CN112480101A - IRAK4 kinase inhibitor and preparation and application thereof - Google Patents

IRAK4 kinase inhibitor and preparation and application thereof Download PDF

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CN112480101A
CN112480101A CN202010962799.8A CN202010962799A CN112480101A CN 112480101 A CN112480101 A CN 112480101A CN 202010962799 A CN202010962799 A CN 202010962799A CN 112480101 A CN112480101 A CN 112480101A
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段文虎
丁健
陈运
谢华
张惠斌
周金培
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Shanghai Institute of Materia Medica of CAS
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Abstract

The invention provides an IRAK4 kinase inhibitor, and preparation and application thereof, and particularly provides a compound shown in formula (I), a stereoisomer, a geometric isomer, a tautomer, a pharmaceutically acceptable salt, a prodrug, a hydrate or a solvate thereof, and a preparation method and application thereof. Compared with the IRAK4 inhibitor in the prior art, the activity of the compound is obviously improved, so that the compound can be used for preparing related diseases such as cancer, inflammatory diseases and autoimmune diseases mediated by IRAK 4.

Description

IRAK4 kinase inhibitor and preparation and application thereof
Technical Field
The invention relates to the field of pharmaceutical chemistry, in particular to a compound with IRAK4 kinase inhibitory activity, and preparation and application thereof.
Background
Interleukin-1 receptor-associated kinase 4(IRAK4) is a serine/threonine kinase located directly downstream of the interleukin-1 (IL-1) receptor family (IL-1, IL-18 and IL-33 receptors) and Toll-like receptors (TLRs, except TLR3) and is a key signaling node for transduction of innate immunity. Upon ligand binding, IL-1 family receptors and TLRs recruit scaffolds to engage the protein myeloid differentiation primary response gene 88(MyD88) via the conserved Toll/IL receptor (TIR) domain. MyD88 in turn recruits IRAK4 using homotypic interactions with death domains (death domains), thereby activating downstream signaling pathways such as NF-. kappa.B and AP-1. IRAK4 has a dual kinase and backbone role, mediating downstream signaling pathways by forming larger signaling complexes-MyD bodies with MyD88 and IRAK 1. IRAK4 is constitutively activated and IL-1 stimulation does not significantly increase its intrinsic activity on Pellino1 in HEK293 cells expressing IL-1 receptors, human THP1 monocytes stimulated by Pam3CSK4 or primary human macrophages. TLR7 and TLR9 mediated IFN- α/β production was abolished in plasma-like dendritic cells with IRAK4 kinase inactivated. It is clear that the kinase activity of IRAK4 is not only essential for the induction of pro-inflammatory cytokines and chemokines by TLRs (2,4,5,7 and 9), but also for TLR7 and TLR9 mediated IFN- α/β induction. Although IRAK4 occupies an extremely important place in innate immunity, studies have shown that the TIR-MyD88-IRAK4 signaling pathway is essential for protective immunity against a minority of purulent bacteria, but is redundant for host defense against most natural infections.
Abnormal activation of the innate immune system is an important feature of many chronic autoimmune diseases. For example, the anti-acidifying antibody immune complex characteristic of Rheumatoid Arthritis (RA) and the anti-nucleic acid immune complex characteristic of Systemic Lupus Erythematosus (SLE) mediate signaling pathways through TLRs. In addition, activation of TLRs can drive B cells to differentiate into antibody-producing plasma cells, which are used to activate the adaptive immune system. Genetically modified mice with IRAK4 deletion or expressing kinase-inactivated IRAK4 exhibited some degree of impaired immune response, such as TNF α and IL-6 induced when induced by bacterial Lipopolysaccharide (LPS). These mice were also resistant to experimentally induced arthritis, atherosclerosis, and MOG-induced encephalomyelitis. IRAK4 kinase inactivated mice were also shown to be resistant to the development of alzheimer's disease, a process thought to be due to reduced IL-1 production and signaling. Small molecule inhibitors of IRAK4 have been used to inhibit TLR-induced inflammatory signaling in vitro and in vivo. Studies have shown that IRAK4 inhibitors can reduce gout-like inflammation in uric acid-induced peritonitis models, ischemia-induced inflammation in 5/6 nephrectomized rats, and a mouse model of lupus erythematosus. Therefore, IRAK4 is considered to be an important pharmacological target for the treatment of chronic inflammatory diseases.
In activated B-cell-like (ABC) diffuse large B-cell lymphoma (DLBCL), leucine mutation at MyD 88265 to proline (L265P) occurred in 29% of cases, being the most prominent activating mutation in ABC DLBCL, with IRAK4 taking over most of the MyD88 function. The compounds ND-2110 and ND-2158 from the company Nimbus have outstanding antitumor effects in a xenografted OCI-Ly10 mouse model, and simultaneously have good combined administration effects with a BTK inhibitor ibrutinib and a BCL-2 inhibitor ABT-199.
Pancreatic ductal carcinoma (PDAC) overall survival rate of no more than 6% for 5 years. Studies have shown that PDAC patients who are pIRAK4 positive have a higher chance of postoperative recurrence and a poorer prognosis than tumor patients that are p-IRAK4 negative. Through shRNA and CRISPR/Cas9n technologies, researchers found that gemcitabine and 5-fluorouracil were more effective in killing pancreatic ductal carcinoma cells after inhibition of IRAK 4. It was also found that IRAK1/4 can drive the production of inflammatory cytokines and chemokines, resulting in the metastasis, invasion and proliferation of cancer-associated fibroblasts (CAFs). Another study showed that by knocking out mouse IRAK4 was effective in reducing mouse lung tumor numbers and did not alter the cellular composition of bronchoalveolar perfusion (BALF).
Disclosure of Invention
The invention aims to provide a compound shown as a formula (I) with IRAK4 inhibitory activity, a stereoisomer, a geometric isomer, a tautomer, a pharmaceutically acceptable salt, a prodrug, a hydrate or a solvate thereof, and a preparation method and application thereof.
In a first aspect of the present invention, there is provided a compound represented by formula (I), a stereoisomer, a geometric isomer, a tautomer, a pharmaceutically acceptable salt thereof, a prodrug thereof, a hydrate thereof, or a solvate thereof:
Figure BDA0002681115790000021
wherein,
x, Y, Z or W is independently selected from CH or N; and when X, Y, Z or W is CH, the H atom may be substituted with a substituent selected from the group consisting of: halogen, C1-C3 alkyl, C2-C6 acyl, C1-C3 alkoxy, trifluoromethoxy, trifluoroethoxy;
ring a is a 3-8 membered saturated heterocyclic ring (including monocyclic, fused or spiro) containing 1-2 heteroatoms selected from N, O and S, optionally substituted with one or more halogen, oxo, carboxy, cyano, hydroxy, substituted or unsubstituted 5-6 membered heterocyclic ring, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C8 cycloalkyl, said substitution being substituted with one or more halogen, C1-C3 alkyl or hydroxy;
R1is selected from-NR2R3Substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C10 cycloalkyl, substituted or unsubstituted 6-10 membered aryl, substituted or unsubstituted 5-10 membered heteroaryl, substituted or unsubstituted 3-11 membered saturated or partially saturated heterocyclyl, said substituents consisting of RaRepresents;
each R isaEach independently selected from halogen, oxo, - (C0-C3 alkyl) -CN, - (C0-C3 alkyl) -OH, - (C0-C3 alkyl) -COOH, - (C0-C3 alkyl) C (═ O) OR5- (C0-C3 alkyl) C (═ O) R5- (C0-C3 alkyl) NR5R6-C (═ O) (C1-C3 alkyl), -C (═ O) NR (C0-C3 alkyl)5R6- (C0-C3 alkyl) S (═ O) NR5R6- (C0-C3 alkyl) S (═ O)2NR5R6- (C0-C3 alkyl) S (═ O)2(C1-C3 alkyl), - (C0-C3 alkyl) OP (═ O) (OC1-C3 alkyl)2Substituted or unsubstituted C1-C3 alkyl, substituted or unsubstituted C3-C8 cycloalkyl, substituted or unsubstituted C1-C6 alkoxy, substituted or unsubstituted C1-C6 alkylthio, said substitution being with one or more halogens;
R2and R3Each independently selected from the group consisting of a hydrogen atom, a substituted or unsubstituted C1-C3 alkyl group, a substituted or unsubstituted C3-C10 cycloalkyl group, - (C0-C3 alkyl) -6-10 membered aryl group, - (C0-C3 alkyl) -5-10 membered heteroaryl group, - (C0-C3 alkyl) -saturated or partially saturated 4-10 membered heterocyclyl group, said substitution being by halogen, hydroxy, amino, cyano or amido;
or R2And R3May form, together with the nitrogen atom to which they are attached, a 4-to 8-membered heterocyclic ring optionally substituted with one or more RaSubstitution;
R5、R6each independently selected from the group consisting of a hydrogen atom, a substituted or unsubstituted C1-C6 alkyl group, a substituted or unsubstituted C3-C10 cycloalkyl group, and a substituted or unsubstituted 4-6 membered heterocyclic ring, said substitution being by one or more of a C1-C3 alkyl group, a hydroxyl group, a halogen, a carboxylic acid, a C2-C6 carboxylic acid ester.
In another preferred embodiment, R2And R3Together with the nitrogen atom to which they are attached form a 4-7 membered heterocyclic ring, optionally substituted with one or more RbSubstitution;
Rbindependently selected from halogen, oxo, cyano, hydroxy, - (C0-C3 alkyl) C (═ O) NHR5R6- (C0-C3 alkyl) -NR5R6Substituted or unsubstituted C1-C3 alkylThe substitution refers to substitution by one or more of halogen, oxo, cyano, hydroxy.
In another preferred embodiment, R1Is a substituted or unsubstituted 6-10 membered aryl, substituted or unsubstituted 5-10 membered heteroaryl, and said aryl or heteroaryl is optionally substituted with one or more RcSubstitution;
Rcindependently selected from halogen, cyano, hydroxy, -COOH, -C (═ O) OR5、-C(＝O)R5-C (═ O) (C1-C3 alkyl), -C0-C3 alkyl) NR5R6- (C0-C3 alkyl) C (═ O) NR5R6- (C0-C3 alkyl) S (═ O) NR5R6- (C0-C3 alkyl) S (═ O)2NR5R6、-S(＝O)2(C1-C3 alkyl), - (C0-C3 alkyl) OP (═ O) (OC1-C3 alkyl)2Substituted or unsubstituted C1-C3 alkyl, substituted or unsubstituted C1-C6 alkoxy, substituted or unsubstituted C1-C6 alkylthio, said substitution being with one or more halogens or hydroxy groups.
In another preferred embodiment, R1Is a substituted or unsubstituted 5-6 membered aromatic heterocycle optionally substituted with one or more RcAnd (4) substitution.
In another preferred embodiment, the ring a is a substituted or unsubstituted group selected from the group consisting of:
Figure BDA0002681115790000031
wherein each p is independently 1, 2 or 3.
In another preferred embodiment, R is1Selected from the group consisting of: -NR2R3Substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted aryl, substituted or unsubstituted 5-to 10-membered heteroaryl, substituted or unsubstituted 3-to 6-membered saturated or partially saturated heterocyclyl, substituted or unsubstituted (3-to 6-membered saturated or partially saturated heterocyclyl) and 5-to 6-membered heteroaryl.
In another preferred embodiment, ring a is selected from the group consisting of:
Figure BDA0002681115790000032
R4selected from the group consisting of a hydrogen atom, a substituted or unsubstituted C1-C6 alkyl group, a substituted or unsubstituted C3-C10 cycloalkyl group, said substitution being by one or more halogens.
In a further preferred embodiment of the method,
Figure BDA0002681115790000041
selected from the group consisting of:
Figure BDA0002681115790000042
in another preferred embodiment, the compound is selected from the compounds shown in table 1.
In a second aspect of the present invention, there is provided a pharmaceutical composition comprising the following components:
1) a therapeutically effective amount of one or more compounds according to the first aspect of the present invention, stereoisomers, geometric isomers, tautomers thereof, pharmaceutically acceptable salts thereof, prodrugs thereof, hydrates thereof or solvates thereof; and
2) a pharmaceutically acceptable carrier or excipient.
In another preferred embodiment, the pharmaceutical composition further comprises one or more active substances selected from the group consisting of: immunosuppressants, glucocorticoids, non-steroidal anti-inflammatory drugs, vinca alkaloids, paclitaxel, DNA damaging agents, Bcl-2 inhibitors, BTK inhibitors, JAK inhibitors, Hsp90 inhibitors, ALK inhibitors, FLT3 inhibitors, PI3K inhibitors, and SYK inhibitors.
In a third aspect of the present invention, there is provided a use of a compound according to the first aspect of the present invention, a stereoisomer, a geometric isomer, a tautomer, a pharmaceutically acceptable salt thereof, a prodrug thereof, a hydrate thereof or a solvate thereof, or a pharmaceutical composition according to the second aspect of the present invention, for a use selected from the group consisting of:
1) for the preparation of a medicament for the prevention and/or treatment of IRAK 4-mediated diseases;
2) preparing a medicament for inhibiting IRAK 4.
In another preferred embodiment, the medicament is further for inhibiting a protein kinase selected from the group consisting of: FLT3, RET, VEGFR, ErbB2, or a combination thereof.
In another preferred embodiment, said IRAK4 mediated disease is selected from the group consisting of: cancer, autoimmune diseases, inflammatory diseases and thromboembolic diseases.
In another preferred embodiment, wherein the cancer is selected from the group consisting of: diffuse large B-cell lymphoma, multiple myeloma, mantle cell lymphoma, Waldenstrom's macroglobulinemia, acute myelogenous leukemia, chronic lymphocytic leukemia, small lymphocytic lymphoma, pancreatic ductal carcinoma.
In another preferred embodiment, wherein the autoimmune and inflammatory diseases are selected from the group consisting of: rheumatoid arthritis, osteoarthritis, juvenile arthritis, chronic obstructive pulmonary disease, multiple sclerosis, systemic lupus erythematosus, psoriasis, psoriatic arthritis, crohn's disease, ulcerative colitis, and irritable bowel syndrome.
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 inventor provides an IRAK4 inhibitor compound with a novel structure through long-term and intensive research, and the compound has obviously improved activity compared with an IRAK4 inhibitor in the prior art, so that the compound can be used for preparing related diseases such as cancer, inflammatory diseases and autoimmune diseases mediated by IRAK 4. Based on the above findings, the inventors have completed the present invention.
Term(s) for
As used herein, the term "heterocyclyl" is a cyclic group having 1, 2,3, 4, or 5 heteroatoms selected from the group consisting of: o, N or S.
In this context, the alkyl group is preferably an aliphatic alkyl group and may be a straight-chain alkyl group, a branched-chain alkyl group, a spiro-cycloalkyl group, a bridged cycloalkyl group, an alkenylalkyl group, an alkynylalkyl group, a cycloalkyl group, a cycloalkenyl group, a cycloalkynyl group, an alkoxyalkyl group, an alkoxyacylalkyl group, a cycloalkylalkyl group, including without limitation: methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, cyclopropane, cyclobutane, cyclopentane, cyclohexane, allyl, propargyl, cyclobutenyl, cyclohexenyl; expressions in the form of "C1-C8" are intended to include the corresponding group having 1, 2,3, 4,5, 6, 7, or 8 carbon atoms, e.g., "C1-C8 alkyl" refers to alkyl having 1, 2,3, 4,5, 6, 7, or 8 carbon atoms, and "C2-C10 alkenyl" refers to alkenyl having 2,3, 4,5, 6, 7, 8, 9, or 10 carbon atoms. In particular, the term "C0 alkyl" as used in the claims of the present invention refers to the case where the groups are chemical bonds.
In this context, the alkenyl group is preferably an ethenyl group, a propenyl group, a butenyl group, a styryl group, a phenylpropenyl group, or the like.
In this context, the cycloalkyl group may be a saturated or partially unsaturated monocyclic or polycyclic cyclic hydrocarbon substituent comprising 3 to 10 carbon atoms, preferably 3 to 8 carbon atoms, and more preferably the cycloalkyl group comprises 3 to 6 carbon atoms. Non-limiting examples of monocyclic cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentenyl, cyclohexyl, cyclooctyl, and the like; polycyclic cycloalkyl groups include spiro, fused and bridged cycloalkyl groups.
The heterocyclic group means a saturated or partially saturated monocyclic or polycyclic cyclic substituent including a 4 to 10-membered heterocyclic group, and the heterocyclic group is a saturated or unsaturated monocyclic, fused, spiro, fused, bridged ring or the like containing one or more hetero atoms (nitrogen, oxygen, sulfur) therein. The heterocyclic group described herein includes, but is not limited to, groups selected from the group consisting of: morpholine rings, piperidine rings, piperazine rings, N-alkyl or acyl substituted piperazine rings, homopiperazine rings, N-alkyl or acyl substituted homopiperazine rings, pyrrole, tetrahydropyrrole, 7H-purine and the like.
The aryl group refers to a 6 to 10 membered all carbon monocyclic or fused polycyclic (i.e., rings which share adjacent pairs of carbon atoms) group, and the group has a conjugated pi-electron system, such as phenyl and naphthyl. The aryl ring may be fused to a heterocyclyl, heteroaryl or cycloalkyl ring, non-limiting examples of which include benzimidazole, benzothiazole, benzoxazole, benzisoxazole, benzopyrazole, quinoline, benzindole, chroman.
The heteroaryl group refers to a heteroaromatic system containing 1 to 4 heteroatoms, 5 to 10 ring atoms, wherein the heteroatoms include oxygen, sulfur, and nitrogen. Heteroaryl is preferably 5-or 6-membered, for example furyl, thienyl, pyridyl, pyrrolyl, N-alkylpyrrolyl, pyrimidinyl, pyrazinyl, imidazolyl, tetrazolyl and the like. The heteroaryl group can be fused to an aryl, heterocyclyl, or cycloalkyl ring, wherein the ring to which the parent structure is attached is a heteroaryl ring.
Unless otherwise specified, the structural formulae depicted herein are intended to include all tautomeric, enantiomeric and stereoisomeric forms (e.g., enantiomers, diastereomers, geometric isomers or conformational isomers): for example, the R, S configuration containing an asymmetric center, the (Z), (E) isomers and the conformational isomers of (Z), (E) of the double bond. Thus, individual stereochemical isomers, tautomers or enantiomers, diastereomers or geometric isomers or conformational isomers or mixtures of tautomers of the compounds of the present invention are within the scope of the present invention.
The term "tautomer" means that structural isomers having different energies may exceed the low energy barrier and thus be converted to each other. For example, proton tautomers (i.e., proton shift) include interconversion by proton shift, such as 1H-indazole and 2H-indazole, 1H-benzo [ d ] imidazole and 3H-benzo [ d ] imidazole, and valence tautomers include interconversion by some bond-forming electron recombination.
Herein, the pharmaceutically acceptable salt is not particularly limited, and preferably includes: inorganic acid salts, organic acid salts, alkylsulfonic acid salts and arylsulfonic acid salts; the inorganic acid salt comprises hydrochloride, hydrobromide, nitrate, sulfate, phosphate and the like; the organic acid salt comprises formate, acetate, propionate, benzoate, maleate, fumarate, succinate, tartrate, citrate and the like; the alkyl sulfonate includes methyl sulfonate, ethyl sulfonate and the like; the aryl sulfonate includes benzene sulfonate, p-toluene sulfonate and the like.
Herein, the pharmaceutically acceptable solvate of the compound represented by the general formula (I) is not particularly limited, and preferably includes: solvates of the compounds represented by the general formula (I) with water, ethanol, isopropanol, ether, acetone, etc.
Compound (I)
The invention provides a compound shown as a formula (I), a stereoisomer, a geometric isomer, a tautomer, a pharmaceutically acceptable salt, a prodrug, a hydrate or a solvate thereof,
Figure BDA0002681115790000061
wherein each group is as defined above.
In another preferred embodiment, among said compounds, X, Y, Z, W, ring A, R1Each of which is a group corresponding to a specific compound described in table 1.
In another preferred embodiment, the compound is preferably the compound prepared in the examples.
In another preferred embodiment, the compound is selected from the compounds listed in table 1.
TABLE 1
Figure BDA0002681115790000062
Figure BDA0002681115790000071
Figure BDA0002681115790000081
Figure BDA0002681115790000091
Figure BDA0002681115790000101
Figure BDA0002681115790000111
Figure BDA0002681115790000121
Figure BDA0002681115790000131
Salt form
As used herein, the term "pharmaceutically acceptable salt" refers to a salt of a compound of the present invention with an acid or base that is suitable for use as a pharmaceutical. Pharmaceutically acceptable salts include inorganic and organic salts. One preferred class of salts is that formed by reacting a compound of the present invention with an acid. Suitable acids for forming the salts include, but are not limited to: inorganic acids such as hydrochloric acid, hydrobromic acid, hydrofluoric acid, sulfuric acid, nitric acid, phosphoric acid, and the like; organic acids such as formic acid, acetic acid, trifluoroacetic acid, propionic acid, oxalic acid, malonic acid, succinic acid, fumaric acid, maleic acid, lactic acid, malic acid, tartaric acid, citric acid, picric acid, benzoic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, benzenesulfonic acid, naphthalenesulfonic acid and the like; and amino acids such as proline, phenylalanine, aspartic acid, glutamic acid, etc.
Another preferred class of salts are those of the compounds of the invention with bases, for example alkali metal salts (e.g., sodium or potassium salts), alkaline earth metal salts (e.g., magnesium or calcium salts), ammonium salts (e.g., lower alkanolammonium salts and other pharmaceutically acceptable amine salts), for example methylamine salts, ethylamine salts, propylamine salts, dimethylamine salts, trimethylamine salts, diethylamine salts, triethylamine salts, tert-butylamine salts, ethylenediamine salts, hydroxyethylamine salts, dihydroxyethylamine salts, triethanolamine salts, and amine salts formed from morpholine, piperazine, lysine, respectively.
