic cell leukemia, acute lymphoblastic leukemia, non-cervical cancer, colorectal cancer, melanoma, endometrial cancer, prostate cancer, leukemia, renal cancer, gastrointestinal stromal tumor, nasopharyngeal carcinoma, thyroid cancer, chronic myelogenous leukemia, chronic cell leukemia, non-malignant lymphoma, chronic myelogenous leukemia, non-malignant lymphoma, multiple myeloma, and multiple myeloma tumors including JAK kinase, multiple myeloma, multiple.

Pharmaceutical process

The pharmaceutical methods of the invention include determining a therapeutically effective dose for a subject in need of a compound of the invention, "therapeutically effective dose" will vary depending on the stage, progression or severity of the disease, the daily dose of the compounds and compositions of the invention will depend on a variety of factors including the condition being treated, the severity of the condition, the potency of the particular compound employed, the particular composition, the age, body weight, general health, sex and diet, the route and schedule of administration, metabolism and/or rate of excretion of the compound, the duration of treatment, etc. furthermore, the desired dose of the compounds of the invention will be administered to humans and other animals after being formulated with a pharmaceutically acceptable carrier, the mode of administration includes oral, rectal, parenteral, intracisternal, intravaginal, intraperitoneal, topical (as by transdermal, powder, ointment, or drop), sublingual, buccal, or nasal spray, etc. the effective dose of the compounds of the invention will generally be metered per kg of the patient, preferably at a daily dose of 0.1 mg/125 mg/body weight, mg/3501, or as a single daily dose of the compound may be administered as a single dose per week or as a multiple dose per unit, or as a combination of the case where the desired dose of the compound may be administered alone, or as a single dose of the case, the case where the desired dose of the invention may vary from a single dose of the case, the invention may be administered, the case may be administered alone, the compound, the case, the invention may be administered alone, as a single dose of the case, the invention may be administered at a single dose of a dosage of the invention, or multiple times of a single dose of the invention may be administered at a dosage of a daily dosage of a therapeutic agent, preferably, the invention, for example, the invention may be administered at a daily dosage of a therapeutic agent, the invention may be administered at a therapeutic agent, the invention may be administered at a dosage of a daily dosage of a therapeutic agent, the invention may be administered at a therapeutic agent, the invention may be administered at a dosage of a daily.

As to compounds

The compounds of the invention may be used in therapy in free form or, where appropriate, in the form of pharmaceutically acceptable salts or other derivatives. As used herein, the term "pharmaceutically acceptable salts" refers to organic and inorganic salts of the compounds of the present invention which are suitable for use in humans and lower animals without undue toxicity, irritation, allergic response and the like, commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable salts of amines, carboxylic acids, phosphonates, and other types of compounds are well known in the art. The salts can be formed by reacting the isolated and purified compounds of the present invention with a suitable free base or acid.

Other pharmaceutically acceptable salts include adipates, alginates, ascorbates, aspartates, benzenesulfonates, benzoates, bisulfates, borates, butyrates, camphorates, citrates, cyclopentanes, digluconates, dodecylsulfates, ethanesulfonates, formates, fumarates, glucoheptonates, glycerophosphates, gluconates, hemisulfates, heptates, hexanoates, hydroiodides, 2-hydroxyethanesulfonates, lactobionates, lactates, laurates, malates, maleates, malonates, methanesulfonates, 2-naphthalenesulfonates, nicotinates, nitrates, oleates, oxalates, palmitates, pamoates, propionates, perpropionates, monopropionates, monophenylalanates, perchlorates, sodium monophenylalanates, perchlorates, ammonium hydrogen sulfate, potassium monopersulfates, ammonium hydrogen sulfate, sodium hydrogen sulfate.

In addition, the term "prodrug" as used herein means that compounds can be converted in vivo to the compounds of formula (I) of the present invention by hydrolysis of the prodrug in the blood or by enzymatic action in the blood or tissue to the parent compound.

Composition comprising a metal oxide and a metal oxide

The compositions described herein consist of compounds (or prodrugs, or pharmaceutically acceptable salts, or other pharmaceutically acceptable derivatives thereof) as described herein, and one or more pharmaceutically acceptable carriers or excipients, these compositions may optionally further comprise one or more additional therapeutic agents the compounds of the invention may be co-administered to a patient in need thereof with one or more additional therapeutic regimens (e.g., administration combinations of Tofacitinib or other kinase inhibitors, interferons, bone marrow transplants, farnesyl transferase inhibitors, bisphosphonates, thalidomide, cancer vaccines, hormonal therapies, antibodies, radiation, etc.).

