CN112830935B - Crystal form containing tricyclic derivative free alkali and pharmaceutical composition thereof - Google Patents

Crystal form containing tricyclic derivative free alkali and pharmaceutical composition thereof Download PDF

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CN112830935B
CN112830935B CN202011293975.XA CN202011293975A CN112830935B CN 112830935 B CN112830935 B CN 112830935B CN 202011293975 A CN202011293975 A CN 202011293975A CN 112830935 B CN112830935 B CN 112830935B
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imidazo
difluoromethyl
dihydrobenzo
degrees
oxazepin
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CN112830935A (en
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詹小兰
呙临松
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Jiangsu Hansoh Pharmaceutical Group Co Ltd
Shanghai Hansoh Biomedical Co Ltd
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Jiangsu Hansoh Pharmaceutical Group Co Ltd
Shanghai Hansoh Biomedical Co Ltd
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    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D498/00—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and oxygen atoms as the only ring hetero atoms
    • C07D498/02—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and oxygen atoms as the only ring hetero atoms in which the condensed system contains two hetero rings
    • C07D498/04—Ortho-condensed systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
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    • A61P25/00—Drugs for disorders of the nervous system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
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    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
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    • A—HUMAN NECESSITIES
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    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D498/00—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and oxygen atoms as the only ring hetero atoms
    • C07D498/12—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and oxygen atoms as the only ring hetero atoms in which the condensed system contains three hetero rings
    • C07D498/14—Ortho-condensed systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D513/00—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for in groups C07D463/00, C07D477/00 or C07D499/00 - C07D507/00
    • C07D513/02—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for in groups C07D463/00, C07D477/00 or C07D499/00 - C07D507/00 in which the condensed system contains two hetero rings
    • C07D513/04—Ortho-condensed systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07B—GENERAL METHODS OF ORGANIC CHEMISTRY; APPARATUS THEREFOR
    • C07B2200/00—Indexing scheme relating to specific properties of organic compounds
    • C07B2200/07—Optical isomers

Abstract

The invention relates to a crystal form containing tricyclic derivative free alkali and a pharmaceutical composition thereof. In particular to a crystal form of a compound free alkali with a general formula (I), a preparation method, a pharmaceutical composition containing the crystal form with a therapeutically effective amount and application in preparing medicines for treating related diseases mediated by PI 3K.

Description

Crystal form containing tricyclic derivative free alkali and pharmaceutical composition thereof
Technical Field
The invention belongs to the field of medicine synthesis, and particularly relates to a crystal form containing a tricyclic derivative free alkali, and a preparation method and application thereof.
Background
The phosphatidylinositol 3 kinase (PI 3K) protein family is divided into four major classes I, II, III and IV, and is involved in the regulation of various cellular functions such as cell growth, proliferation, differentiation, survival and glucose metabolism. The four types of PI3K proteins have different structures and functions, wherein the most widely studied type I PI3K is divided into four subtypes of PI3K alpha, PI3K beta, PI3K delta and PI3K gamma, and the PI3K alpha is subjected to activation mutation and amplification in various tumors and is closely related to the occurrence and development of the tumors. It has been reported that PI3K beta activates platelets and plays an important role in the development of diseases such as thrombosis. Pi3kδ and pi3kγ are mainly expressed in the blood system, are closely related to the immune system and the occurrence of inflammation, and pi3kγ is closely related to blood pressure stabilization and smooth muscle contraction.
PI3K alpha is a driving factor for the development of tumorigenesis by activating mutations and amplifications in a variety of tumors. PI3kα is a heterodimer consisting of a p110 catalytic subunit and a p85 regulatory subunit. PI3kα is activated by Receptor Tyrosine Kinases (RTKs) and G protein-coupled receptors (GPCRs), which upon activation catalyze the production of phosphatidylinositol 3 phosphate (PIP 3) from phosphatidylinositol 2 phosphate (PIP 2), PIP3 further activates protein kinase B (PKB, also known as AKT) and its downstream signaling pathway. Various cell growth factors such as Epidermal Growth Factor (EGF), fibroblast Growth Factor (FGF), vascular Endothelial Growth Factor (VEGF), hepatocyte Growth Factor (HGF) and insulin activate PI3K alpha, thereby activating the downstream proliferation signaling pathway of cells, and abnormal activation of PI3K alpha can lead to rapid proliferation of cells, thereby causing tumorigenesis.
PI3kα has been an important target for development of tumor drugs, but most of the compounds are broad-spectrum inhibitors of PI3Ks, so that clinical research has a large side effect, and development of PI3Ks inhibitors is severely limited. Studies have now established that the majority of adverse effects of broad-spectrum inhibitors of PI3Ks are due to inhibition of PI3kβ, pi3kδ and pi3kγ subtypes, with pi3kδ inhibition playing an important role in thrombocytopenia and thrombotic adverse mechanisms, pi3kδ inhibition leading to immune system abnormalities, autoimmune and infectious toxicities such as pneumonia, hepatitis and diarrhea/enteritis being closely related to inhibition of pi3kδ targets, pi3kγ being closely related to blood pressure stabilization and smooth muscle contraction being the primary targets leading to hypertensive adverse effects. Therefore, the development of highly active and highly selective PI3K alpha inhibitors can further improve the anti-tumor effect of PI3K alpha inhibitors and reduce or eliminate the serious side effects of inhibiting various inflammations, thrombocytopenia, hypertension and the like caused by other subtypes.
Pi3K alpha selective inhibitor BYL-719 developed by Nohua is currently in clinical phase III research, pi3K alpha selective inhibitor MLN1117 developed by Wuta-tsu enters phase II clinical research, and selective inhibitor GDC-0077 developed by Gentec is also in clinical phase I research.
International applications WO2010029082A1 and WO2011022439A1 report compounds related to PI3K alpha selective inhibitors, but later studies indicate that the cell activity of the compounds is not high, and the clinical anti-tumor effect is affected. Therefore, there is an urgent need to develop PI3kα selective inhibitors with high activity and high selectivity. The PI3K alpha selective inhibitor can be used for treating various multiple tumors with PI3K alpha activating mutation or amplification, and has great clinical application value.
PCT patents (application numbers: PCT/CN2019/088788 and PCT/CN 2019/104558) of Jiangsu haoshen pharmaceutical industry group limited disclose a series of structures containing tricyclic derivative inhibitors, and in subsequent research and development, the free alkali of the above substances is comprehensively studied in an effort to obtain the most suitable crystal form in order to facilitate the easy handling, filtration and drying of the product, convenient storage, long-term stability of the product, high bioavailability, and the like.
Disclosure of Invention
All that is referred to in patent PCT/CN2019/088788 and PCT/CN2019/104558 are incorporated by reference into the present invention.
The invention aims to provide a crystal form of a compound shown in a general formula (I), and the structure of the crystal form is shown in the general formula (I):
wherein:
w is selected from-O-, -S-or-NR aa -；
G is selected from-O-, -S-, -CR aa R bb -or-NR aa -；
R 1 And R is 1 ' selected from hydrogen, deuterium, cyano, halogen, nitro, amino, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, C 1-6 Hydroxyalkyl-cyano-substituted C 1-6 Alkyl, C 3-8 Cycloalkyl, 3-8 membered heterocyclyl, C 6-10 Aryl, 5-10 membered heteroaryl, - (CH) 2 ) n R cc 、-(CH 2 ) n OR cc or-CR aa R bb OR cc ；
Or, R 1 And R is 1 ' join to form a C 3-8 Cycloalkyl or 3-8 membered heterocyclyl, wherein said C 3-8 Cycloalkyl or 3-8 membered heterocyclyl, optionally further substituted by hydrogen, deuterium, cyano, halogen, nitro, amino, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, C 1-6 Hydroxyalkyl, C 3-8 Cycloalkyl, 3-8 membered heterocyclyl, C 6-10 One or more substituents in aryl and 5-10 membered heteroaryl;
R 2 selected from hydrogen, deuterium, cyano, halogenNitro, amino, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, C 1-6 Hydroxyalkyl, C 3-8 Cycloalkyl, 3-8 membered heterocyclyl, C 6-10 Aryl or 5-to 10-membered heteroaryl or- (CH) 2 ) n OR cc ；
Or, any two of R's attached 2 Form a C 3-8 Cycloalkyl or 3-8 membered heterocyclyl, wherein said C 3-8 Cycloalkyl or 3-8 membered heterocyclyl, optionally further substituted by hydrogen, deuterium, cyano, halogen, nitro, amino, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, C 1-6 Hydroxyalkyl, C 3-8 Cycloalkyl, 3-8 membered heterocyclyl, C 6-10 One or more substituents in aryl and 5-10 membered heteroaryl;
R 3 and R is 3 ' selected from hydrogen, deuterium, cyano, halogen, nitro, amino, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, C 1-6 Hydroxyalkyl, C 3-8 Cycloalkyl, 3-8 membered heterocyclyl, C 6-10 Aryl or 5-10 membered heteroaryl;
or, R 3 And R is 3 ' linking to form an oxo group, C 3-8 Cycloalkyl or 3-8 membered heterocyclyl, wherein said C 3-8 Cycloalkyl or 3-8 membered heterocyclyl, optionally further substituted by hydrogen, deuterium, cyano, halogen, nitro, amino, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, C 1-6 Hydroxyalkyl, C 3-8 Cycloalkyl, 3-8 membered heterocyclyl, C 6-10 One or more substituents in aryl and 5-10 membered heteroaryl;
R 4 selected from hydrogen, deuterium, cyano, halogen, nitro, amino, and C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, C 1-6 Hydroxyalkyl, C 3-8 Cycloalkyl, 3-8 membered heterocyclyl, C 6-10 Aryl or 5-10 membered heteroaryl;
R 5 selected from hydrogen, deuterium, C 1-6 Alkyl or C 1-6 A haloalkyl group;
or, R 1 Or R is 1 ' and R 5 To form a 3-8 membered heterocyclic group, wherein said 3-8 membered heterocyclic group is optionally further substituted with hydrogen, deuterium, cyano, halogen, nitro, amino, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, C 1-6 Hydroxyalkyl, C 3-8 Cycloalkyl, 3-8 membered heterocyclyl, C 6-10 One or more substituents in aryl and 5-10 membered heteroaryl;
R aa 、R bb and R is cc Each independently selected from hydrogen, deuterium, cyano, halogen, nitro, amino, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, C 1-6 Hydroxyalkyl, C 3-8 Cycloalkyl, 3-8 membered heterocyclyl, C 6-10 Aryl or 5-10 membered heteroaryl;
n is 0, 1, 2 or 3; and is also provided with
x is 0, 1, 2 or 3.
In a further preferred embodiment of the invention, R 1 And R is 1 ' selected from hydrogen, C 1-6 Alkyl, C 1-6 Hydroxy-substituted alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, 3-8 membered heterocyclyl, - (CH) 2 ) n OR cc or-CR aa R bb OR cc Preferably hydrogen, C 1-3 Alkyl, C 1-3 Hydroxyalkyl, C 1-3 Haloalkyl, C 1-3 Alkoxy, 3-6 membered heterocyclyl, - (CH) 2 ) n OR cc or-CR aa R bb OR cc More preferably hydrogen, methyl, ethyl, propyl, isopropyl, methoxy, ethoxy, propoxy, fluoromethyl, fluoroethyl, fluoropropyl, chloromethyl, chloroethyl, chloropropyl, hydroxymethyl, hydroxyethyl, hydroxypropyl, oxetanyl, aziridinyl, azetidinyl, - (CH) 2 )OCH 3 、-(CH 2 ) 2 OCH 3 、-CH(CH 3 )OCH 3 or-C (CH) 3 ) 2 OCH 3 Further preferred are hydrogen, methyl, methoxy, isopropyl, fluoromethyl and hydroxyMethyl, oxetanyl, - (CH) 2 )OCH 3 or-CH (CH) 3 )OCH 3 ；
R 2 Selected from hydrogen, C 1-6 Alkyl, halogen, cyano or- (CH) 2 ) n OR cc Preferably hydrogen, C 1-3 Alkyl, halogen, cyano or- (CH) 2 ) n OR cc More preferably hydrogen, methyl, ethyl, propyl, methoxy, ethoxy, propoxy, fluoro, chloro, bromo or cyano, still more preferably hydrogen, fluoro, methyl, methoxy or cyano;
or, any two of R's attached 2 To form a substituted or unsubstituted C 3-6 Cycloalkyl or substituted or unsubstituted 3-6 membered heterocyclyl, preferably substituted or unsubstituted C 3-6 Cycloalkyl or a substituted or unsubstituted 3-6 membered heterocyclic group containing 1 to 3N, O or S atoms, more preferably cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, oxetanyl, oxcyclohexyl, aziridinyl, azetidinyl, further preferably cyclobutyl, cyclopentyl, 1, 3-dioxanyl or 1, 3-dioxanyl;
R 3 and R is 3 ' selected from hydrogen, C 1-6 Alkyl, halogen, cyano or C 1-6 Alkoxy, preferably hydrogen, C 1-3 Alkyl, halogen, cyano or C 1-3 Alkoxy, more preferably hydrogen, methyl, ethyl, propyl, fluoro, chloro, bromo, cyano, methoxy, ethoxy or propoxy, still more preferably hydrogen, fluoro, methyl, methoxy or cyano;
Or, R 3 And R is 3 ' linking to form an oxo group, C 3-6 Cycloalkyl or 3-6 membered heterocyclyl, preferably oxo, C 3-6 Cycloalkyl or a 3-6 membered heterocyclic group containing 1 to 3N, O or S atoms, more preferably oxo, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, oxetanyl, cyclohexyl, aziridinyl, azetidinyl or azetidinyl, still more preferably oxo, cyclopropyl or oxetanyl;
R 4 selected from hydrogen、C 1-6 Alkyl, halogen, cyano, C 1-6 Haloalkyl or C 3-8 Cycloalkyl, preferably hydrogen, C 1-3 Alkyl, halogen, cyano, C 1-3 Haloalkyl or C 3-6 Cycloalkyl, more preferably hydrogen, methyl, ethyl, propyl, fluoro, chloro, bromo, cyano, fluoromethyl, fluoroethyl, chloromethyl, chloroethyl, trifluoromethyl, trifluoroethyl, trichloromethyl, trichloroethyl, cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl, further preferably hydrogen, fluoro, chloro, methyl, trifluoromethyl, cyano or cyclopropyl;
R 5 selected from hydrogen, C 1-6 Alkyl or C 1-6 Haloalkyl, preferably hydrogen, C 1-3 Alkyl or C 1-3 Haloalkyl, more preferably hydrogen, methyl, ethyl, propyl, fluoromethyl, fluoroethyl, fluoropropyl, chloromethyl, chloroethyl or chloropropyl, still more preferably hydrogen or methyl;
Or, R 1 Or R is 1 ' and R 5 And are connected to form a 3-6 membered heterocyclic group, wherein the 3-6 membered heterocyclic group is optionally substituted by one or more substituents selected from fluorine, chlorine, bromine, methyl, ethyl or propyl; preferably aziridinyl, azetidinyl, fluoro-substituted aziridinyl, fluoro-substituted azetidinyl, methyl-substituted aziridinyl, methyl-substituted azetidinyl, methylpyrrolidinyl or methyl-substituted azetidinyl, further preferably azetidinyl, azetidinyl or methylpyrrolidinyl;
R aa 、R bb and R is cc Each independently selected from hydrogen, C 1-6 Alkyl, C 1-6 Alkoxy, C 3-8 Cycloalkyl or 3-8 membered heterocyclyl, preferably hydrogen, C 1-3 Alkyl, C 1-3 Alkoxy, C 3-6 Cycloalkyl or 3-to 6-membered heterocyclyl having 1 to 3N, O or S atoms, more preferably hydrogen, methyl, ethyl, propyl, methoxy, ethoxy, propoxy, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, oxetanyl or oxetanyl, still more preferably hydrogen, methyl, ethyl, isopropylA group, methoxy, cyclopropyl or oxetanyl group.
Preferably, W is O, further G is O or S, further R 5 Is hydrogen, further R 1 ' and R 3 ' is hydrogen.
In a further preferred embodiment of the present invention, the compound has the structure shown in formula (II-A) or (II-B):
the invention also provides a method for preparing the crystal form of the compound shown in the general formula (I), which specifically comprises the following steps:
1) Weighing a proper amount of A substance, and adding a proper amount of L-alanine, a solvent and an oxidant for reaction;
2) Adding a catalyst into the solution in the step 1), and adding a solvent and a reactant D into the solution to react after the reaction is completed;
3) Adding amine salt into the solution in the step 2) to react;
4) Adding a solvent to the solution of step 3) and separating the organic phase;
5) Adding a solvent to step 4);
6) Optionally seeding in step 5);
7) Cooling and crystallizing to obtain a substance C;
preferably, the oxidizing agent in step 1) is potassium phosphate;
preferably, the catalyst in step 2) is cuprous oxide;
preferably, the solvent in step 2) is NH 3 MeOH solution of (b);
preferably, the reactive substance D is NH 4 Cl, HOSu and DIPEA;
preferably, the amine salt in step 3) is EDCl;
W、G、R 2 、R 3 、R 3 ’、R 4 and X is as described in formula (I).
The invention also provides a method for preparing the crystal form of the compound shown in the general formula (I), which specifically comprises the following steps:
1) Weighing a proper amount of free alkali or a crystal form thereof, and suspending with a poor solvent;
2) Shaking the suspension obtained above;
3) Centrifuging the suspension, removing supernatant, and drying the rest solid in a vacuum drying oven until the weight is constant to obtain a target product;
preferably, the poor solvent is selected from one or more of water, ketones, alcohols, esters, ethers, benzenes, amides or acetonitrile, preferably one or more of water, acetone, 2-butanone, ethanol, N-propanol, isopropanol, N-butanol, isobutanol, tert-butanol, ethyl acetate, tetrahydrofuran, 1, 4-dioxane, benzene, toluene, N-dimethylformamide, N-dimethylacetamide or acetonitrile, preferably one or more of water, tetrahydrofuran, acetone or 1, 4-dioxane, more preferably one or more of water, tetrahydrofuran, 88% acetone or 1, 4-dioxane.