The term "solvate" refers to a complex of a compound of the present invention coordinated to solvent molecules in a specific ratio. "hydrate" refers to a complex formed by the coordination of a compound of the present invention with water.
The term "prodrug" includes a class of compounds which are biologically active or inactive in nature and which undergo metabolic or chemical reactions in the body to become transformed, or a salt or solution of the compound, when administered by an appropriate method. The prodrugs include, but are not limited to, carboxylate, carbonate, phosphate, nitrate, sulfate, sulfone, sulfoxide, amide, carbamate, azo, phosphoramide, glucoside, ether, acetal forms of the compounds.
Preparation of the Compounds of the invention
The following reaction schemes illustrate the preparation of the compounds of the present invention.
Reaction scheme I
Figure BDA0002681115790000141
The general synthetic method of the compound is shown as a reaction scheme I. The method comprises the following steps:
1. the compound 1a/1b and methyl magnesium bromide are subjected to Grignard reaction at-78 ℃ to obtain a compound 2a/2 b;
2. performing a ring closure reaction on the compound 2a/2b under the condition of potassium tert-butoxide to obtain a compound 3a/3 b;
3. carrying out nitration reaction on the compound 3a/3b under the action of nitric acid to obtain a compound 4a/4b
4. The compound 4a is subjected to Suzuki coupling reaction under the action of a palladium catalyst, and the compound 4b is subjected to substitution reaction under the action of alkali, so that a different form of a formula 5 can be obtained;
5. reducing the compound 5 under palladium carbon/hydrogen or reducing the compound under the condition of iron powder/ammonium chloride to obtain a compound 6;
6. the compound 6 reacts under the conditions of 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride and 1-hydroxybenzotriazole condensing agent to obtain a target compound 7.
Reaction scheme II
Figure BDA0002681115790000142
The general synthetic method of the compound is shown as a reaction scheme II. The method comprises the following steps:
1. the compound II-1 is debrominated under the action of n-butyllithium and reacts with dimethylcyclopropane to obtain a compound II-2;
2. the compound II-2 is subjected to ring closure under the action of potassium tert-butoxide to obtain a compound II-3;
3. carrying out nitration reaction on the compound II-3 under the action of acetic anhydride/copper nitrate to obtain a compound II-4;
4. carrying out substitution reaction on the compound II-4 under the action of organic base to obtain a compound II-5;
5. reducing the compound II-5 under palladium carbon/hydrogen to obtain a compound II-6;
6. and reacting the compound II-6 under the conditions of 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride and 1-hydroxybenzotriazole condensing agent to obtain a target compound II-7.
Pharmaceutical compositions containing compounds of formula (I)
The invention also relates to a pharmaceutical composition, which comprises one or more compounds selected from the compounds shown in the formula (I), pharmaceutically acceptable salts thereof, prodrugs thereof, hydrates thereof and solvates thereof in a therapeutically effective amount and optionally a pharmaceutically acceptable carrier, and can be used for treating diseases related to the activity or expression amount of IRAK 4. The pharmaceutical composition may be prepared in various forms according to different administration routes.
Because the compound has excellent inhibitory activity on IRAK4, the compound and various crystal forms, pharmaceutically acceptable inorganic or organic salts, hydrates or solvates thereof, and a pharmaceutical composition containing the compound as a main active ingredient can be used for treating, preventing and relieving diseases related to the activity or expression amount of IRAK4, such as diseases related to the abnormal expression of IRAK4 signal pathway. According to the prior art, the compounds of the invention are useful for the treatment of the following diseases: cancer, autoimmune diseases, inflammatory diseases and thromboembolic diseases, preferably, the compounds can be used for the treatment of the following diseases: diffuse large B-cell lymphoma, multiple myeloma, mantle cell lymphoma, fahrenheit macroglobulinemia, acute myelogenous leukemia, chronic lymphocytic leukemia, small lymphocytic lymphoma, pancreatic ductal carcinoma, rheumatoid arthritis, osteoarthritis, juvenile arthritis, chronic obstructive pulmonary disease, multiple sclerosis, systemic lupus erythematosus, psoriasis, psoriatic arthritis, crohn's disease, ulcerative colitis, and irritable bowel syndrome.
The pharmaceutical composition of the present invention comprises the compound of the present invention or a pharmacologically acceptable salt thereof in a safe and effective amount range and a pharmacologically 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, 5-200mg of a compound of the invention per dose. Preferably, said "dose" is a capsule or tablet.
"pharmaceutically acceptable carrier" refers to: one or more compatible solid or liquid fillers or gel substances 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)Oil, etc.), polyol (such as propylene glycol, glycerin, mannitol, sorbitol, etc.), emulsifier (such as
Figure BDA0002681115790000151
) 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, intratumoral, rectal, parenteral (intravenous, intramuscular or subcutaneous) and topical administration.
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.
Dosage forms for topical administration of the compounds of the present invention include ointments, powders, patches, sprays, and inhalants. The active ingredient is mixed under sterile conditions with a physiologically acceptable carrier and any preservatives, buffers, or propellants which may be required if necessary.
The compounds of the present invention may be administered alone or in combination with other pharmaceutically acceptable compounds.
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 5 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 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 by weight.
Intermediate 1
6-fluoro-2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran (prepared according to the method reported in WO 2017/108723)
Figure BDA0002681115790000161
Step 1: preparation of methyl 2- (2, 4-difluorophenyl) acetate
Figure BDA0002681115790000162
Dissolving 2- (2, 4-difluorophenyl) acetic acid (5g, 29.0mmol) in 100mL of methanol, adding 5mL of concentrated sulfuric acid, refluxing for reaction overnight, concentrating under reduced pressure, dissolving the residue in ethyl acetate, washing with water, washing with saturated sodium bicarbonate solution, washing with saturated salt water, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure to dryness to obtain 5g of light yellow oily substance, and directly putting into the next reaction.
Step 2: 1- (2, 4-difluorophenyl) -2-methylpropan-2-ol
Figure BDA0002681115790000171
Dissolving methyl 2- (2, 4-difluorophenyl) acetate (5g, 26.8mmol) in anhydrous tetrahydrofuran (60mL), replacing with argon for protection, dropwise adding methyl magnesium bromide solution (27mL of 3M diethyl ether solution, 80.5mmol) at-78 ℃, reacting at room temperature for 1 hour after 10 minutes, and detecting by TLCAnd finishing. The mixture was quenched by dropwise addition of a saturated ammonium chloride solution in an ice bath, extracted with ethyl acetate (150mL) and water, and the ethyl acetate layer was washed with saturated brine and dried over anhydrous sodium sulfate. Vacuum concentrating to obtain light yellow oily matter, and directly adding into the next reaction.1H NMR(300MHz,Chloroform-d)δ7.24–7.15(m,1H),6.90–6.73(m,2H),2.77(d,J＝1.6Hz,2H),1.23(s,6H).
And step 3: preparation of 6-fluoro-2, 2-dimethyl-2, 3-dihydrobenzofuran
Figure BDA0002681115790000172
Dissolving 1- (2, 4-difluorophenyl) -2-methylpropan-2-ol (5.41g, 29.0mmol) in anhydrous tetrahydrofuran (120mL), adding potassium tert-butoxide (8.15g, 72.6mmol), refluxing for 2 hours, detecting by TLC after the reaction is finished, concentrating under reduced pressure, adding ethyl acetate (150mL) and water for extraction, washing an ethyl acetate layer with saturated common salt water, drying with anhydrous sodium sulfate, filtering, and performing silica gel column chromatography (ethyl acetate/petroleum ether ═ 1% elution) to obtain 2.62g of colorless oily substance. The yield of the three-step reaction is as follows: 49 percent.1H NMR(400MHz,Chloroform-d)δ7.02(ddt,J＝8.1,5.8,1.1Hz,1H),6.57–6.37(m,2H),2.95(t,J＝1.4Hz,2H),1.47(s,6H).
And 4, step 4: preparation of 6-fluoro-2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran
Figure BDA0002681115790000173
Dissolving 6-fluoro-2, 2-dimethyl-2, 3-dihydrobenzofuran (2.62g, 15.7mmol) in 50mL dichloromethane, dropwise adding nitric acid (2.4mL) in ice bath, reacting at room temperature until TLC detection reaction is finished, extracting with ethyl acetate (150mL) and water, washing an ethyl acetate layer with saturated sodium bicarbonate solution, washing with saturated common salt water, drying with anhydrous sodium sulfate, filtering, and performing silica gel column chromatography (gradient elution with ethyl acetate/petroleum ether being 3% -10%) to obtain 2.29g of orange-yellow solid. Yield: and 69 percent.1H NMR(300MHz,DMSO-d6)δ8.04(dt,J＝8.2,1.3Hz,1H),6.97(d,J＝12.5Hz,1H),3.06(d,J＝1.4Hz,2H),1.45(s,6H).
Intermediate 2
6-chloro-2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran (prepared according to the method reported in WO 2017/108723)
Figure BDA0002681115790000174
Step 1: preparation of methyl 2- (4-chloro-2-fluorophenyl) acetate
Figure BDA0002681115790000175
Dissolving 2- (4-chloro-2-fluorophenyl) acetic acid (5g, 26.5mmol) in 50mL of methanol, adding 3mL of concentrated sulfuric acid, refluxing for overnight reaction, concentrating under reduced pressure, dissolving the residue in ethyl acetate, washing with water, washing with saturated sodium bicarbonate solution, washing with saturated common salt water, drying with anhydrous sodium sulfate, filtering, and concentrating under reduced pressure to dryness to obtain 5g of light yellow oily substance.1H NMR(400MHz,Chloroform-d)δ7.20(t,J＝8.2Hz,1H),7.10(dt,J＝9.8,2.3Hz,2H),3.71(s,3H),3.64(d,J＝1.4Hz,2H).
Step 2: preparation of 1- (4-chloro-2-fluorophenyl) -2-methylpropan-2-ol
Figure BDA0002681115790000181
Methyl 2- (4-chloro-2-fluorophenyl) acetate (5.37g, 26.5mmol) was dissolved in anhydrous tetrahydrofuran (60mL) and under argon replacement protection, methyl magnesium bromide solution (26.5mL 3M diethyl ether solution) was added dropwise at-78 ℃ and after dropping, the reaction was allowed to reach room temperature after 15 minutes for 1 hour and TLC detection was complete. The mixture was quenched by dropwise addition of a saturated ammonium chloride solution in an ice bath, extracted with ethyl acetate (200mL) and water, and the ethyl acetate layer was washed with saturated brine and dried over anhydrous sodium sulfate. Vacuum concentrating to dry to obtain light yellow oil.1H NMR(400MHz,Chloroform-d)δ7.20(dd,J＝9.2,7.3Hz,1H),7.13–7.05(m,2H),2.78(d,J＝1.6Hz,2H),1.24(d,J＝1.0Hz,6H).
And step 3: preparation of 6-chloro-2, 2-dimethyl-2, 3-dihydrobenzofuran
Figure BDA0002681115790000182
Dissolving 1- (4-chloro-2-fluorophenyl) -2-methylpropan-2-ol (5.37g, 26.5mmol) in anhydrous tetrahydrofuran (125mL), adding potassium tert-butoxide (7.43g, 66.2mmol), refluxing overnight, detecting by TLC after reaction, concentrating under reduced pressure, adding ethyl acetate (150mL) and water for extraction, washing the ethyl acetate layer with saturated saline, drying over anhydrous sodium sulfate, filtering, and performing silica gel column chromatography (ethyl acetate/petroleum ether ═ 1% elution) to obtain 3.57g of colorless oily substance. The yield of the three-step reaction is as follows: 74 percent.1H NMR(400MHz,Chloroform-d)δ7.05(dt,J＝7.9,1.1Hz,1H),6.81(dd,J＝7.9,1.9Hz,1H),6.74(d,J＝1.8Hz,1H),2.98(s,2H),1.49(s,6H).
And 4, step 4: preparation of 6-chloro-2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran
Figure BDA0002681115790000183
Dissolving 6-fluoro-2, 2-dimethyl-2, 3-dihydrobenzofuran (3.57g, 19.5mmol) in 40mL dichloromethane, dropwise adding nitric acid (2.4mL) at room temperature, reacting at room temperature until TLC detection reaction is finished, extracting with ethyl acetate (150mL) and water, washing an ethyl acetate layer with a saturated sodium bicarbonate solution, washing with a saturated common salt solution, drying with anhydrous sodium sulfate, filtering, performing silica gel column chromatography (gradient elution with ethyl acetate/petroleum ether of 1-10%) to obtain 4.45g of an orange solid, and recrystallizing with petroleum ether to obtain 2.02g of a white needle-shaped solid. Yield: 45 percent.1H NMR(400MHz,Chloroform-d)δ7.84(s,1H),6.82(s,1H),3.04(d,J＝1.3Hz,2H),1.52(s,6H).
Intermediate 3
2- (2- ((tert-butoxycarbonyl) amino) pyridin-4-yl) oxazole-4-carboxylic acid
Figure BDA0002681115790000184
Step 1: preparation of tert-butyl (4-bromopyridin-2-yl) carbamate
Figure BDA0002681115790000191
2-amino-4-bromopyridine (0.4g, 2.31mmol), di-tert-butyl dicarbonate (605mg, 2.77mmol) and 4-dimethylaminopyridine (423mg, 3.47mmol) were dissolved in dichloromethane (12mL) and reacted overnight at room temperature, after completion of the TLC detection reaction,
silica gel column chromatography (gradient elution with ethyl acetate/petroleum ether of 3% -20%) is carried out to obtain 354mg of white solid. Yield: 56 percent.1H NMR(300MHz,Chloroform-d)δ8.22(d,J＝1.8Hz,1H),8.10–8.04(m,1H),7.97(s,1H),7.11(dd,J＝5.4,1.7Hz,1H),1.53(s,9H).
Step 2: preparation of tert-butyl (4- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-2-yl) carbamate
Figure BDA0002681115790000192
Tert-butyl (4-bromopyridin-2-yl) carbamate (0.2g, 732. mu. mol), pinacol diboron (223mg, 878. mu. mol) and potassium acetate (215mg, 2.20mmol) were dissolved in tetrahydrofuran (12mL) and protected by argon displacement, and [1,1' -bis (diphenylphosphino) ferrocene ] dichloropalladium (21mg, 29.2. mu. mol) was added thereto for reflux reaction overnight, after the TLC detection reaction was completed, followed by suction filtration with celite and concentration, and then directly fed to the next reaction.
And step 3: preparation of ethyl 2- (2- ((tert-butoxycarbonyl) amino) pyridin-4-yl) oxazole-4-carboxylate
Figure BDA0002681115790000193
Ethyl 2-chlorooxazole-4-carboxylate (110mg, 626. mu. mol), (tert-butyl 4- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) pyridin-2-yl) carbamate (240mg, 751. mu. mol), sodium carbonate (199mg, 1.88. mu. mol) and [1,1' -bis (diphenylphosphino) ferrocene]Palladium dichloride (23mg, 31. mu. mol)) Ethylene glycol dimethyl ether/water (10mL/1mL) was added to the reaction flask, replaced with argon several times, and reacted at 90 ℃ overnight. TLC detection reaction is finished, and the reaction is filtered by suction through kieselguhr, extracted by ethyl acetate (30mL) and water, washed by saturated salt water and dried by anhydrous sodium sulfate. Filtration and silica gel column chromatography (ethyl acetate/petroleum ether: 1:5 elution) gave 121mg of a white solid. Yield: 58 percent.1H NMR(300MHz,DMSO-d6)δ10.15(s,1H),9.06(d,J＝1.2Hz,1H),8.49–8.40(m,2H),7.56(dd,J＝5.2,1.5Hz,1H),4.34(q,J＝7.0Hz,2H),1.50(d,J＝1.1Hz,9H),1.32(t,J＝7.1Hz,3H).
And 4, step 4: preparation of 2- (2- ((tert-butoxycarbonyl) amino) pyridin-4-yl) oxazole-4-carboxylic acid
Figure BDA0002681115790000201
Ethyl 2- (2- ((tert-butoxycarbonyl) amino) pyridin-4-yl) oxazole-4-carboxylate (118mg, 356. mu. mol) was dissolved in tetrahydrofuran/methanol/water (4mL/2mL/1mL), lithium hydroxide monohydrate (45mg, 1.07mmol) was added to the solution, the reaction was carried out at room temperature for 20 minutes, after completion of the TLC detection reaction, the mixture was concentrated, the pH was adjusted to weak acidity with 1N hydrochloric acid solution, and a solid was precipitated, which was filtered off with suction and dried to obtain 83mg of a white solid. Yield: 77 percent.1H NMR(300MHz,DMSO-d6)δ8.93(s,1H),8.49–8.39(m,2H),7.54(dd,J＝5.2,1.5Hz,1H),1.50(s,9H).
Example 1
2- (2-Aminopyridin-4-yl) -N- (6- (4- (hydroxymethyl) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-1)
Figure BDA0002681115790000202
Step 1: preparation of (1- (2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran-6-yl) piperidin-4-yl) methanol
Figure BDA0002681115790000203
6-fluoro-2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran (200mg, 94.7 mu mol) and 4-hydroxymethylpiperidine (218mg, 1.89mmol) are dissolved in N, N-dimethylformamide (5mL), potassium carbonate (262mg, 1.89mmol) is added, reaction is carried out for two hours at 40 ℃, TLC detection reaction is finished, ethyl acetate (25mL) and water are added for extraction, an ethyl acetate layer is washed with water, saturated salt solution is washed with water, anhydrous sodium sulfate is dried, suction filtration is carried out, concentration is carried out till dryness, and the mixture is directly put into the next reaction.
Step 2: preparation of (1- (5-amino-2, 2-dimethyl-2, 3-dihydrobenzofuran-6-yl) piperidin-4-yl) methanol
Figure BDA0002681115790000204
Dissolving the reaction product in methanol (30mL), adding 10% palladium carbon (42mg), reacting at room temperature under hydrogen atmosphere for 6h, detecting by TLC, filtering with diatomaceous earth, concentrating, and performing silica gel column chromatography (gradient elution with dichloromethane/methanol of 1:100 to 1: 50) to obtain 208mg of yellow-white solid. The two-step yield: 79 percent.1H NMR(300MHz,DMSO-d6)δ6.49(s,1H),6.30(s,1H),4.46(t,J＝5.3Hz,1H),4.15(s,2H),3.27(t,J＝5.8Hz,2H),2.96(d,J＝11.3Hz,2H),2.80(s,2H),2.39(t,J＝11.2Hz,2H),1.70(d,J＝12.3Hz,2H),1.47-1.35(m，1H)1.31(s,6H),1.22(td,J＝12.4,11.1,3.3Hz,2H).
And step 3: preparation of tert-butyl (4- (4- ((6- (4- (hydroxymethyl) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) carbamoyl) oxazol-2-yl) pyridin-2-yl) carbamate
Figure BDA0002681115790000211
2- (2- ((tert-butoxycarbonyl) amino) pyridin-4-yl) oxazole-4-carboxylic acid (80mg, 263. mu. mol), 1- (5-amino-2, 2-dimethyl-2, 3-dihydrobenzofuran-6-yl) piperidin-4-yl) methanol (69mg, 251. mu. mol), 1-ethyl- (3-dimethylaminopropyl) carbonyl diimine hydrochloride (58mg, 301. mu. mol) and 1-hydroxybenzotriazole (41mg, 301. mu. mol) were dissolved in N, N-dimethylformamide (5mL), and N, N-diisopropyl was addedPhenylethylamine (104 μ L, 627 μmol) was reacted at room temperature for 4 hours, after completion of the TLC detection reaction, ethyl acetate (25mL) and water were added for extraction, the ethyl acetate layer was washed with water, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and subjected to silica gel column chromatography (dichloromethane/methanol ═ 2% elution) to obtain 80mg of a pale yellow solid. Yield: 56 percent.1H NMR(300MHz,DMSO-d6)δ10.19(s,1H),9.98(s,1H),8.99(s,1H),8.49(s,1H),8.45(d,J＝5.2Hz,1H),8.18(s,1H),7.67–7.59(m,1H),6.69(s,1H),4.53(t,J＝5.1Hz,1H),2.96(d,J＝27.2Hz,4H),2.65(t,J＝10.3Hz,2H),1.80(d,J＝10.0Hz,2H),1.49(s,9H),1.42-1.35(m,6H).
Step 4: preparation of 2- (2-aminopyridin-4-yl) -N- (6- (4- (hydroxymethyl) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide
Figure BDA0002681115790000212
Tert-butyl (4- (4- ((6- (4- (hydroxymethyl) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) carbamoyl) oxazol-2-yl) pyridin-2-yl) carbamate (35mg, 62. mu. mol) was dissolved in dichloromethane (4.5mL), trifluoroacetic acid (1.5mL) was added and the reaction was allowed to react for two hours at room temperature, TLC was performed to detect completion of the reaction, and the mixture was concentrated, extracted with saturated sodium bicarbonate and dichloromethane, dried over anhydrous sodium sulfate in the dichloromethane layer, filtered, concentrated, and purified by preparative thin layer chromatography (dichloromethane/methanol ═ 16:1 as a developing reagent) to give 26mg of a yellow solid.1H NMR(300MHz,DMSO-d6)δ9.97(s,1H),8.92(s,1H),8.22(s,1H),8.09(d,J＝5.3Hz,1H),7.16–6.98(m,2H),6.69(s,1H),6.39(s,2H),4.73(t,J＝5.3Hz,1H),3.43(s,2H),2.96(d,J＝23.8Hz,4H),2.67(t,J＝11.0Hz,2H),1.80(d,J＝11.4Hz,2H),1.73–1.52(m,3H),1.40(s,6H).