As described herein, the compositions of the present invention comprise a compound of the present invention in combination with a pharmaceutically acceptable carrier and/or excipient, including any and all solvents, diluents or other carriers, dispersing or suspending aids, surfactants, isotonicity agents, thickeners or emulsifiers, preservatives, solid binders, lubricants and the like, as appropriate to the particular dosage form desired examples of such pharmaceutically acceptable carrier materials include, but are not limited to, sugars such as lactose, glucose and sucrose, starches such as corn and potato starch, celluloses and derivatives thereof such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate, powdered tragacanth, malt, gelatin, talc, excipients such as cocoa butter and suppository waxes, oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil, glycols such as propylene glycol, esters such as ethyl oleate and ethyl laurate, agar, buffers such as magnesium hydroxide and aluminum hydroxide, alginic acid, pyrogen-free water, isotonic saline, ringer's solution, ethanol, and phosphate buffer solutions, and other lubricant compatibility agents such as sodium lauryl sulfate and magnesium stearate, and flavoring agents, as well as nontoxic coloring agents, preservatives, coating agents and flavoring agents may also be present in the non-toxic compositions.

Formulation of

The present invention also encompasses -class compositions (collectively referred to herein as "carrier" materials) of the active compounds of the present invention in combination with or more pharmaceutically acceptable carriers and/or diluents and/or adjuvants, and if desired, other active ingredients the active compounds of the present invention may be administered by any suitable route, preferably in the form of a pharmaceutical composition suitable for such route of administration, at an effective dose as required for the intended treatment, the administration of the compounds and compositions of the present invention may be in the form of oral, mucosal, topical, rectal, pulmonary (e.g., by inhalation spray), or parenteral, including intravascular, intravenous, intraperitoneal, subcutaneous, intramuscular, intrasternal, and infusion techniques.

The active compounds of the present invention generally constitute routes of administration with one or more adjuvants, excipients, or carriers if administered orally, the compounds may be mixed with lactose, sucrose, starch powder, cellulose alkanoates, cellulose alkyl esters, talc, stearic acid, magnesium stearate, magnesium oxide, sodium and calcium salts of phosphoric and sulfuric acids, gelatin, acacia, sodium alginate, polyvinylpyrrolidone, and/or polyvinyl alcohol, and then tableted or encapsulated for convenient administration.such capsules or tablets may comprise controlled release formulations, the active compounds may be dispersed in hydroxypropylmethylcellulose to provide suitable formulations for topical administration including liquid or semi-liquid formulations suitable for penetration through the skin (such as liniments, ointments, lotions, creams, or pastes) and drops suitable for administration to the eye, ear, or nose. suitable topical administration dosages of the compounds of the present invention are from 0.1 to 150 milligrams, to four times per day, preferably 1 to 2 times per day.

The active ingredient may also be administered by injection, in combination with a suitable carrier including saline, dextrose or water, or with cyclodextrin (Captisol), co-solvent solubilization (i.e., propylene glycol), or micelle solubilization (i.e., tween 80). The formulations may also be presented as sterile injectable solutions or suspensions in a non-toxic parenterally-acceptable diluent or solvent, for example as 1, 3-butanediol. Solvents which may be used are water, ringer's solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. Any bland fixed oil may be employed for this purpose including synthetic mono-or diglycerides.

For pulmonary administration, the pharmaceutical compositions may be administered in the form of an aerosol or with an inhaler, including a dry powder aerosol. Suppositories for rectal administration of the drug can be prepared by mixing the drug with a suitable non-irritating excipient, such as cocoa butter and polyethylene glycols which are solid at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum and release the drug. The pharmaceutical compositions may be formulated with conventional pharmaceutical procedures such as sterilization and/or may contain conventional adjuvants such as preservatives, stabilizers, wetting agents, emulsifiers, buffers and the like in tablets and pills or may be formulated with enteric coatings. Such compositions may also contain adjuvants such as wetting agents, sweetening, flavoring and perfuming agents.

The pharmaceutical compositions of the invention comprise a compound of formula (I) as described herein or a pharmaceutically acceptable salt thereof, a kinase inhibitor (small molecule, polypeptide, antibody, etc.), an immunosuppressant, an anticancer agent, an antiviral agent, an anti-inflammatory agent, an antifungal agent, an antibiotic or an anti-vascular hyperproliferative compound, in addition to any pharmaceutically acceptable carrier, adjuvant or excipient.