Preferably, the suspension density of the suspension in step 1) is 50-100 mg/mL;
preferably, the temperature in step 2) is from 25 to 40 ℃, preferably from 25 to 30 ℃, for a period of from 1 to 10 days, preferably from 5 to 10 days, more preferably 10 days;
preferably, the vacuum drying temperature in step 3) is 40 ℃.
The invention also provides a method for preparing the crystal form of the compound shown in the general formula (I), which specifically comprises the following steps:
1) Weighing a proper amount of salt of free alkali, and suspending with water;
2) Shaking the suspension obtained above;
3) And (3) rapidly centrifuging the suspension, removing the supernatant, and drying the residual solid in vacuum to constant weight to obtain the target product.
Preferably, the method comprises the steps of, the salt form is selected from hydrochloride, sulfate, nitrate, hydrobromide, hydrofluoric acid, hydroiodide, phosphate, 2, 5-dihydroxybenzoate, 1-hydroxy-2-naphthoate, acetate, dichloroacetate, trichloroacetate, acetohydroxamate, adipate, benzenesulfonate, 4-chlorobenzenesulfonate, benzoate, 4-acetaminobenzoate, 4-aminobenzoate, caprate, caproate, caprylate, cinnamate, citrate, cyclohexanaminosulfonate, camphorsulfonate, aspartate, camphorate, gluconate, glucarate, glutamate, isoascorbate, lactate, malate, mandelate, pyroglutamate, D-tartrate, pamoate, dodecylsulfate dibenzoyltartrate, ethanesulfonate, ethane-1, 2-disulfonate, ethanesulfonate, formate, fumarate, galactarate, gentisate, glutarate, 2-ketoglutarate, glycolate, hippurate, isethionate, lactobionate, ascorbate, aspartate, laurate, campholate, maleate, malonate, methanesulfonate, 1, 5-naphthalenedisulfonate, naphthalene-2-sulfonate, nicotinate, oleate, orotate, oxalate, palmitate, pamoate, propionate, salicylate, 4-aminosalicylate, sebacate, stearate, succinate, thiocyanate, undecylenate, trifluoroacetate, benzenesulfonate, p-toluenesulfonate or L-malate, preferably phosphate, sulfate, benzenesulfonate, methanesulfonate, ethanesulfonate, fumarate, isethionate, maleate, salicylate, benzenesulfonate or ethanesulfonate, more preferably methanesulfonate, sulfate or ethanesulfonate.
Preferably, the suspension density of the suspension in step 1) is 50-100 mg/mL;
preferably, the temperature in step 2) is from 25 to 40 ℃, preferably from 25 to 30 ℃, for a period of from 1 to 10 days, preferably from 5 to 10 days, more preferably 10 days;
preferably, the vacuum drying temperature in step 3) is 40 ℃.
In a further preferred embodiment of the present invention, the compound of formula (I), crystalline form (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide, form A, has a X-ray powder diffraction pattern with diffraction peaks at 2θ of 5.7, 11.3, 19.5 and 19.8; preferably, it further comprises diffraction peaks at 20.4, 21.8 and 24.3 in 2 theta; more preferably, it further comprises diffraction peaks at 2θ of 16.9, 18.5, 27.4 and 33.9; it is further preferred that there are diffraction peaks at 9.8, 15.9, 18.0 and 23.7 for 2 theta, each 2 theta having an error of about + -0.2 deg..
The X-ray characteristic diffraction peaks expressed in terms of 2 theta angle and interplanar spacing d values using Cu-K alpha radiation are shown in Table 1.
TABLE 1
The invention relates to a crystal form A of a compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide shown in a general formula (I), and the X-ray powder diffraction pattern of the crystal form A is basically shown in the figure 1; the TGA diagram is basically shown in figure 2; the DSC spectrum is basically shown in figure 3.
In a further preferred embodiment of the present invention, the compound of formula (I), crystalline form (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide, form B, has a X-ray powder diffraction pattern with diffraction peaks at 2 theta of 7.9, 8.3, 13.1 and 23.6; preferably, it further comprises diffraction peaks at 2θ of 7.6, 14.2 and 19.5; more preferably, it further comprises diffraction peaks at 14.7, 15.7, 21.8 and 24.9 of 2 theta, each 2 theta having an error of about + -0.2 deg..
The X-ray characteristic diffraction peaks expressed in terms of 2 theta angle and interplanar spacing d values using Cu-K alpha radiation are shown in Table 2.
TABLE 2
The invention relates to a crystal form B of a compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide shown in a general formula (I), and the X-ray powder diffraction pattern of the crystal form B is basically shown in figure 4; the DSC spectrum is basically shown in figure 5.
In a further preferred embodiment of the present invention, the compound of formula (I), crystalline form (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide, form C, has a X-ray powder diffraction pattern with diffraction peaks at 2 theta of 7.7, 19.4, 19.7 and 22.3; preferably, it further comprises diffraction peaks at 2θ of 13.0, 15.3 and 17.7; more preferably, it further comprises diffraction peaks at 20.4, 23.2, 24.1 and 25.3 of 2 theta, each 2 theta having an error of about + -0.2 deg..
The X-ray characteristic diffraction peaks expressed in terms of 2 theta angle and interplanar spacing d values using Cu-K alpha radiation are shown in Table 3.
TABLE 3 Table 3
The invention relates to a crystal form C of a compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide shown in a general formula (I), and the X-ray powder diffraction pattern of the crystal form C is basically shown in figure 6; the TGA diagram is basically shown in figure 7; the DSC spectrum is basically shown in figure 8.
In a further preferred embodiment of the present invention, the compound of formula (I), crystalline form (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-D ] [1,4] oxazepin-9-yl) amino) propanamide, form D, has a X-ray powder diffraction pattern with diffraction peaks at 2 theta of 6.6, 10.1, 13.8 and 18.9; preferably, it further comprises diffraction peaks at 2θ of 11.4, 13.4, 18.6 and 19.5; more preferably, it further comprises diffraction peaks at 2 theta of 16.2, 20.4, 21.9, 24.1 and 26.1, each 2 theta having an error of about + -0.2 deg..
The X-ray characteristic diffraction peaks expressed in terms of 2 theta angle and interplanar spacing d values using Cu-K alpha radiation are shown in Table 4.
TABLE 4 Table 4
The invention relates to a crystal form D of a compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-D ] [1,4] oxazepin-9-yl) amino) propanamide shown in a general formula (I), and the X-ray powder diffraction pattern of the crystal form D is basically shown in figure 9; the TGA diagram is basically shown in figure 10; the DSC spectrum is basically shown in figure 11.
In a further preferred embodiment of the present invention, the compound of formula (I) is a crystalline form of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide, form E, having a X-ray powder diffraction pattern with diffraction peaks at 77.7, 17.6, 19.4 and 22.1 in 2. Theta; preferably, it further comprises diffraction peaks at 2θ of 15.4, 23.1, 23.9 and 25.1; more preferably, it further comprises diffraction peaks at 2θ of 12.8, 20.5 and 25.9, each 2θ having an error of about ±0.2°.
The X-ray characteristic diffraction peaks expressed in terms of 2 theta angle and interplanar spacing d values using Cu-K alpha radiation are shown in Table 5.
TABLE 5
The invention relates to a crystal form E of a compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide shown in a general formula (I), and the X-ray powder diffraction pattern of the crystal form E is basically shown in figure 12; the TGA spectrum is basically shown in figure 13; the DSC spectrum is basically shown in figure 14.
In a further preferred embodiment of the present invention, the compound of formula (I), crystalline form (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ F ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide, form F, has a X-ray powder diffraction pattern with diffraction peaks at 2 theta of 6.5, 10.1, 19.1 and 24.2; preferably, further comprising diffraction peaks at 2θ of 13.7, 17.3, and 26.2; more preferably, it further comprises diffraction peaks at 2 theta of 13.4, 20.1, 25.1 and 27.7, each 2 theta having an error of about + -0.2 deg..
The X-ray characteristic diffraction peaks expressed in terms of 2 theta angle and interplanar spacing d values using Cu-K alpha radiation are shown in Table 6.
TABLE 6
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The X-ray powder diffraction pattern of the compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ F ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide crystal form F shown in the general formula (I) is basically shown in the figure 15; the TGA diagram is basically shown in figure 16; the DSC spectrum is basically shown in figure 17.
The invention also aims to provide a pharmaceutical composition which contains a therapeutically effective amount of the crystal form of the compound shown in the general formula (I) and one or more pharmaceutically acceptable carriers, diluents or excipients.
The invention also aims to provide the application of the crystal form of the general formula compound shown in the formula (I) and the pharmaceutical composition in the preparation of PI3K inhibitor drugs, preferably in the preparation of PI3K alpha inhibitor drugs.
The invention also aims to provide the crystal forms of the compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide shown in the general formula (I), and one or more pharmaceutically acceptable carriers, diluents or excipients.
The invention also aims to provide the crystal form of the compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide shown in the general formula (I), and application of the pharmaceutical composition in preparation of PI3K inhibitor medicines, preferably PI3K alpha inhibitor medicines.
The use as described above, wherein the use is in the manufacture of a medicament for the treatment of cancer, bone disease, inflammatory disease, immune disease, neurological disease, metabolic disease, respiratory disease and heart disease; wherein the cancer is selected from the group consisting of breast cancer, pancreatic cancer, non-small cell lung cancer, thyroid cancer, seminoma, melanoma, bladder cancer, liver cancer, kidney cancer, myelodysplastic syndrome, acute myelogenous leukemia, and colorectal cancer.
Drawings
FIG. 1 is an XRPD representation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form A.
FIG. 2 is a TGA graphic representation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form A.
FIG. 3 is a DSC illustration of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form A.
FIG. 4 is an XRPD representation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form B.
FIG. 5 is a DSC illustration of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form B.
FIG. 6 is an XRPD representation of crystalline form C of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide.
FIG. 7 is a TGA graphic representation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form C.
FIG. 8 is a DSC illustration of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form C.
FIG. 9 is an XRPD representation of crystalline form D of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-D ] [1,4] oxazepin-9-yl) amino) propanamide.
FIG. 10 is a TGA graphic representation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-D ] [1,4] oxazepin-9-yl) amino) propanamide form D.
FIG. 11 is a DSC plot of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-D ] [1,4] oxazepin-9-yl) amino) propanamide form D.
FIG. 12 is an XRPD representation of crystalline form E of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide.
FIG. 13 is a TGA graphic representation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form E.
FIG. 14 is a DSC illustration of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form E.
FIG. 15 is an XRPD representation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ F ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form F.
FIG. 16 is a TGA graphic representation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ F ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form F.
FIG. 17 is a DSC plot of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ F ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form F.
Detailed Description
Unless stated to the contrary, the terms used in the specification and claims have the following meanings.
The term "alkyl" refers to a saturated aliphatic hydrocarbon group which is a straight or branched chain group containing from 1 to 20 carbon atoms, preferably an alkyl group containing from 1 to 8 carbon atoms, more preferably an alkyl group containing from 1 to 6 carbon atoms, and most preferably an alkyl group containing from 1 to 3 carbon atoms. Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1-dimethylpropyl, 1, 2-dimethylpropyl, 2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1, 2-trimethylpropyl, 1-dimethylbutyl, 1, 2-dimethylbutyl, 2-dimethylbutyl, 1, 3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2, 3-dimethylbutyl, n-heptyl, 2-methylhexyl, 3-methylhexyl 4-methylhexyl, 5-methylhexyl, 2, 3-dimethylpentyl, 2, 4-dimethylpentyl, 2-dimethylpentyl, 3-dimethylpentyl, 2-ethylpentyl, 3-ethylpentyl, n-octyl, 2, 3-dimethylhexyl, 2, 4-dimethylhexyl, 2, 5-dimethylhexyl, 2-dimethylhexyl, 3-dimethylhexyl 4, 4-dimethylhexyl, 2-ethylhexyl, 3-ethylhexyl, 4-ethylhexyl, 2-methyl-2-ethylpentyl, 2-methyl-3-ethylpentyl, n-nonyl, 2-methyl-2-ethylhexyl, 2-methyl-3-ethylhexyl, 2-diethylpentyl, n-decyl, 3-diethylhexyl, 2, 2-diethylhexyl group and various branched isomers thereof, and the like. The alkyl group may be substituted or unsubstituted, and when substituted, the substituent may be substituted at any available point of attachment, preferably one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, oxo, carboxy or carboxylate, with methyl, ethyl, isopropyl, t-butyl, haloalkyl, deuteroalkyl, alkoxy-substituted alkyl, hydroxy-substituted alkyl and cyano-substituted alkyl being preferred.
The term "alkylene" means that one hydrogen atom of the alkyl group is further substituted, for example: "methylene" means-CH 2 - "ethylene" means- (CH) 2 ) 2 - "propylene" means- (CH) 2 ) 3 "butylene" means- (CH) 2 ) 4 -and the like, the above substituents may be linked to different carbon atoms to form a carbon chain, or may be linked to one carbon atom to form a cycloalkyl group. The term "alkenyl" refers to an alkyl group as defined above consisting of at least two carbon atoms and at least one carbon-carbon double bond, such as vinyl, 1-propenyl, 2-propenyl, 1-, 2-, or 3-butenyl, and the like. Alkenyl groups may be substituted or unsubstituted, and when substituted, the substituents are preferably one or more groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio.
The term "cycloalkyl" refers to a saturated or partially unsaturated monocyclic or polycyclic cyclic hydrocarbon substituent, the cycloalkyl ring containing from 3 to 20 carbon atoms, preferably from 3 to 8 carbon atoms, more preferably from 3 to 6 carbon atoms. Non-limiting examples of monocyclic cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptatrienyl, cyclooctyl, and the like; polycyclic cycloalkyl groups include spiro, fused and bridged cycloalkyl groups, preferably cyclopropyl, cyclobutyl, cyclohexyl, cyclopentyl and cycloheptyl.
The term "heterocyclyl" refers to a saturated or partially unsaturated monocyclic or polycyclic cyclic hydrocarbon substituent containing from 3 to 20 ring atoms in which one or more ring atoms are selected from nitrogen, oxygen or S (O) m (wherein m is an integer from 0 to 2), but does not include a ring moiety of-O-O-, -O-S-, or-S-S-, and the remaining ring atoms are carbon. Preferably containing 3 to 12 ring atoms, of which 1 to 4 are heteroatoms; more preferably 3 to 8 ring atoms; most preferably containing 3 to 8 ring atoms. Non-limiting examples of monocyclic heterocyclyl groups include oxetanyl, pyrrolidinyl, pyrrolidinonyl, imidazolidinyl, tetrahydrofuranyl, tetrahydrothienyl, dihydroimidazolyl, dihydrofuranyl, dihydropyrazolyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl, homopiperazinyl, pyranyl, and the like, with oxetanyl, pyrrolidinonyl, tetrahydrofuranyl, pyrazolidinyl, morpholinyl, piperazinyl, and pyranyl being preferred. Polycyclic heterocyclyl groups include spiro, fused and bridged heterocyclic groups; the heterocyclic groups of the spiro ring, the condensed ring and the bridged ring are optionally connected with other groups through single bonds, or are further connected with other cycloalkyl groups, heterocyclic groups, aryl groups and heteroaryl groups through any two or more atoms on the ring in a parallel ring mode.
The term "alkoxy" refers to-O- (alkyl) and-O- (unsubstituted cycloalkyl), wherein alkyl is as defined above. Alkoxy groups having 1 to 8 carbon atoms are preferred, alkoxy groups having 1 to 6 carbon atoms are more preferred, and alkoxy groups having 1 to 3 carbon atoms are most preferred. Non-limiting examples of alkoxy groups include: methoxy, ethoxy, propoxy, butoxy, cyclopropoxy, cyclobutoxy, cyclopentoxy, cyclohexyloxy. The alkoxy groups may be optionally substituted or unsubstituted, and when substituted, the substituents are preferably one or more groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, carboxyl, or carboxylate groups.
"haloalkyl" refers to an alkyl group substituted with one or more halogens, where alkyl is as defined above.
"haloalkoxy" refers to an alkoxy group substituted with one or more halogens, wherein the alkoxy group is as defined above.
"hydroxyalkyl" refers to an alkyl group substituted with a hydroxy group, wherein alkyl is as defined above.
"hydroxy" refers to an-OH group.
"halogen" means fluorine, chlorine, bromine or iodine.
"amino" means-NH 2 。
"cyano" refers to-CN.
"nitro" means-NO 2 。
"THF" refers to tetrahydrofuran.
"EtOAc" refers to ethyl acetate.
"DMSO" refers to dimethyl sulfoxide.
"LDA" refers to lithium diisopropylamide.
"DMAP" refers to 4-dimethylaminopyridine.
"EtMgBr" refers to ethylmagnesium bromide.
"HOSu" refers to N-hydroxysuccinimide.
"EDCl" refers to 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride.
"IPA" refers to isopropanol.
"MeOH" refers to methanol.
"EtOH" refers to ethanol.
"DMF" refers to N, N-dimethylformamide.
"DIPEA" refers to N, N-diisopropylethylamine.
"HEPES" refers to 4-hydroxyethylpiperazine ethanesulfonic acid.
The terms "X is selected from A, B or C", "X is selected from A, B and C", "X is A, B or C", "X is A, B and C", etc. all express the same meaning, that is, X may be any one or several of A, B, C.