Example 2
(R) -N- (6- (3-aminopiperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) -2- (2-aminopyridin-4-yl) oxazole-4-carboxamide trifluoroacetate (I-2)
Figure BDA0002681115790000221
Step 1: preparation of R) - (tert-butyl 4- (4- ((6- (3- ((tert-butoxycarbonyl) amino) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl-tert-butyl) carbamoyl) oxazol-2-yl) pyridin-2-yl) carbamate
Figure BDA0002681115790000222
Tert-butyl (R) - (4- (4- ((6- (3- ((tert-butoxycarbonyl) amino) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl tert-butyl) carbamoyl) oxazol-2-yl) pyridin-2-yl) carbamate can be prepared by a similar method as in step 4 of example 1.1H NMR(400MHz,Chloroform-d)δ10.15(s,1H),8.65(s,1H),8.41(d,J＝5.2Hz,1H),8.37(s,1H),8.31(s,1H),7.99(s,1H),7.63(dd,J＝5.2,1.5Hz,1H),6.60(s,1H),3.14(s,1H),3.02(s,2H),2.77(s,3H),1.90(s,1H),1.67(s,3H),1.55(s,9H),1.47(s,6H),1.19(s,9H).
Step 2: preparation of (R) -N- (6- (3-aminopiperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) -2- (2-aminopyridin-4-yl) oxazole-4-carboxamide trifluoroacetate
(R) - (4- (4- ((6- (3- ((tert-butoxycarbonyl) amino) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl-tert-butyl) carbamoyl) oxazol-2-yl) pyridin-2-yl) carbamic acid tert-butyl ester was dissolved in dichloromethane (4mL), trifluoroacetic acid (1mL) was added thereto and reacted for two hours at room temperature, and after completion of the reaction by TLC, the mixture was concentrated to dryness under reduced pressure, slurried with methanol/ether and suction-filtered to give 34mg of a pale yellow solid.1H NMR(400MHz,DMSO-d6)δ9.69(s,1H),9.07(s,1H),8.15(d,J＝6.1Hz,1H),8.06(s,4H),7.75(s,2H),7.33(s,1H),7.19(dd,J＝6.1,1.6Hz,1H),6.67(s,1H),3.46–3.33(m,2H),3.13(d,J＝10.8Hz,1H),3.01(s,2H),2.85(d,J＝11.9Hz,1H),2.77–2.61(m,2H),2.08(s,1H),1.91(d,J＝13.3Hz,1H),1.80(d,J＝11.3Hz,1H),1.50(d,J＝8.6Hz,1H),1.42(d,J＝4.1Hz,6H).
Example 3
N- (6- (4- (aminomethyl) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) -2- (2-aminopyridin-4-yl) oxazole-4-carboxamide trifluoroacetate (I-3)
Figure BDA0002681115790000231
The trifluoroacetate salt of N- (6- (4- (aminomethyl) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) -2- (2-aminopyridin-4-yl) oxazole-4-carboxamide may be prepared by a similar method as in example 2.1H NMR(400MHz,DMSO-d6)δ9.88(s,1H),9.05(s,1H),8.21–8.09(m,2H),7.86(s,5H),7.32(s,1H),7.18(dd,J＝6.1,1.6Hz,1H),6.74(s,1H),2.98(d,J＝24.2Hz,4H),2.81(t,J＝6.4Hz,2H),2.68(t,J＝11.5Hz,2H),1.91(d,J＝12.5Hz,2H),1.74(s,1H),1.49(q,J＝11.0Hz,2H),1.41(s,6H).
Example 4
2- (2-Aminopyridin-4-yl) -N- (6- (4-cyanopiperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-4)
Figure BDA0002681115790000232
2- (2-Aminopyridin-4-yl) -N- (6- (4-cyanopiperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that described in example 1.1H NMR(400MHz,Chloroform-d)δ9.93(s,1H),8.35(s,2H),8.23(d,J＝5.3Hz,1H),7.20(dd,J＝5.3,1.4Hz,1H),7.17–7.11(m,1H),6.55(s,1H),3.49(s,2H),3.12(d,J＝12.2Hz,2H),3.08–2.98(m,2H),2.75(t,J＝11.9Hz,3H),2.44–2.23(m,4H),1.48(s,6H).
Example 5
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6-morpholino-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-5)
Figure BDA0002681115790000233
2- (2-Aminopyrazine) may be prepared by a similar procedure as in example 1Pyridin-4-yl) -N- (2, 2-dimethyl-6-morpholino-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide.1H NMR(400MHz,Chloroform-d)δ10.16(s,1H),8.34(d,J＝6.7Hz,2H),8.19(d,J＝5.4Hz,1H),7.13(d,J＝5.3Hz,1H),6.97(s,1H),6.63(s,1H),4.90(s,2H),3.25(t,J＝6.0Hz,4H),3.04(s,2H),2.84(t,J＝6.0Hz,4H),1.48(s,6H).
Example 6
2- (2-Aminopyridin-4-yl) -N- (6- (3- (hydroxymethyl) pyrrolidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-6)
Figure BDA0002681115790000241
2- (2-Aminopyridin-4-yl) -N- (6- (3- (hydroxymethyl) pyrrolidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method as in example 1.1H NMR(400MHz,Chloroform-d)δ9.71(s,1H),8.33(s,1H),8.24(s,1H),8.17(d,J＝5.4Hz,1H),7.27(d,J＝1.2Hz,1H),7.19(dd,J＝5.4,1.4Hz,1H),6.63(s,1H),5.14(s,2H),3.98–3.81(m,2H),3.26(td,J＝8.7,3.2Hz,1H),3.13–2.96(m,4H),2.87(q,J＝8.5Hz,1H),2.58(d,J＝7.2Hz,1H),1.99–1.85(m,1H),1.47(s,6H).
Example 7
2- (2-Aminopyridin-4-yl) -N- (6- (4- (2, 2-difluoroethyl) piperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-7)
Figure BDA0002681115790000242
Step 1: preparation of 1- (2, 2-difluoroethyl) piperazine hydrochloride
Figure BDA0002681115790000243
1-tert-Butyloxycarbonylpiperazine (1g, 5.37mmol), 2, 2-difluoroethanol (528mg, 6.44mmol) were dissolved in anhydrous dichloromethane, triethylamine, argon were addedUnder the protection of gas, dropwise adding trifluoromethanesulfonic anhydride slowly under ice bath, reacting for 30 minutes in ice bath, and transferring to room temperature for reaction overnight. After TLC detection reaction, dichloromethane and water extraction, dichloromethane layer water washing, saturated salt water washing, anhydrous sodium sulfate drying, filtration, concentration to dryness, dissolution in dioxane (10mL), ice bath, addition of dioxane hydrochloride solution (10mL, 4.5mol/L), reaction at room temperature for three hours, after reaction, ice bath, suction filtration while cold to obtain 123mg of yellow brown solid. Yield: 12 percent.1H NMR(400MHz,DMSO-d6)δ9.78(s,2H),6.45(tt,J＝54.5,4.2Hz,1H),3.44(td,J＝15.3,4.1Hz,2H),3.36–3.17(m,8H).
Step 2: preparation of 6- (4- (2, 2-difluoroethyl) piperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-amine
Figure BDA0002681115790000251
6- (4- (2, 2-difluoroethyl) piperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-amine may be prepared by a method analogous to that of example 1, steps 1 and 2.1H NMR(400MHz,Chloroform-d)δ6.58(s,1H),6.49(s,1H),5.92(tt,J＝55.9,4.4Hz,1H),3.48(s,6H),2.94–2.85(m,6H),1.43(s,6H).
And step 3: preparation of 2- (2-aminopyridin-4-yl) -N- (6- (4- (2, 2-difluoroethyl) piperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide
Figure BDA0002681115790000252
2- (2-Aminopyridin-4-yl) -N- (6- (4- (2, 2-difluoroethyl) piperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide may be prepared from 6- (4- (2, 2-difluoroethyl) piperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-amine by a reaction analogous to example 1, steps 3, 4.1H NMR(400MHz,Chloroform-d)δ9.96(s,1H),8.34(s,1H),8.31(s,1H),7.31(dd,J＝5.3,1.4Hz,1H),7.14(t,J＝1.0Hz,1H),6.63(s,1H),5.99(tt,J＝55.7,4.2Hz,1H),4.72(s,2H),3.49(s,2H),3.06–2.99(m,2H),2.97–2.91(m,8H),1.48(s,6H).
Example 8
2- (2-Aminopyridin-4-yl) -N- (6- (4- (2-hydroxypropan-2-yl) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-8)
Figure BDA0002681115790000253
2- (2-Aminopyridin-4-yl) -N- (6- (4- (2-hydroxypropan-2-yl) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 1.1H NMR(300MHz,DMSO-d6)δ10.06(s,1H),8.89(q,J＝1.5Hz,1H),8.27(s,1H),8.11(d,J＝5.3Hz,1H),7.60(s,1H),7.28(s,1H),7.16–6.94(m,2H),6.72(d,J＝2.5Hz,1H),6.48(s,2H),2.97(d,J＝18.3Hz,4H),2.70(t,J＝11.6Hz,2H),2.34(d,J＝11.5Hz,1H),2.07(q,J＝12.2,11.8Hz,2H),1.85(d,J＝12.6Hz,2H),1.40(d,J＝2.6Hz,6H).
Example 9
2- (2-Aminopyridin-4-yl) -N- (6- (4-hydroxypiperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-9)
Figure BDA0002681115790000261
2- (2-Aminopyridin-4-yl) -N- (6- (4-hydroxypiperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that described in example 1.1H NMR(400MHz,DMSO-d6)δ10.14(s,1H),8.91(s,1H),8.18(s,1H),8.10(d,J＝5.3Hz,1H),7.24–7.04(m,2H),6.68(s,1H),6.30(s,2H),5.02(s,1H),3.72(s,1H),3.00(s,2H),2.95–2.84(m,2H),2.72(t,J＝10.7Hz,2H),1.97(d,J＝12.0Hz,2H),1.81(q,J＝10.2,9.1Hz,2H),1.40(s,6H).
Example 10
2- (2-Aminopyridin-4-yl) -N- (6- (4- (cyanomethyl) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-10)
Figure BDA0002681115790000262
2- (2-Aminopyridin-4-yl) -N- (6- (4- (cyanomethyl) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method as in example 1.1H NMR(400MHz,Chloroform-d)δ9.58(s,1H),8.35(d,J＝8.1Hz,2H),8.19(d,J＝5.4Hz,1H),7.27(s,1H),7.19(dd,J＝5.3,1.5Hz,1H),6.61(s,1H),5.09(s,2H),3.15(d,J＝11.8Hz,2H),3.03(s,2H),2.70(t,J＝11.5Hz,2H),2.54(d,J＝4.2Hz,2H),2.03–1.81(m,5H),1.48(s,6H).
Example 11
2- (2-Aminopyridin-4-yl) -N- (6- (4- (2-hydroxyethyl) piperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-11)
Figure BDA0002681115790000271
2- (2-Aminopyridin-4-yl) -N- (6- (4- (2-hydroxyethyl) piperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method as in example 1.1H NMR(400MHz,DMSO-d6)δ9.91(s,1H),8.92(s,1H),8.19(s,1H),8.11(d,J＝5.3Hz,1H),7.14(d,J＝5.3Hz,1H),7.05(s,1H),6.72(s,1H),6.37(s,2H),4.65(s,1H),3.62(s,2H),3.00(s,3H),2.97–2.55(m,10H),1.41(s,6H).
Example 12
N- (6- (4- (2-amino-2-oxoethyl) piperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) -2- (2-aminopyridin-4-yl) oxazole-4-carboxylic acid amide (I-12)
Figure BDA0002681115790000272
Step 1: preparation of 4- (2-amino-2-oxoethyl) piperazine-1-carboxylic acid tert-butyl ester
Figure BDA0002681115790000273
1-tert-butyloxycarbonyl piperazine (550mg, 2.95mmol) and bromoacetamide (611mg, 4.43mmol) were dissolved in tetrahydrofuran (20mL), N-diisopropylethylamine (763mg, 5.91mmol) was added and reacted at room temperature overnight, the reaction was detected by TLC, extraction was performed with ethyl acetate and water, the ethyl acetate layer was washed with saturated brine, dried over anhydrous sodium sulfate and concentrated to dryness to give 464mg of a white solid. The yield thereof was found to be 64%.1H NMR(400MHz,DMSO-d6)δ7.17(d,J＝30.9Hz,2H),2.86(s,2H),2.36(t,J＝4.9Hz,4H),1.39(s,9H).
Step 2: preparation of 2- (piperazin-1-yl) acetamide trifluoroacetate
Figure BDA0002681115790000274
Tert-butyl 4- (2-amino-2-oxoethyl) piperazine-1-carboxylate (150mg, 616. mu. mol) was dissolved in dichloromethane (8mL), trifluoroacetic acid (1.5mL) was added and the reaction was allowed to proceed at room temperature for two hours, after completion of TLC detection, concentrated to dryness, ether/methanol was added and the mixture was slurried and filtered to obtain 209mg of a white solid. Yield: 91 percent.1H NMR(400MHz,DMSO-d6)δ9.01(s,2H),7.55(d,J＝79.8Hz,2H),3.47(s,2H),3.26(t,J＝5.1Hz,4H),3.06(s,4H).
Step preparation of 32- (4- (5-amino-2, 2-dimethyl-2, 3-dihydrobenzofuran-6-yl) piperazin-1-yl) acetamide
Figure BDA0002681115790000281
2- (4- (5-amino-2, 2-dimethyl-2, 3-dihydrobenzofuran-6-yl) piperazin-1-yl) acetamide may be prepared by a similar method to that of example 1, steps 1 and 2.1H NMR(400MHz,DMSO-d6)δ7.16(d,J＝16.7Hz,2H),6.52(s,1H),6.32(s,1H),4.18(s,2H),2.90(s,2H),2.83(s,2H),2.77(s,4H),2.57(s,4H),1.33(s,6H).
And 4, step 4: preparation of N- (6- (4- (2-amino-2-oxoethyl) piperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) -2- (2-aminopyridin-4-yl) oxazole-4-carboxamide
Figure BDA0002681115790000282
N- (6- (4- (2-amino-2-oxoethyl) piperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) -2- (2-aminopyridin-4-yl) oxazole-4-carboxamide can be prepared from 2- (4- (5-amino-2, 2-dimethyl-2, 3-dihydrobenzofuran-6-yl) piperazin-1-yl) acetamide by a similar reaction as in example 1, steps 3, 4.1H NMR(400MHz,DMSO-d6)δ9.91(s,1H),8.91(s,1H),8.21(s,1H),8.14(d,J＝5.3Hz,1H),7.30(s,1H),7.17(s,1H),7.12(dd,J＝5.3,1.4Hz,1H),7.06(s,1H),6.70(s,1H),6.41(s,2H),3.04(s,2H),3.01(s,2H),2.86(d,J＝4.7Hz,4H),2.78(s,4H),1.41(s,6H).
Example 13
2- (2-Aminopyridin-4-yl) -N- (6- (4-carbamoylpiperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-13)
Figure BDA0002681115790000283
2- (2-Aminopyridin-4-yl) -N- (6- (4-carbamoylpiperazin-1-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide may be prepared by a similar method to that described in example 1.1H NMR(300MHz,DMSO-d6)δ9.81(s,1H),8.91(d,J＝2.7Hz,1H),8.25(s,1H),8.04(d,J＝4.9Hz,1H),7.10(t,J＝3.2Hz,2H),6.69(d,J＝2.8Hz,1H),6.35(s,2H),4.40(s,1H),3.00(s,3H),2.62(t,J＝11.5Hz,2H),1.95–1.55(m,4H),1.40(d,J＝3.8Hz,6H),1.14(d,J＝3.6Hz,6H).
Example 14
2- (2-Aminopyridin-4-yl) -N- (2- (hydroxymethyl) -6- (4- (hydroxymethyl) piperidin-1-yl) -2-methyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-14)
Figure BDA0002681115790000291
Step 1: preparation of methyl 3- (2, 4-difluorophenyl) -2-hydroxy-2-methylpropionate
Figure BDA0002681115790000292
Adding magnesium chips (2.47g, 104mmol) and iodine simple substance (147mg, 579 mu mol) into a two-necked bottle, replacing argon, adding 30mL of anhydrous ether, heating to keep the solution slightly boiling, slowly adding 1- (bromomethyl) -2, 4-difluorobenzene (6g, 28.9mmol), after the addition is finished, keeping boiling and stirring for half an hour to obtain (2, 4-difluorobenzyl) magnesium bromide for later use. Dissolving methyl 2-oxopropionate (2.96g, 28.9mmol) in anhydrous ether, stirring at-78 deg.C for 30 min under the protection of argon gas, slowly adding freshly prepared (2, 4-difluorobenzyl) magnesium bromide, reacting at room temperature for 2hr, detecting by TLC, adding ammonium chloride solution, quenching, filtering off insoluble substances, extracting the filtrate with ethyl acetate and water, washing ethyl acetate layer with saturated salt water, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, and performing gel column chromatography (ethyl acetate/petroleum ether elution) to obtain 2.88g yellow oily substance. Yield: and 43 percent.1H NMR(400MHz,Chloroform-d)δ7.24(dd,J＝8.6,6.5Hz,1H),6.88–6.75(m,2H),3.79(s,3H),3.18–3.07(m,2H),2.94(dd,J＝13.9,1.2Hz,1H),1.50(s,3H).
Step 2: preparation of 6-fluoro-2-methyl-2, 3-dihydrobenzofuran-2-carboxylic acid
Figure BDA0002681115790000293
Methyl 3- (2, 4-difluorophenyl) -2-hydroxy-2-methylpropionate (2.88g, 12.51mmol) was dissolved in anhydrous tetrahydrofuran (75mL), potassium tert-butoxide (3.51g, 31.28mmol) was added and the reaction was allowed to proceed overnight at 50 deg.C, after completion of the TLC detection, ethyl acetate (100mL) was added to separate the organic layer, dried over anhydrous sodium sulfate, filtered and concentrated to dryness to give a tan solid.1H NMR(400MHz,DMSO-d6)δ12.99(s,1H),7.21–7.13(m,1H),6.77–6.51(m,2H),3.46(d,J＝16.0Hz,1H),3.11(d,J＝16.0Hz,1H),1.59(s,3H).
And step 3: 6-fluoro-2-methyl-2, 3-dihydrobenzofuran-2-carboxylic acid methyl ester
Figure BDA0002681115790000294
6-fluoro-2-methyl-2, 3-dihydrobenzofuran-2-carboxylic acid (500mg, 2.55mmol) was dissolved in N, N-dimethylformamide (10mL), cesium carbonate (765mg, 3.82mmol) was added, iodomethane (434mg, 3.06mmol) was added with stirring, and the reaction was carried out at room temperature for 5 hours, after TLC detection. Extracting with ethyl acetate and water, washing with saturated sodium chloride, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, and performing gel column chromatography (eluting with ethyl acetate/petroleum ether at a ratio of 1: 50) to obtain 444mg of pale yellow oily substance. Yield: 83 percent.
And 4, step 4: preparation of methyl 6-fluoro-2-methyl-5-nitro-2, 3-dihydrobenzofuran-2-carboxylate
Figure BDA0002681115790000301
Dissolving 6-fluoro-2-methyl-2, 3-dihydrobenzofuran-2-carboxylic acid (444mg, 2.11mmol) in dichloromethane (8mL), slowly adding nitric acid (0.5mL) dropwise at room temperature, reacting at room temperature until TLC detection reaction is finished, adding water dichloromethane and water for extraction, separating dichloromethane layer, washing with saturated common salt water for three times, and drying with anhydrous sodium sulfate. Filtering, concentrating under reduced pressure, and performing gel column chromatography (gradient elution with ethyl acetate/petroleum ether at a ratio of 1: 20-1: 8) to obtain 227mg of light yellow oily substance. Yield: 42 percent.1H NMR(300MHz,Chloroform-d)δ7.93(d,J＝7.5Hz,1H),6.70(d,J＝11.2Hz,1H),3.81(d,J＝1.5Hz,3H),3.65(d,J＝16.3Hz,1H),3.17(d,J＝16.3Hz,1H),1.76(s,2H).
And 5: preparation of (6-fluoro-2-methyl-5-nitro-2, 3-dihydrobenzofuran-2-yl) methanol
Figure BDA0002681115790000302
Dissolving 6-fluoro-2-methyl-5-nitro-2, 3-dihydrobenzofuran-2-carboxylic acid methyl ester (227mg, 889 μmol) in tetrahydrofuran/ethanol (8mL/2mL), adding sodium borohydride (101mg, 2.67mmol) and lithium chloride (113mg, 2.67mmol) under ice bath, reacting at room temperature for 3 hours, detecting by TLC, quenching with saturated ammonium chloride solution, extracting with dichloromethane, separating an organic layer, washing with saturated common salt water, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, and performing gel column chromatography (gradient elution with ethyl acetate/petroleum ether ═ 1: 1) to obtain 200mg of brown yellow oil. Yield: 99 percent.1H NMR(300MHz,DMSO-d6)δ8.03(d,J＝8.1Hz,1H),6.95(dt,J＝11.0,2.4Hz,1H),5.19(dq,J＝8.0,4.9,4.1Hz,1H),3.60–3.40(m,2H),2.92(d,J＝16.2Hz,1H),1.38(q,J＝2.4Hz,3H).
Step 6: preparation of (1- (2- (hydroxymethyl) -2-methyl-5-nitro-2, 3-dihydrobenzofuran-6-yl) piperidin-4-yl) methanol
Figure BDA0002681115790000303
Dissolving (6-fluoro-2-methyl-5-nitro-2, 3-dihydrobenzofuran-2-yl) methanol and 4-hydroxymethylpiperidine (29mg, 248 mu mol) in DMF (4mL), adding potassium carbonate (57mg, 413 mu mol), reacting at room temperature overnight, detecting by TLC after the reaction is finished, adding ethyl acetate (20mL), washing twice with water, washing with saturated common salt water, drying with anhydrous sodium sulfate, filtering, concentrating to dryness, and directly putting into the next reaction without purification.
And 7: preparation of (1- (5-amino-2- (hydroxymethyl) -2-methyl-2, 3-dihydrobenzofuran-6-yl) piperidin-4-yl) methanol
Figure BDA0002681115790000311
Dissolving the product in methanol (18mL), adding 5% palladium carbon (20mg), reacting for 6 hours in hydrogen atmosphere, detecting by TLC after the reaction is finished, filtering by using kieselguhr, concentrating under reduced pressure to dryness, and directly putting the product into the next step without purification.