The term "pharmaceutically acceptable carrier or adjuvant" refers to carriers or adjuvants that may be administered to a patient with compound of the present invention and which do not destroy pharmaceutical activity and are non-toxic at dosages sufficient to deliver a therapeutic amount for administration.pharmaceutically acceptable carriers, adjuvants and excipients may be used in the pharmaceutical compositions of the present invention, including but not limited to, ion exchangers, alumina, aluminum stearate, lecithin, Self Emulsifying Drug Delivery Systems (SEDDS), such as d-atocopHerol polyethylene glycol 1000 succinate, surfactants used in pharmaceutical dosage forms, such as tweens or other similar polymeric delivery matrices, serum proteins such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, silicon dioxide, magnesium trisilicate, polyvinyl pyrrolidone, cellulose-based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, polyethylene-polyoxypropylene block polymers, polyethylene glycol-polyoxypropylene block polymers, and polyethylene glycol-cyclodextrin block polymers, and other solubilizing agents useful for oral formulations, such as are also useful in the art of the aqueous suspension of the aqueous or aqueous dispersions and/aqueous dispersions of the pharmaceutical formulations described herein, including but also useful for the oral suspension of the pharmaceutical formulations of the present invention, including, and other pharmaceutical formulations, such as the aqueous dispersions and the use of the aqueous dispersions and the suspending agents may be formulated in the art of pharmaceutical formulations of the art of which may be used to enhance the use of the oral formulation of the use of the pharmaceutical formulations of the pharmaceutical excipients, including, the aqueous dispersions and/or modified, including, the aqueous dispersions of the pharmaceutical excipients, the aqueous dispersions and/or modified, the aqueous dispersions of the pharmaceutical excipients, the aqueous dispersions of the aqueous dispersions and/or modified, the aqueous dispersions of the pharmaceutical excipients, the aqueous dispersions of the pharmaceutical excipients, the adjuvants, the aqueous dispersions of the adjuvants, the adjuvants.

Combination drug

The compounds of the present invention may be used alone, or in combination with or more other compounds of the present invention or with or more other agents when administered in combination, the therapeutic agents may be formulated for simultaneous administration or sequential administration at different times, or the therapeutic agents may be administered as a single composition so-called "combination therapy" refers to the use of the compounds of the present invention with another agent by simultaneous co-administration of each agent or sequential administration of each agent, whichever is the case in order to achieve the best effect of the drug.

The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents and improvements made within the spirit and principle of the present invention are intended to be included within the scope of the present invention.

Claims

A compound of structural formula (I), its isomer, hydrate, solvate, pharmaceutically acceptable salt or prodrug:

wherein the content of the first and second substances,

x is N;

R¹is composed of

n₁Is an integer of 0 to 8, n₂Is an integer of 0 to 1, n₃Is an integer of 0 to 8, and n₁、n₂、n₃The sum is 10 or less;

L₁is composed of

R⁴、R⁵Each independently is H, C₁-C₃Alkyl radical, R⁴、R⁵The same or different, or a combination thereof,

R³is unsubstituted or C₁-C₃Alkyl substituted C₂-C₈Alkenyl or notSubstituted or C₁-C₃Alkyl substituted C₄-C₈Cycloalkenyl group of (a);

R²is composed of

n₄Is an integer of 0 to 8, n₅Is an integer of 0 to 1, n₆Is an integer of 0 to 8, and n₄、n₅、n₆The sum is 10 or less;

L₂is composed of

R⁷、R⁸Each independently is H, C₁-C₃Alkyl radical, R⁷、R⁸The same or different, or a combination thereof,

R⁶is-H, hydroxyalkyl, C₁-C₅Alkyl radical, C₃-C₈Cycloalkyl radical, C₁-C₅Alkoxyalkyl group, C₁-C₅Alkylthioalkyl, 5-6 membered heterocyclyl, or-NR 'R ", wherein R', R" are each independently H, C₁-C₃Alkyl group of (1).
2. The compound of claim 1, an isomer, hydrate, solvate, pharmaceutically acceptable salt or prodrug thereof,

wherein R is³Is composed of

R⁹、R¹⁰、R¹¹、R¹²、R¹³Each independently is H, C₁-C₃An alkyl group, m is an integer of 0 to 2,