"optional" or "optionally" means that the subsequently described event or circumstance may but need not occur, and that the description includes instances where the event or circumstance occurs or does not.
"substituted" means that one or more hydrogen atoms, preferably up to 5, more preferably 1 to 3 hydrogen atoms in the group are independently substituted with a corresponding number of substituents. It goes without saying that substituents are only in their possible chemical positions, and that the person skilled in the art is able to determine (by experiment or theory) possible or impossible substitutions without undue effort. For example, amino or hydroxyl groups having free hydrogen may be unstable when bound to carbon atoms having unsaturated (e.g., olefinic) bonds.
"stereoisomers" include three classes of geometric (cis-trans) isomerism, optical isomerism, conformational isomerism.
The hydrogen atoms of the invention can be replaced by the isotope deuterium thereof, and any hydrogen atom in the compound of the embodiment of the invention can be replaced by deuterium atoms.
"pharmaceutical composition" means a mixture comprising one or more of the compounds described herein or a physiologically/pharmaceutically acceptable salt or prodrug thereof, and other chemical components, such as physiologically/pharmaceutically acceptable carriers and excipients. The purpose of the pharmaceutical composition is to promote the administration to organisms, facilitate the absorption of active ingredients and thus exert biological activity.
By "pharmaceutically acceptable salts" is meant salts of the compounds of the present invention which are safe and effective when used in a mammal, and which possess the desired biological activity.
"TGA" refers to thermogravimetric analysis (TGA) experiments.
"DSC" refers to Differential Scanning Calorimetry (DSC) experiments.
"XRPD" refers to X-ray powder diffraction (XRPD) experiments.
"HPLC" refers to High Performance Liquid Chromatography (HPLC) experiments.
"PK" refers to Pharmacokinetic (PK) experiments.
As described herein, the new crystalline forms can be identified by powder X-ray diffraction patterns. However, those skilled in the art will appreciate that the peak intensity and/or peak condition of powder X-ray diffraction may vary from experimental condition to experimental condition, such as different diffraction test conditions and/or preferential orientations, etc. Also, due to the different accuracy of the different instruments, the measured 2 theta values may have an error of about + -0.2, and the individual peaks may have an error of about + -0.3 or + -0.4. However, it is known that the relative intensity value of a peak is more dependent on certain properties of the sample being measured than the position of the peak, such as the size of the crystals in the sample, the orientation of the crystals and the purity of the material being analyzed, and therefore variations in the intensity of the peaks shown are likely to occur over a range of about + -20% or more.
The invention is further described below in connection with examples, which are not intended to limit the scope of the invention.
1. Preparation of the Compounds
Examples
The structure of the compounds of the present invention is determined by Nuclear Magnetic Resonance (NMR) or/and liquid chromatography-mass spectrometry (LC-MS). NMR chemical shifts (δ) are given in parts per million (ppm). NMR was performed using Bruker AVANCE-400 nuclear magnetic resonance apparatus with deuterated dimethyl sulfoxide (DMSO-d) 6 ) Deuterated methanol (CD) 3 OD) and deuterated chloroform (CDCl) 3 ) The internal standard is Tetramethylsilane (TMS).
An Agilent 1200 affinity Series mass spectrometer was used for LC-MS measurement. HPLC was performed using Agilent 1200DAD high pressure liquid chromatography (Sunfire C18X 4.6mm column) and Waters 2695-2996 high pressure liquid chromatography (Gimini C18X 4.6mm column).
The thin layer chromatography silica gel plate uses a smoke table yellow sea HSGF254 or Qingdao GF254 silica gel plate, the specification adopted by TLC is 0.15 mm-0.20 mm, and the specification adopted by the thin layer chromatography separation and purification product is 0.4 mm-0.5 mm. Column chromatography generally uses yellow sea silica gel with 200-300 meshes as a carrier.
The starting materials in the examples of the present invention are known and commercially available or may be synthesized according to methods known in the art.
All reactions of the invention were carried out under continuous magnetic stirring under dry nitrogen or argon atmosphere, with the solvent being a dry solvent and the reaction temperature being in degrees celsius, without specific explanation.
Intermediate 1
(S) -4- (difluoromethyl) oxazolidin-2-one
The first step: preparation of (R) -3-benzyl-4- (hydroxymethyl) oxazolidin-2-one
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(R) -Oxopropan-2-ylmethanol (3.7 g,50.0 mmol) and (isocyanatomethyl) benzene (6.66 g,50.0 mmol) were mixed in dichloromethane (50 mL) and warmed to 45℃under nitrogen and stirred overnight. After cooling, 100mL of saturated aqueous sodium bicarbonate solution was added and extracted with methylene chloride (100 mL. Times.2). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure, followed by column chromatography to give the title compound (R) -3-benzyl-4- (hydroxymethyl) oxazolidin-2-one (4.14 g, 40%).
MS m/z(ESI):208.2[M+H] + .
And a second step of: preparation of (S) -3-benzyl-4- (dihydroxymethyl) oxazolidin-2-one
(R) -3-benzyl-4- (hydroxymethyl) oxazolidin-2-one (4.14 g,20.0 mmol), IBX (16.8 g,60.0 mmol) were mixed in ethyl acetate (100 mL) and stirred at 85deg.C for 3h under nitrogen. After cooling, filtration was carried out, and the filtrate was concentrated under reduced pressure to give 4.46g of crude (S) -3-benzyl-4- (dihydroxymethyl) oxazolidin-2-one, which was directly used for the next reaction.
MS m/z(ESI):224.2[M+H] + .
And a third step of: preparation of (S) -3-benzyl-4- (difluoromethyl) oxazolidin-2-one
(S) -3-benzyl-4- (dimethylol) oxazolidin-2-one (4.46 g,20.0 mmol) was dissolved in dichloromethane (100 mL), and DAST (6.45 g,40.0 mmol) was added dropwise to the reaction solution under nitrogen protection in an ice bath, and the reaction was carried out at room temperature for 3h. The reaction solution was slowly added dropwise to a pre-cooled saturated aqueous sodium bicarbonate solution. Dichloromethane (200 mL x 2) was extracted, and the organic phases were combined, concentrated under reduced pressure, and then column chromatographed to give the title compound (S) -3-benzyl-4- (difluoromethyl) oxazolidin-2-one (1.82 g, 40% yield in two steps).
MS m/z(ESI):228.2[M+H] + .
Fourth step: preparation of (S) -4- (difluoromethyl) oxazolidin-2-one
(S) -3-benzyl-4- (difluoromethyl) oxazolidin-2-one (1.82 g,8 mmol) was dissolved in ethanol (100 mL) and Pd (OH) was added 2 C (300 mg) under hydrogen atmosphere, stirring overnight at 70 ℃. Cooled, filtered, and the filtrate concentrated under reduced pressure to give the title compound (S) -4- (difluoromethyl) oxazolidin-2-one (0.88 g, 80%).
1 H NMR(400MHz,CDCl 3 )δ4.05-4.18(m,1H),4.39-4.45(m,1H),4.54(t,J＝9.3Hz,1H),5.78(td,J＝55.3,4.7Hz,1H),6.07(s,1H)；
MS m/z(ESI):138.1[M+H] + .
Intermediate 2
9-bromo-2-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
The first step: preparation of 5-bromo-2- (1H-imidazol-2-yl) phenol
To a methanol solution (250 mL) of 4-bromo-2-hydroxybenzaldehyde (24.0 g,119 mmol) was added an aqueous glyoxal solution (40 wt.%,87g,597 mmol), and then aqueous ammonia (28 wt.%,121g,860 mmol) was slowly added dropwise with stirring in a water bath for 30 minutes, and the reaction solution temperature was controlled to not exceed 40 ℃. Then, the mixture was stirred at 35℃for two days, cooled, and the organic solvent was removed under reduced pressure to give crude 5-bromo-2- (1H-imidazol-2-yl) phenol which was directly used in the next reaction.
MS m/z(ESI):239.0[M+H] + .
And a second step of: preparation of 9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
Crude 5-bromo-2- (1H-imidazol-2-yl) phenol (about 29g,119 mmol), cesium carbonate (158 g, 480 mmol), 1, 2-dibromoethane (42 mL, 480 mmol) were mixed in DMF (250 mL), stirred overnight at 85 ℃, cooled, and diluted with a large amount of ethyl acetate. The organic phase was washed with saturated brine several times, then dried over anhydrous sodium sulfate, concentrated, and column chromatographed to give the title compound 9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (12.5 g, two-step yield: 38%).
MS m/z(ESI):265.0[M+H] + .
And a third step of: preparation of 9-bromo-2, 3-diiodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
NIS (29.8 g,132 mmol) was added in portions to a DMF solution (150 mL) of 9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (11.7 g,44.1 mmol) at room temperature, followed by stirring overnight at 60 ℃. Cooling, adding water, and separating out solid. After filtration, the solid was dissolved in ethyl acetate, washed with 1M aqueous NaOH and saturated brine in this order, dried over anhydrous sodium sulfate, and concentrated to give the title compound 9-bromo-2, 3-diiodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (22.5 g, yield: 98.7%).
MS m/z(ESI):516.7[M+H] + .
Fourth step: preparation of 9-bromo-2-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
EtMgBr (1.0M in THF, 60.9mL,60.9 mmol) was slowly added dropwise to a solution of 9-bromo-2, 3-diiodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (21.0 g,40.6 mmol) in THF (140 mL) at-20 ℃. After the completion of the dropwise addition, the mixture was stirred at-15℃for 3 hours. Slowly warmed to room temperature, then saturated aqueous ammonium chloride was added dropwise, stirred for 15 minutes, and extracted with ethyl acetate multiple times. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, concentrated, and column chromatographed to give the title compound 9-bromo-2-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (12.5 g, yield 79%).
MS m/z(ESI):390.9[M+H] + .
Fifth step: preparation of (S) -3- (9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one
9-bromo-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d][1,4]Oxazepine (300 mg,0.77 mmol), (S) -4- (difluoromethyl) oxazolidin-2-one (105 mg,0.77 mmol), (1R, 2R) -N 1 ,N 2 Dimethylcyclohexane-1, 2-diamine (43 mg,0.30 mmol), copper acetate (27 mg,0.15 mmol), cesium carbonate (4819 mg,1.5 mmol), mixed in 2-methyltetrahydrofuran (6 mL), The reaction was allowed to react with nitrogen instead of tertiary air at 78℃for 22 hours, the reaction mixture was cooled to room temperature, 15% aqueous ammonia was added thereto, and the mixture was stirred for 5 minutes, followed by extraction with EtOAc three times. The organic phases were combined, then the organic phase was washed with saturated aqueous sodium chloride, the filtrate was dried over anhydrous sodium sulfate, and the organic solvent was concentrated under reduced pressure and then separated by column chromatography to give the title compound (S) -3- (9-bromo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d][1,4]Oxazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one (186 mg, 61%).
1 H NMR(400MHz,CDCl 3 )δ4.35-4.41(m,2H),4.44-4.52(m,2H),4.53-4.55(m,1H),4.73-4.76(m,1H),4.89-4.91(m,1H),6.62-6.71(m,1H),7.19-7.28(m,2H),7.30(s,1H),8.21(d,J＝8.6Hz,1H)；
MS m/z(ESI):400.1[M+H] + .
Example 1
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The first step: preparation of 9-bromo-3-fluoro-2-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
To a solution of LDA (1.28 mL,2.56 mmol) in tetrahydrofuran (10 mL) at-78deg.C was added dropwise a solution of 9-bromo-2-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (500 mg,1.28 mmol) in tetrahydrofuran (10 mL), after the addition was completed, stirring was continued at-78deg.C for 30 minutes, and after stirring a solution of N-fluorobenzenesulfonamide (806 mg,2.56 mmol) in tetrahydrofuran (9 mL) was added dropwise and stirring was continued at this temperature for 30 minutes. The reaction solution was quenched with saturated aqueous ammonium chloride, extracted with dichloromethane (100 mL. Times.2), the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated and then column chromatographed to give the title compound 9-bromo-3-fluoro-2-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (150 mg, 29%).
1 H NMR(400MHz,DMSO-d 6 )δ4.31-4.34(m,2H),4.43-4.48(m,2H),7.19-7.34(m,2H),8.17(d,J＝8.6Hz,1H)；
MS m/z(ESI):408.9[M+H] + .
And a second step of: preparation of (S) -3- (9-bromo-3-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one
9-bromo-3-fluoro-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d][1,4]Oxazepine (100 mg,0.24 mmol), (S) -4- (difluoromethyl) oxazolidin-2-one (33.5 mg,0.24 mmol), (1R, 2R) -N 1 ,N 2 Dimethylcyclohexane-1, 2-diamine (35 mg,0.24 mmol), cuprous iodide (46 mg,0.24 mmol), potassium phosphate (155 mg,0.73 mmol), and dimethyl sulfoxide (10 mL) were mixed and reacted at 130℃for 3 hours. The reaction was cooled to room temperature, 15% aqueous ammonia was added, stirred for 5 minutes, and extracted three times with EtOAc. The organic phases were combined, then the organic phase was washed with saturated sodium chloride, dried over anhydrous sodium sulfate, concentrated under reduced pressure and then column chromatographed to give the title compound (S) -3- (9-bromo-3-fluoro-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d][1,4]Oxazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one (21 mg, 20%).
1 H NMR(400MHz,CDCl 3 )δ4.25-4.29(m,1H),4.42-4.50(m,2H),4.56-4.69(m,4H),6.16-6.35(m,1H),7.20-7.25(m,2H),8.15(d,J＝8.4Hz,1H)；
MS m/z(ESI):417.9[M+H] + .
And a third step of: preparation of (2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) -L-alanine
((S) -3- (9-bromo-3-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one (21 mg,0.05 mmol), L-alanine (13.5 mg,0.15 mmol), cuprous iodide (4.8 mg,0.025 mmol), potassium phosphate (21 mg,0.1 mmol) were mixed in dimethyl sulfoxide (3 mL), the reaction system was replaced with nitrogen for three times with air, and the mixture was cooled to room temperature and used directly in the next reaction without any treatment.
MS m/z(ESI):427.1[M+H] + .
Fourth step: preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
To the crude reaction solution of the above step (2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) -L-alanine was added ammonium chloride (16 mg,0.29 mmol), triethylamine (76 mg,0.75 mmol), stirred for 5 minutes, 2- (7-benzotriazol-oxide) -N, N' -tetramethylurea hexafluorophosphate (166 mg,0.44 mmol), stirred at room temperature for 2 hours, the reaction solution was filtered, and a saturated aqueous sodium hydrogen carbonate solution was added to the filtrate, followed by extraction with ethyl acetate three times. The organic phases were combined, dried over anhydrous sodium sulfate and the organic solvent was concentrated under reduced pressure and then separated by column chromatography to give the title compound (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide (8.5 mg, 39%).
1 H NMR(400MHz,CDCl 3 )δ1.55(d,J＝7.0Hz,3H),3.70-3.87(m,1H),4.21(d,J＝3.6Hz,2H),4.43(d,J＝5.2Hz,2H),4.57-4.66(m,2H),5.35(s,1H),6.10-6.27(m,2H),6.37-6.50(m,2H),8.07(d,J＝8.6Hz,1H)；
MS m/z(ESI):426.1[M+H] + .
Example 2
Preparation of (S) -1- (2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) pyrrolidine-2-carboxamide
The preparation of (S) -1- (2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) pyrrolidine-2-carboxamide is described in example 1.
MS m/z(ESI):452.1[M+H] + .
Example 3
Preparation of (S) -2- ((3-chloro-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The preparation of (S) -2- ((3-chloro-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide is described in example 1.
1 H NMR(400MHz,CD 3 OD)δ1.46(d,J＝7.0Hz,3H),3.80-3.86(m,1H),4.29-4.32(m,2H),4.43-4.46(m,2H),4.57-4.67(m,3H),6.07-6.31(m,2H),6.43-6.46(m,1H),7.98(d,J＝8.8Hz,1H)；
MS m/z(ESI):442.1[M+H] + .
Example 4
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The first step: preparation of 5-bromo-2- (5-methyl-1H-imidazol-2-yl) phenol
To a methanol solution (100 mL) of 4-bromo-2-hydroxybenzaldehyde (5 g,119 mmol) was added an aqueous methylglyoxal solution (40 wt.%,80 mL), and then aqueous ammonia (28 wt.%,40 g) was slowly added dropwise with stirring in a water bath for 30 minutes, with the solution temperature controlled to not exceed 40 ℃. Then, the reaction solution was stirred at 75℃for 2 hours, cooled to room temperature, and then a solid was precipitated and filtered to give the title compound, 5-bromo-2- (5-methyl-1H-imidazol-2-yl) phenol (3.6 g, 57%).
MS m/z(ESI):253.0[M+H] + .
And a second step of: preparation of 9-bromo-3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
5-bromo-2- (5-methyl-1H-imidazol-2-yl) phenol (2.5 g,9.8 mmol), cesium carbonate (12.2 g,37.5 mmol), 1, 2-dibromoethane (42.0 mL,37.5 mmol) were mixed in DMF (30 mL) and stirred overnight at 85 ℃. After the reaction was cooled to room temperature, a large amount of ethyl acetate was added for dilution. The organic phase was washed with saturated brine several times, then dried over sodium sulfate, concentrated and column chromatographed to give the title compound 9-bromo-3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (0.92 g, 33%).
1 H NMR(400MHz,CDCl 3 )δ2.25(s,3H),4.12-4.29(m,2H),4.40-4.53(m,2H),6.94(s,1H),7.14-7.18(m,1H),7.20-7.22(m,1H),8.37(d,J＝8.6Hz,1H)；
MS m/z(ESI):279.1[M+H] + .
And a third step of: preparation of 9-bromo-2-iodo-3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
The preparation of 9-bromo-2-iodo-3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine is described in example 1.