And 8: preparation of 2- (2-aminopyridin-4-yl) -N- (2- (hydroxymethyl) -6- (4- (hydroxymethyl) piperidin-1-yl) -2-methyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide
Figure BDA0002681115790000312
2- (2-aminopyridin-4-yl) -N- (2- (hydroxymethyl) -6- (4- (hydroxymethyl) piperidin-1-yl) -2-methyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared from (1- (5-amino-2- (hydroxymethyl) -2-methyl-2, 3-dihydrobenzofuran-6-yl) piperidin-4-yl) methanol by a similar method as in steps 3,4 of example 1.1H NMR(300MHz,DMSO-d6)δ9.97(s,1H),8.90(s,1H),8.21(s,1H),8.10(d,J＝5.2Hz,1H),7.09(dd,J＝5.3,1.5Hz,1H),7.06(s,1H),6.67(s,1H),6.34(s,2H),5.04(t,J＝5.8Hz,1H),4.70(t,J＝5.4Hz,1H),3.43(dd,J＝5.8,3.5Hz,4H),3.25–3.16(m,1H),2.94(d,J＝11.3Hz,2H),2.83(d,J＝15.8Hz,1H),2.67(t,J＝11.0Hz,2H),1.81(d,J＝11.3Hz,2H),1.72–1.47(m,3H),1.33(s,3H).
Example 15
2- (2-Aminopyridin-4-yl) -N- (2- (hydroxymethyl) -2-methyl-6-morpholino-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-15)
Figure BDA0002681115790000313
2- (2-Aminopyridin-4-yl) -N- (2- (hydroxymethyl) -2-methyl-6-morpholino-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method as in example 14.1H NMR(300MHz,DMSO-d6)δ9.86(s,1H),8.92(s,1H),8.15(d,J＝10.2Hz,2H),7.03(s,2H),6.72(s,1H),6.39(s,2H),5.05(s,1H),3.86(s,4H),3.43(s,2H),3.20(d,J＝16.2Hz,1H),2.82(d,J＝5.3Hz,4H),1.34(s,3H).
Example 16
2- (2-Aminopyridin-4-yl) -N- (7-morpholino-3-oxo-3, 4-dihydro-2H-benzo [ b ] [1,4] oxazin-6-yl) oxazole-4-carboxamide (I-16)
Figure BDA0002681115790000321
Step 1: preparation of 7-fluoro-6-nitro-2H-benzo [ b ] [1,4] oxazin-3 (4H) -one
Figure BDA0002681115790000322
The 7-fluoro-2H-1, 4-benzoxazine-3 (4H) -ketone (1g, 5.98mmol) and 80% sulfuric acid (10mL) are subjected to an ice salt bath to-10 ℃, 65% nitric acid (580mg, 5.98mmol) and 80% sulfuric acid (1mL) are slowly added into the reaction solution, the reaction solution reacts for 30 minutes at room temperature, TLC detection is finished, the reaction solution is poured into crushed ice, stirred and filtered to obtain g of yellow solid.1H NMR(300MHz,DMSO-d6)δ11.03(s,1H),7.64(d,J＝7.5Hz,1H),7.27(d,J＝12.3Hz,1H),4.79(s,2H),4.03(q,J＝7.1Hz,2H),1.99(s,3H),1.18(t,J＝7.1Hz,3H).
Step 2: 7-morpholino-6-nitro-2H-benzo [ b ] [1,4] oxazin-3 (4H) -one
Figure BDA0002681115790000323
Reacting 7-fluoro-6-nitro-2H-benzo [ b][1,4]Dissolving oxazine-3 (4H) -ketone (100mg, 471 mu mol) in N, N-dimethylformamide (1mL) and morpholine (1mL), reacting at 50 ℃ for 2 hours, detecting by TLC after the reaction is finished, adding ethyl acetate, washing twice, washing with saturated salt water, drying with anhydrous sodium sulfate, filtering, concentrating to dryness, and directly putting into the next step for reaction.1H NMR(300MHz,Chloroform-d)δ7.30–7.24(m,1H),6.61(d,J＝5.3Hz,1H),4.33(q,J＝5.0Hz,2H),3.86(q,J＝4.9Hz,4H),3.44(s,2H),2.97(q,J＝4.8Hz,4H).
And step 3: 6-amino-7-morpholino-2H-benzo [ b ] [1,4] oxazin-3 (4H) -one
Figure BDA0002681115790000324
The product of the previous step was dissolved in methanol (10mL), 5% palladium on carbon (20mg), hydrogen was addedReacting for 6 hours in the atmosphere, detecting by TLC, filtering by using diatomite, concentrating under reduced pressure until the diatomite is dry, and directly putting the diatomite into the next reaction.1H NMR(300MHz,DMSO-d6)δ10.42(s,1H),6.56(d,J＝1.8Hz,1H),6.29(d,J＝1.7Hz,1H),4.64(s,2H),4.37(d,J＝1.8Hz,2H),3.71(t,J＝4.4Hz,4H),2.71(t,J＝4.5Hz,4H).
And 4, step 4: preparation of 2- (2-aminopyridin-4-yl) -N- (7-morpholino-3-oxo-3, 4-dihydro-2H-benzo [ b ] [1,4] oxazin-6-yl) oxazole-4-carboxamide
Figure BDA0002681115790000331
By analogy with steps 3,4 of example 1 starting from 6-amino-7-morpholino-2H-benzo [ b ]][1,4]Preparation of 2- (2-aminopyridin-4-yl) -N- (7-morpholino-3-oxo-3, 4-dihydro-2H-benzo [ b ] using oxazin-3 (4H) -one][1,4]Oxazin-6-yl) oxazole-4-carboxamide.1H NMR(300MHz,DMSO-d6)δ10.76(s,1H),9.95(s,1H),9.03(s,1H),8.17(d,J＝5.7Hz,1H),8.08(s,1H),7.19(s,1H),7.13(d,J＝5.9Hz,1H),7.00(s,1H),4.55(s,2H),2.84(s,4H).
Example 17
2- (2-Aminopyridin-4-yl) -N- (7- (4- (hydroxymethyl) piperidin-1-yl) -3-oxo-3, 4-dihydro-2H-benzo [ b ] [1,4] oxazin-6-yl) oxazole-4-carboxamide (I-16)
Figure BDA0002681115790000332
2- (2-Aminopyridin-4-yl) -N- (7- (4- (hydroxymethyl) piperidin-1-yl) -3-oxo-3, 4-dihydro-2H-benzo [ b ] can be prepared by a similar procedure to example 16][1,4]Oxazin-6-yl) oxazole-4-carboxamide.1H NMR(300MHz,DMSO-d6)δ10.72(s,1H),10.06(s,1H),9.00(d,J＝2.1Hz,1H),8.10(d,J＝5.1Hz,2H),7.19(d,J＝8.5Hz,2H),6.93(s,1H),4.74(s,1H),4.53(d,J＝2.1Hz,2H),2.93(d,J＝11.0Hz,2H),2.69(t,J＝11.0Hz,2H),1.79(d,J＝11.3Hz,2H),1.64(d,J＝11.9Hz,2H),1.58(s,1H).
Example 18
2- (2-Aminopyridin-4-yl) -N- (7-morpholino-3, 4-dihydro-2H-benzo [ b ] [1,4] oxazin-6-yl) oxazole-4-carboxamide (I-18)
Figure BDA0002681115790000333
Can be prepared by a method analogous to that of example 16 starting from 7-fluoro-3, 4-dihydro-2H-benzo [ b ]][1,4]Preparation of 2- (2-aminopyridin-4-yl) -N- (7-morpholino-3, 4-dihydro-2H-benzo [ b ] oxazines][1,4]Oxazin-6-yl) oxazole-4-carboxamide.1H NMR(300MHz,DMSO-d6)δ10.01(s,1H),8.91(d,J＝1.8Hz,1H),8.19–8.07(m,1H),7.72(s,1H),7.03(s,2H),6.68(d,J＝1.9Hz,1H),6.39(s,2H),5.88(s,1H),4.10(s,2H),3.84(s,4H),2.76(s,4H).
Example 19
2- (2-Aminopyridin-4-yl) -N- (7- (4- (hydroxymethyl) piperidin-1-yl) -3, 4-dihydro-2H-benzo [ b ] [1,4] oxazin-6-yl) oxazole-4-carboxamide (I-19)
Figure BDA0002681115790000341
Can be prepared by a method analogous to that of example 26 from 7-fluoro-3, 4-dihydro-2H-benzo [ b ]][1,4]Preparation of 2- (2-aminopyridin-4-yl) -N- (7- (4- (hydroxymethyl) piperidin-1-yl) -3, 4-dihydro-2H-benzo [ b ] oxazines][1,4]Oxazin-6-yl) oxazole-4-carboxamide.1H NMR(300MHz,DMSO-d6)δ10.13(s,1H),8.89(d,J＝2.1Hz,1H),8.09(d,J＝5.3Hz,1H),7.73(s,1H),7.14–7.02(m,2H),6.62(s,1H),6.37(s,2H),5.82(s,1H),4.71(s,1H),4.09(s,2H),2.86(d,J＝11.1Hz,2H),2.65(d,J＝11.1Hz,2H),1.77(s,2H),1.55(s,1H),1.26(d,J＝19.2Hz,2H).
Example 20
2- (2-Aminopyridin-4-yl) -N- (2-isopropyl-6-morpholino-1-oxoisoindol-5-yl) oxazole-4-carboxamide (I-20)
Figure BDA0002681115790000342
Step 1: preparation of 6-chloro-2-isopropyl-5-nitroisoindol-1-one
Figure BDA0002681115790000343
Methyl 2- (bromomethyl) -5-chloro-4-nitrobenzoate (prepared according to the method reported in WO 2013/079505) (200mg, 648. mu. mol) was dissolved in methanol (8mL), triethylamine (108. mu.L, 777. mu. mol) and isopropylamine (66. mu.L, 777. mu. mol) were added, the reaction was allowed to react at 70 ℃ for 6 hours, TLC was complete, ethyl acetate was extracted with 1mol/L aqueous hydrochloric acid, the aqueous layer was extracted twice more with ethyl acetate and dried over anhydrous sodium sulfate. Concentrating to dryness. Directly putting into the next reaction.1H NMR(300MHz,DMSO-d6)δ8.34(s,1H),8.00(s,1H),4.54(s,2H),4.47–4.35(m,1H),1.26–1.22(m,6H).
Step 2: 2-isopropyl-6-morpholino-5-nitroisoindol-1-one
Figure BDA0002681115790000351
6-chloro-2-isopropyl-5-nitroisoindol-1-one (100mg, 392. mu. mol) and morpholine (69. mu.L, 785. mu. mol), N, N-diisopropylethylamine (129. mu.L, 785. mu. mol) were dissolved in dimethyl sulfoxide (2mL), reacted at 90 ℃ overnight, cooled to room temperature, extracted with ethyl acetate and water, the ethyl acetate layer was washed with water, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and subjected to gel column chromatography (ethyl acetate/petroleum ether elution) to give 32mg of an orange oil. Yield: 26 percent.1H NMR(300MHz,Chloroform-d)δ7.81(s,1H),7.65(d,J＝4.3Hz,1H),4.68(s,1H),4.37(d,J＝5.0Hz,2H),3.85(s,4H),3.08(s,4H),1.36–1.25(m,6H).
And step 3: 5-amino-2-isopropyl-6-morpholinoisoindol-1-one
Figure BDA0002681115790000352
Preparation from 2-isopropyl-6-morpholino-5-nitroisoindol-1-one by a similar procedure as in step 2 of example 15-amino-2-isopropyl-6-morpholinoisoindol-1-one.1H NMR(300MHz,Chloroform-d)δ7.54(s,1H),6.80(s,1H),4.66(d,J＝7.0Hz,1H),4.24(s,2H),3.89(d,J＝5.4Hz,4H),2.96(d,J＝4.8Hz,4H),1.31–1.27(m,6H).
And 4, step 4: preparation of 2- (2-aminopyridin-4-yl) -N- (2-isopropyl-6-morpholino-1-oxoisoindol-5-yl) oxazole-4-carboxamide
Figure BDA0002681115790000353
2- (2-Aminopyridin-4-yl) -N- (2-isopropyl-6-morpholino-1-oxoisoindol-5-yl) oxazole-4-carboxamide can be prepared from 5-amino-2-isopropyl-6-morpholinoisoindol-1-one by a similar method to Steps 3,4 of example 1.1H NMR(300MHz,DMSO-d6)δ10.29(s,1H),9.02(s,1H),8.59(s,1H),8.14(d,J＝5.4Hz,1H),7.57(s,1H),7.04(d,J＝9.1Hz,2H),6.39(s,2H),4.42(s,2H),3.90(s,4H),2.92(s,4H),1.21(d,J＝6.8Hz,6H).
Example 21
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6-morpholino-2, 3-dihydrofuro [2,3-b ] pyridin-5-yl) oxazole-4-carboxamide (I-21)
Figure BDA0002681115790000361
Step 1: preparation of 3-bromo-2, 6-difluoropyridine
Figure BDA0002681115790000362
3-bromo-2, 6-dichloropyridine (2.8g, 12.3mmol) and cesium fluoride (7.5g, 49.3mmol) were dissolved in dimethyl sulfoxide (35mL), reacted at 80 ℃ for 8 hours, extracted with ethyl acetate and water, the ethyl acetate layer was washed twice with water, with saturated brine and dried over anhydrous sodium sulfate. Filtration, concentration under reduced pressure and gel column chromatography (petroleum ether elution) gave 1.01g of a colorless oil.1H NMR(400MHz,Chloroform-d)δ8.15–7.97(m,1H),6.79(dq,J＝8.3,2.7,2.2Hz,1H).
Step 2: preparation of 1- (2, 6-difluoropyridin-3-yl) -2-methylpropan-2-ol
Figure BDA0002681115790000363
Dissolving 3-bromo-2, 6-difluoropyridine (1.01g, 5.21mmol) in anhydrous tetrahydrofuran (60mL), replacing with argon for protection, adding n-butyllithium (2.2mL, 6.25mmol) at-78 ℃, adding dimethyl propylene oxide (0.53mL, 6.25mmol) and boron trifluoride diethyl etherate (0.75mL, 6.25mmol) in sequence after 30 minutes, turning to room temperature for reaction for 1 hour, adding saturated ammonium chloride for quenching, extracting with ethyl acetate, washing with saturated common salt water, and drying with anhydrous sodium sulfate. Filtering, decompressing, concentrating, and carrying out gel column chromatography (gradient elution with ethyl acetate/petroleum ether of 10% -15%) to obtain 146mg of yellow oily matter.1H NMR(400MHz,Chloroform-d)δ7.82(dt,J＝9.3,7.9Hz,1H),6.79(ddd,J＝8.0,2.9,0.6Hz,1H),2.78(s,2H),1.26(d,J＝0.8Hz,7H).
And step 3: preparation of 6-fluoro-2, 2-dimethyl-2, 3-dihydrofuro [2,3-b ] pyridine
Figure BDA0002681115790000364
1- (2, 6-Difluoropyridin-3-yl) -2-methylpropan-2-ol (146mg, 779. mu. mol) and potassium tert-butoxide (218mg, 1.95mmol) were dissolved in anhydrous tetrahydrofuran (6mL) and reacted at room temperature for 1 hour, TLC detection was completed, and extraction was performed with ethyl acetate and water, the ethyl acetate layer was washed with water, washed with saturated brine and dried over anhydrous sodium sulfate. Filtration, concentration under reduced pressure and gel column chromatography (ethyl acetate/petroleum ether ═ 5% elution) gave 67mg of a colorless oil. Yield: 51 percent.1H NMR(400MHz,Chloroform-d)δ7.45(tt,J＝7.8,1.2Hz,1H),6.35(dd,J＝7.8,1.5Hz,1H),2.99(dd,J＝2.0,1.2Hz,2H),1.51(s,6H).
And 4, step 4: preparation of 6-fluoro-2, 2-dimethyl-5-nitro-2, 3-dihydrofuro [2,3-b ] pyridine
Figure BDA0002681115790000365
Reacting 6-fluoro-2, 2-dimethyl-2, 3-dihydrofuro [2,3-b ]]Pyridine (136mg, 813. mu. mol) was dissolved in acetic acid/acetic anhydride (4mL/4mL), copper nitrate (457mg, 2.44mmol) was added under ice bath, the mixture was allowed to warm to room temperature for reaction overnight, TLC detection showed that the reaction was substantially complete, ethyl acetate and water were extracted, the ethyl acetate layer was washed with water, washed with saturated brine, and dried over anhydrous sodium sulfate. Filtering, decompressing, concentrating, and silica gel column chromatography (5% -15% ethyl acetate/petroleum ether gradient elution) to obtain 80mg colorless oil. Yield: 46 percent.1H NMR(400MHz,Chloroform-d)δ8.28(dt,J＝8.8,1.5Hz,1H),3.12(t,J＝1.7Hz,2H),1.59(s,6H).
And 5: preparation of 2, 2-dimethyl-6-morpholino-5-nitro-2, 3-dihydrofuro [2,3-b ] pyridine
Figure BDA0002681115790000371
Reacting 6-fluoro-2, 2-dimethyl-5-nitro-2, 3-dihydrofuro [2,3-b ]]Pyridine (40mg, 188. mu. mol) was dissolved in morpholine (1mL) and N, N-dimethylformamide (1mL) and reacted at room temperature for 2 hours, TLC detected completion of the reaction, extracted with ethyl acetate and water, the ethyl acetate layer was washed with water, washed with saturated brine and dried over anhydrous sodium sulfate. Filtering, concentrating under reduced pressure to dryness, and directly putting into the next reaction.1H NMR(400MHz,DMSO-d6)δ8.18(s,1H),3.65(t,J＝4.5Hz,4H),2.99(s,2H),1.46(s,6H).
Step 6: preparation of 2, 2-dimethyl-6-morpholino-2, 3-dihydrofuro [2,3-b ] pyridin-5-amine
Figure BDA0002681115790000372
Prepared by a method analogous to that of example 1, step 2 from 2, 2-dimethyl-6-morpholino-5-nitro-2, 3-dihydrofuro [2,3-b ]]Preparation of 2, 2-dimethyl-6-morpholino-2, 3-dihydrofuro [2,3-b ] using pyridine]Pyridin-5-amine.1H NMR(400MHz,Chloroform-d)δ6.89(t,J＝1.1Hz,1H),3.88–3.80(m,4H),3.45(s,2H),3.13–3.04(m,4H),2.93(d,J＝1.1Hz,2H).
And 7: preparation of 2- (2-aminopyridin-4-yl) -N- (2, 2-dimethyl-6-morpholino-2, 3-dihydrofuro [2,3-b ] pyridin-5-yl) oxazole-4-carboxamide
Figure BDA0002681115790000373
2, 2-dimethyl-6-morpholino-2, 3-dihydrofuro [2,3-b ] can be prepared by a method similar to steps 3 and 4 in example 1]Preparation of 2- (2-aminopyridin-4-yl) -N- (2, 2-dimethyl-6-morpholino-2, 3-dihydrofuro [2,3-b ] using pyridin-5-amine]Pyridin-5-yl) oxazole-4-carboxamide.1H NMR(400MHz,Chloroform-d)δ9.36(s,1H),8.50(s,1H),8.33(s,1H),8.23(d,J＝5.3Hz,1H),7.24(s,1H),7.09(s,1H),4.72(s,2H),3.95(s,4H),3.09(t,J＝4.2Hz,4H),3.05(s,2H),1.50(s,6H).
Example 22
2- (2-Aminopyridin-4-yl) -N- (6- (4- (hydroxymethyl) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrofuro [2,3-b ] pyridin-5-yl) oxazole-4-carboxamide (I-22)
Figure BDA0002681115790000381
2- (2-Aminopyridin-4-yl) -N- (6- (4- (hydroxymethyl) piperidin-1-yl) -2, 2-dimethyl-2, 3-dihydrofuro [2,3-b ] can be prepared by a similar method to that in example 31]Pyridin-5-yl) oxazole-4-carboxamide.1H NMR(400MHz,DMSO-d6)δ9.46(s,1H),8.97(s,1H),8.41(d,J＝2.9Hz,1H),8.16–8.04(m,1H),7.09(d,J＝3.5Hz,2H),6.50(s,2H),4.70(s,1H),3.41(s,2H),3.16(d,J＝11.6Hz,2H),3.03(s,2H),2.72(t,J＝11.2Hz,2H),1.78(d,J＝10.4Hz,2H),1.57(t,J＝10.1Hz,3H),1.43(s,6H).
Example 23
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (1-methyl-1H-pyrazol-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-23)
Figure BDA0002681115790000382
Step 1: preparation of 4- (2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran-6-yl) -1-methyl-1H-pyrazole
Figure BDA0002681115790000383
6-chloro-2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran (0.1g, 0.439mmol), 1-methylpyrazole-4-boronic acid pinacol ester (0.183g, 0.878mmol) and cesium carbonate (0.286g, 0.878mmol) were dissolved in dioxane/water (4mL/0.5mL), and [1,1' -bis (diphenylphosphino) ferrocene was added under an argon stream]Palladium dichloride (33mg, 0.043mmol), reaction at 100 ℃ overnight, TLC detection after the reaction, suction filtration with celite, dilution with ethyl acetate (15mL), washing with water and saturated brine once, drying over anhydrous sodium sulfate, filtration, concentration under reduced pressure, and silica gel column chromatography (gradient elution with ethyl acetate/petroleum ether ═ 0-5%) to give 119mg of a yellow oil. Yield: 99 percent.1H NMR(400MHz,Chloroform-d)δ7.71(s,1H),7.54(d,J＝10.6Hz,2H),6.69(s,1H),3.93(s,3H),3.06(s,2H),1.52(s,6H).