m₁is an integer of 0 to 5, m₂Is an integer of 0 to 5, and m₁,m₂The sum is less than or equal to 5.
3. The compound of claim 1, an isomer, hydrate, solvate, pharmaceutically acceptable salt or prodrug thereof,

wherein R is³Is composed of

R⁹、R¹⁰Each independently is H, C₁-C₃Alkyl radical, R¹¹、R¹²、R¹³Zhongren is C₄-C₆Alkyl, the remainder are each independently H, C₁-C₃Alkyl, m is an integer of 0 to 2.
4. The compound, isomer, hydrate, solvate, pharmaceutically acceptable salt or prodrug thereof according to claim 2, wherein n is₁Is an integer of 0 to 2, n₂Is an integer of 0 to 1, n₃Is an integer of 0 to 3, R⁴、R⁵Each independently is H, methyl, R⁴、R⁵The same or different;

R⁹、R¹⁰、R¹¹、R¹²、R¹³each independently is H, methyl or ethyl, and m is 0 or 1.
5. The compound, isomer, hydrate, solvate, pharmaceutically acceptable salt or prodrug thereof according to claim 4, wherein R is³Is composed of

R⁹、R¹¹、R¹²、R¹³Each independently is H, methyl or ethyl.
6. The compound, isomer, hydrate, solvate, pharmaceutically acceptable salt or prodrug thereof according to claim 1, wherein n is₄Is an integer of 0 to 3, n₅Is an integer of 0 to 1, n₆Is an integer of 0 to 5, R⁷、R⁸Each independently is H, methyl, R⁷、R⁸The same or different;

R⁶is-H, hydroxyethyl, hydroxypropyl, C₁-C₅Alkyl radical, C₃-C₈Cycloalkyl radical, C₁-C₅Alkoxyethyl radical, C₁-C₅Alkoxypropyl group, C₁-C₅An alkylthioethyl group, a 5-6 membered heterocyclyl group, or-NR 'R ", wherein R', R" are each independently H, C₁-C₃Alkyl group of (1).
7. The compound, isomer, hydrate, solvate, pharmaceutically acceptable salt or prodrug thereof according to claim 6, wherein R⁶Is H, hydroxyethyl, hydroxypropyl, methyl, ethyl, propyl, isopropyl, t-butyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, methoxyethyl, ethoxyethyl, propoxyethyl, isopropoxyethyl, methoxypropyl, ethoxypropyl, propoxypropyl, isopropoxypropyl, methylthioethyl, ethylthioethyl, propylthioethyl, isopropylthioethyl, a 5-6 membered heterocyclic group, or-NR 'R ", wherein R', R" are each independently H, methyl, ethyl;

the 5-6 membered heterocyclic group is unsubstituted or substituted by 1-2 groups selected from hydroxy, C₁-C₃Alkyl radical, C₁-C₃Acyl substituents substituted heterocyclyl containing 1-2 heteroatoms selected from N, O, S.
8. The compound, an isomer, a hydrate, a solvate, a pharmaceutically acceptable salt or a prodrug thereof according to claim 7, wherein the heterocyclic group containing 1 to 2 heteroatoms selected from N, O, S is any selected from:

wherein R is¹⁴H, methyl, ethyl, propyl and isopropyl.
9. The compound of claim 1, isomers, hydrates, solvates, pharmaceutically acceptable salts or prodrugs thereof, wherein the compound of formula (I) is selected from:

4- (butyl-3-en-1-ylamino) -2- ((1- (tert-butyl) -1H-pyrazol-4-yl) amino ] pyrimidine-5-carboxamide;

2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((2-methylallyl) amino) pyrimidine-5-carboxamide;

2- ((1- (tert-butyl) -1H-pyrazol-4-yl) amino) -4- ((3-methylbutyl-2-en-1-yl) amino) pyrimidine-5-carboxamide;

2- ((1- (tert-butyl) -1H-pyrazol-4-yl) amino) -4- (pentyl-4-en-1-ylamino) pyrimidine-5-carboxamide;

4- ((2-methylallyl) amino) -2- ((1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-yl) amino) pyrimidine-5-carboxamide;

4- ((3-methylbutyl-2-en-1-yl) amino) -2- ((1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-yl) amino) pyrimidine-5-carboxamide;

4- (butyl-3-en-1-ylamino) -2- ((1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-yl) amino) pyrimidine-5-carboxamide;