MS m/z(ESI):404.9[M+H] + .
Fourth step: preparation of (S) -3- (9-bromo-3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one
The preparation of (S) -3- (9-bromo-3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one is described in example 1.
MS m/z(ESI):414.0[M+H] + .
Fifth step: preparation of (2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) -L-alanine
The preparation of (2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) -L-alanine is described in example 1.
MS m/z(ESI):423.1[M+H] + .
Sixth step: synthesis of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide is described in example 1.
1 H NMR(400MHz,CD 3 OD)δ1.37(d,J＝7.0Hz,3H),2.08(s,3H),3.68-3.75(m,1H),4.18-4.24(m,2H),4.32-4.35(m,2H),4.45-4.61(m,3H),6.10(m,2H),6.34(d,J＝8.8Hz,1H),7.83(d,J＝8.8Hz,1H).
MS m/z(ESI):422.2[M+H] + .
Example 5
Preparation of (S) -1- (2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) pyrrolidine-2-carboxamide
The preparation of (S) -1- (2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) pyrrolidine-2-carboxamide is described in example 4.
MS m/z(ESI):448.2[M+H] + .
Example 6
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3- (trifluoromethyl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -3- (trifluoromethyl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide is described in example 4.
MS m/z(ESI):476.1[M+H] + .
Example 7
Preparation of (S) -2- ((3-cyano-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The preparation of (S) -2- ((3-cyano-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide is described in example 4.
MS m/z(ESI):433.1[M+H] + .
Example 8
Preparation of (S) -1- (3-cyano-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) pyrrolidine-2-carboxamide
The preparation of (S) -1- (3-cyano-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) pyrrolidine-2-carboxamide is described in example 4.
MS m/z(ESI):459.2[M+H] + .
Example 9
Preparation of (S) -2- ((3-cyclopropyl-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The preparation of (S) -2- ((3-cyclopropyl-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide is described in example 4.
MS m/z(ESI):448.2[M+H] + .
Example 10
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5,6,10,11-tetrahydrocyclobutano [5,6] benzo [1,2-f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The first step: preparation of 1- (bicyclo [4.2.0] oct-1 (6), 2, 4-trien-3-yl) ethan-1-one
AlCl 3 (3.33 g,25 mmol) in nitromethane (25 mL), N 2 In the atmosphere, ice-bath and drop-adding bicyclo [4.2.0]]A solution of octyl-1 (6), 2, 4-triene (2.08 g,20 mmol) and acetyl chloride (1.73 g,22 mmol) in nitromethane (25 mL) was allowed to react overnight at room temperature. The reaction mixture was added to 200mL of ice-water, extracted with DCM (200 mL. Times.2), and the combined organic phases concentrated under reduced pressure and then chromatographed to give the title compound 1- (bicyclo [ 4.2.0)]Oct-1 (6), 2, 4-trien-3-yl) ethan-1-one (800 mg, 27%).
And a second step of: preparation of 1- (5-bromobicyclo [4.2.0] oct-1 (6), 2, 4-trien-3-yl) ethan-1-one
1- (bicyclo [ 4.2.0)]Octyl-1 (6), 2, 4-trien-3-yl) ethan-1-one (735 mg,5 mmol) was dissolved in acetic acid (20 mL), N 2 Bromine (878.9 mg,5.5 mmol) was added dropwise thereto under an atmosphere, and the mixture was reacted at room temperature for 3 hours. The reaction mixture was concentrated, DCM was separated from saturated aqueous sodium bicarbonate, and the organic phase was concentrated under reduced pressure and then column chromatographed to give the title compound 1- (5-bromobicyclo [ 4.2.0) ]Oct-1 (6), 2, 4-trien-3-yl) ethan-1-one (900 mg, 80%).
And a third step of: preparation of 5-bromobicyclo [4.2.0] oct-1 (6), 2, 4-trien-3-yl acetate
1- (5-bromobicyclo [ 4.2.0)]Octyl-1 (6), 2, 4-trien-3-yl) ethan-1-one (900 mg,4 mmol) and m-CPBA (75%, 2.30g,10 mmol) were mixed in DCM (20 mL), N 2 The reaction was refluxed overnight under an atmosphere. Cooling to room temperature, filtering to remove insoluble substances, washing the reaction solution with saturated sodium bicarbonate water solution, concentrating the organic phase under reduced pressure, and performing column chromatography to obtain the title compound 5-bromobicyclo [4.2.0]]Oct-1 (6), 2, 4-trien-3-yl acetate (323 mg, 75%).
Fourth step: preparation of 5-bromobicyclo [4.2.0] oct-1 (6), 2, 4-trien-3-ol
5-bromobicyclo [4.2.0] oct-1 (6), 2, 4-trien-3-yl acetate (323 mg,3 mmol) was dissolved in methanol (20 mL), and a 5N aqueous solution of sodium hydroxide (3 mL) was added to the solution, followed by reaction at room temperature overnight. 50mL of water, 1N hydrochloric acid was added to adjust the pH of the reaction to 5, the mixture was extracted with DCM (50 mL. Times.2), and the combined organic phases were concentrated under reduced pressure and then column chromatographed to give the title compound 5-bromobicyclo [4.2.0] oct-1 (6), 2, 4-trien-3-ol (567 mg, 95%).
Fifth step: preparation of 5-bromo-3-hydroxybicyclo [4.2.0] oct-1 (6), 2, 4-triene-2-carbaldehyde
5-bromobicyclo [4.2.0] oct-1 (6), 2, 4-trien-3-ol (567.2 mg,2.85 mmol), magnesium chloride (407 mg,4.28 mmol) and TEA (1.15 g,11.4 mmol) were added to acetonitrile (5 mL), and reacted at a temperature of 40℃for 30min, paraformaldehyde (770 mg,8.55 mmol) was added, and reacted at 80℃overnight. Cooled to room temperature, water 50mL,4n hydrochloric acid was added to adjust the pH of the reaction to 5, dcm (50 ml×2) was added, the combined organic phases were concentrated under reduced pressure and column chromatography gave the title compound 5-bromo-3-hydroxybicyclo [4.2.0] oct-1 (6), 2, 4-triene-2-carbaldehyde (517.6 mg, 80%).
Sixth step: preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5,6,10,11-tetrahydrocyclobutano [5,6] benzo [1,2-f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5,6,10,11-tetrahydrocyclobutano [5,6] benzo [1,2-f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference example 1.
MS m/z(ESI):434.2[M+H] + .
Example 11
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5,6,10,11-tetrahydrocyclobutano [5,6] benzo [1,2-f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -2-methoxyacetamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5,6,10,11-tetrahydrocyclobutano [5,6] benzo [1,2-f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -2-methoxyacetamide reference example 10.
MS m/z(ESI):450.1[M+H] + .
Example 12
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5,6,11, 12-tetrahydro-10H-imidazo [1,2-d ] indeno [4,5-f ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5,6,11, 12-tetrahydro-10H-imidazo [1,2-d ] indeno [4,5-f ] [1,4] oxazepin-9-yl) amino) propanamide reference example 10.
MS m/z(ESI):448.1[M+H] + .
Example 13
Preparation of (S) -2- ((11- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -7, 8-dihydro- [1,3] dioxa [4',5':5,6] benzo [1,2-f ] imidazo [1,2-d ] [1,4] oxazepin-4-yl) amino) propanamide
Preparation of (S) -2- ((11- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -7, 8-dihydro- [1,3] dioxino [4',5':5,6] benzo [1,2-f ] imidazo [1,2-d ] [1,4] oxazepin-4-yl) amino) propanamide reference example 10.
MS m/z(ESI):452.1[M+H] + .
Example 14
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methylbutanamide
The preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methylbutanamide is described in example 1.
1 H NMR(400MHz,CD 3 OD)δ1.09(t,J＝6.1Hz,6H),2.13(d,J＝7.0Hz,1H),3.60(d,J＝6.4Hz,1H),4.38(d,J＝19.3Hz,4H),4.68-4.60(m,3H),6.27(s,1H),6.43-6.78(m,2H),7.17(s,1H),8.06(d,J＝8.7Hz,1H)；
MS m/z(ESI):436.1[M+H] + .
Example 15
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -2-methoxyacetamide
The preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -2-methoxyacetamide is described in example 1.
MS m/z(ESI):424.1[M+H] + .
Example 16
Preparation of (R) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-fluoropropionamide
The preparation of (R) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-fluoropropionamide is described in example 1.
MS m/z(ESI):426.1[M+H] + .
Example 17
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -2- (oxetan-3-yl) acetamide
The preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -2- (oxetan-3-yl) acetamide is described in example 1.
1 H NMR(400MHz,CD 3 OD)δ3.26-3.33(m,2H),4.08(d,J＝9.6Hz,1H),4.22-4.25(m,2H),4.29-4.31(m,2H),4.40-4.50(m,5H),4.61-4.69(m,1H),6.18(d,J＝2.2Hz,1H),6.44-6.50(m,2H),7.06(s,1H),7.97(d,J＝8.8Hz,1H)；
MS m/z(ESI):450.1[M+H] + .
Example 18
Preparation of (S) -2- ((2- (4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -2-methylpropanamide
The preparation of (S) -2- ((2- (4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -2-methylpropanamide is described in example 1.
1 H NMR(400MHz,CD 3 OD)δ1.50(s,6H),4.31-4.36(m,2H),4.38-4.43(m,2H),4.61-4.65(m,2H),4.95(d,J＝10.6Hz,1H),6.19(d,J＝2.2Hz,1H),6.64-6.81(m,2H),7.17(s,1H),8.05(d,J＝8.8Hz,1H)；
MS m/z(ESI):422.1[M+H] + .
Example 19
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) (methyl) amino) propanamide
The preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) (methyl) amino) propanamide is described in example 1.
1 H NMR(400MHz,CD 3 OD):δ1.40(d,J＝6.8Hz,3H),2.90(s,3H),4.37-4.64(m,7H),4.96(m,1H),6.41(s,1H),6.46-6.74(m,2H),7.16(s,1H),8.13(d,J＝9.2Hz,1H)；
MS m/z(ESI):422.1[M+H] + .
Example 20
Preparation of (S) -3- ((2- (4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) oxabutylen-3-carboxamide
The preparation of (S) -3- ((2- (4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) oxabutylen-3-carboxamide is described in example 1.
1 H NMR(400MHz,CD 3 OD)δ4.35(m,4H),4.63(m,4H),4.90(m,1H),5.10(d,J＝8.0Hz,2H),5.90(s,1H),6.29(d,J＝8.0Hz,1H),6.59(t,J＝56Hz,1H),7.16(s,1H),8.10(d,J＝8.0Hz,1H)；
MS m/z(ESI):436.1[M+H] + .
Example 21
Preparation of (S) -1- (2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) azetidine-2-carboxamide
The preparation of (S) -1- (2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) azetidine-2-carboxamide is described in example 1.
1 H NMR(400MHz,CD 3 OD)δ2.30-2.40(m,1H),2.52-2.58(m,1H),3.66-3.72(m,1H),3.91-3.96(m,1H),4.22-4.27(m,2H),4.28-4.34(m,2H),4.48-4.59(m,2H),4.79-4.85(m,2H),6.00(d,J＝2.2Hz,1H),6.20-6.22(m,1H),6.37-6.65(m,1H),7.08(s,1H),8.06(d,J＝8.7Hz,1H).
MS m/z(ESI):420.1[M+H] + .
Example 22
Preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The first step: preparation of (S) -3- (9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) oxazolidine-2-thione
To a toluene solution (10 mL) of (S) -3- (10-bromo-6, 7-dihydro-5H-benzo [ b ] imidazo [2,1-d ] [1,5] oxazocine-2-yl) -4- (difluoromethyl) oxazolidin-2-one (100 mg,0.25 mmol) was added Lawson reagent (1.01 g,2.5 mmol) and reacted at microwave 140℃for three hours. After cooling to room temperature, the filter cake was washed with EtOAc (20 mL), the filtrate was dried over anhydrous sodium sulfate, concentrated under reduced pressure and column chromatographed to give the title compound (S) -3- (9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-thione (42 mg, 40%).
1 H NMR(400MHz,DMSO-d 6 )δ4.43-4.52(m,4H),4.79-4.86(m,2H),5.24-5.35(m,1H),6.57-6.85(m,1H),7.23-7.38(m,2H),8.10(s,1H),8.26(d,J＝8.6Hz,1H)；
MS m/z(ESI):416.1[M+H] + .
And a second step of: preparation of (R) -3- (9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) thiazolidin-2-one
To a toluene solution (1 mL) of (S) -3- (10-bromo-6, 7-dihydro-5H-benzo [ b ] imidazo [2,1-d ] [1,5] oxazocine-2-yl) -4- (difluoromethyl) oxazolidine-2-thione (33 mg,0.079 mmol) was added dichloro (p-cymene) ruthenium (II) dimer (14.7 mg,0.024 mmol), and 2-dicyclohexylphosphine-2 ',6' -dimethoxybiphenyl (9.7 mg,0.024 mmol), and the mixture was reacted at 110℃for 12 hours under an air atmosphere. The reaction was cooled to room temperature, diluted with EtOAc, and the organic phase was washed with saturated aqueous sodium chloride, dried over anhydrous sodium sulfate, and concentrated under reduced pressure then column chromatography to give the title compound (R) -3- (9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) thiazolidin-2-one (26 mg, 79%).
1 H NMR(400MHz,CDCl 3 )δ3.57-3.72(m,2H),4.28-4.41(m,2H),4.44-4.47(m,2H)5.14-5.24(m,1H),6.29-6.67(m,1H),7.14-7.25(m,2H),7.42(s,1H),8.21(d,J＝8.8Hz,1H)；
MS m/z(ESI):416.1[M+H] + .
And a third step of: preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
(R) -3- (9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) thiazolidin-2-one (26 mg,0.062 mmol), L-alanine (19.5 mg,0.22 mmol), cuprous iodide (6 mg,0.03 mmol), potassium phosphate (40 mg,0.19 mmol), and dimethyl sulfoxide (3 mL) were mixed, the reaction system was reacted three times with nitrogen with air replaced at 100℃for 12 hours, cooled to room temperature, ammonium chloride (20 mg,0.37 mmol), triethylamine (95 mg,0.94 mmol) were added, stirring was performed for 5 minutes, 2- (7-oxybenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate (212 mg,0.56 mmol) was added, stirring was performed at room temperature for 2 hours, filtration was performed, a saturated aqueous solution was added, and ethyl acetate was extracted three times. The organic phases were combined, dried over anhydrous sodium sulfate and concentrated under reduced pressure followed by column chromatography to give the title compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonylthiazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide (15 mg, 56%).
1 H NMR(400MHz,CD 3 OD)δ1.37(d,J＝7.2Hz,3H),3.57-3.61(m,1H),3.83-3.87(m,2H),4.33-4.41(m,4H),5.12-5.19(m,1H),6.15-6.17(m,1H),6.47-6.52(m,2H),7.28(s,1H),8.10(d,J＝8.8Hz,1H)；
MS m/z(ESI):424.1[M+H] + .
Example 23
Preparation of (S) -2- ((2- ((S) -5- (difluoromethyl) -2-carbonylimidazolidin-1-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -5- (difluoromethyl) -2-carbonylimidazolidin-1-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide the procedure of example 22 is referred to.
MS m/z(ESI):407.2[M+H] + .
Example 24
Preparation of (S) -2- ((2- ((S) -5- (difluoromethyl) -3-methyl-2-carbonylimidazolidin-1-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -5- (difluoromethyl) -3-methyl-2-carbonylimidazolidin-1-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 22.
1 H NMR(400MHz,CD 3 OD)δ1.46(d,J＝7.0Hz,3H),2.85(s,3H),3.62-3.68(m,2H),3.79-3.85(m,1H),4.27-4.30(m,2H),4.35-4.37(m,2H),4.63-4.69(m,1H),6.17(d,J＝2.0Hz,1H),6.34-6.62(m,2H),7.05(s,1H),8.01(d,J＝8.8Hz,1H)；
MS m/z(ESI):421.2[M+H] + .
Example 25
Preparation of (S) -2- ((2- ((4S, 5R) -4- (difluoromethyl) -5-methyl-2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The first step: preparation of methyl (4S, 5R) -5-methyl-2-carbonyl oxazolidine-4-carboxylate
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Methyl L-threonine hydrochloride (500 mg,2.95 mmol) was dissolved in dichloromethane (15 mL), cooled to 0deg.C with an ice-water bath, triphosgene (289 mg,0.97 mmol) was added, a solution of triethylamine (895 mg,8.84 mmol) in dichloromethane (2 mL) was added dropwise, after the addition, the reaction was carried out at 0deg.C for 1 hour, water was added, extraction was performed with dichloromethane, the organic phase was dried over anhydrous sodium sulfate, the organic solvent was concentrated under reduced pressure, and crude product was purified by column chromatography to give the title compound methyl (4S, 5R) -5-methyl-2-carbonyloxazolidine-4-carboxylate (251 mg, 53%).
MS m/z(ESI):160.1[M+H] + .
And a second step of: preparation of methyl (4 s,5 r) -3-benzyl-5-methyl-2-carbonyloxazolidine-4-carboxylate
Methyl (4S, 5R) -5-methyl-2-carbonyloxazolidine-4-carboxylate (200 mg,1.26 mmol) was dissolved in DMF (5 mL), cooled to-15℃and NaH (60% kerosene, 50mg,1.26 mmol) was added and stirred at this temperature for one hour, benzyl bromide (322 mg,1.89 mmol) was added and stirring was continued for 2 hours, water quench reaction was added, extraction with dichloromethane, the organic phase dried over anhydrous sodium sulfate, and crude column chromatography purification of the title compound methyl (4S, 5R) -3-benzyl-5-methyl-2-carbonyloxazolidine-4-carboxylate (260 mg, 83%) was obtained after concentrating the organic solvent under reduced pressure.