Step 2: preparation of 2, 2-dimethyl-6- (1-methyl-1H-pyrazol-4-yl) -2, 3-dihydrobenzofuran-5-amine
Figure BDA0002681115790000391
Dissolving 4- (2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran-6-yl) -1-methyl-1H-pyrazole (101mg, 0.369mmol) in methanol (10mL), adding palladium/carbon (30mg), reacting in water bath at 30 ℃ for 6 hours under hydrogen atmosphere, detecting by TLC after the reaction is finished, filtering by using kieselguhr, concentrating under reduced pressure, and directly putting into the next reaction without further purification.1H NMR(400MHz,Chloroform-d)δ7.65(s,1H),7.53(s,1H),6.60(d,J＝9.9Hz,2H),3.94(s,3H),2.95(s,2H),1.46(s,6H).
And step 3: preparation of 2- (2-aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (1-methyl-1H-pyrazol-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide
Figure BDA0002681115790000392
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (1-methyl-1H-pyrazol-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared from 2, 2-dimethyl-6- (1-methyl-1H-pyrazol-4-yl) -2, 3-dihydrobenzofuran-5-amine by a similar method to Steps 3,4 of example 1.1H NMR(400MHz,DMSO-d6)δ9.58(s,1H),8.88(s,1H),8.11(d,J＝5.1Hz,1H),7.97(s,1H),7.70(s,1H),7.50(d,J＝10.9Hz,1H),7.02(d,J＝4.6Hz,2H),6.82(s,1H),6.36(d,J＝4.6Hz,1H),3.85(s,3H),3.03(s,2H),1.44(s,6H).
Example 24
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-24)
Figure BDA0002681115790000393
Step 1: preparation of 4- (2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran-6-yl) pyridine
Figure BDA0002681115790000394
4- (2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran-6-yl) pyridine can be prepared from 6-chloro-2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran by a similar method to step 1 in example 23.1H NMR(400MHz,Chloroform-d)δ8.70–8.58(m,2H),7.93(t,J＝1.3Hz,1H),7.24–7.14(m,2H),6.60(s,1H),3.12(d,J＝1.3Hz,2H),1.55(s,6H).MS(ESI,m/z):[M+H]+:271.2。
Step 2: preparation of 2, 2-dimethyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-5-amine
4- (2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran-6-yl) pyridine (0.1g, 0.370mmol) was dissolved in ethanol/water (6mL/2mL) and addedReduced iron powder (0.124g, 2.20mmol) and ammonium chloride (0.119g, 2.20mmol) react for 1.5 hours in an oil bath at 50 ℃, TLC detects that the reaction is finished, diatomite performs suction filtration, concentration under reduced pressure, ethyl acetate dissolution (15mL), 1mol/L diluted hydrochloric acid extraction (15mL), water layer collection, saturated sodium carbonate solution is adjusted to be alkaline, ethyl acetate (15mL 2) extraction, anhydrous sodium sulfate drying, filtration and concentration under reduced pressure to obtain 75mg of light yellow solid. Yield: 84 percent.1H NMR(400MHz,Chloroform-d)δ8.72–8.59(m,2H),7.46–7.36(m,2H),6.64(s,1H),6.53(s,1H),3.32(brs,2H),2.99(s,2H),1.47(s,6H).
And step 3: preparation of 2- (2-aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide
Figure BDA0002681115790000401
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared from 2, 2-dimethyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-5-amine in a similar manner to Steps 3,4 of example 1.1H NMR(400MHz,DMSO-d6)δ9.72(s,1H),8.76(s,1H),8.64–8.52(m,2H),8.08(d,J＝5.2Hz,1H),7.52–7.34(m,3H),7.04–6.89(m,2H),6.77(s,1H),6.35(s,2H),3.09(s,2H),1.46(s,6H).
Example 25
2- (2-Aminopyridin-4-yl) -N- (6- (furan-3-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-25)
Figure BDA0002681115790000402
2- (2-Aminopyridin-4-yl) -N- (6- (furan-3-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,DMSO-d6)δ9.61(s,1H),8.87(s,1H),8.20–8.06(m,1H),7.95(t,J＝1.2Hz,1H),7.77(t,J＝1.7Hz,1H),7.50(s,1H),7.12–6.97(m,2H),6.86(s,1H),6.82(dd,J＝1.9,0.9Hz,1H),6.38(s,2H),3.05(s,2H),1.44(s,6H).
Example 26
2- (2-Aminopyridin-4-yl) -N- (6- (3, 5-dimethylisoxazol-4-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-26)
Figure BDA0002681115790000411
2- (2-Aminopyridin-4-yl) -N- (6- (3, 5-dimethylisoxazol-4-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,Chloroform-d)δ8.68(s,1H),8.29(d,J＝2.6Hz,2H),8.20(d,J＝5.4Hz,1H),7.13(dd,J＝5.4,1.4Hz,1H),7.03(s,1H),6.55(s,1H),4.88(s,2H),3.11(s,2H),2.33(d,J＝2.0Hz,3H),2.20(d,J＝2.0Hz,3H),1.52(s,6H).
Example 27
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (pyridin-3-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-27)
Figure BDA0002681115790000412
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (pyridin-3-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,Chloroform-d)δ8.73(s,1H),8.68(d,J＝4.6Hz,2H),8.27(s,1H),8.17(d,J＝5.3Hz,1H),8.08(s,1H),7.77(d,J＝7.8Hz,1H),7.41(dd,J＝7.8,4.8Hz,1H),7.06(d,J＝5.3Hz,1H),6.97(s,1H),6.69(s,1H),4.76(s,2H),3.11(s,2H),1.53(s,6H).
Example 28
2- (2-Aminopyridin-4-yl) -N- (6-morpholino-3H-spiro [ benzofuran-2, 4' -piperidin ] -5-yl) oxazole-4-carboxamide (I-28)
Figure BDA0002681115790000413
Step 1: preparation of 2,2, 2-trifluoro-1- (6-fluoro-5-nitro-3H-spiro [ benzofuran-2, 4 '-piperidin ] -1' -yl) ethan-1-one
Figure BDA0002681115790000414
6-fluoro-5-nitro-3H-spiro-tert-butyl [ benzofuran-2, 4' -piperidine ] can be prepared by a method similar to that of Steps 1 to 5 of example 29]-1' -carboxylic acid tert-butyl ester.1H NMR(400MHz,Chloroform-d)δ7.96(d,J＝7.6Hz,1H),6.65(d,J＝11.2Hz,1H),4.39(d,J＝13.7Hz,1H),3.92(d,J＝14.4Hz,1H),3.64(t,J＝12.9Hz,1H),3.36(t,J＝12.7Hz,1H),3.08(s,2H),2.09(d,J＝14.1Hz,2H),1.93–1.76(m,2H).
Step 2: preparation of 2,2, 2-trifluoro-1- (6-morpholino-5-nitro-3H-spiro [ benzofuran-2, 4 '-piperidin ] -1' -yl) ethan-1-one
Figure BDA0002681115790000421
2,2, 2-trifluoro-1- (6-fluoro-5-nitro-3H-spiro [ benzofuran-2, 4' -piperidine)]-1' -yl) ethan-1-one (259mg,0.744mmol) was dissolved in morpholine (5mL) and reacted at 50 ℃ for 45 minutes, water (25mL) was added to precipitate a solid, which was filtered and dried in vacuo to give 303mg of a yellow solid. Yield: 98 percent.1H NMR(400MHz,Chloroform-d)δ7.85(s,1H),6.51(s,1H),4.37(d,J＝13.9Hz,1H),3.87(d,J＝5.1Hz,5H),3.64(t,J＝12.7Hz,1H),3.36(t,J＝12.8Hz,1H),3.03(d,J＝6.0Hz,6H),2.06(d,J＝14.1Hz,2H),1.84(d,J＝13.4Hz,2H).
And step 3: preparation of 1- (5-amino-6-morpholino-3H-spiro [ benzofuran-2, 4 '-piperidin ] -1' -yl) -2,2, 2-trifluoroethane-1-one
Figure BDA0002681115790000422
2,2, 2-trifluoro-1- (6-morpholino-5-nitro-3H-spiro [ benzofuran-2, 4' -piperidine)]-1' -Yl) ethan-1-one (303mg, 0.729mmol) inEthanol/water (24mL/8mL), reduced iron powder (244mg, 4.38mmol) and ammonium chloride (234mg, 4.38mmol) were added, the reaction was carried out in an oil bath at 50 ℃ for 1.5 hours, TLC detection was complete, filtration was carried out with celite, concentration was carried out under reduced pressure, ethyl acetate was dissolved (100mL), washing with saturated brine, drying over anhydrous sodium sulfate, filtration and concentration under reduced pressure were carried out to obtain 277mg of a brown-yellow solid. Yield: 99 percent.1H NMR(400MHz,Chloroform-d)δ6.60(s,1H),6.53(s,1H),4.33(d,J＝13.2Hz,1H),3.84(t,J＝4.6Hz,5H),3.65(t,J＝12.8Hz,1H),3.36(t,J＝12.5Hz,1H),2.93(s,2H),2.87(t,J＝4.6Hz,4H),2.03(d,J＝13.6Hz,2H),1.73(t,J＝13.0Hz,3H).
And 4, step 4: preparation of 2- (2-aminopyridin-4-yl) -N- (6- (pyridin-4-yl) -1'- (2,2, 2-trifluoroacetyl) -3H-spiro [ benzofuran-2, 4' -piperidin ] -5-yl) oxazole-4-carboxamide
Figure BDA0002681115790000423
2- (2-Aminopyridin-4-yl) -N- (6- (pyridin-4-yl) -1'- (2,2, 2-trifluoroacetyl) -3H-spiro [ benzofuran-2, 4' -piperidine ] -5-yl) oxazole-4-carboxamide can be prepared from 1- (5-amino-6-morpholino-3H-spiro [ benzofuran-2, 4 '-piperidin ] -1' -yl) -2,2, 2-trifluoroethan-1-one by a similar method to that described in example 1, Steps 3, 4. MS (ESI) 573.3[ M + H ].
And 5: preparation of 2- (2-aminopyridin-4-yl) -N- (6-morpholino-3H-spiro [ benzofuran-2, 4' -piperidin ] -5-yl) oxazole-4-carboxamide
Figure BDA0002681115790000431
2- (2-Aminopyridin-4-yl) -N- (6-morpholino-3H-spiro [ benzofuran-2, 4' -piperidine) can be prepared by a similar method to step 6 in example 29]-5-yl) oxazole-4-carboxamide.1H NMR(400MHz,DMSO-d6)δ9.86(s,1H),8.92(s,1H),8.18(s,1H),8.14(d,J＝5.2Hz,1H),7.12–6.98(m,2H),6.78(s,1H),6.38(s,2H),3.95–3.80(m,4H),2.99(s,2H),2.86(dq,J＝18.6,4.6Hz,6H),2.71–2.59(m,2H),1.68(dq,J＝15.5,10.5,9.4Hz,4H).
Example 29
2- (2-Aminopyridin-4-yl) -N- (6- (pyridin-4-yl) -3H-spiro [ benzofuran-2, 4' -piperidin ] -5-yl) oxazole-4-carboxamide trifluoroacetate (I-29)
Figure BDA0002681115790000432
Step 1: preparation of 4- (4-chloro-2-fluorobenzyl) -4-hydroxypiperidine-1-carboxylic acid tert-butyl ester
Figure BDA0002681115790000433
Adding magnesium chips (2.18g,35.8mmol) and iodine granules (151mg) into a two-necked bottle, adding anhydrous ether (25mL) under the protection of argon, heating to reflux, slowly dropwise adding 1- (bromomethyl) -4-chloro-2-fluorobenzene (8.0g,35.8mmol), and keeping reflux reaction for 30 minutes after dropwise adding is finished, thus preparing (4-chloro-2-fluorobenzyl) magnesium bromide. Tert-butyl 4-oxopiperidine-1-carboxylate (5.94g,29.8mmol) was dissolved in diethyl ether (190mL), and the resulting 4-chloro-2-fluorobenzyl magnesium bromide was added dropwise to the solution in a low-temperature bath, and the reaction was allowed to proceed at room temperature for 2 hours. Water and ethyl acetate (200mL) were added, the organic layer was separated, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and subjected to silica gel column chromatography (methanol/dichloromethane ═ 5% elution) to give 5.18g of a pale yellow solid. Yield: 51 percent.1H NMR(400MHz,Chloroform-d)δ7.16(t,J＝8.0Hz,1H),7.11(d,J＝1.9Hz,1H),7.08(q,J＝2.0,1.6Hz,1H),3.85(s,2H),3.09(t,J＝12.5Hz,2H),2.78(d,J＝1.6Hz,2H),1.49(s,2H),1.45(s,9H).
Step 2: preparation of 6-chloro-3H-spiro [ benzofuran-2, 4 '-piperidine ] -1' -carboxylic acid tert-butyl ester
Figure BDA0002681115790000441
4- (4-chloro-2-fluorobenzyl) -4-hydroxypiperidine-1-carboxylic acid tert-butyl ester (2.91g,8.46mmol) was dissolved in anhydrous tetrahydrofuran (100mL) and reacted at 65 ℃ for 3 hours, water and ethyl acetate were added, the organic layer was separated, dried over anhydrous sodium sulfate, filtered,vacuum concentrating, and silica gel column chromatography (gradient elution with ethyl acetate/petroleum ether of 0-10%) to obtain 1.10g of white solid. Yield: 40 percent.1H NMR(400MHz,Chloroform-d)δ7.03(d,J＝7.9Hz,1H),6.80(dd,J＝7.9,1.9Hz,1H),6.75(d,J＝1.8Hz,1H),3.74(s,2H),3.52–3.31(m,2H),2.94(s,2H),1.89(d,J＝13.4Hz,2H),1.69(td,J＝13.9,12.0,4.4Hz,2H),1.47(s,9H).
And step 3: preparation of 6-chloro-3H-spiro [ benzofuran-2, 4' -piperidine ] trifluoroacetate
Figure BDA0002681115790000442
Reacting 6-chloro-3H-spiro [ benzofuran-2, 4' -piperidine]Tert-butyl (300mg,0.926mmol) of the (E) -1' -carboxylate was dissolved in methylene chloride (10mL), and trifluoroacetic acid (3mL) was added to the solution to react at room temperature for 1.5 hours. Concentrated under reduced pressure, slurried with diethyl ether, and filtered to obtain 280mg of a white solid. Yield: 89 percent.1H NMR(400MHz,DMSO-d6)δ8.70(d,J＝38.2Hz,2H),7.22(d,J＝7.8Hz,1H),6.89(d,J＝9.3Hz,2H),3.20(s,4H),3.08(s,2H),2.08–1.86(m,4H).
And 4, step 4: preparation of 1- (6-chloro-3H-spiro [ benzofuran-2, 4 '-piperidin ] -1' -yl) -2,2, 2-trifluoroethane-1-one
Figure BDA0002681115790000443
Reacting 6-chloro-3H-spiro [ benzofuran-2, 4' -piperidine]Adding trifluoroacetate (200mg) into a two-necked bottle, adding dichloromethane (8mL) and N, N-diisopropylethylamine (294 mu 9,1.78mmol) under the protection of argon, adding trifluoroacetic anhydride after ice bath for 10 minutes, reacting at room temperature for 3 hours, adding water and dichloromethane, separating an organic layer, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, drying in vacuum to obtain 200mg of oily matter, and directly putting into the next reaction without further purification.1H NMR(400MHz,Chloroform-d)δ7.06(dd,J＝7.9,1.3Hz,1H),6.84(dd,J＝7.9,1.9Hz,1H),6.78(d,J＝1.9Hz,1H),4.35(d,J＝13.3Hz,1H),3.88(d,J＝14.2Hz,1H),3.65(ddd,J＝14.3,12.0,2.7Hz,1H),3.45–3.29(m,1H),3.08–2.92(m,2H),2.14–1.98(m,2H),1.89–1.70(m,2H).
And 5: preparation of 1- (6-chloro-5-nitro-3H-spiro [ benzofuran-2, 4 '-piperidin ] -1' -yl) -2,2, 2-trifluoroethane-1-one
Figure BDA0002681115790000444
1- (6-chloro-3H-spiro [ benzofuran-2, 4' -piperidine)]-1' -yl) -2,2, 2-trifluoroethane-1-one (72mg,0.225mmol) was dissolved in dichloromethane (3mL), concentrated nitric acid (0.5mL) was added for reaction for 2 hours, water and dichloromethane were added, the organic layer was separated, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by preparative TLC to give 46mg of a pale yellow oil. Yield: 56 percent.1H NMR(400MHz,Chloroform-d)δ7.86(t,J＝1.3Hz,1H),6.92(s,1H),4.39(d,J＝13.6Hz,1H),3.92(d,J＝14.2Hz,1H),3.64(t,J＝12.8Hz,1H),3.35(t,J＝12.7Hz,1H),3.08(d,J＝1.3Hz,2H),2.20–2.01(m,2H),1.96–1.76(m,2H).
Step 6: preparation of 6-chloro-5-nitro-3H-spiro [ benzofuran-2, 4' -piperidine ]
Figure BDA0002681115790000451
1- (6-chloro-5-nitro-3H-spiro [ benzofuran-2, 4 '-piperidin ] -1' -yl) -2,2, 2-trifluoroethane-1-one (46mg,0.126mmol) was dissolved in methanol, potassium carbonate (52mg) was added and reacted at room temperature for 40 minutes, water and methylene chloride (containing 5% methanol) were added, the organic layer was separated, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and directly charged into the next reaction without further purification.
And 7: preparation of 6-chloro-5-nitro-3H-spiro [ benzofuran-2, 4 '-piperidine ] -1' -carboxylic acid tert-butyl ester
Figure BDA0002681115790000452
Dissolving the product in dioxane (3mL), adding di-tert-butyl dicarbonate (35mg,0.152mmol), reacting at room temperature for 2 hours, adding water and ethyl acetate, and separatingThe organic layer was separated, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 56mg of a yellow oil.1H NMR(400MHz,Chloroform-d)δ7.84(t,J＝1.2Hz,1H),6.89(s,1H),3.93–3.74(m,2H),3.37(t,J＝11.9Hz,2H),3.03(d,J＝1.4Hz,2H),1.92(d,J＝13.6Hz,2H),1.74(td,J＝14.1,12.2,4.5Hz,2H),1.48(s,9H).
And 8: preparation of 5-nitro-6- (pyridin-4-yl) -3H-spiro [ benzofuran-2, 4 '-piperidine ] -1' -carboxylic acid tert-butyl ester
Figure BDA0002681115790000453
Reacting 6-chloro-5-nitro-3H-spiro [ benzofuran-2, 4' -piperidine]-1 '-Carboxylic acid tert-butyl ester (405mg,1.10mmol), pyridine-4-boronic acid (540mg,4.39mmol) and cesium carbonate (716mg,2.20mmol) were dissolved in dioxane/water (20mL/2mL), and [1,1' -bis (diphenylphosphino) ferrocene ] was added under argon flow]Palladium dichloride (80mg, 0.110mmol), reacted overnight at 100 ℃, after TLC detection, filtered through celite, diluted with ethyl acetate (100mL), washed once with water and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and chromatographed on silica gel (ethyl acetate/petroleum ether ═ 30% elution) to give 291mg of a yellow solid. Yield: 76 percent.1H NMR(400MHz,Chloroform-d)δ8.71–8.61(m,2H),7.99–7.90(m,1H),7.24–7.16(m,2H),6.67(s,1H),3.81(s,2H),3.39(t,J＝11.9Hz,2H),3.11(s,2H),1.96(d,J＝13.4Hz,2H),1.78(td,J＝14.1,12.3,4.4Hz,3H),1.48(s,9H).
And step 9: preparation of 5-amino-6- (pyridin-4-yl) -3H-spiro [ benzofuran-2, 4 '-piperidine ] -1' -carboxylic acid tert-butyl ester
Figure BDA0002681115790000454
Reacting 5-nitro-6- (pyridin-4-yl) -3H-spiro [ benzofuran-2, 4' -piperidine]-1' -Carboxylic acid tert-butyl ester (42mg, 0.102mmol) was dissolved in ethanol/water (1.5mL/0.5mL), reduced iron powder (34mg, 0.612mmol) and ammonium chloride (33mg, 0.612mmol) were added, the reaction was oil-bathed at 50 ℃ for 1.5 hours, TLC was performed, the reaction was completed, filtered through celite, concentrated under reduced pressure, ethyl acetateDissolved (15mL), washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 38mg of a yellow solid. Yield: 98 percent.1H NMR(400MHz,Chloroform-d)δ8.71–8.60(m,2H),7.46–7.35(m,2H),6.64(s,1H),6.57(s,1H),3.75(s,2H),3.39(t,J＝11.9Hz,2H),2.97(d,J＝1.1Hz,2H),1.91(d,J＝13.6Hz,2H),1.78–1.63(m,4H),1.47(s,9H).
Step 10: preparation of 2- (2-aminopyridin-4-yl) -N- (6- (pyridin-4-yl) -3H-spiro [ benzofuran-2, 4' -piperidin ] -5-yl) oxazole-4-carboxamide trifluoroacetate
Figure BDA0002681115790000461
Can be prepared by a method analogous to steps 3,4 of example 1 from 5-amino-6- (pyridin-4-yl) -3H-spiro [ benzofuran-2, 4' -piperidine]Preparation of 2- (2-aminopyridin-4-yl) -N- (6- (pyridin-4-yl) -3H-spiro [ benzofuran-2, 4 '-piperidine) from tert-butyl (E) -1' -carboxylate]-5-yl) oxazole-4-carboxamide trifluoroacetate salt.1H NMR(400MHz,DMSO-d6)δ10.03(s,1H),8.91(s,1H),8.83–8.66(m,3H),8.63(s,1H),8.12(d,J＝6.5Hz,3H),7.67(d,J＝5.5Hz,2H),7.41(s,1H),7.39(d,J＝1.6Hz,1H),7.21(dd,J＝6.4,1.6Hz,1H),6.94(s,1H),3.23(s,6H),2.03(dt,J＝18.4,12.3Hz,4H).