2- ((1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-yl) amino) -4- (pentyl-4-en-1-ylamino) pyrimidine-5-carboxamide.
10. The compound of claim 1, isomers, hydrates, solvates, pharmaceutically acceptable salts or prodrugs thereof, wherein the compound of formula (I) is selected from:

2- ((1- (tert-butyl) -1H-pyrazol-4-yl) amino) -4- ((cyclopentyl-3-en-1-ylmethyl) amino) pyrimidine-5-carboxamide;

4- ((cyclopentyl-3-en-1-ylmethyl) amino) -2- ((1- (tetrahydro-2H-pyran-4-yl) -1H-pyrazol-4-yl) amino) pyrimidine-5-carboxamide;

4- (allylamino) -2- ((1- (tert-butyl) -1H-pyrazol-4-yl) amino) pyrimidine-5-carboxamide;

(Z) -2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((pentan-2-en-1-yl) amino) pyrimidine-5-carboxamide;

(E) -2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((4-methylpentan-2-en-1-yl) amino) pyrimidine-5-carboxamide;

(E) -2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((pentan-3-en-1-yl) amino) pyrimidine-5-carboxamide;

2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((3-methylbutyl-3-en-1-yl) amino) pyrimidine-5-carboxamide;

2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((4-methylpentan-3-en-1-yl) amino) pyrimidine-5-carboxamide;

2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((4-methylpentan-4-en-1-yl) amino) pyrimidine-5-carboxamide;

2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((5-methylhexan-4-en-1-yl) amino) pyrimidine-5-carboxamide;

(E) -2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- ((5-methylhexan-2-en-1-yl) amino) pyrimidine-5-carboxamide;

2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- (((2E,4E) -hexane-2, 4-dien-1-yl) amino) pyrimidine-5-carboxamide;

2- ((1-tert-butyl-1H-pyrazol-4-yl) amino) -4- (((2E,4E) -heptane-2, 4-dien-1-yl) amino) pyrimidine-5-carboxamide.
11. The compound of any , or an isomer, hydrate, solvate, pharmaceutically acceptable salt, or prodrug thereof, wherein the pharmaceutically acceptable salt of the compound is or more selected from hydrochloride, hydrobromide, hydroiodide, perchlorate, sulfate, nitrate, phosphate, formate, acetate, propionate, glycolate, lactate, succinate, maleate, tartrate, malate, citrate, fumarate, gluconate, benzoate, mandelate, methanesulfonate, isethionate, benzenesulfonate, oxalate, palmitate, 2-naphthalenesulfonate, p-toluenesulfonate, cyclamate, salicylate, hexonate, trifluoroacetate, aluminum salt, calcium salt, chloroprocaine salt, choline salt, diethanolamine salt, ethylenediamine salt, lithium salt, magnesium salt, potassium salt, sodium salt, and zinc salt of the compound.
12. Use of the compound of , its isomer, hydrate, solvate, pharmaceutically acceptable salt or prodrug thereof in the manufacture of a medicament for the treatment of autoimmune diseases and cancers associated with tyrosine kinases JAK1, JAK2, JAK3, TYK2, wherein the autoimmune diseases and cancers associated with tyrosine kinases JAK1, JAK2, JAK3, TYK2 include ocular fundus disease, dry eye, psoriasis, vitiligo, dermatitis, alopecia areata, rheumatoid arthritis, colitis, multiple sclerosis, systemic lupus erythematosus, crohn's disease, atheroma, pulmonary fibrosis, hepatic fibrosis, myelofibrosis, non-small cell lung cancer, breast cancer, pancreatic cancer, glioma, glioblastoma, ovarian cancer, cervical cancer, colorectal cancer, melanoma, endometrial cancer, prostate cancer, bladder cancer, leukemia, gastric cancer, liver cancer, gastrointestinal tumor, thyroid cancer, mesenchymal leukemia, acute myelogenous leukemia, non-hodgkin's cell lymphoma, nasopharyngeal carcinoma, lymphoma, brain cell lymphoma, T-cell lymphoma, multiple myeloma, cholangiocarcinoma, multiple myeloma, and cholangiocarcinoma.
A pharmaceutical composition of comprising the compound of any of of claims 1-10, an isomer, hydrate, solvate, pharmaceutically acceptable salt or prodrug thereof, and or more pharmaceutically acceptable carriers or excipients.
14. The pharmaceutical composition of claim 13, wherein the pharmaceutical composition further comprises one or more additional therapeutic agents.