MS m/z(ESI):250.1[M+H] + .
And a third step of: preparation of (4R, 5R) -3-benzyl-4- (hydroxymethyl) -5-methyl oxazolidin-2-one
(4S, 5R) -3-benzyl-5-methyl-2-carbonyloxazolidine-4-carboxylate (260 mg,1.0 mmol) was dissolved in methanol (5 mL), cooled to 0deg.C in ice water bath, sodium borohydride (11 mg,3.1 mmol) was added in portions, gradually warmed to room temperature and reacted for 2 hours, after concentrating the reaction solution, the crude product was purified by column chromatography to give the title compound (4R, 5R) -3-benzyl-4- (hydroxymethyl) -5-methyloxazolid-2-one (180 mg, 78%).
MS m/z(ESI):222.1[M+H] + .
Fourth step: preparation of (4 s,5 r) -3-benzyl-5-methyl-2-carbonyl oxazolidine-4-carbaldehyde
(4R, 5R) -3-benzyl-4- (hydroxymethyl) -5-methyl oxazolidin-2-one (180 mg,0.81 mmol), IBX (683 mg,2.44 mmol) were mixed in ethyl acetate (10 mL) and reacted for 3h at 85℃under nitrogen. After cooling, the reaction solution was filtered, and the reaction solution was concentrated under reduced pressure to obtain 178mg of crude (4 s,5 r) -3-benzyl-5-methyl-2-carbonyloxazolidine-4-carbaldehyde, which was directly used in the next reaction.
MS m/z(ESI):220.2[M+H] + .
Fifth step: preparation of (4S, 5R) -3-benzyl-4- (difluoromethyl) -5-methyl-oxazolidin-2-one
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(4S, 5R) -3-benzyl-5-methyl-2-carbonyl oxazolidine-4-formaldehyde (178 mg,0.81 mmol) is dissolved in methylene chloride (10 mL), cooled to 0 ℃ in an ice-water bath under the protection of nitrogen, DAST (262 mg,1.62 mmol) is added dropwise to the reaction solution, and the temperature is naturally raised to room temperature for reaction for 3h. The reaction solution was slowly added dropwise to a pre-cooled saturated aqueous sodium bicarbonate solution. Extraction with dichloromethane (20 mL x 2), combining the organic phases and drying over anhydrous sodium sulfate, concentration of the organic solvent under reduced pressure followed by column chromatography gave the title compound (4 s,5 r) -3-benzyl-4- (difluoromethyl) -5-methyl-oxazolidin-2-one (110 mg, two step yield 56%).
1 H NMR(400MHz,CDCl 3 )δ1.33(d,J＝6.4Hz,3H),3.27-3.33(m,1H),4.16-4.20(m,1H),4.41-4.64(m,1H),4.91(d,J＝15.0Hz,1H),5.56-5.88(m,1H),7.27-7.44(m,5H)；
MS m/z(ESI):242.1[M+H] + .
Sixth step: preparation of (4S, 5R) -4- (difluoromethyl) -5-methyl-oxazolidin-2-one
(4S, 5R) -3-benzyl-4- (difluoromethyl) -5-methyl-oxazolidin-2-one (110 mg,0.46 mmol) was dissolved in mesitylene (2 mL), methanesulfonic acid (438 mg,4.56 mmol) was added, and the mixture was heated to 135℃for 5 hours. The reaction was cooled to room temperature, and the reaction solution was slowly added dropwise to a pre-cooled saturated aqueous sodium bicarbonate solution. Extraction with dichloromethane (20 mL x 2), drying of the combined organic phases over anhydrous sodium sulfate, concentration of the organic solvent under reduced pressure followed by column chromatography gave 68mg of crude title compound (4 s,5 r) -4- (difluoromethyl) -5-methyl-oxazolidin-2-one, which was used directly in the next reaction.
MS m/z(ESI):152.1[M+H] + .
Seventh step: preparation of (4S, 5R) -3- (9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) -5-methyl oxazolidin-2-one
9-bromo-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d][1,4]Oxazepin (100 mg,0.25 mmol), (4S, 5R) -4- (difluoromethyl) -5-methyl-oxazolidin-2-one (38.5 mg,0.25 mmol), (1R, 2R) -N 1 ,N 2 Dimethylcyclohexane-1, 2-diamine (22 mg,0.15 mmol), cuprous iodide (14 mg,0.08 mmol), potassium phosphate (108 mg,051 mmol), and dimethyl sulfoxide (3 mL) were mixed and reacted at 130℃for 3 hours. The reaction was cooled to room temperature, 15% aqueous ammonia (5 mL) was added, stirred for 5 minutes, and extracted three times with ethyl acetate. The organic phases were combined, then the organic phase was washed with saturated sodium chloride, dried over anhydrous sodium sulfate, and the organic solvent was concentrated under reduced pressure to give a column layerThe title compound (S) -3- (9-bromo-3-fluoro-5, 6-dihydrobenzo [ f) is obtained by chromatography]Imidazo [1,2-d][1,4]Oxazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one (61 mg, 57%).
MS m/z(ESI):414.2[M+H] + .
Eighth step: preparation of (S) -2- ((2- ((4S, 5R) -4- (difluoromethyl) -5-methyl-2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
(4S, 5R) -3- (9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) -5-methyl oxazolidin-2-one (61 mg,0.15 mmol), L-alanine (39 mg,0.44 mmol), cuprous iodide (14 mg,0.07 mmol), potassium phosphate (94 mg,0.44 mmol), and mixed in dimethyl sulfoxide (5 mL), the reaction system was reacted with nitrogen gas three times with air replacement, at 100℃for 5 hours, cooled to room temperature, ammonium chloride (47 mg,0.88 mmol), triethylamine (223 mg,2.21 mmol), 2- (7-benzotriazol oxide) -N, N, N ', N' -tetramethylurea hexafluorophosphate (505 mg,1.33 mmol) was added, stirred at room temperature for 2 hours, filtered, saturated aqueous solution was added, and ethyl acetate was extracted three times. The organic phases were combined, dried over anhydrous sodium sulfate, and the organic solvent was concentrated under reduced pressure and separated by column chromatography to give the title compound (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -6, 7-dihydro-5H-benzo [ b ] imidazo [2,1-d ] [1,5] oxazocine-10-yl) amino) propanamide (33 mg, 53%).
1 H NMR(400MHz,CD 3 OD)δ1.46(d,J＝6.8Hz,3H),1.53(d,J＝6.2Hz,3H),3.79-3.85(m,1H),4.32-4.39(m,4H),4.46-4.55(m,1H),4.93-4.95(m,1H),6.17(s,1H),6.39-6.72(m,2H),7.14(s,1H),8.03(d,J＝8.6Hz,1H)；
MS m/z(ESI):422.1[M+H] + .
Example 26
Preparation of (R) -2- ((2- ((4S, 5R) -4- (difluoromethyl) -5-methyl-2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (R) -2- ((2- ((4S, 5R) -4- (difluoromethyl) -5-methyl-2-carbonyloxazolid-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide the procedure of example 25 is referred to.
MS m/z(ESI):422.2[M+H] + .
Example 27
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -5, 5-dimethyl-2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -5, 5-dimethyl-2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide is described in example 25.
MS m/z(ESI):436.2[M+H] + .
Example 28
Preparation of (S) -2- ((2- ((S) -7- (difluoromethyl) -5-carbonyl-4-oxa-6-azaspiro [2.4] heptan-6-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -7- (difluoromethyl) -5-oxo-4-oxa-6-azaspiro [2.4] heptan-6-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 25.
MS m/z(ESI):434.2[M+H] + .
Example 29
Preparation of (S) -2- ((2- ((S) -8- (difluoromethyl) -6-carbonyl-2, 5-dioxa-7-azaspiro [3.4] oct-7-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -8- (difluoromethyl) -6-carbonyl-2, 5-dioxa-7-azaspiro [3.4] oct-7-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide the procedure described in example 25.
MS m/z(ESI):450.2[M+H]+.
Example 30
Preparation of (S) -2- ((2- ((S) -5- (difluoromethyl) -3-methyl-2, 4-dicarbonyl imidazolidin-1-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The preparation of (S) -2- ((2- ((S) -5- (difluoromethyl) -3-methyl-2, 4-dicarbonyimidazolidin-1-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide is described in example 1.
MS m/z(ESI):435.2[M+H] + .
Example 31
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The first step: preparation of 4-bromo-3-fluoro-2-methoxybenzaldehyde
To a solution of 4-bromo-2, 3-difluorobenzaldehyde (2.0 g,9.05 mmol) in methanol (25 mL) at room temperature was added sodium methoxide (733 mg,13.56 mmol), and the mixture was reacted at 65℃for 2 hours, concentrated, and purified by column chromatography to give 4-bromo-3-fluoro-2-methoxybenzaldehyde (1.78 g, 85%).
MS m/z(ESI):233.0[M+H] + 。
And a second step of: preparation of 4-bromo-3-fluoro-2-hydroxybenzaldehyde
To a solution of 4-bromo-3-fluoro-2-methoxybenzaldehyde (1.78 g,7.67 mmol) in acetic acid (15 mL) at room temperature was added hydrobromic acid (8.7 mL, 48%), the mixture was reacted at 120℃for 16 hours, the reaction mixture was cooled and concentrated under reduced pressure, then water and ethyl acetate were added to the reaction flask, the organic phase was dried over anhydrous sodium sulfate, and after concentrating the organic solvent under reduced pressure, the 4-bromo-3-fluoro-2-hydroxybenzaldehyde (1.12 g, 67%) was isolated and purified by column chromatography.
MS m/z(ESI):219.0[M+H] + 。
And a third step of: preparation of 3-bromo-2-fluoro-6- (1H-imidazol-2-yl) phenol
To a methanol solution (12 mL) of 4-bromo-3-fluoro-2-hydroxybenzaldehyde (1.12 g,5.14 mmol) was added an aqueous glyoxal solution (40 wt.%,3.73g,25.7 mmol), and then aqueous ammonia (28 wt.%,5.14g,51.4 mmol) was slowly added dropwise with stirring in a water bath for 30 minutes, and the temperature of the reaction solution was controlled to not exceed 40 ℃. The mixture was then stirred at 35℃for two days, cooled, and the organic solvent was removed under reduced pressure, and purified by column chromatography to give 3-bromo-2-fluoro-6- (1H-imidazol-2-yl) phenol (1.31 g, 100%).
MS m/z(ESI):257.0[M+H] + .
Fourth step: preparation of 9-bromo-8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
3-bromo-2-fluoro-6- (1H-imidazol-2-yl) phenol (1.31 g,5.14 mmol), cesium carbonate (6.3 g,19.53 mmol), 1, 2-dibromoethane (3.6 g,19.12 mmol) were mixed in DMF (12 mL), stirred at 85℃overnight, and the reaction solution was cooled and diluted with ethyl acetate. The organic phase was washed with saturated brine several times, then dried over anhydrous sodium sulfate, concentrated in vacuo and purified by column chromatography to give the title compound 9-bromo-8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (995 mg, 69%).
MS m/z(ESI):283.0[M+H] + .
Fifth step: preparation of 9-bromo-8-fluoro-2, 3-diiodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
NIS (2.23 g,9.88 mmol) was added to a solution of 9-bromo-8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (995 mg,3.53 mmol) in DMF (8 mL) at room temperature, followed by stirring overnight at 60 ℃. Cooling, adding water, and separating out solid. After filtration, the solid was dissolved in ethyl acetate, washed with 1M aqueous NaOH and saturated brine in this order, dried over anhydrous sodium sulfate, and concentrated to give the title compound 9-bromo-8-fluoro-2, 3-diiodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxaazepine (1.79 g, 94%).
MS m/z(ESI):534.7[M+H] + .
Sixth step: preparation of 9-bromo-8-fluoro-2-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
EtMgBr (1.0M in THF, 1.23mL,3.69 mmol) was slowly added dropwise to a solution of 9-bromo-8-fluoro-2, 3-diiodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (1.79 g,3.35 mmol) in THF (10 mL) at-20deg.C. After the completion of the dropwise addition, the mixture was stirred at-15℃for 3 hours. Slowly warmed to room temperature, then saturated aqueous ammonium chloride was added dropwise, stirred for 15 minutes, and extracted with ethyl acetate multiple times. The organic phases were combined, washed with saturated brine, separated, dried over anhydrous sodium sulfate, and concentrated in vacuo to give the title compound 9-bromo-8-fluoro-2-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (610 mg, 45%) by column chromatography.
MS m/z(ESI):408.9[M+H] + .
Seventh step: preparation of (S) -4- (difluoromethyl) -3- (8-fluoro-9-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) oxazolidin-2-one
9-bromo-8-fluoro-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d][1,4]Oxazepine (300 mg,0.74 mmol), (S) -4- (difluoromethyl) oxazolidin-2-one (102 mg,0.74 mmol), (1R, 2R) -N 1 ,N 2 Dimethylcyclohexane-1, 2-diamine (42 mg,0.30 mmol), cuprous iodide (28 mg,0.15 mmol), potassium carbonate (205 mg,1.5 mmol), and mixed in 1, 4-dioxane (6 mL), the reaction system was reacted with nitrogen for 5 hours at 105℃with replacement of tertiary air, cooled to room temperature, 15% aqueous ammonia was added, stirred for 5 minutes, and extracted three times with EtOAc. The organic phases were combined, then the organic phase was washed with saturated aqueous sodium chloride, dried over anhydrous sodium sulfate, concentrated under reduced pressure and then column chromatographed to give the title compound (S) -4- (difluoromethyl) -3- (8-fluoro-9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d][1,4]Oxazepin-2-yl) oxazolidin-2-one (225 mg, 65%).
MS m/z(ESI):466.0[M+H] + 。
The subsequent preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide is described in example 1.
1 H NMR(400MHz,CD 3 OD)δ1.50(d,J＝7.0Hz,3H),3.95-4.01(m,1H),4.36-4.41(m,2H),4.47-4.53(m,2H),4.57-4.67(m,2H),4.93-4.98(m,1H),6.37-6.42(m,1H),6.44-6.73(m,1H),7.20(s,1H),7.87-7.91(m,1H)；
MS m/z(ESI):426.1[M+H] + .
Example 32
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -11-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -11-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 31.
1 H NMR(400MHz,CD 3 OD)δ1.46(d,J＝4.0Hz,3H),3.84(m,1H),4.24(m,2H),4.49(m,2H),4.60(m,3H),6.19(s,1H),6.28(d,J＝8.0Hz,1H),6.49(t,J＝56Hz,1H),7.30(s,1H)；
MS m/z(ESI):426.1[M+H] + .
Example 33
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -10-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -10-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 31.
1 H NMR(400MHz,CD 3 OD):δ1.52(d,J＝6.8Hz,3H),3.86-3.96(m,1H),4.30-4.42(m,4H),4.60-4.69(m,3H),4.91-5.00(m,1H),6.19-6.25(m,1H),6.46-6.76(m,1H),7.18(s,1H),8.04(d,J＝13.4Hz,1H).
MS m/z(ESI):426.1[M+H] + .
Example 34
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -8-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -8-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 31.
1 H NMR(400MHz,CD 3 OD)δ1.51(d,J＝6.9Hz,3H),2.15(s,3H),3.99-4.02(m,1H),4.33-4.37(m,2H),4.43-4.47(m,2H),4.55-4.68(m,2H),4.93-4.97(m,1H),6.36(d,J＝8.9Hz,1H),6.43-6.71(m,1H),7.19(s,1H),7.94(d,J＝8.8Hz,1H)；
MS m/z(ESI):422.1[M+H] + .
Example 35
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -11-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -11-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 31.
MS m/z(ESI):422.1[M+H] + .
Example 36
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -10-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -10-methyl-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 31.
1 H NMR(400MHz,CD 3 OD):δ1.52(d,J＝6.9Hz,3H),2.19(s,3H),3.85-3.93(m,1H),4.25-4.36(m,4H),4.55-4.67(m,2H),4.92-4.96(m,1H),6.09(s,1H),6.43-6.71(m,1H),7.12(s,1H),7.90(s,1H).
MS m/z(ESI):422.1[M+H] + .
Example 37
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -8-methoxy-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -8-methoxy-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 31.
MS m/z(ESI):438.1[M+H] + .
Example 38
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -11-methoxy-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -11-methoxy-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 31.
MS m/z(ESI):438.1[M+H] + .
Example 39
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -10-methoxy-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -10-methoxy-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 31.
MS m/z(ESI):438.1[M+H] + .
Example 40
Preparation of (S) -2- ((8-cyano-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((8-cyano-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 31.
MS m/z(ESI):433.1[M+H] + .
Example 41
Preparation of (S) -2- ((11-cyano-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((11-cyano-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 31.
MS m/z(ESI):433.1[M+H] + .
Example 42
Preparation of (S) -2- ((10-cyano-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((10-cyano-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide reference is made to example 31.
MS m/z(ESI):433.1[M+H] + .
Example 43
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methoxypropionamide
The preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methoxypropionamide is described in example 1.
1 H NMR(400MHz,CD 3 OD)δ3.39(s,3H),3.67-3.76(m,2H),3.94-3.98(m,1H),4.30-4.34(m,2H),4.37-4.41(m,2H),4.57-4.66(m,2H),4.91-4.96(m,1H),6.21-6.25(m,1H),6.43-6.46(m,1H),6.48-6.73(m,1H),7.15(s,1H),8.06(d,J＝8.8Hz,1H)；
MS m/z(ESI):438.2[M+H] + .
Example 44
Preparation of (2S, 3 r) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methoxybutyramide
The preparation method of (2S, 3 r) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methoxybutyramide is described in example 1.