Example 30
2- (2-Aminopyridin-4-yl) -N- (6- (1-methyl-1H-pyrazol-4-yl) -3H-spiro [ benzofuran-2, 4' -piperidin ] -5-yl) oxazole-4-carboxamide trifluoroacetate (I-30)
Figure BDA0002681115790000462
2- (2-Aminopyridin-4-yl) -N- (6- (1-methyl-1H-pyrazol-4-yl) -3H-spiro [ benzofuran-2, 4' -piperidine) can be prepared by a similar method to that in example 29]-5-yl) oxazole-4-carboxamide trifluoroacetate salt.1H NMR(400MHz,DMSO-d6)δ9.71(s,1H),9.01(s,1H),8.68(d,J＝42.0Hz,2H),8.16(d,J＝6.4Hz,1H),7.98(s,3H),7.72(s,1H),7.47(s,1H),7.40(s,1H),7.24(d,J＝6.2Hz,1H),6.93(s,1H),3.84(s,3H),3.23(s,4H),3.15(s,2H),2.12–1.88(m,4H).
Example 31
2- (2-Aminopyridin-4-yl) -N- (6- (1-isopropyl-1H-pyrazol-4-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-31)
Figure BDA0002681115790000471
2- (2-Aminopyridin-4-yl) -N- (6- (1-isopropyl-1H-pyrazol-4-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,DMSO-d6)δ9.55(s,1H),8.89(s,1H),8.09(d,J＝5.2Hz,1H),8.06(s,1H),7.71(s,1H),7.50(s,1H),7.02(d,J＝5.2Hz,2H),6.86(s,1H),6.35(s,2H),4.47(hept,J＝6.7Hz,1H),3.04(s,2H),1.44(s,6H),1.37(d,J＝6.7Hz,6H).
Example 32
2- (2-Aminopyridin-4-yl) -N- (6- (4-chloro-2-fluorophenyl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-32)
Figure BDA0002681115790000472
2- (2-Aminopyridin-4-yl) -N- (6- (4-chloro-2-fluorophenyl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,Chloroform-d)δ8.67(s,1H),8.26(s,1H),8.20(d,J＝5.3Hz,1H),8.14(s,1H),7.31(q,J＝9.3,8.0Hz,3H),7.08(d,J＝5.4Hz,1H),6.90(s,1H),6.67(s,1H),4.79(s,2H),3.11(s,2H),1.52(s,6H).
Example 33
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (2-methylpyridin-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-33)
Figure BDA0002681115790000473
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (2-methylpyridin-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that described in example 24.1H NMR(400MHz,Chloroform-d)δ8.89(s,1H),8.65(d,J＝5.1Hz,1H),8.26(d,J＝4.0Hz,2H),8.17(d,J＝5.3Hz,1H),7.22(d,J＝5.2Hz,1H),7.04(d,J＝5.3Hz,1H),6.85(s,1H),6.68(s,1H),5.05(s,2H),3.11(s,2H),2.63(s,3H),1.52(s,6H).
Example 34
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (6-methylpyridin-3-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-34)
Figure BDA0002681115790000481
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (6-methylpyridin-3-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that described in example 24.1H NMR(400MHz,DMSO-d6)δ9.62(s,1H),8.77(s,1H),8.46(d,J＝2.3Hz,1H),8.08(d,J＝5.3Hz,1H),7.71(dd,J＝8.0,2.4Hz,1H),7.43(s,1H),7.28(d,J＝8.0Hz,1H),6.96(s,1H),6.94(dd,J＝5.3,1.5Hz,1H),6.73(s,1H),6.34(s,2H),3.08(s,2H),2.46(s,3H),1.46(s,6H).
Example 35
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (6- (trifluoromethyl) pyridin-3-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-35)
Figure BDA0002681115790000482
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (6- (trifluoromethyl) pyridin-3-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,DMSO-d6)δ9.85(s,1H),8.78(d,J＝2.1Hz,1H),8.75(s,1H),8.17–8.01(m,2H),7.93(d,J＝8.1Hz,1H),7.37(s,1H),6.98(s,1H),6.96(dd,J＝5.3,1.4Hz,1H),6.86(s,1H),6.34(s,2H),3.10(s,2H),1.47(s,6H).
Example 36
2- (2-Aminopyridin-4-yl) -N- (6- (2-methoxypyridin-4-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-36)
Figure BDA0002681115790000491
2- (2-Aminopyridin-4-yl) -N- (6- (2-methoxypyridin-4-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,Chloroform-d)δ8.98(s,1H),8.31(d,J＝5.3Hz,1H),8.27(s,1H),8.25(s,1H),8.18(d,J＝5.4Hz,1H),7.10(d,J＝5.4Hz,1H),7.02–6.95(m,1H),6.93(s,1H),6.86(s,1H),6.68(s,1H),4.86(s,2H),4.00(s,3H),3.11(s,2H),1.52(s,6H).
Example 37
2- (2-Aminopyridin-4-yl) -N- (6-fluoro-2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-37)
Figure BDA0002681115790000492
Step 1: preparation of 6-fluoro-2, 2-dimethyl-2, 3-dihydrobenzofuran-5-amine
Figure BDA0002681115790000493
Dissolving 6-fluoro-2, 2-dimethyl-5-nitro-2, 3-dihydrobenzofuran (60mg, 0.284mmol) in ethanol/water (6mL/2mL), adding reduced iron powder (95mg, 1.70mmol) and ammonium chloride (91mg, 1.70mmol), carrying out oil bath reaction at 50 ℃ for 2 hours, detecting the reaction by TLC, carrying out suction filtration on kieselguhr, carrying out reduced pressure concentration, dissolving ethyl acetate (15mL), extracting 1mol/L diluted hydrochloric acid (15mL), collecting a water layer, adjusting the saturated sodium carbonate solution to be alkaline, extracting ethyl acetate (15mL & gt2), drying anhydrous sodium sulfate, filtering, and carrying out reduced pressure concentration to obtain 42mg of brown oily substance. Yield: 82％。1H NMR(400MHz,Chloroform-d)δ6.61(dt,J＝9.1,1.2Hz,1H),6.44(d,J＝11.1Hz,1H),3.35(s,2H),2.90(t,J＝1.4Hz,2H),1.44(s,6H).
Step 2: preparation of 2- (2-aminopyridin-4-yl) -N- (6-fluoro-2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide
Figure BDA0002681115790000494
2- (2-Aminopyridin-4-yl) -N- (6-fluoro-2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared from 6-fluoro-2, 2-dimethyl-2, 3-dihydrobenzofuran-5-amine by a method similar to steps 3,4 in example 1.1H NMR(400MHz,DMSO-d6)δ9.69(s,1H),8.90(s,1H),8.18–8.02(m,1H),7.41(d,J＝8.0Hz,1H),7.05(d,J＝4.1Hz,2H),6.73(d,J＝10.9Hz,1H),6.36(s,2H),3.00(s,2H),1.43(s,6H).
Example 38
2- (2-Aminopyridin-4-yl) -N- (6-chloro-2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-38)
Figure BDA0002681115790000501
2- (2-Aminopyridin-4-yl) -N- (6-chloro-2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 37.1H NMR(400MHz,DMSO-d6)δ9.63(s,1H),8.92(s,1H),8.10(dd,J＝5.1,1.0Hz,1H),7.65(s,1H),7.05(dd,J＝6.5,1.3Hz,2H),6.94(s,1H),6.34(s,2H),3.04(s,2H),1.43(s,6H).
Example 39
2- (2-Aminopyridin-4-yl) -N- (6- (6-cyanopyridin-3-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-39)
Figure BDA0002681115790000502
2- (2-Aminopyridin-4-yl) -N- (6- (6-cyanopyridin-3-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,DMSO-d6)δ9.90(s,1H),8.77(s,1H),8.73(s,1H),8.08(d,J＝5.9Hz,1H),8.05(s,2H),7.33(s,1H),6.98(d,J＝4.2Hz,2H),6.85(s,1H),6.33(s,2H),3.10(s,2H),1.47(s,6H).
Example 40
2- (2-Aminopyridin-4-yl) -N- (6- (6-methoxypyridin-3-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-40)
Figure BDA0002681115790000503
2- (2-Aminopyridin-4-yl) -N- (6- (6-methoxypyridin-3-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,DMSO-d6)δ9.54(s,1H),8.76(s,1H),8.19(d,J＝2.4Hz,1H),8.07(d,J＝8.5Hz,1H),7.75(dd,J＝8.5,2.5Hz,1H),7.47(s,1H),6.97(t,J＝1.1Hz,1H),6.93(dd,J＝5.2,1.5Hz,1H),6.89–6.81(m,1H),6.72(s,1H),6.31(s,2H),3.86(s,3H),3.08(s,2H),1.46(s,6H).
EXAMPLE 41
2- (2-Aminopyridin-4-yl) -N- (6- (1-ethyl-1H-pyrazol-4-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-41)
Figure BDA0002681115790000511
2- (2-Aminopyridin-4-yl) -N- (6- (1-ethyl-1H-pyrazol-4-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,DMSO-d6)δ9.52(s,1H),8.87(s,1H),8.17–8.07(m,1H),8.02(s,1H),7.71(s,1H),7.51(s,1H),7.10–6.96(m,2H),6.83(s,1H),6.33(s,2H),4.13(q,J＝7.3Hz,2H),3.04(s,2H),1.44(s,6H),1.35(t,J＝7.3Hz,3H).
Example 42
(R) -2- (2-Aminopyridin-4-yl) -N- (2- (2-fluoro-3-hydroxy-3-methylbutyl) -1-oxo-6- (pyridin-4-yl) isoindolin-5-yl) oxazole-4-carboxylic acid amide (I-42)
Figure BDA0002681115790000512
Step 1: preparation of (R) -6-chloro-2- (2-fluoro-3-hydroxy-3-methylbutyl) -5-nitroisoindol-1-one
Figure BDA0002681115790000513
Methyl 2- (bromomethyl) -5-chloro-4-nitrobenzoate (prepared according to the method reported in WO 2013/079505) (975mg, 3.16mmol) and (R) -4-amino-3-fluoro-2-methylbutan-2-ol (prepared according to the method reported in WO 2015/103453) (459mg,3.79mmol) were dissolved in methanol (30mL) and placed in a sealed tube, triethylamine (527 μ 5) was added, 707 was reacted for 4 hours, concentrated under reduced pressure, dissolved by adding ethyl acetate (50mL), washed with 1mol/L HCl solution, the aqueous layer was extracted with ethyl acetate (15mL × 2), the organic layers were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 903mg of a pale yellow solid. Yield: 90 percent.1H NMR(400MHz,DMSO-d6)δ8.01(s,1H),7.91(s,1H),4.72(d,J＝17.9Hz,1H),4.65–4.41(m,2H),4.27(ddd,J＝37.0,15.1,2.2Hz,1H),3.64(ddd,J＝16.3,15.0,9.1Hz,1H),1.34(d,J＝1.8Hz,6H).
Step 2: preparation of (R) -2- (2-fluoro-3-hydroxy-3-methylbutyl) -5-nitro-6- (pyridin-4-yl) isoindol-1-one
Figure BDA0002681115790000521
(R) -6-chloro-2- (2-fluoro-3-hydroxy-3-methylbutyl) -5-nitroisoindol-1-one (300mg, 0.947mmol), pyridine-4-boronic acid (349mg, 2.84mmol) and cesium carbonate (617mg, 1.89mmol) were dissolved in dioxane/water (10mL/1mL), and [1,1' -bis (diphenyl) was added under an argon streamPhosphino) ferrocene]Palladium dichloride (69mg, 0.095mmol) is reacted at 95 ℃ overnight, after the TLC detection reaction is finished, the mixture is filtered by suction through diatomite, diluted by ethyl acetate (50mL), washed once by water and saturated saline, dried by anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and subjected to silica gel column chromatography (methanol/dichloromethane is 0-3% of gradient elution) to obtain 191mg of yellow solid. Yield: 56 percent.1H NMR(400MHz,Chloroform-d)δ8.83–8.63(m,2H),8.04(s,1H),7.90(s,1H),7.27(d,J＝1.7Hz,1H),4.81(d,J＝18.0Hz,1H),4.67–4.46(m,2H),4.30(ddd,J＝36.8,15.1,2.2Hz,1H),3.78–3.60(m,1H),1.35(d,J＝1.7Hz,6H).
And step 3: preparation of (R) -5-amino-2- (2-fluoro-3-hydroxy-3-methylbutyl) -6- (pyridin-4-yl) isoindolin-1-one
Figure BDA0002681115790000522
Dissolving (R) -2- (2-fluoro-3-hydroxy-3-methylbutyl) -5-nitro-6- (pyridin-4-yl) isoindol-1-one (188mg, 0.523mmol) in ethanol/water (10mL/3.3mL), adding reduced iron powder (175mg, 3.14mmol) and ammonium chloride (168mg, 3.14mmol), carrying out oil bath reaction at 50 ℃ for 1 hour, detecting the reaction by TLC, carrying out suction filtration on kieselguhr, carrying out reduced pressure concentration, dissolving the mixture in ethyl acetate (45mL), washing with saturated common salt water, drying with anhydrous sodium sulfate, filtering, and carrying out reduced pressure concentration to obtain 65mg of yellow solid. Yield: 38 percent.1H NMR(400MHz,Methanol-d4)δ8.68–8.53(m,2H),7.62–7.49(m,2H),7.46(s,1H),6.91(s,1H),4.64–4.36(m,3H),4.17–4.00(m,1H),3.68(td,J＝15.2,9.3Hz,1H),1.28(d,J＝1.7Hz,6H).
And 4, step 4: preparation of (R) -2- (2-aminopyridin-4-yl) -N- (2- (2-fluoro-3-hydroxy-3-methylbutyl) -1-oxo-6- (pyridin-4-yl) isoindolin-5-yl) oxazole-4-carboxamide
Figure BDA0002681115790000523
Can be prepared from (R) -5-amino-2- (2-fluoro-3-hydroxy-3-methylbutyl) -6- (pyridin-4-yl) isoindoline by methods analogous to Steps 3,4 of example 1-1-keto preparation of (R) -2- (2-aminopyridin-4-yl) -N- (2- (2-fluoro-3-hydroxy-3-methylbutyl) -1-oxo-6- (pyridin-4-yl) isoindolin-5-yl) oxazole-4-carboxamide.1H NMR(400MHz,DMSO-d6)δ9.91(s,1H),8.89(s,1H),8.81–8.60(m,2H),8.19(s,1H),8.09(d,J＝5.2Hz,1H),7.68(s,1H),7.57(d,J＝5.0Hz,2H),6.95(s,1H),6.90(d,J＝5.1Hz,1H),6.36(s,2H),4.95(s,1H),4.76–4.59(m,2H),4.48(dd,J＝49.5,9.1Hz,1H),3.99(dd,J＝38.6,14.9Hz,1H),3.74(q,J＝15.0,14.5Hz,1H),1.21(dd,J＝14.2,3.8Hz,6H).
Example 43
2- (2-Aminopyridin-4-yl) -N- (6- (5-fluoropyridin-3-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-43)
Figure BDA0002681115790000531
2- (2-Aminopyridin-4-yl) -N- (6- (5-fluoropyridin-3-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,DMSO-d6)δ8.68(s,1H),8.54(t,J＝2.2Hz,2H),8.29(s,1H),8.18(d,J＝5.3Hz,1H),8.04(s,1H),7.60–7.49(m,1H),7.10(dd,J＝5.4,1.4Hz,1H),7.04–6.95(m,1H),6.69(s,1H),4.75(s,2H),3.22–3.04(m,2H),1.53(s,6H).
Example 44
2- (2-Aminopyridin-4-yl) -N- (6- (3-fluoropyridin-4-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-44)
Figure BDA0002681115790000532
2- (2-Aminopyridin-4-yl) -N- (6- (3-fluoropyridin-4-yl) -2, 2-dimethyl-2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,DMSO-d6)δ8.64(s,1H),8.62(d,J＝1.6Hz,1H),8.54(d,J＝4.9Hz,1H),8.26(s,1H),8.18(d,J＝5.3Hz,1H),8.00(s,1H),7.37(t,J＝5.6Hz,1H),7.08(dd,J＝5.3,1.4Hz,1H),6.90(s,1H),6.70(s,1H),4.88(s,2H),3.12(s,2H),1.53(s,6H).
Example 45
4- (6- (2- (2-aminopyridin-4-yl) oxazole-4-carboxamido) -5-ethoxy-1-methyl-1H-benzo [ d ] imidazol-2-yl) -2, 2-dimethylbutyric acid methyl ester (I-45)
Figure BDA0002681115790000541
Step 1: preparation of 5-chloro-2-ethoxy-4-nitroaniline
Figure BDA0002681115790000542
2-amino-4-chloro-5-nitrophenol (2.0g,10.6mmol) was dissolved in N, N-dimethylformamide (40mL), potassium carbonate (2.2g,15.9mmol) and bromoethane (1.39g,12.7mmol) were added, reaction was carried out overnight at room temperature, water and ethyl acetate (150mL) were added, the organic layer was separated, water and saturated brine were washed once each, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 2.0g of a brown solid. Yield: 87 percent.1H NMR(400MHz,DMSO-d6)δ7.54(s,1H),6.73(s,1H),6.49(s,2H),4.10(q,J＝6.9Hz,2H),1.36(t,J＝6.9Hz,3H).
Step 2: preparation of tert-butyl (5-chloro-2-ethoxy-4-nitrophenyl) carbamate
Figure BDA0002681115790000543
5-chloro-2-ethoxy-4-nitroaniline (2.0g,9.23mmol) was dissolved in dichloromethane (30mL), di-tert-butyl dicarbonate (2.42g,11.1mmol) and 4-dimethylaminopyridine (11mg) were added, the reaction was refluxed for 3 hours, concentrated under reduced pressure, and subjected to silica gel column chromatography (elution with ethyl acetate/petroleum ether ═ 1%) to give 1.68g of a gold-yellow solid. Yield: 57 percent.1H NMR(400MHz,Chloroform-d)δ8.38(s,1H),7.51(s,1H),4.17(q,J＝7.0Hz,2H),1.53(d,J＝14.0Hz,12H).
And step 3: preparation of tert-butyl (2-ethoxy-5- (methylamino) -4-nitrophenyl) carbamate
Figure BDA0002681115790000544
Adding tert-butyl (5-chloro-2-ethoxy-4-nitrophenyl) carbamate (895mg,2.83mmol) and methylamine alcohol solution (40mL) into a sealed tube, reacting for 36 hours in a 70-tube, concentrating under reduced pressure, and performing silica gel column chromatography (gradient elution with ethyl acetate/petroleum ether being 5% -10%) to obtain 600mg of orange solid. Yield: 68 percent.1H NMR(400MHz,DMSO-d6)δ8.35(q,J＝4.9Hz,1H),8.20(s,1H),7.62(s,1H),7.48(s,1H),4.06(q,J＝6.9Hz,2H),2.93(d,J＝4.9Hz,3H),1.50(s,9H),1.36(t,J＝6.9Hz,3H).
And 4, step 4: preparation of tert-butyl (4-amino-2-ethoxy-5- (methylamino) phenyl) carbamate
Figure BDA0002681115790000551
Dissolving tert-butyl (2-ethoxy-5- (methylamino) -4-nitrophenyl) carbamate (565mg,1.81mmol) in methanol (25mL), adding 10% palladium carbon (200mg) and ammonium formate (686mg,10.9mmol), reacting at 60 ℃ for 2 hours, detecting by TLC after the reaction is finished, leaching by using kieselguhr, concentrating under reduced pressure, dissolving ethyl acetate (45mL), washing with saturated salt water, drying by using anhydrous sodium sulfate, filtering, concentrating under reduced pressure to obtain 485mg of blue-violet oil, and directly putting into the next reaction without further purification.1H NMR(400MHz,DMSO-d6)δ7.42(s,1H),6.63(s,1H),6.31(s,1H),4.37(s,2H),4.22(s,1H),3.85(s,2H),2.64(s,3H),1.43(s,9H),1.26(t,J＝6.9Hz,3H).
And 5: preparation of methyl 4- (6- ((tert-butoxycarbonyl) amino) -5-ethoxy-1-methyl-1H-benzo [ d ] imidazol-2-yl) -2, 2-dimethylbutyrate
Figure BDA0002681115790000552
Reacting (4-amino-2-ethoxy-5- (methylamino) phenyl) Tert-butyl carbamate (485mg,1.72mmol) and 5-methoxy-4, 4-dimethyl-5-oxopentanoic acid (315mg,1.81mmol) were dissolved in N, N-dimethylformamide (15mL), 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethyluronium hexafluorophosphate (983mg,2.59mmol) and N, N-diisopropylethylamine (712. mu.7, 4.31mmol) were added, the reaction was allowed to proceed overnight at room temperature, ethyl acetate (60mL) was diluted, water and saturated brine were washed once each, anhydrous sodium sulfate was dried, filtered, and concentrated under reduced pressure to give a residue which was dissolved in acetic acid (15mL), reacted at room temperature for 12 hours, water and ethyl acetate (60mL) were added, the organic layer was separated, saturated sodium bicarbonate and saturated brine were washed, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, silica gel column chromatography (gradient elution with ethyl acetate/petroleum ether 20% -40%) to obtain 391mg of white solid. Yield: 54 percent.1H NMR(400MHz,DMSO-d6)δ7.79(s,2H),7.14(s,1H),4.06(q,J＝7.1Hz,2H),3.65(d,J＝1.4Hz,3H),3.59(d,J＝1.4Hz,3H),2.74(t,J＝8.4Hz,2H),2.03–1.92(m,2H),1.48(d,J＝1.5Hz,9H),1.37(td,J＝6.9,1.4Hz,3H),1.21(d,J＝1.5Hz,6H).