1 H NMR(400MHz,CD 3 OD):δ1.23-1.27(d,J＝6.9Hz,3H),3.39(s,3H),3.75-3.80(m,1H),3.88-3.93(m,1H),4.29-4.43(m,4H),4.56-4.68(m,2H),4.89-4.98(m,1H),6.22-6.25(m,1H),6.43-6.74(m,2H),7.15(s,1H),8.03-8.08(d,J＝8.8Hz,1H)；
MS m/z(ESI):452.2[M+H] + .
Example 45
Preparation of (2S, 3S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methoxybutyramide
The preparation of (2S, 3S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methoxybutyramide is described in example 1.
MS m/z(ESI):452.2[M+H] + .
Example 46
Preparation of (S) -2- ((2- ((S) -2- (difluoromethyl) -5-carbonyl pyrrolidin-1-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The preparation of (S) -2- ((2- ((S) -2- (difluoromethyl) -5-carbonyl pyrrolidin-1-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide is described in example 1.
1 H NMR(400MHz,DMSO-d 6 )δ1.30(d,J＝8.0Hz,3H),2.20-2.45(m,3H),3.31(d,J＝8.0Hz,1H),3.76(t,J＝7.6Hz,1H),4.32-4.36(m,4H),4.69-4.78(m,1H),6.08(s,1H),6.15(d,J＝8.0Hz,1H),6.41(d,J＝8.0Hz,1H),6.66(t,J＝56Hz,1H),7.00(s,1H),7.38(d,J＝8.0Hz,1H),7.40(s,1H),8.00(d,J＝8.0Hz,1H)；
MS m/z(ESI):406.2[M+H] + .
Example 47
Preparation of (S) -2- ((2- ((3S, 5S) -5- (difluoromethyl) -3-methoxy-2-carbonyl pyrrolidin-1-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The preparation of (S) -2- ((2- ((3S, 5S) -5- (difluoromethyl) -3-methoxy-2-carbonyl pyrrolidin-1-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide is described in example 1.
1 H NMR(400MHz,CD 3 OD)δ1.46(d,J＝7.0Hz,3H),2.10-2.20(m,1H),2.74-2.84(m,1H),3.57(s,3H),3.81(q,J＝7.0Hz,1H),4.25-4.40(m,5H),4.71-4.84(m,1H),6.13-6.18(m,1H),6.37-6.70(m,2H),7.38(s,1H),8.04(d,J＝8.8Hz,1H)；
MS m/z(ESI):436.2[M+H] + .
Example 48
Preparation of (S) -2- ((2- ((3R, 5S) -5- (difluoromethyl) -3-methoxy-2-carbonyl pyrrolidin-1-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The preparation of (S) -2- ((2- ((3 r, 5S) -5- (difluoromethyl) -3-methoxy-2-carbonyl pyrrolidin-1-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide is described in example 1.
MS m/z(ESI):436.2[M+H] + .
Example 49
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-hydroxypropionamide
The preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-hydroxypropionamide is described in example 1.
1 H NMR(400MHz,CD 3 OD)δ3.87(s,2H),4.34(d,J＝4.3Hz,2H),4.37-4.43(m,2H),4.62(m,4H),6.23(d,J＝2.6Hz,1H),6.41-6.62(m,2H),7.16(s,1H),8.06(d,J＝8.8Hz,1H)；
MS m/z(ESI):424.1[M+H] + .
Example 50
Preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
The first step: preparation of 4-bromo-3-fluoro-2-methoxybenzaldehyde
To a solution of 4-bromo-2, 3-difluorobenzaldehyde (2.0 g,9.05 mmol) in methanol (25 mL) at room temperature was added sodium methoxide (733 mg,13.56 mmol), and the mixture was reacted at 65℃for 2 hours, concentrated, and purified by column chromatography to give 4-bromo-3-fluoro-2-methoxybenzaldehyde (1.78 g, 85%).
MS m/z(ESI):233.0[M+H] + 。
And a second step of: preparation of 4-bromo-3-fluoro-2-hydroxybenzaldehyde
To a solution of 4-bromo-3-fluoro-2-methoxybenzaldehyde (1.78 g,7.67 mmol) in acetic acid (15 mL) at room temperature was added hydrobromic acid (8.7 mL, 48%), the mixture was reacted at 120℃for 16 hours, the reaction mixture was cooled and concentrated under reduced pressure, then water and ethyl acetate were added to the reaction flask, the organic phase was dried over anhydrous sodium sulfate, and after concentrating the organic solvent under reduced pressure, the 4-bromo-3-fluoro-2-hydroxybenzaldehyde (1.12 g, 67%) was isolated and purified by column chromatography.
MS m/z(ESI):219.0[M+H] + 。
And a third step of: preparation of 3-bromo-2-fluoro-6- (1H-imidazol-2-yl) phenol
To a methanol solution (12 mL) of 4-bromo-3-fluoro-2-hydroxybenzaldehyde (1.12 g,5.14 mmol) was added an aqueous glyoxal solution (40 wt.%,3.73g,25.7 mmol), and then aqueous ammonia (28 wt.%,5.14g,51.4 mmol) was slowly added dropwise with stirring in a water bath for 30 minutes, and the temperature of the reaction solution was controlled to not exceed 40 ℃. The mixture was then stirred at 35℃for two days, cooled, and the organic solvent was removed under reduced pressure, and purified by column chromatography to give 3-bromo-2-fluoro-6- (1H-imidazol-2-yl) phenol (1.31 g, 100%).
MS m/z(ESI):257.0[M+H] + .
Fourth step: preparation of 9-bromo-8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
3-bromo-2-fluoro-6- (1H-imidazol-2-yl) phenol (1.31 g,5.14 mmol), cesium carbonate (6.3 g,19.53 mmol), 1, 2-dibromoethane (3.6 g,19.12 mmol) were mixed in DMF (12 mL), stirred at 85℃overnight, and the reaction solution was cooled and diluted with ethyl acetate. The organic phase was washed with saturated brine several times, then dried over anhydrous sodium sulfate, concentrated in vacuo and purified by column chromatography to give the title compound 9-bromo-8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (995 mg, 69%).
MS m/z(ESI):283.0[M+H] + .
Fifth step: preparation of 9-bromo-8-fluoro-2, 3-diiodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
NIS (2.23 g,9.88 mmol) was added to a solution of 9-bromo-8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (995 mg,3.53 mmol) in DMF (8 mL) at room temperature, followed by stirring overnight at 60 ℃. Cooling, adding water, and separating out solid. After filtration, the solid was dissolved in ethyl acetate, washed with 1M aqueous NaOH and saturated brine in this order, dried over anhydrous sodium sulfate, and concentrated to give the title compound 9-bromo-8-fluoro-2, 3-diiodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxaazepine (1.79 g, 94%).
MS m/z(ESI):534.7[M+H] + .
Sixth step: preparation of 9-bromo-8-fluoro-2-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine
EtMgBr (1.0M in THF, 1.23mL,3.69 mmol) was slowly added dropwise to a solution of 9-bromo-8-fluoro-2, 3-diiodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (1.79 g,3.35 mmol) in THF (10 mL) at-20deg.C. After the completion of the dropwise addition, the mixture was stirred at-15℃for 3 hours. Slowly warmed to room temperature, then saturated aqueous ammonium chloride was added dropwise, stirred for 15 minutes, and extracted with ethyl acetate multiple times. The organic phases were combined, washed with saturated brine, separated, dried over anhydrous sodium sulfate, and concentrated in vacuo to give the title compound 9-bromo-8-fluoro-2-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepine (610 mg, 45%) by column chromatography.
MS m/z(ESI):408.9[M+H] + .
Seventh step: preparation of (S) -4- (difluoromethyl) -3- (8-fluoro-9-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) oxazolidin-2-one
9-bromo-8-fluoro-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d][1,4]Oxazepine (300 mg,0.74 mmol), (S) -4- (difluoromethyl) oxazolidin-2-one (102 mg,0.74 mmol), (1R, 2R) -N 1 ,N 2 Dimethylcyclohexane-1, 2-diamine (42 mg,0.30 mmol), cuprous iodide (28 mg,0.15 mmol), potassium carbonate (205 mg,1.5 mmol), and mixed in 1, 4-dioxane (6 mL), the reaction system was reacted with nitrogen for 5 hours at 105℃with replacement of tertiary air, cooled to room temperature, 15% aqueous ammonia was added, stirred for 5 minutes, and extracted three times with EtOAc. The organic phases were combined, then the organic phase was washed with saturated aqueous sodium chloride, dried over anhydrous sodium sulfate, concentrated under reduced pressure and then column chromatographed to give the title compound (S) -4- (difluoromethyl) -3- (8-fluoro-9-iodo-5, 6-dihydrobenzo [ f)]Imidazo [1,2-d][1,4]Oxazepin-2-yl) oxazolidin-2-one (225 mg, 65%).
MS m/z(ESI):466.0[M+H] + 。
Eighth step: preparation of (S) -4- (difluoromethyl) -3- (8-fluoro-9-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) oxazolidin-2-thione
To a toluene solution (20 mL) of (S) -4- (difluoromethyl) -3- (8-fluoro-9-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) oxazolidin-2-one (220 mg,0.47 mmol) was added Lawson reagent (1.92 g,4.73 mmol), and the mixture was reacted at 145℃for 6 hours. After cooling to room temperature, the filter cake was washed with EtOAc (20 mL), the filtrate was dried over anhydrous sodium sulfate, concentrated under reduced pressure and column chromatographed to give the title compound (S) -3- (9-bromo-8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-thione (105 mg, 46%).
MS m/z(ESI):482.1[M+H] + .
Ninth step: preparation of (R) -4- (difluoromethyl) -3- (8-fluoro-9-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) thiazolidin-2-one
To a toluene solution (3 mL) of (S) -3- (9-bromo-8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) oxazolidine-2-thione (105 mg,0.22 mmol) was added dichloro (p-cymene) ruthenium (II) dimer (27 mg,0.045 mmol), and 2-dicyclohexylphosphine-2 ',6' -dimethoxybiphenyl (27 mg,0.065 mmol) and the mixture was reacted at 115℃for 16 hours under an air atmosphere. The reaction was cooled to room temperature, diluted with EtOAc, and the organic phase was washed with saturated aqueous sodium chloride, dried over anhydrous sodium sulfate, and concentrated under reduced pressure then column chromatography to give the title compound (R) -4- (difluoromethyl) -3- (8-fluoro-9-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) thiazolidin-2-one (55 mg, 52%).
MS m/z(ESI):482.1[M+H]+.
Tenth step: preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
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(R) -4- (difluoromethyl) -3- (8-fluoro-9-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) thiazolidin-2-one (40 mg,0.083 mmol), L-alanine (15 mg,0.17 mmol), cuprous iodide (6.3 mg,0.033 mmol), potassium phosphate (53 mg,0.25 mmol), were mixed in dimethyl sulfoxide (3 mL), the reaction system was reacted with nitrogen gas three times with air replacement, at 125℃for 1.5 hours, cooled to room temperature, ammonium chloride (27 mg,0.5 mmol) was added, DMAP (161 mg,1.25 mmol) was stirred for 5 minutes, 2- (7-benzotriazol-oxide) -N, N, N ', N' -tetramethylurea hexafluorophosphate (284 mg,0.75 mmol) was added, stirred at room temperature for 2 hours, filtered, and ethyl acetate was added three times to extract. The organic phases were combined, dried over anhydrous sodium sulfate and concentrated under reduced pressure before column chromatography to give the title compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl-thiazolidine-3-yl) -8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide (7.9 mg, 22%).
1 H NMR(400MHz，CD 3 OD)δ1.49(d,J＝7.0Hz,3H),3.54-3.60(m,1H)，3.76-3.93(m,1H),3.95-4.00(m,1H),4.36-4.40(m,2H),4.47-4.52(m,2H),5.10-5.20(m,1H),6.32-6.62(m,2H),7.32(s,1H),7.85-7.91(m,1H)；
MS m/z(ESI):442.1[M+H] + .
Example 51
Preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methoxypropionamide
Preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -8-fluoro-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methoxypropionamide reference example 50.
1 H NMR(400MHz，CD 3 OD)δ3.40(s,3H),3.53-3.60(m,1H),3.69-3.83(m,3H),4.06-4.13(m,1H),4.35-4.41(m，2H),4.47-4.52(m,2H),5.10-5.21(m,1H),6.30-6.60(m,2H),7.32(s,1H),7.89(d,J＝8.5Hz,1H)；
MS m/z(ESI):472.1[M+H] + .
Example 52
Preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methoxypropionamide
The first step: preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methoxypropionamide
(R) -3- (9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-2-yl) -4- (difluoromethyl) thiazolidin-2-one (26 mg,0.062 mmol), O-methyl-L-serine (22 mg,0.18 mmol), cuprous iodide (6.0 mg,0.03 mmol), potassium phosphate (40 mg,0.19 mmol), and dimethyl sulfoxide (3 mL) were mixed, the reaction system was reacted with nitrogen for 12 hours at 100℃with air three times, cooled to room temperature, ammonium chloride (20 mg,0.37 mmol), triethylamine (95 mg,0.94 mmol) was added, stirring was performed for 5 minutes, 2- (7-benzotriazol oxide) -N, N, N ', N' -tetramethylurea hexafluorophosphate (212 mg,0.56 mmol) was added, stirring was performed at room temperature for 2 hours, the reaction solution was filtered and saturated aqueous solution was added thereto, and ethyl acetate was extracted three times. The organic phases were combined, dried over anhydrous sodium sulfate, and the organic solvent was concentrated under reduced pressure and separated by column chromatography to give the title compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) -3-methoxypropionamide (13 mg, 46%).
1 H NMR(400MHz,CD 3 OD)δ3.39(s,3H),3.53-3.57(m,1H),3.62-3.76(m,3H),3.93-3.98(m,1H),4.16-4.30(m,4H),5.06-5.16(m,1H),6.21-6.23(m,1H),6.28-6.52(m,2H),7.23(s,1H),8.02(d,J＝8.8Hz,1H)；
MS m/z(ESI):454.1[M+H] + .
Example 53
Preparation of (S) -1- (2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) pyrrolidine-2-carboxamide
The preparation of (S) -1- (2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) pyrrolidine-2-carboxamide was described in example 22.
1 H NMR(400MHz,DMSO-d 6 )δ1.83-1.92(m,2H),2.09-2.15(m,1H),3.72-3.81(m,4H),4.25-4.32(m,4H),5.07-5.15(m,1H),5.93-5.97(m,1H),6.22-6.28(m,1H),6.35-6.65(s,1H),7.00(s,1H),7.26(s,1H),7.35(s,1H),7.99(d,J＝8.6Hz,1H)；
MS m/z(ESI):450.1[M+H] + .
Example 54
Preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) (methyl) amino) propanamide
The preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) (methyl) amino) propanamide is described in example 22.
1 H NMR(400MHz,CD 3 OD)δ1.40(d,J＝7.0Hz,3H),2.90(s,3H),3.53-3.58(m,1H),3.75-3.80(m,1H),4.30-4.44(m,4H),4.46-4.51(m,1H),5.08-5.18(m,1H),6.22-6.41(m,2H),6.51-6.73(m,1H),7.28(s,1H),8.11(d,J＝9.0Hz,1H)；
MS m/z(ESI):438.1[M+H] + .
Example 55
Preparation of (2S, 3 r) -1- (2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) -3-methylpyrrolidine-2-carboxamide
Preparation of (2S, 3 r) -1- (2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) -3-methylpyrrolidine-2-carboxamide reference example 1.
MS m/z(ESI):448.1[M+H] + .
Example 56
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -7-methyl-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-9-yl) amino) propanamide
The first step: preparation of 5-bromo-2- (1H-imidazol-2-yl) aniline
To a methanol solution (50 mL) of 2-amino-4-bromobenzene (methylaldehyde) (4.9 g,24.6 mmol) was added an aqueous glyoxal solution (40 wt.%,18g,124 mmol), and then aqueous ammonia (28 wt.%,24g,172 mmol) was slowly added dropwise with stirring in a water bath for 30 minutes, with the reaction solution temperature controlled to be not more than 40 ℃. The mixture was then stirred overnight at 35℃and cooled, after which the title compound was obtained by column chromatography (3.5 g, yield: 60%) of 5-bromo-2- (1H-imidazol-2-yl) aniline.
MS m/z(ESI):238.0[M+H] + .
And a second step of: preparation of 10-bromo-5, 6,7, 8-tetrahydrobenzo [ c ] imidazo [1,2-a ] [1,5] diazepine Xin Yin
5-bromo-2- (1H-imidazol-2-yl) aniline (3.3 g,14 mmol), 1, 2-dibromoethane (1.38 mL,15.9 mmol), cesium carbonate (10.4 g,31.8 mmol) were mixed in N, N-dimethylformamide (50 mL), and stirred at room temperature for 1.5 hours. Water was added, stirred for 5 min, and extracted three times with EtOAc. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure, followed by column chromatography to give the title compound 10-bromo-5, 6,7, 8-tetrahydrobenzo [ c ] imidazo [1,2-a ] [1,5] diazocine (1.55 g, yield: 40%).
MS m/z(ESI):278.0[M+H] + .
And a third step of: preparation of 9-bromo-2, 3-diiodo-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepine
NIS (3.8 g,16.8 mmol) was added in portions to a solution of 10-bromo-5, 6,7, 8-tetrahydrobenzo [ c ] imidazo [1,2-a ] [1,5] diazocine (1.55 g,5.6 mmol) in DMF (30 mL) at room temperature, followed by stirring overnight at 60 ℃. Cooling, adding water, and separating out solid. After filtration, the solid was dissolved in ethyl acetate, washed with 1M aqueous NaOH and saturated brine in this order, dried over anhydrous sodium sulfate, and concentrated to give the title compound 9-bromo-2, 3-diiodo-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepine (2.6 g, yield: 90.2%).