Step 6: preparation of methyl 4- (6-amino-5-ethoxy-1-methyl-1H-benzo [ d ] imidazol-2-yl) -2, 2-dimethylbutyrate
Figure BDA0002681115790000553
Reacting 4- (6- ((tert-butoxycarbonyl) amino) -5-ethoxy-1-methyl-1H-benzo [ d]Imidazol-2-yl) -2, 2-dimethylbutanoic acid methyl ester (387mg,0.922mmol) was dissolved in dichloromethane (20mL), trifluoroacetic acid (5mL) was added to react at room temperature for 1 hour, concentrated under reduced pressure, ethyl acetate and saturated sodium bicarbonate solution were added to extract, the organic layer was separated, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 283mg of a brown solid. Yield: 96 percent.1H NMR(400MHz,DMSO-d6)δ6.93(s,1H),6.69–6.56(m,1H),4.65(s,2H),3.98(q,J＝6.9Hz,2H),3.59(d,J＝1.5Hz,3H),3.55(d,J＝1.4Hz,3H),2.66(t,J＝8.5Hz,2H),1.93(t,J＝8.4Hz,2H),1.42–1.30(m,3H),1.21(s,6H).
And 7: preparation of methyl 4- (6- (2- (2-aminopyridin-4-yl) oxazole-4-carboxamido) -5-ethoxy-1-methyl-1H-benzo [ d ] imidazol-2-yl) -2, 2-dimethylbutyrate
Figure BDA0002681115790000561
Can be prepared by a method similar to steps 3 and 4 of example 1 from 4- (6-amino-5-ethoxy-1-methyl-1H-benzo [ d]Preparation of 4- (6- (2- (2-aminopyridin-4-yl) oxazole-4-carboxamido) -5-ethoxy-1-methyl-1H-benzo [ d ] c-b-enz-2-yl) -2, 2-dimethylbutyric acid methyl ester]Imidazol-2-yl) -2, 2-dimethylbutyric acid methyl ester.1H NMR(400MHz,DMSO-d6)δ9.66(s,1H),8.96(s,1H),8.41(s,1H),8.12(d,J＝5.2Hz,1H),7.29(s,1H),7.03(d,J＝5.7Hz,2H),6.39(s,2H),4.20(q,J＝6.9Hz,2H),3.69(s,3H),3.60(s,3H),2.85–2.71(m,2H),2.08–1.94(m,2H),1.50(t,J＝6.9Hz,3H),1.22(s,6H).
Example 46
2- (2-Aminopyridin-4-yl) -N- (6-methoxy-2- (piperidin-4-yl) -2H-indazol-5-yl) oxazole-4-carboxamide trifluoroacetate salt (I-46)
Figure BDA0002681115790000562
Step 1: preparation of 2-fluoro-4-methoxy-5-nitrobenzaldehyde
Figure BDA0002681115790000563
The concentrated sulfuric acid (1.3mL) is bathed in ice salt to-15 ℃, 2-fluoro-4-methoxybenzaldehyde (3.0g,19.5mmol) is added, concentrated nitric acid is slowly dripped, the temperature of the reaction liquid is controlled below-10 ℃, the reaction is kept for 45 minutes, the reaction liquid is poured into crushed ice, a large amount of solid is separated out, the solid is filtered and dried in vacuum, and the solid obtained is pulped with petroleum ether, filtered and dried in vacuum to obtain 3.49g of yellow white solid. Yield: 90 percent.1H NMR(400MHz,Chloroform-d)δ10.21(s,1H),8.46(d,J＝7.2,1H),6.87(d,J＝11.5Hz,1H),4.06(s,3H).
Step 2: preparation of 2-azido-4-methoxy-5-nitrobenzaldehyde
Figure BDA0002681115790000564
2-fluoro-4-methoxy-5-nitrobenzaldehyde (2.0g,10.0mmol) was dissolved in dimethyl sulfoxide (10mL), sodium azide (1.31g,20.1mmol) was added to the solution to react at room temperature for 30 minutes, water and ethyl acetate (250mL) were added to the solution, and the ethyl acetate layer was washed once with water and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 2.13g of a brown solid. Yield: 95 percent.1H NMR(400MHz,Chloroform-d)δ10.19(s,1H),8.45(s,1H),6.80(s,1H),4.09(s,3H).
And step 3: preparation of 4- (6-methoxy-5-nitro-2H-indazol-2-yl) piperidine-1-carboxylic acid tert-butyl ester
Figure BDA0002681115790000571
2-azido-4-methoxy-5-nitrobenzaldehyde (1.0g,4.50mmol) was dissolved in dichloromethane (20mL), tert-butyl 4-aminopiperidine-1-carboxylate (902mg,4.50mmol) was dissolved in dichloromethane, and the reaction mixture was added to the solution
Figure BDA0002681115790000572
Molecular sieves (2g) reacted at room temperature for 1.5 hours and then supplemented with tert-butyl 4-aminopiperidine-1-carboxylate (270mg,1.35mmol) and
Figure BDA0002681115790000573
molecular sieve (2g), reacting for 14 hours at room temperature, filtering with diatomite, concentrating under reduced pressure, dissolving in anhydrous toluene (30mL), reacting for 1 hour at 120 ℃, concentrating under reduced pressure, and performing silica gel column chromatography (gradient elution with ethyl acetate/petroleum ether being 20-40%) to obtain 1.46g of yellow solid. Yield: 86 percent.1H NMR(400MHz,Chloroform-d)δ8.21(s,1H),8.07(s,1H),7.10(s,1H),4.52(tt,J＝11.6,4.0Hz,1H),4.33(s,2H),3.96(s,3H),2.94(s,2H),2.32–2.18(m,2H),2.08(qd,J＝12.2,4.6Hz,2H),1.48(s,9H).
And 4, step 4: preparation of 4- (5-amino-6-methoxy-2H-indazol-2-yl) piperidine-1-carboxylic acid tert-butyl ester
Figure BDA0002681115790000574
Dissolving 4- (6-methoxy-5-nitro-2H-indazol-2-yl) piperidine-1-carboxylic acid tert-butyl ester (233mg, 0.675mmol) in ethanol/water (12mL/4mL), adding reduced iron powder (226mg, 4.05mmol) and ammonium chloride (217mg, 4.05mmol), reacting in an oil bath at 50 ℃ for 6 hours, detecting the completion of the reaction by TLC, carrying out suction filtration on kieselguhr, carrying out reduced pressure concentration, dissolving ethyl acetate (60mL), washing with water and saturated saline water once, drying with anhydrous sodium sulfate, filtering, carrying out reduced pressure concentration, and directly putting into the next step for reaction without further purification.1H NMR(400MHz,Chloroform-d)δ7.61(d,J＝0.9Hz,1H),6.93(s,1H),6.75(s,1H),4.42(tt,J＝11.7,4.1Hz,1H),4.25(d,J＝31.8Hz,2H),3.91(s,3H),2.91(s,2H),2.19(d,J＝12.9Hz,2H),2.04(s,3H),1.48(s,9H).
And 5: preparation of 2- (2-aminopyridin-4-yl) -N- (6-methoxy-2- (piperidin-4-yl) -2H-indazol-5-yl) oxazole-4-carboxamide trifluoroacetate salt
Figure BDA0002681115790000575
2- (2-Aminopyridin-4-yl) -N- (6-methoxy-2- (piperidin-4-yl) -2H-indazol-5-yl) oxazole-4-carboxamide trifluoroacetate can be prepared from tert-butyl 4- (5-amino-6-methoxy-2H-indazol-2-yl) piperidine-1-carboxylate by a similar method as in example 2.1H NMR(400MHz,DMSO-d6)δ9.52(s,1H),9.11(s,1H),8.81(d,J＝11.1Hz,1H),8.57(s,2H),8.37(s,1H),8.14(d,J＝6.3Hz,1H),7.84(s,2H),7.41(s,1H),7.27(d,J＝6.2Hz,1H),7.17(s,1H),4.82–4.69(m,1H),4.01(s,3H),3.46(d,J＝12.6Hz,2H),3.22–3.06(m,2H),2.42–2.10(m,4H).
Example 47
2- (2-Aminopyridin-4-yl) -N- (2- (hydroxymethyl) -2-methyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-47)
Figure BDA0002681115790000581
Step 1: preparation of (6-chloro-2-methyl-5-nitro-2, 3-dihydrobenzofuran-2-yl) methanol
Figure BDA0002681115790000582
(6-chloro-2-methyl-5-nitro-2, 3-dihydrobenzofuran-2-yl) methanol can be prepared by a method similar to steps 1 to 5 in example 14.1H NMR(400MHz,Chloroform-d)δ7.85(d,J＝1.6Hz,1H),6.86(s,1H),3.76(d,J＝5.6Hz,1H),3.66(d,J＝7.3Hz,1H),3.40–3.31(m,1H),2.99–2.90(m,1H),1.84(dd,J＝7.3,5.7Hz,1H),1.48(s,3H),1.26(dd,J＝7.5,6.8Hz,1H).
Step 2: preparation of (2-methyl-5-nitro-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-2-yl) methanol
Figure BDA0002681115790000583
6-chloro-2-methyl-5-nitro-2, 3-dihydrobenzofuran-2-yl) methanol (58mg, 0.238mmol), pyridine-4-boronic acid (88mg, 0.714mmol) and cesium carbonate (155mg, 0.476mmol) were dissolved in dioxane/water (5mL/0.5mL) and [1,1' -bis (diphenylphosphino) ferrocene was added under argon flow]Palladium dichloride (17mg, 0.024mmol), reacted at 100 ℃ overnight, after completion of TLC detection, filtered through celite, diluted with ethyl acetate (30mL), washed once with water and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and chromatographed on silica gel (ethyl acetate/petroleum ether ═ 60% elution) to give 58mg of a pale yellow solid. Yield: 85 percent.1H NMR(400MHz,Chloroform-d)δ8.68–8.57(m,2H),7.94(t,J＝1.3Hz,1H),7.20–7.14(m,2H),6.54(s,1H),3.82(d,J＝12.0Hz,1H),3.67(d,J＝12.1Hz,1H),3.45(dd,J＝16.2,1.3Hz,1H),3.01(dd,J＝16.2,1.2Hz,1H),2.52(s,1H),1.51(s,3H).
And step 3: preparation of (5-amino-2-methyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-2-yl) methanol
Figure BDA0002681115790000584
Dissolving (2-methyl-5-nitro-6- (pyridine-4-yl) -2, 3-dihydrobenzofuran-2-yl) methanol (50mg, 0.175mmol) in ethanol/water (6mL/2mL), adding reduced iron powder (58mg, 1.05mmol) and ammonium chloride (56mg, 1.05mmol), performing oil bath reaction at 50 ℃ for 1 hour, detecting the reaction by TLC, performing suction filtration by using kieselguhr, concentrating under reduced pressure, dissolving ethyl acetate (20mL), washing with saturated salt water, drying with anhydrous sodium sulfate, filtering, and concentrating under reduced pressure to obtain 38mg of yellow solid. Yield: 85 percent.1H NMR(400MHz,Chloroform-d)δ8.70–8.60(m,2H),7.44–7.36(m,2H),6.65(s,1H),6.54(s,1H),3.75–3.61(m,2H),3.25(d,J＝16.2Hz,1H),2.88(d,J＝15.9Hz,1H),1.44(s,3H).
And 4, step 4: preparation of 2- (2-aminopyridin-4-yl) -N- (2- (hydroxymethyl) -2-methyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide
Figure BDA0002681115790000591
2- (2-Aminopyridin-4-yl) -N- (2- (hydroxymethyl) -2-methyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared from (5-amino-2-methyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-2-yl) methanol by a similar method as that described in example 1, Steps 3, 4.1H NMR(400MHz,DMSO-d6)δ9.70(s,1H),8.75(s,1H),8.65–8.51(m,2H),8.08(d,J＝5.3Hz,1H),7.47–7.40(m,2H),7.39(s,1H),7.03–6.91(m,2H),6.75(s,1H),6.33(s,2H),5.13(t,J＝5.8Hz,1H),3.49(qd,J＝11.5,5.8Hz,2H),3.28(s,1H),3.17(d,J＝5.3Hz,1H),2.92(d,J＝16.3Hz,1H),1.39(s,3H).
Example 48
2- (2-Aminopyrimidin-4-yl) -N- (2, 2-dimethyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-48)
Figure BDA0002681115790000592
2- (2-Aminopyrimidin-4-yl) -N- (2, 2-dimethyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method as in example 24.1H NMR(400MHz,Methanol-d4)δ8.56(s,1H),8.55–8.50(m,2H),8.45(d,J＝5.2Hz,1H),7.55–7.48(m,2H),7.45(s,1H),7.28(d,J＝5.1Hz,1H),6.77(s,1H),1.51(s,6H).
Example 49
2- (6-Aminopyrimidin-4-yl) -N- (2, 2-dimethyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-49)
Figure BDA0002681115790000601
2- (6-Aminopyrimidin-4-yl) -N- (2, 2-dimethyl-6- (pyridin-4-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method as in example 24.1H NMR(400MHz,Chloroform-d)δ8.82–8.69(m,3H),8.65(d,J＝1.2Hz,1H),8.34(s,1H),8.13(s,1H),7.42(d,J＝5.5Hz,2H),6.86(d,J＝1.2Hz,1H),6.70(s,1H),5.59(s,2H),3.12(d,J＝1.2Hz,2H),1.53(s,6H).
Example 50
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (pyrimidin-5-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxylic acid amide (I-50)
Figure BDA0002681115790000602
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (pyrimidin-5-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that described in example 24.1H NMR(400MHz,Chloroform-d)δ9.25(s,1H),8.85(s,2H),8.53(s,1H),8.28(s,1H),8.18(d,J＝5.4Hz,1H),7.87(s,1H),7.12(d,J＝5.9Hz,1H),7.01(s,1H),6.70(s,1H),4.72(s,2H),3.12(s,2H),1.53(s,6H).
Example 51
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (2-methylpyrimidin-5-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide (I-51)
Figure BDA0002681115790000603
2- (2-Aminopyridin-4-yl) -N- (2, 2-dimethyl-6- (2-methylpyrimidin-5-yl) -2, 3-dihydrobenzofuran-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that described in example 24.1H NMR(400MHz,DMSO-d6)δ9.84(s,1H),8.75(s,1H),8.70(s,2H),8.08(d,J＝5.3Hz,1H),7.32(s,1H),6.99(d,J＝4.7Hz,2H),6.83(s,1H),6.31(s,2H),3.09(s,2H),2.60(s,3H),1.47(s,6H).
Example 52
(R) -2- (2-Aminopyridin-4-yl) -N- (2- (2-fluoro-3-hydroxy-3-methylbutyl) -1-oxo-6- (pyridin-3-yl) isoindolin-5-yl) oxazole-4-carboxylic acid amide (I-52)
Figure BDA0002681115790000611
(R) -2- (2-Aminopyridin-4-yl) -N- (2- (2-fluoro-3-hydroxy-3-methylbutyl) -1-oxo-6- (pyridin-3-yl) isoindolin-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that described in example 24.1H NMR(400MHz,DMSO-d6)δ9.82(s,1H),8.88(s,1H),8.70(d,J＝2.3Hz,1H),8.67(dd,J＝4.8,1.6Hz,1H),8.20(s,1H),8.09(d,J＝5.2Hz,1H),7.96(dt,J＝7.8,2.0Hz,1H),7.67(s,1H),7.56(dd,J＝7.9,4.8Hz,1H),6.93(t,J＝1.1Hz,1H),6.89(dd,J＝5.3,1.4Hz,1H),6.34(s,2H),4.94(s,1H),4.75–4.59(m,2H),4.58–4.39(m,1H),4.08–3.91(m,1H),3.74(td,J＝15.6,9.4Hz,1H),1.19(dd,J＝4.5,1.6Hz,6H).
Example 53
(R) -2- (2-Aminopyridin-4-yl) -N- (2- (2-fluoro-3-hydroxy-3-methylbutyl) -6- (1-methyl-1H-pyrazol-4-yl) -1-oxoisoindol-5-yl) oxazole-4-carboxamide (I-53)
Figure BDA0002681115790000612
(R) -2- (2-Aminopyridin-4-yl) -N- (2- (2-fluoro-3-hydroxy-3-methylbutyl) -6- (1-methyl-1H-pyrazol-4-yl) -1-oxoisoindol-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,DMSO-d6)δ9.82(s,1H),8.99(s,1H),8.30(s,1H),8.20(s,1H),8.13(d,J＝5.5Hz,1H),7.85(s,1H),7.72(s,1H),7.00(d,J＝3.3Hz,2H),6.37(s,2H),4.93(s,1H),4.69–4.55(m,2H),4.47(dd,J＝49.8,9.8Hz,1H),4.09–3.86(m,4H),3.72(td,J＝15.4,9.2Hz,1H),1.19(d,J＝4.2Hz,6H).
Example 54
(R) -2- (2-Aminopyridin-4-yl) -N- (2- (2-fluoro-3-hydroxy-3-methylbutyl) -6- (6-methylpyridin-3-yl) -1-oxoisoindol-5-yl) oxazole-4-carboxylic acid amide (I-54)
Figure BDA0002681115790000621
(R) -2- (2-Aminopyridin-4-yl) -N- (2- (2-fluoro-3-hydroxy-3-methylbutyl) -6- (6-methylpyridin-3-yl) -1-oxoisoindol-5-yl) oxazole-4-carboxamide can be prepared by a similar method to that in example 24.1H NMR(400MHz,Methanol-d4)δ8.60(d,J＝2.3Hz,1H),8.57(s,1H),8.42(s,1H),8.04(d,J＝5.5Hz,1H),7.89(dd,J＝8.0,2.4Hz,1H),7.78(s,1H),7.52(d,J＝8.1Hz,1H),7.10(s,1H),7.02(dd,J＝5.5,1.6Hz,1H),4.78–4.65(m,2H),4.62–4.48(m,1H),4.16(dd,J＝37.9,14.8Hz,1H),3.80(td,J＝15.2,9.6Hz,1H),2.66(s,3H),1.31(s,6H).
Example 55
In this example, the following comparative compounds 1-4 were synthesized using a similar method.
Figure BDA0002681115790000622
Comparative Compound 1 is identical to Compound I-5, except that the ortho position to the pyridine N atom is a methyl group.1H NMR(400MHz,DMSO-d6)δ9.99(s,1H),9.02(s,1H),8.72(d,J＝5.1Hz,1H),8.19(s,1H),7.89–7.81(m,1H),7.75(dd,J＝5.2,1.6Hz,1H),6.76(s,1H),3.89(dd,J＝5.7,3.2Hz,4H),3.01(s,2H),2.84(t,J＝4.5Hz,4H),2.59(s,3H),1.41(s,6H).
Figure BDA0002681115790000623
Comparative Compound 2 is identical to Compound I-1, except that the pyridine ortho to the N atom is a methyl group.1H NMR(300MHz,DMSO-d6)δ10.01(s,1H),9.00(s,1H),8.68(d,J＝5.0Hz,1H),8.22(s,1H),7.80(d,J＝6.7Hz,2H),6.70(s,1H),4.63(t,J＝5.1Hz,1H),3.43(d,J＝5.3Hz,2H),3.00(s,2H),2.94(d,J＝11.1Hz,2H),2.68(t,J＝10.7Hz,2H),2.60(s,3H),1.91–1.77(m,2H),1.68–1.51(m,3H),1.41(s,6H).
Figure BDA0002681115790000624
Comparative compound 3 is identical to compound I-5 except that the pyridine N-atom is unsubstituted in the ortho position.1H NMR(400MHz,Chloroform-d)δ10.00(s,1H),8.95–8.76(m,2H),8.36(d,J＝25.4Hz,2H),8.01–7.86(m,2H),6.65(s,1H),4.01(t,J＝4.5Hz,4H),3.04(s,2H),2.93(dd,J＝5.5,3.3Hz,4H),1.49(s,6H).
Figure BDA0002681115790000631
Comparative Compound 4 is identical to Compound I-5, except that the amino group ortho to the pyridine N atom is substituted with an acetyl group.1H NMR(400MHz,DMSO-d6)δ10.81(s,1H),9.91(s,1H),9.01(s,1H),8.81(s,1H),8.57(dd,J＝5.1,0.8Hz,1H),8.20(s,1H),7.64(dd,J＝5.2,1.6Hz,1H),6.75(s,1H),3.87(dd,J＝5.9,3.2Hz,4H),3.01(s,2H),2.83(dd,J＝5.4,3.3Hz,4H),2.14(s,3H),1.41(s,6H)。
Test example 1
Inhibition of IRAK4 kinase Activity by Compounds
1. Test method
Using Z' -LYTETMThe kinase kit detects the kinase inhibitory activity of the compound. The method mainly comprises the following steps: compound solutions or solvent controls at different concentrations were added at 2.5 μ L/well in 384-well plates, with two replicate wells per concentration. The solvent control components were three: 0% phosphorylation, 0% inhibition and 100% phosphorylation. The enzyme and substrate were diluted with kinase buffer to a mixture of 1.5 ng/well and 2. mu.M/well, and the mixture was used in the test compound group and the 0% phosphorylated and 0% inhibited groups, with 5. mu.L per well. In the 100% phosphorylation group, 5. mu.L of the phosphorylated substrate solution was added to each well at a concentration of 2. mu.M/well. The test compound group and the 0% inhibition group were reacted by adding 2.5. mu.L of ATP (final concentration: 400. mu.M), and the 384-well plate was placed in an oven at 27 ℃ for 1 hour. Adding a secondary reaction reagent, adding 5 mu L of the secondary reaction reagent into each well, performing oven reaction at 27 ℃ for 1h, adding 5 mu L of the termination reagent to terminate the reaction, and reading the plate by using a fluorescence detection microplate reader (Synergy 2, BIOTEK) to respectively read the emitted light at the wavelengths of 445nM and 520nM under the excitation light of 400 nM. Inhibition rate and half inhibition amount IC50The ratio of the two emitted lights and the percentage of phosphorylation were determined.
2. Results of the experiment
As can be seen from Table 2, the compound of the invention has obvious inhibitory effect on IRAK4 kinase, wherein the inhibitory activity of compound I-5 on IRAK4 is obviously improved compared with that of comparative compounds 1,3 and 4, and the inhibitory activity of compound I-1 is obviously improved compared with that of comparative compound 2, which shows that the amino group at the ortho position of pyridine N atom has important effect on the inhibitory activity of IRAK4, and the compound activity is obviously deteriorated after the substituent group at the ortho position of pyridine N atom is replaced.
TABLE 2 Compound assay for IRAK4 kinase Activity
Figure BDA0002681115790000632
Figure BDA0002681115790000641
Test example 2
Growth inhibition assay of OCI-LY-10 cell line (containing MYD 88L 265P mutant cell) with Compounds
The Cell proliferation inhibition effect of the compound was examined by CCK-8(Cell Counting Kit-8) staining. The CCK-8 dyeing method comprises the following steps: OCI-LY-10 cells in logarithmic growth phase are inoculated into 96-well culture plates according to the growth rate of the cells at different concentrations in 90 mu L/well, after the cells grow overnight, compounds with different concentrations in 10 mu L/well are added, each concentration is provided with three multiple wells, and corresponding solvent control wells and blank control wells without cells are provided. Cells are placed in an incubator for compound action for 72h, then cell culture fluid is discarded, and 10 mu L/hole of CCK-8 reagent is added. And (3) putting the 96-well plate back to the cell culture box for reaction for 2h, and then placing the 96-well plate under a 450nm wavelength of an enzyme labeling instrument for determining the OD value. The inhibition rate of the compound on cell growth is calculated by the formula: the inhibition ratio%. Half the inhibition IC50The values were determined by fitting inhibition curves to the microplate reader with software using a four-parameter method. EXAMPLE 23(I-23) Compound IC50The value was 0.291. + -. 0.096. mu.M; EXAMPLE 24(I-24) Compound IC50The value was 1.586. + -. 0.280. mu.M.
Test example 3
Growth inhibition assay of compounds on U2932 cell line (without MYD 88L 265P mutant cells)
The Cell proliferation inhibition effect of the compound was examined by CCK-8(Cell Counting Kit-8) staining. The CCK-8 dyeing method comprises the following steps: OCI-LY-10 cells in logarithmic growth phase are inoculated into 96-well culture plates according to the growth rate of the cells at different concentrations in 90 mu L/well, after the cells grow overnight, compounds with different concentrations in 10 mu L/well are added, each concentration is provided with three multiple wells, and corresponding solvent control wells and blank control wells without cells are provided. Cells are placed in an incubator for compound action for 72h, then cell culture fluid is discarded, and 10 mu L/hole of CCK-8 reagent is added. And (3) putting the 96-well plate back to the cell culture box for reaction for 2h, and then placing the 96-well plate under a 450nm wavelength of an enzyme labeling instrument for determining the OD value. The inhibition rate of the compound on cell growth is calculated by the formula: the inhibition ratio%. Half the inhibition IC50The values are obtained by fitting an inhibition curve by a four-parameter method through software attached to a microplate readerAnd (5) obtaining the product. EXAMPLE 24(I-24) Compound IC50Values greater than 50 μ M indicate that the compound of example 24(I-24) has selective inhibition of MYD 88L 265P mutant diffuse large B-cell lymphoma.
Test example 4
Test of inhibition of tyrosine kinase Activity by Compounds
The inhibition of a series of kinase activities by the compounds was examined by ELISA. This series of kinases includes VEGFR-1, VEGFR-2, VEGFR-3, PDGFR-alpha, PDGFR-beta, RET, C-Kit, FLT3, EGFR, ErbB2, ErbB4, Src, Abl, EPH-A2, IGF1R, IR, FGFR1, FGFR2, FGFR3, FGFR4, BTK, FAK, CSF1R and ITK, all available from Europhins Inc.
The ELISA main steps are as follows: enzyme reaction substrate Poly (Glu, Tyr) was treated with potassium ion-free PBS4:1Reacting at 37 ℃ for 12-16h, coating the enzyme label plate, and then discarding the liquid in the hole. And washing the plate with T-PBS for three times, and drying the ELISA plate in a 37 ℃ oven for 1-2h for later use. A diluted ATP (final concentration 5mM) solution, compound or solvent control in reaction buffer was added to each well, followed by addition of kinase to initiate the reaction, followed by shaking reaction at 37 ℃ for 1 h. The plate was washed three times with T-PBS and reacted for 0.5h at 37 ℃ with the addition of the antibody PY 99. After washing the plate with T-PBS for three times, adding horseradish peroxidase labeled goat anti-mouse IgG for shaking table reaction at 37 ℃ for 0.5 h. Discarding the liquid in the hole, washing the plate again, adding 100 mu L/hole of OPD color development liquid, and reacting for 1-10min at 25 ℃ in a dark place. Addition of H2SO4The reaction is stopped, and the reading is carried out by using a microplate reader with adjustable wavelength, wherein the wavelength is 490 nm. The inhibition rate calculation formula is as follows: inhibition [% 1- (compound OD value-no enzyme control OD value)/(negative control OD value-no enzyme control OD value) ]]×100％。IC50The values were determined by fitting inhibition curves with a four-parameter method using a microplate reader with software.
As can be seen from Table 3, the compounds of example 23(I-23) and example (I-24) have better selectivity on tyrosine kinase spectrum and simultaneously have inhibitory effect on VEGFR-3, RET, FLT3 and ErbB 2.
TABLE 3 detection of the Activity of Compounds on the series of tyrosine kinases
Figure BDA0002681115790000661
Test example 5
The compounds FLT3 kinase and FLT3-ITD, FLT3D835YMutant kinase activity assay
The inhibition of the kinase activity by the compound was calculated by measuring the ability of the kinase to phosphorylate substrates by Enzyme-Linked Immunosorbent Assay (ELISA). The kinase adopts Flt-3 and Flt-3ITDAnd Flt-3D835Y(available from Eurofins). The ELISA main steps are as follows: enzyme reaction substrate Poly (Glu, Tyr)4:1Diluting to 2.5 mu g/hole with PBS without potassium ions, reacting at 37 ℃ for 12-16h, and coating an enzyme label plate for later use. Reaction buffer (50mM HEPES pH 7.4, 20mM MgCl) was added to each well2，0.1mM MnCl2，0.2mM Na3VO41mM DTT) was added to a compound or solvent control, followed by addition of kinase to initiate the reaction, followed by shaking at 37 ℃ for 1 h. The plate was washed three times with T-PBS and shaken at 37 ℃ for 0.5h with the addition of the PY99 antibody. After washing the plate with T-PBS, horseradish peroxidase-labeled goat anti-mouse IgG was added and the mixture was subjected to shake reaction at 37 ℃ for 0.5 hour. After washing the plate again, 0.03% H was added2O2And 2mg/mL OPD developing solution, and reacting for 1-10min at 25 ℃ in a dark place. 2M H was added2SO4The reaction was stopped and read using a tunable wavelength microplate reader (SpectraMax Plus384, Molecular Devices) at a wavelength of 490 nm. IC (integrated circuit)50Values were obtained from the inhibition curves.
As can be seen from the results in Table 4, example 24(I-24) is shown for FLT3 kinase, FLT3-ITD and FLT3D835YThe mutant kinases all have high inhibitory activity.
TABLE 4 Compounds FLT3 kinase and FLT3-ITD, FLT3D835YMutant kinase activity assay
Compound (I) Flt-3 IC50(nM) Flt-3 ITD IC50(nM) FLT3D835Y IC50(nM)
I-24 1.7±0.1 3.5±2.1 0.3±0.1
Quizartinib 3.8±0.8 2.6±0.4 278.0±68.4
Test example 6
Effect of Compounds on cell proliferation of leukemia cell lines
1. Experimental Material
Leukemia cell lines: acute myeloid leukemia cell line MV4-11 (expression FLT3-ITD mutant gene)
MOLM-3 (expressing FLT3-ITD mutant gene and wild type FLT3 gene)
2. Test method
Leukemia cells in logarithmic growth phase were seeded at appropriate density into 96-well culture plates at 90. mu.L per well, after overnight incubation, compounds at different concentrations (1. mu.M for initial concentration, 5-fold dilution) were added for 72hr, and a solvent control group (negative control) was set. After the compound acts on cells for 72 hours, the influence of the compound on cell proliferation is detected by a CCK-8 cell counting kit (Shanghai Liji Biotech), 10 mu L of CCK-8 reagent is added into each hole, the hole is placed in an incubator at 37 ℃ for 2 to 4 hours, then a SpectraMax 190 reading is carried out by a full-wavelength micro-hole plate enzyme-labeling instrument, and the measurement wavelength is 450 nm. By usingThe inhibition (%) of tumor cell growth by the compound was calculated by the following formula: the inhibition ratio (%) (OD negative control well-OD administration well)/OD negative control well × 100%. IC (integrated circuit)50The values were determined by regression with a four parameter method using a microplate reader random plus software.
3. Results of the experiment
As can be seen from the results in Table 5, example 24(I-24) showed significant inhibition of cell proliferation activity of acute myeloid leukemia cell strain MV4-11 and MOLM-3 expressing the FLT3-ITD mutation.
TABLE 5 Effect of Compounds on MV4-11 and MOLM-3 cell proliferation
Compound (I) MV4-11 IC50(nM) MOLM-13IC50(nM)
I-24 <3.9 6.0±1.9
Quizartinib 4.8±2.7 9.8±4.0
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 (15)

1. A compound of formula (I), a stereoisomer, a geometric isomer, a tautomer thereof, a pharmaceutically acceptable salt thereof, a prodrug thereof, a hydrate thereof, or a solvate thereof:
Figure FDA0002681115780000011
wherein,
x, Y, Z or W is independently selected from CH or N; and when X, Y, Z or W is CH, the H atom may be substituted with a substituent selected from the group consisting of: halogen, C1-C3 alkyl, C2-C6 acyl, C1-C3 alkoxy, trifluoromethoxy, trifluoroethoxy;
ring a is a 3-8 membered saturated heterocyclic ring (including monocyclic, fused or spiro ring) containing 1-2 heteroatoms selected from N, O and S, a 5-8 membered aromatic heterocyclic ring containing 1-2 heteroatoms selected from N, O and S, said saturated heterocyclic ring being optionally substituted with one or more halogen, oxo, carboxyl, cyano, hydroxyl, substituted or unsubstituted 5-6 membered heterocyclic ring, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C8 cycloalkyl, said substitution being substituted with one or more substituents selected from the group consisting of: halogen, C1-C3 alkyl, hydroxy, C2-C6 acyl, 3-8 membered saturated or partially saturated heterocyclyl;
R1is selected from-NR2R3Substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C1-C6 alkoxy, substituted or unsubstituted C3-C10 cycloalkyl, substituted or unsubstituted 6-10 membered aryl, substituted or unsubstituted 5-10 membered heteroaryl, substituted or unsubstituted 3-11 membered saturated or partially saturated heterocyclic group, the substituents being represented by RaRepresents;
each R isaEach independently selected from halogen, oxo, - (C0-C3 alkyl) -CN, - (C0-C3 alkyl) -OH, - (C0-C3 alkyl) -COOH, - (C0-C3 alkyl) C (═ O) OR5- (C0-C3 alkyl) C (═ O) R5- (C0-C3 alkyl) NR5R6-C (═ O) (C1-C3 alkyl), -C (═ O) NR (C0-C3 alkyl)5R6- (C0-C3 alkyl) S (═ O) NR5R6- (C0-C3 alkyl) S (═ O)2NR5R6- (C0-C3 alkyl) S (═ O)2(C1-C3 alkyl), - (C0-C3 alkyl) OP (═ O) (OC1-C3 alkyl)2Substituted or unsubstituted C1-C3 alkyl, substituted or unsubstituted C3-C8 cycloalkyl, substituted or unsubstituted C1-C6 alkoxy, substituted or unsubstituted C1-C6 alkylthio, said substitution being with one or more halogens;
R2and R3Each independently selected from the group consisting of a hydrogen atom, a substituted or unsubstituted C1-C3 alkyl group, a substituted or unsubstituted C3-C10 cycloalkyl group, - (C0-C3 alkyl) -6-10 membered aryl group, - (C0-C3 alkyl) -5-10 membered heteroaryl group, - (C0-C3 alkyl) -saturated or partially saturated 4-10 membered heterocyclyl group, said substitution being by halogen, hydroxy, amino, cyano or amido;
or R2And R3May form, together with the nitrogen atom to which they are attached, a 4-to 8-membered heterocyclic ring optionally substituted with one or more RaSubstitution;
R5、R6independently selected from hydrogen atom, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C10 cycloalkyl, and substituted or unsubstituted 4-6 membered heterocycle, wherein the substitution refers to substitution by one or more C1-C3 alkyl, hydroxyl, halogen, carboxylic acid group, C2-C6 carboxylic acid ester group.
2. The compound, stereoisomer, geometric isomer, tautomer, pharmaceutically acceptable salt thereof, prodrug thereof, hydrate thereof, or solvate thereof according to claim 1, wherein X, Y, Z or W are each independently selected from CH or N;
ring a is a 3-8 membered saturated heterocyclic ring (including monocyclic, fused or spiro) containing 1-2 heteroatoms selected from N, O and S, optionally substituted with one or more halogen, oxo, carboxy, cyano, hydroxy, substituted or unsubstituted 5-6 membered heterocyclic ring, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C8 cycloalkyl, said substitution being substituted with one or more halogen, C1-C3 alkyl or hydroxy;
R1is selected from-NR2R3Substituted or notSubstituted C1-C6 alkyl, substituted or unsubstituted C3-C10 cycloalkyl, substituted or unsubstituted 6-10 membered aryl, substituted or unsubstituted 5-10 membered heteroaryl, substituted or unsubstituted 3-11 membered saturated or partially saturated heterocyclyl, said substituents being represented by RaRepresents;
each R isaEach independently selected from halogen, oxo, - (C0-C3 alkyl) -CN, - (C0-C3 alkyl) -OH, - (C0-C3 alkyl) -COOH, - (C0-C3 alkyl) C (═ O) OR5- (C0-C3 alkyl) C (═ O) R5- (C0-C3 alkyl) NR5R6-C (═ O) (C1-C3 alkyl), -C (═ O) NR (C0-C3 alkyl)5R6- (C0-C3 alkyl) S (═ O) NR5R6- (C0-C3 alkyl) S (═ O)2NR5R6- (C0-C3 alkyl) S (═ O)2(C1-C3 alkyl), - (C0-C3 alkyl) OP (═ O) (OC1-C3 alkyl)2Substituted or unsubstituted C1-C3 alkyl, substituted or unsubstituted C3-C8 cycloalkyl, substituted or unsubstituted C1-C6 alkoxy, substituted or unsubstituted C1-C6 alkylthio, said substitution being with one or more halogens;
R2and R3Each independently selected from the group consisting of a hydrogen atom, a substituted or unsubstituted C1-C3 alkyl group, a substituted or unsubstituted C3-C10 cycloalkyl group, - (C0-C3 alkyl) -6-10 membered aryl group, - (C0-C3 alkyl) -5-10 membered heteroaryl group, - (C0-C3 alkyl) -saturated or partially saturated 4-10 membered heterocyclyl group, said substitution being by halogen, hydroxy, amino, cyano or amido;
or R2And R3May form, together with the nitrogen atom to which they are attached, a 4-to 8-membered heterocyclic ring optionally substituted with one or more RaSubstitution;
R5、R6each independently selected from hydrogen atom, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C10 cycloalkyl, and substituted or unsubstituted 4-6 membered heterocycle, wherein the substitution refers to substitution by one or more C1-C3 alkyl, hydroxyl and halogen.
3. The compound of claim 1, stereoisomers, geometric isomers, tautomers, pharmaceutically acceptable salts thereof, pro-forms thereof, and pharmaceutically acceptable salts thereofA drug, a hydrate or solvate thereof, characterized in that R2And R3Together with the nitrogen atom to which they are attached form a 4-7 membered heterocyclic ring, optionally substituted with one or more RbSubstitution;
Rbindependently selected from halogen, oxo, cyano, hydroxy, - (C0-C3 alkyl) C (═ O) NHR5R6- (C0-C3 alkyl) -NR5R6Substituted or unsubstituted C1-C3 alkyl, said substitution being by one or more of halogen, oxo, cyano, hydroxy.
4. The compound, its stereoisomers, geometric isomers, tautomers, its pharmaceutically acceptable salts, its prodrugs, its hydrates or solvates according to claim 1, wherein R is1Is a substituted or unsubstituted 6-10 membered aryl, substituted or unsubstituted 5-10 membered heteroaryl, and said aryl or heteroaryl is optionally substituted with one or more RcSubstitution;
Rcindependently selected from halogen, cyano, hydroxy, -COOH, -C (═ O) OR5、-C(＝O)R5-C (═ O) (C1-C3 alkyl), -C0-C3 alkyl) NR5R6- (C0-C3 alkyl) C (═ O) NR5R6- (C0-C3 alkyl) S (═ O) NR5R6- (C0-C3 alkyl) S (═ O)2NR5R6、-S(＝O)2(C1-C3 alkyl), - (C0-C3 alkyl) OP (═ O) (OC1-C3 alkyl)2Substituted or unsubstituted C1-C3 alkyl, substituted or unsubstituted C1-C6 alkoxy, substituted or unsubstituted C1-C6 alkylthio, said substitution being with one or more halogens or hydroxy groups.
5. The compound, its stereoisomers, geometric isomers, tautomers, its pharmaceutically acceptable salts, its prodrugs, its hydrates or solvates according to claim 1, wherein R is1Is a substituted or unsubstituted 5-6 membered aromatic heterocycle optionally substituted with one or more RcAnd (4) substitution.
6. The compound, its stereoisomers, geometric isomers, tautomers, its pharmaceutically acceptable salts, its prodrugs, its hydrates or solvates according to claim 1, wherein ring a is selected from the group consisting of:
Figure FDA0002681115780000031
R4selected from the group consisting of a hydrogen atom, a substituted or unsubstituted C1-C6 alkyl group, a substituted or unsubstituted C3-C10 cycloalkyl group, said substitution being by one or more halogens.
7. The compound, its stereoisomers, geometric isomers, tautomers, its pharmaceutically acceptable salts, its prodrugs, its hydrates or solvates according to claim 1, characterized in that:
Figure FDA0002681115780000032
selected from the group consisting of:
Figure FDA0002681115780000033
8. the compound, its stereoisomers, geometric isomers, tautomers, pharmaceutically acceptable salts thereof, its prodrugs, its hydrates or solvates according to claim 1, wherein said compound is selected from the group consisting of:
Figure FDA0002681115780000034
Figure FDA0002681115780000041
Figure FDA0002681115780000051
Figure FDA0002681115780000061
Figure FDA0002681115780000071
Figure FDA0002681115780000081
Figure FDA0002681115780000091
9. a pharmaceutical composition comprising the following components:
1) a therapeutically effective amount of one or more compounds of claim 1, stereoisomers, geometric isomers, tautomers, pharmaceutically acceptable salts thereof, prodrugs thereof, hydrates thereof, or solvates thereof; and
2) a pharmaceutically acceptable carrier or excipient.
10. The pharmaceutical composition of claim 9, further comprising one or more active agents selected from the group consisting of: immunosuppressants, glucocorticoids, non-steroidal anti-inflammatory drugs, vinca alkaloids, paclitaxel, DNA damaging agents, Bcl-2 inhibitors, BTK inhibitors, JAK inhibitors, Hsp90 inhibitors, ALK inhibitors, FLT3 inhibitors, PI3K inhibitors, and SYK inhibitors.
11. Use of a compound according to claim 1, its stereoisomers, geometric isomers, tautomers, pharmaceutically acceptable salts, prodrugs, hydrates or solvates thereof, for the preparation of a pharmaceutical composition for the prevention or treatment of indications mediated by FLT3, FLT3-ITD or other FLT3 mutations.
12. The use of claim 11, wherein the indication is selected from the group consisting of: myelodysplastic syndrome, acute myeloid leukemia, neurofibroma type I, multiple myeloma, glioblastoma, non-small cell lung cancer, liver cancer, hepatocellular carcinoma, cervical cancer, lymphoma, bone metastasis cancer, hormone refractory prostate cancer, hormone dependent prostate cancer, thyroid tumor, medullary thyroid cancer, mesothelioma, glioblastoma, anaplastic bone metastasis cancer, Merkel cell cancer, genitourinary tract tumor, Mercker cell cancer, bladder cancer, papillary thyroid cancer, breast cancer, soft tissue sarcoma, glioma, neuroendocrine tumor, renal cell carcinoma, late stage solid tumor, undifferentiated astrocytoma, gastrointestinal stromal tumor, von Willebrand syndrome, small cell lung cancer, pancreatic endocrine cancer, central nervous system tumor, metastatic renal cancer, endometrioid carcinoma, endometrioid adenocarcinoma, colon cancer, bladder cancer, lung cancer, colorectal cancer, ovarian cancer, rhabdomyosarcoma, melanoma, retinoblastoma, tumors of the central and peripheral nervous system, acute leukemia, chronic leukemia, bile duct cancer, bronchial cancer, esophageal cancer, testicular cancer, skin cancer, oral cancer, neuroblastoma, anaplastic large cell lymphoma.
13. Use of a compound according to claim 1, a stereoisomer, a geometric isomer, a tautomer, a pharmaceutically acceptable salt thereof, a prodrug thereof, a hydrate or a solvate thereof, or a pharmaceutical composition according to claim 9, for the preparation of a medicament for the prophylaxis and/or treatment of IRAK 4-mediated diseases.
14. Use of a compound according to claim 1, its stereoisomers, geometric isomers, tautomers, pharmaceutically acceptable salts, prodrugs, hydrates or solvates thereof, for the preparation of a pharmaceutical composition for the treatment or prevention of indications co-mediated by IRAK4 and FLT3, FLT3-ITD or other FLT3 mutations.
15. The use of claim 14, wherein the indication is selected from the group consisting of: myelodysplastic syndrome, acute myeloid leukemia, cancer.
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