MS m/z(ESI):515.8[M+H] + .
Fourth step: preparation of 9-bromo-2-iodo-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepine
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EtMgBr (1.0M in THF, 10mL,10 mmol) was slowly added dropwise to a solution of 9-bromo-2, 3-diiodo-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepine (2.52 g,4.9 mmol) in THF (20 mL) at-20deg.C. After the completion of the dropwise addition, the mixture was stirred at-15℃for 3 hours. Slowly heating to room temperature, dropwise adding saturated ammonium chloride aqueous solution, stirring for 15 min, and extracting with ethyl acetate for 3 times. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated and then column-chromatographed to give the title compound 9-bromo-2-iodo-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepine (1.52 g, yield: 80%).
MS m/z(ESI):389.9[M+H] + .
Fifth step: preparation of (S) -3- (9-bromo-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one
9-bromo-2-iodo-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepine (178 mg,0.46 mmol), (S) -4- (difluoromethyl) oxazolidin-2-one (63 mg,0.46 mmol), (1R, 2R) -N1, N2-dimethylcyclohexane-1, 2-diamine (28.4 mg,0.2 mmol), cuprous iodide (19.0 mg,0.1 mmol) and potassium carbonate (138 mg,1.0 mmol) were mixed in 1, 4-dioxane (4 mL), heated to 100deg.C for 5 hours, cooled to room temperature, 14% aqueous ammonia was added, stirred for 5 minutes, and extracted three times with EtOAc. The organic phases were combined, washed with saturated aqueous sodium chloride, dried over anhydrous sodium sulfate, concentrated under reduced pressure and then column chromatographed to give the title compound (S) -3- (9-bromo-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one (111 mg, yield: 60%).
MS m/z(ESI):399.1[M+H] + 。
Sixth step: preparation of (S) -3- (9-bromo-7-methyl-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one
(S) -3- (9-bromo-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one (111 mg,0.28 mmol) was dissolved in methanol (5 mL), catalytic amounts of acetic acid and aqueous formaldehyde (37% in water, 50mg,0.62 mmol) were added, stirred at room temperature for 30 min, sodium cyanoborohydride (39 mg,0.62 mmol) was added, the reaction was stopped at room temperature for 3 hours, and the reaction was quenched with saturated aqueous ammonium chloride solution and extracted three times with EtOAc. The organic phases were combined, washed with saturated aqueous sodium chloride, dried over anhydrous sodium sulfate, concentrated under reduced pressure and then column chromatographed to give the title compound (S) -3- (9-bromo-7-methyl-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one (81 mg, yield: 70%).
MS m/z(ESI):413.1[M+H] + .
Seventh step: preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -7-methyl-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-9-yl) amino) propanamide
(S) -3- (9-bromo-7-methyl-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one (49.4 mg,0.12 mmol), L-alanine (21.4 mg,0.24 mmol), cuprous iodide (4.6 mg,0.024 mmol) and potassium phosphate (51.5 mg,0.24 mmol) were mixed in dimethyl sulfoxide (2 mL), reacted at 100℃for 5 hours, cooled to room temperature, ammonium chloride (39 mg,0.72 mmol), triethylamine (184 mg,1.8 mmol) was added, stirring was performed for 5 minutes, 2- (7-benzotriazol-oxide) -N, N, N ', N' -tetramethylurea hexafluorophosphate (418 mg,1.1 mmol) was added, stirring was performed at room temperature for 2 hours, filtration, saturated aqueous sodium bicarbonate was added, and ethyl acetate was extracted three times. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure, followed by column chromatography to give the title compound (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -7-methyl-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-9-yl) amino) propanamide (20 mg, yield: 40%).
1 H NMR(400MHz,CD 3 OD)δ1.47(d,J＝7.0Hz,3H),2.95(s,3H),3.43-3.50(m,2H),3.86(q,J＝7.0Hz,1H),4.15(t,J＝5.2Hz,2H),4.54-4.67(m,2H),4.90-4.95(m,1H),6.18(d,J＝2.2Hz,1H),6.27(dd,J＝8.7,2.2Hz,1H),6.35-6.68(m,1H),7.16(s,1H),7.84(d,J＝8.7Hz,1H)；
MS m/z(ESI):421.1[M+H] + .
Example 57
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-9-yl) amino) propanamide reference is made to example 56.
1 H NMR(400MHz,CD 3 OD)δ1.45(d,J＝7.0Hz,3H),3.42-3.49(m,2H),3.78(q,J＝7.0Hz,1H),4.12-4.18(m,2H),4.54-4.67(m,2H),4.90-4.96(m,1H),5.86(d,J＝2.3Hz,1H),6.17(dd,J＝8.8,2.3Hz,1H),6.32-6.62(m,1H),7.05(s,1H),7.91(d,J＝8.8Hz,1H)；
MS m/z(ESI):407.1[M+H] + .
Example 58
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -7-ethyl-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazoheptin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -7-ethyl-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-9-yl) amino) propanamide the procedure of example 56 is described.
MS m/z(ESI):435.1[M+H] + .
Example 59
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -7-isopropyl-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazoheptin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -7-isopropyl-6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-9-yl) amino) propanamide the procedure of example 56 is described.
MS m/z(ESI):449.1[M+H] + .
Example 60
Preparation of (S) -2- ((7-cyclopropyl-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-9-yl) amino) propanamide
Preparation of (S) -2- ((7-cyclopropyl-2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-9-yl) amino) propanamide the procedure of example 56 is described.
MS m/z(ESI):447.1[M+H] + .
Example 61
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -7- (oxetan-3-yl) -6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazo heptin-9-yl) amino) propanamide
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -7- (oxetan-3-yl) -6, 7-dihydro-5H-benzo [ f ] imidazo [1,2-d ] [1,4] diazepin-9-yl) amino) propanamide reference is made to example 56.
MS m/z(ESI):463.1[M+H] + .
Example 62
Preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepin-9-yl) amino) propanamide
The first step: preparation of 2- (5-bromo-2-fluorophenyl) -1H-imidazole
5-bromo-2-fluorobenzaldehyde (5.0 g,24.6 mmol) was dissolved in isopropanol/water (25 mL/25 mL) at room temperature, ammonium acetate (17.6 g,221.7 mmol) was added, glyoxal (4.5 mL,221.7 mmol) was added dropwise, and stirring was continued overnight. Isopropanol was added for dilution, the reaction solution was filtered, then concentrated under reduced pressure, the concentrated solution was concentrated with methylene chloride and water, the organic phases were combined, then dried over anhydrous sodium sulfate, and after concentration under reduced pressure, column chromatography gave the title compound 2- (5-bromo-2-fluorophenyl) -1H-imidazole (3.3 g, yield: 56%).
1 H NMR(400MHz，DMSO-d6)δ8.16-8.10(m,1H),7.60-7.56(m,1H),7.38-7.33(m,1H),7.27-7.18(m,2H).
MS m/z(ESI):241.0[M+H] + .
And a second step of: preparation of 9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepine
2- (5-bromo-2-fluorophenyl) -1H-imidazole (2.0 g,8.4 mmol) was dissolved in N, N-dimethylformamide (10 mL), sodium hydride (442 mg,9.2 mmol) was added to the reaction under ice-water bath cooling, stirring was performed for 10 minutes, thiirane (612 mg,10.2 mmol) was added, and stirring was performed for 6 hours at 95 ℃. Cooled to room temperature, saturated aqueous ammonium chloride solution was added to the reaction flask, and the mixture was extracted three times with methylene chloride. The organic phases were combined, then the organic phase was dried over anhydrous sodium sulfate, concentrated under reduced pressure and then column chromatographed to give the title compound 9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepine (1.0 g, yield: 43%).
MS m/z(ESI):281.0[M+H] + .
Third step, preparation of 9-bromo-2, 3-diiodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepine
NIS (2.4 g,10.5 mmol) was added in portions to a DMF solution (20 mL) of 9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepine (480 mg,3.5 mmol) at room temperature, followed by stirring overnight at 60 ℃. Cooling, adding water, and separating out solid. After filtration, the solid was dissolved in ethyl acetate, washed with 1M aqueous NaOH and saturated brine in this order, dried over anhydrous sodium sulfate, and concentrated to give the title compound 9-bromo-2, 3-diiodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepine (1.6 g, yield: 86%).
MS m/z(ESI):532.8[M+H] + .
Fourth step, preparation of 9-bromo-2-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepine
EtMgBr (1.0M in THF, 3.3mL,3.3 mmol) was slowly added dropwise to a solution of 9-bromo-2, 3-diiodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepine (1.6 g,3.0 mmol) in THF (10 mL) at-20deg.C. After the completion of the dropwise addition, the mixture was stirred at-15℃for 3 hours. Slowly heating to room temperature, dropwise adding saturated ammonium chloride aqueous solution, stirring for 15 min, and extracting with ethyl acetate for 3 times. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated and then column-chromatographed to give the title compound 9-bromo-2-iodo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepine (1.03 g, yield: 85%).
MS m/z(ESI):406.9[M+H] + .
Fifth step (S) -preparation of 3- (9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one
9-bromo-2-iodo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d][1,4]Thiazepine (186.7 mg,0.46 mmol), (S) -4- (difluoromethyl) oxazolidin-2-one (63 mg,0.46 mmol), (1R, 2R) -N 1 ,N 2 Dimethylcyclohexane-1, 2-diamine (28.4 mg,0.2 mmol), cuprous iodide (19.0 mg,0.1 mmol), potassium carbonate (138 mg,1.0 mmol), and mixed in 1, 4-dioxane (4 mL), reacted at 100℃for 5 hours, cooled to room temperature, added with 14% ammonia water, stirred for 5 minutes, and extracted three times with EtOAc. The organic phases were combined, washed with saturated aqueous sodium chloride, dried over anhydrous sodium sulfate, concentrated under reduced pressure and then column chromatographed to give the title compound (S) -3- (9-bromo-5, 6-dihydrobenzo [ f) ]Imidazo [1,2-d][1,4]Thiazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one (124 mg, yield: 65%).
MS m/z(ESI):416.0[M+H] + .
Sixth step preparation of (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyl oxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepin-9-yl) amino) propanamide
(S) -3- (9-bromo-5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepin-2-yl) -4- (difluoromethyl) oxazolidin-2-one (49.8 mg,0.12 mmol), L-alanine (21.4 mg,0.24 mmol), cuprous iodide (4.6 mg,0.024 mmol), potassium phosphate (51.5 mg,0.24 mmol), mixed in dimethyl sulfoxide (2 mL), reacted for 5 hours at 100 ℃, cooled to room temperature, ammonium chloride (39 mg,0.72 mmol), triethylamine (184 mg,1.8 mmol) were added, stirred for 5 minutes, 2- (7-benzotriazol-oxide) -N, N, N ', N' -tetramethylurea hexafluorophosphate (418 mg,1.1 mmol) was added, stirred for 2 hours at room temperature, filtered, saturated aqueous sodium bicarbonate was added, and ethyl acetate was extracted three times. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure, followed by column chromatography to give the title compound (S) -2- ((2- ((S) -4- (difluoromethyl) -2-carbonyloxazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] thiazepin-9-yl) amino) propanamide (18 mg, yield: 35%).
1 H NMR(400MHz,CDCl 3 )δ1.56(d,J＝7.0Hz,3H),3.44-3.52(m,2H),3.84-3.92(m,1H),4.12-4.21(m,2H),4.48-4.56(m,1H),4.68-4.74(m,1H),4.88-5.02(m,1H),5.36(s,1H),6.40(s,1H),6.45-6.77(m,2H),6.83-6.88(m,1H),7.33(s,1H),7.61(d,J＝8.4Hz,1H)；
MS m/z(ESI):424.1[M+H] + .
2. Compound biological assay evaluation
The invention is further illustrated below in conjunction with test examples, which do not limit the scope of the invention.
1. Assay of inhibition of PI3K alpha/beta/gamma/delta kinase Activity by Compounds of the examples of this invention
1.1 Experimental purposes:
the purpose of this test example was to test the activity of the compounds of the example against inhibition of pi3kα/β/γ/δ kinase activity.
1.2 laboratory apparatus:
centrifuge (5810R) was purchased from Eppendorf corporation
Pipettes were purchased from Eppendorf or Rainin Inc
The enzyme-labeled instrument was purchased from BioTek company of America
Full-functional enzyme-labeled instrument with model number of SynergyH1
1.3 experimental method:
the experiment uses ADP-Glo lipid kinase assay method (Promega#V9102) from Promega company, lipid kinase PI3K alpha/beta/gamma/delta on substrate PIP2: catalytic reaction of 3PS with ATP to generate ADP, characterization of lipid kinase activity by measuring ADP content in the reaction, and obtaining half inhibition concentration IC of compound to inhibition of PI3K alpha/beta/gamma/delta kinase activity 50 。
The specific experimental operation is as follows:
the kinase reaction was performed in a white 384 well plate (Perkin Elmer # 6007299) with 2. Mu.L of ddH in 1% DMSO per well 2 O diluted compounds at different concentrations, positive control wells were added with 2. Mu.L ddH containing 1% DMSO 2 O, then 2. Mu.L of 5 Xkinase buffer (HEPES 250mM, mgCl) was added to each well 2 15mM,NaCl 250mM,BSA0.05%) diluted 0.1-2 nM PI3K kinase solution, 2. Mu.L of 5 Xkinase buffer was added to the negative control wells and 4. Mu.L of 10 Xdilution buffer and ddH was added to all wells 2 O formulated 50 μm substrate PIP2:3PS (Promega#V1701), and finally adding 2. Mu.L of 50-100. Mu.M ATP solution diluted with water to start the reaction, and after 90-120 minutes at room temperature, adding 10. Mu.L of ADP-Glo Reagent (containing 10mM MgCl) per well 2 ) After 60 minutes at room temperature to remove the excess adenosine triphosphate ATP, 20. Mu. L Kinase Detection Reagent was added to each well, and the reaction was carried out at room temperature for 20 minutes in the dark, and the chemiluminescent value was measured by a BioTek reaction H1 microplate reader.
Enzyme name Goods number Concentration of enzyme reaction Enzyme reaction time ATP concentration
PI3Kα Promega#V1721 0.1nM 120min 50μM
PI3Kβ Carna#11-102 0.4nM 90min 100μM
PI3Kγ Thermofisher#PV4786 0.4nM 120min 50μM
PI3Kδ Carna#11-103 0.1nM 90min 100μM
The experimental data processing method comprises the following steps:
percent inhibition data {% inhibition = 100- [ (test compound value-negative control value) for wells treated with compound were calculated by positive control wells (DMSO control wells) and negative control wells (no kinase added) on plates]/(positive control value-negative control value) ×100}. IC is calculated by using GraphPad prism to fit different concentration and corresponding percent inhibition rate data to a four-parameter nonlinear logic formula 50 Values.
1.4 experimental conclusion:
from the above schemes, it was concluded that the example compounds according to the invention show the biological activity in the PI3K alpha/beta/gamma/delta kinase activity assay as shown in table 7 below.
TABLE 7
The above data show that the compounds of the examples of the present invention have good activity and selectivity in terms of PI3K alpha/beta/gamma/delta kinase activity.
2. Determination of proliferation inhibition of PI3K alpha mutant cancer cells by Compounds of the examples of this invention
2.1 Experimental purposes:
the purpose of this test example was to test the proliferation inhibitory activity of the example compounds against PI3kα mutant cancer cells HCC1954 (H1047R), HGC-27 (E542K) and MKN1 (E545K).
2.2 laboratory apparatus:
centrifuge (5702R) was purchased from Eppendorf Corp
Carbon dioxide incubator is available from Thermo company
Biological safety cabinets are purchased from Shanghai Bo Highway Co
Pipettes were purchased from Eppendorf or Rainin Inc
The enzyme-labeled instrument was purchased from BioTek company of America
Full-functional enzyme-labeled instrument with model number of SynergyH1
2.3 experimental method:
proliferation inhibition of PI3K alpha mutant cancer Cell lines (HCC 1954, HGC-27 and MKN 1) by the example compounds was examined by the method of Cell Titer-Glo Cell lines were cultured in RPMI 1640 medium (Gibco # 22400089) containing 10% FBS (Gibco # 10091148) and 1% P/S (Hyclone # SV 30010) at 37℃with 5% CO 2 Is cultured under the condition of (2). Collecting cells before experiment, regulating cell density after cell count, seeding cells at 1000-10000 cells/well in white 96-well plate (Corning # 3610), placing into 37deg.C, 5% CO 2 After the culture in the incubator overnight, compound solutions with different concentrations are prepared, corresponding solvent controls are arranged, and the incubator is continuously put into 37 ℃ and 5% CO 2 After culturing in an incubator for 48-96 hours, the Cell plate and its contents are equilibrated to room temperature, 20-100 μl of Cell Titer-Glo solution (Promega#G7573) is added to each well, and after shaking and mixing, incubated at room temperature for 5-30 minutes in the absence of light, the cells are incubated withThe chemiluminescent values were detected by a BioTek's synergy H1 microplate reader.
2.4 experimental data processing method:
percent inhibition data {% inhibition = 100- (test compound value/vehicle control value) ×100} for wells treated with the compound of the example were calculated by vehicle control wells on plates, IC was calculated using GraphPad prism to fit different concentrations and corresponding percent inhibition data to a four parameter nonlinear logic formula 50 Values.
2.5 experimental conclusion:
the activity test of the compounds of the examples shown in the present invention against proliferation inhibition of PI3kα mutant cancer cells HCC1954 (H1047R), HGC-27 (E542K) and MKN1 (E545K) was derived from the above protocol, and showed the biological activity as shown in table 8 below.
TABLE 8
The above data shows that the compounds of the examples of the present invention have good activity in inhibiting proliferation of pi3kα mutant cancer cells HCC1954 (H1047R), HGC-27 (E542K) and MKN1 (E545K).
3. Toxicity test of SD rat by repeated gastric lavage for 7 days
3.1 purpose of experiment
The purpose of this study was to examine the toxic response that may occur 7 days after repeated gavage administration of GDC-0077, example 22 and example 62 to SD rats and to compare the differences in toxicity of GDC-0077, example 22 and example 62.
3.2 Experimental materials and instruments
3.2.1 test pieces
Test article 1: GDC-0077
Test article 2: example 22 and example 62
3.2.2 solvent
Name: 20% aqueous SBE-beta-CD (Captisol)
3.2.3 animal information
Species & strain: sprague-Dawley (SD) rats
Animal grade: SPF stage
Animal number and sex: 160 rats, male and female halves.
3.2.4 instruments
2120 series of fully automatic hematology analyzers for blood cell count;
the SYSMEX CA-500 coagulometer is used for detecting coagulation function indexes;
the TBA-120FR full-automatic biochemical analyzer is used for detecting biochemical indexes of blood;
the easy electrolyte analyzer is used for electrolyte detection;
liquid phase mass spectrum detector model API4000;
Electrospray Source (ESI) positive ion mode and chromatographic column model Agilent ZORBAX XDB-C18 (3.5 μm, 2.1X150 mm) for bioanalytical detection of plasma samples;
3.3 Experimental methods
1) In the test, 160 rats (80/sex) are divided into 20 groups according to weight, 100 rats are used as toxicity study (1-10 groups, 5/sex/group) and 60 rats are used as toxicity study (11-20 groups, 3/sex/group);
2) Animals of groups 1 and 11 were gavaged with 20% aqueous SBE-beta-CD (Captisol) as vehicle control;
3) Groups 2 and 12, 3 and 13, 4 and 14 animals were given 10, 30, 60mg/kg GDC-0077 by gavage, respectively;
4) Animals of groups 5 and 15, 6 and 16, 7 and 17 were given 10, 30, 60mg/kg of example 22 by gavage, respectively.
5) Animals of groups 8 and 18, 9 and 19, 10 and 20 were given 10, 30, 60mg/kg of example 62 by gavage, respectively.
5) Animals were dosed 1 time a day for 7 consecutive days (animals of groups 7, 17, 10, 20 were dosed for 6 consecutive days).
6) The administration volume was 10mL/kg.
7) During the test, the items of clinical observation, weight, food intake, clinical pathological indexes (blood cell count, blood coagulation function, blood biochemistry), toxicology and the like are examined.
8) All animals were euthanized at d8 (animals of groups 7, 10, 17, 20 were euthanized after d 6).
9) During the test period, gross anatomical observations were made on animals from groups 1 to 10, animals from groups 17 and 20, and dead animals (including toxin study animals), and histopathological examination was performed on abnormal tissues, gastrointestinal tissues (e.g., colon, cecum), and immune tissues (e.g., thymus).
3.4 conclusion of experiments
At a dose of 30mg, the average systemic exposure AUC (male: 11400h ng/mL, female: 159700 h ng/mL) after the last administration of example 22 was about 2.4-3.8 times that of GDC-0077 (male: 3000h ng/mL, female: 6510h ng/mL) at the same dose, similar to that of GDC-0077 (male: 15400h ng/mL, female: 22800h ng/mL) after the first administration at a dose of 60 mg/kg.
At a dose of 10mg, the average systemic exposure AUC (male: 2110h ng/mL, female: 3170h ng/mL) after the last administration of example 22 was about 1.4-2.5 times that of GDC-0077 (male: 845h ng/mL, female: 2250h ng/mL).
Thus, at the same dose, the system exposure of example 22 was significantly higher than GDC-0077.
Under the test conditions, the GDC-0077 and the test sample of example 22 were administered to SD rats repeatedly at 10, 30, 60mg/kg doses for 7 days (1 day), the lethal dose of GDC-0077 and example 22 was 60mg/kg, the Maximum Tolerated Dose (MTD) was 30mg/kg, and the C of example 22 was administered at a dose of 30mg/kg max And AUC (0-24h) Significantly higher than GDC-0077, example 22 has better tolerability than GDC-0077.
Under the test conditions, the GDC-0077, example 22 and example 62 samples were repeatedly given to SD rats at 10, 30, 60mg/kg doses for 7 days (1 time/day), and the C of example 22 and example 62 were repeatedly given by gastric lavage max And AUC (0-24h) Is significantly higher than GDC-0077 and has better tolerance than GDC-0077.
4. In vivo efficacy test of Compounds of the examples of the invention
4.1 purpose of experiment
The compounds with obvious efficacy and small toxic and side effects are screened out through in vivo efficacy experiments.
4.2 Experimental major instruments and materials
4.2.1 instruments:
1. biological safety cabinet (BSC-1300 II A2, shanghai Bo Xie medical equipment factory)
2. Ultra clean bench (CJ-2F, von Willebrand laboratory animal Co., ltd.)
3、CO 2 Incubator (Thermo-311)
4. Centrifuge (Centrifuge 5702R, eppendorf)
5. Full-automatic cell counter (Countess II, life)
6. Pipettor (10-20. Mu.L, eppendorf)
7. Microscope (TS 2, nikang)
8. Vernier caliper (CD-6' AX, sanfeng Japan)
9. Cell culture bottle (T75/T225, corning)
10. Electronic balance (CPA 2202S, sidoris)
4.2.2 reagents:
1. RPMI-1640 medium (22400-089, gibco)
2. Fetal Bovine Serum (FBS) (10091-148, gibco)
3. 0.25% trypsin (25200-056, gibco)
4. Green streptomycin double antibody (15140-122, gibco)
5. Phosphate Buffered Saline (PBS) (10010-023, gibco)
6. Matrigel Matrix (356234, corning)
4.2.3 animals:
BALB/c nude mice, 6-8 weeks, male, purchased from Shanghai Sipule-BiKai laboratory animal Co.
4.3 Experimental procedures
4.3.1 cell culture and preparation of cell suspension
a, removing a strain of HCC1954 cells from the cell bank, resuscitating the fine cells with RPMI-1640 medium (RPMI-1640+10% FBS+1% SP)Placing cells after recovery in a cell culture flask (cell type, date, cultured person name, etc. are marked on the flask wall), placing in CO 2 Culturing in incubator (temperature of incubator is 37deg.C, CO) 2 Concentration 5%).
b, after the cells are spread at 80-90% of the bottom of the culture flask, passaging, and placing the cells in CO continuously after passaging 2 Culturing in an incubator. This process is repeated until the number of cells meets the in vivo pharmacodynamic requirements.
c, collecting cultured cells, counting with a fully automatic cell counter, and re-suspending cells with PBS and matrigel according to the counting result to obtain cell suspension (density 5×10) 7 /mL), placed in an ice bin for use.
4.3.2 cell seeding
a, marking nude mice with disposable universal ear tags for large and small mice before inoculation
b, mixing the cell suspension during inoculation, extracting 0.1-1 mL of the cell suspension by using a 1mL syringe, removing bubbles, and then placing the syringe on an ice bag for standby.
c, the left hand is kept to be a nude mouse, the right shoulder position (inoculation position) of the right back of the nude mouse is sterilized by 75% alcohol, and the inoculation is started after 30 seconds.
d, the test nude mice were inoculated sequentially (0.1 mL cell suspension was inoculated per mouse).
4.3.3 tumor-bearing mice were metered, grouped and dosed
a, tumor is measured on the 14 th to 18 th days after inoculation according to the growth condition of the tumor, and the tumor size is calculated.
Tumor volume calculation: tumor volume (mm) 3 ) =length (mm) ×width (mm)/2
And b, grouping according to the weight of the tumor-bearing mice and the size of the tumors by adopting a random grouping method.
c, according to the grouping result, starting to administer the test medicine (administration mode: oral administration; administration dosage: 10mg/kg; administration volume: 10mL/kg; administration frequency: 1 time/day; administration period: 21 days; vehicle: 0.5% CMC/1% Tween 80).
d, tumor twice weekly after starting to administer test drug, and weighing.
e, euthanized animals after the end of the experiment.
f, processing the data by Excel and other software. Calculation of compound tumor inhibition rate TGI (%): when there was no tumor regression, TGI (%) = [ (1- (mean tumor volume at the end of the treatment group-mean tumor volume at the beginning of the treatment group))/(mean tumor volume at the end of the treatment with the solvent control group-mean tumor volume at the beginning of the treatment with the solvent control group) ]%100%. When there was regression of the tumor, TGI (%) = [1- (mean tumor volume at the end of dosing of a treatment group-mean tumor volume at the beginning of dosing of the treatment group)/mean tumor volume at the beginning of dosing of the treatment group ] ×100%.
4.4 test data are shown in Table 9 below:
TABLE 9
Grouping Number of animals (Only) Days of administration (Tian) Tumor inhibition rate
Blank control 5 21 -
Example 22 5 19 132％
Example 25 5 21 120％
Example 50 5 21 96％
Example 52 5 21 98％
Example 56 5 21 122％
Example 62 5 21 147％
4.5 experimental results
From the above results, it can be seen that the above compounds of the present patent have a better tumor inhibition rate.
5. Pharmacokinetic PK experiments of Compounds of the examples of the invention on mice
The pharmacokinetic experiments in mice of the compounds of the preferred embodiments of the present invention were performed using Balb/c male mice (Shanghai Jieshijie laboratory animals Co., ltd.).
5.1 modes of administration: single gastric lavage administration.
5.2 dosing dose: 5 mg/10 ml/kg (body weight).
5.3 formulation recipe: 0.5% CMC-Na, dissolved ultrasonically, formulated as a clear solution or homogeneous suspension.
5.4 sampling points: 0.5, 1, 2, 4, 6, 8 and 24 hours after administration.
5.5 sample treatment:
1) The orbit is sampled by 0.1mL and placed in K 2 In EDTA test tube, the plasma is separated by centrifugation at 1000-3000 Xg for 5-20 min and stored at-80 deg.C.
2) The plasma sample 40uL was precipitated by adding 160uL acetonitrile, and after mixing, it was centrifuged at 500 to 2000 Xg for 5 to 20 minutes.
3) 100uL of the treated supernatant was subjected to LC/MS/MS analysis for the concentration of the test example.
5.6LC-MS/MS analysis:
liquid phase conditions: shimadzu LC-20AD pump
Mass spectrometry conditions: AB Sciex API 4000 mass spectrometer
Chromatographic column: phenomenex Gemiu 5um C 18 50×4.6mm
Mobile phase: solution A is 0.1% formic acid water solution, solution B is acetonitrile
Flow rate: 0.8mL/min
Elution time: gradient elution for 0-4 min
5.7 pharmacokinetics:
the main parameters were calculated with WinNonlin 6.1 and the results of the mouse drug substitution experiments are shown in table 10 below:
table 10
The results of the mouse-over experiments in the table show that the compounds of the examples shown in the invention show good metabolic properties, plasma exposure AUC and maximum blood concentration C max All perform well.
3. Crystal form study of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide
1.1 laboratory apparatus
1.1.1 some parameters of the physicochemical detection Instrument
1.2 Instrument and conditions for liquid phase analysis
1.2.1 instruments and apparatus
Instrument name Model number
Analytical balance Sartorius BSA224S-CW
Water purifier Milli-Q Plus，Millipore
High performance liquid chromatograph Agilent1260
Pump with a pump body Agilent G1311B
Sample injector G1329B
Column temperature box G1316A
Detector for detecting a target object G1315D
1.2.2 chromatographic conditions
Chromatographic column: ZORBAX (SB-C8, 3.5 μm, 4.6X75 mm)
Flow rate: 1mL/min
Column temperature: 40 DEG C
Detection wavelength: 220/328nm
Sample injection volume: 5.0 mu L
Run time: 12min
A diluent: ACN-Water (v/v, 1:1)
Mobile phase: a: water (0.05% trifluoroacetic acid); b: acetonitrile (0.05% trifluoroacetic acid)
1. Preparation of different crystalline forms of the compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide of formula (I)
1.1 Preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form A
The first step: preparation of (2- ((R) -4- (difluoromethyl) -2-oxothiazolidin-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazol-9-yl) -l-alanine amine
25.5g of (R) -3- (9-bromo-5, 6-dihydrobenzo [ f) are weighed out]Imidazo [1,2-d][1,4]Oxazepin-2-yl) -4- (difluoromethyl) thiazolidin-2-one in a reaction kettle, 350mL DMSO, 16.8. 16.8g L-alanine, 39.5g potassium phosphate were added to the reaction kettle. After bubbling with nitrogen for 40min, 1.32g was addedCu 2 O is added into the reaction kettle. Bubbling with nitrogen was continued for 15min, vacuum was pulled for 15min, and nitrogen was used to replace 3 times. The temperature of the reaction kettle is raised to 95-100 ℃, and the reaction kettle is stirred for 16h with heat preservation. Then cooling to 10-20 ℃, detecting liquid phase, and adding 400mL of water and 300mLDCM after the reaction is finished. After stirring, the solution was separated and the aqueous phase was washed twice with 300mL portions of DCM. 600mL of THF was added and 200mL of 20% aqueous sodium bisulfate solution was added dropwise with a temperature control of less than 25 ℃. The solution was separated and the aqueous phase was extracted once with 600mL THF. The organic phases were combined and washed twice with 300mL portions of 15% aqueous sodium chloride. 12g of sulfur silica gel, N, are added 2 Bubbling for 40min and stirring for 4h. Anhydrous sodium sulfate was added to dry. Filtered, rinsed with 50ml anhydrous THF. The mixture was swirled under reduced pressure to about 400mL of organic solvent, heated to 50℃and 9mL of 7M NH was added dropwise 3 MeOH, adding a small amount of seed crystal, precipitating a white solid, and continuing to dropwise add 18mL 7M NH 3 MeOH, drop Bi Jiaoban for 0.5h, and gradually cooling the reaction solution to 10-15 ℃. Stirring for 12h, filtering, rinsing with anhydrous THF, and vacuum drying to obtain 17.0g of ammonium (2- ((R) -4- (difluoromethyl) -2-oxo-thiazolidine-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d][1,4]Oxazol-9-yl) -l-alanine, wherein (R) -3- (9-bromo-5, 6-dihydrobenzo [ f]Imidazo [1,2-d][1,4]Oxazepin-2-yl) -4- (difluoromethyl) thiazolidin-2-one was prepared as described in example 22.
And a second step of: preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form A
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17g of ammonium (2- ((R) -4- (difluoromethyl) -2-oxothiazolidin-3-yl) -5, 6-dihydrobenzo [ f) were weighed out]Imidazo [1,2-d][1,4]Oxazopan-9-yl) -l-alanine was placed in a reaction vessel, and 170mL of DMSO and 12.36g of NH were added to the reaction vessel 4 Cl, 5.76g HOSu and 39.8g DIPEA. The mixture was bubbled with nitrogen for 30min, and 20.7g of EDCl was added at 10 to 20 ℃. Then, the whole system is stirred at 15-25 ℃ for reaction for 24 hours, 300mL of THF is added, 150mL of 15% sodium chloride aqueous solution is added, and stirring is carried out for 10 minutes. The whole system is layered, and liquid and water are separated by a separating funnel The phase was extracted once with 200mL THF. The organic phases were combined, washed once with 150mL of 15% aqueous sodium chloride solution, twice with 125mL of 5% citric acid solution, twice with a mixture of 20% aqueous ammonia and 15% aqueous sodium chloride solution, 125mL each time. The organic phase is rotated to 200mL under reduced pressure, 100mL of ethanol is added, the mixture is rotated to 200mL of ethanol, 100mL of ethanol is added, a small amount of seed crystal is added, the mixture is rotated to 200mL of ethanol for 3 times, 100mL of ethanol is used for each time, the mixture is finally rotated to 190mL of ethanol, 25mL of water is added dropwise at 50 ℃, stirring is carried out for 0.5h, cooling is carried out slowly to 10-15 ℃ and stirring is carried out for 12h. Finally, filtering, and using ethanol for filter cakes: water = 7:1, the solid was placed in a vacuum dried at 40 ℃ to constant temperature to give 11.5g of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonylthiazolidin-3-yl) -5, 6-dihydrobenzo [ f) as a white solid]Imidazo [1,2-d][1,4]Oxazepin-9-yl) amino) propanamide form a. Upon detection analysis, it was (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d][1,4]Oxazepan-9-yl) amino) propanamide form a having an XRPD pattern as shown in figure 1, a TGA pattern as shown in figure 2, and a DSC pattern as shown in figure 3.
1.2 Preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form B
30mg of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form A is weighed into a 2mL glass bottle, 200 μl of acetone is added to form a suspension, after shaking for 10 days at room temperature, the supernatant is removed, and the remaining solid is dried to constant weight in a vacuum oven at 40 ℃ to obtain (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form B. A crystalline form B of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide, having an XRPD pattern as shown in figure 4, a DSC pattern as shown in figure 5, as determined by detection analysis.
1.3 Preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form C
30mg of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form A is weighed into a 2mL glass bottle, 200 μl of tetrahydrofuran is added to form a suspension, after shaking for 10 days at room temperature, the supernatant is removed, and the remaining solid is dried to constant weight in a vacuum oven at 40 ℃ to obtain (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form C. A crystalline form C of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide, having an XRPD pattern as shown in figure 6, a TGA pattern as shown in figure 7, and a DSC pattern as shown in figure 8, as determined by detection analysis.
1.4 Preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-D ] [1,4] oxazepin-9-yl) amino) propanamide form D
30mg of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-D ] [1,4] oxazepin-9-yl) amino) propanamide form A is weighed into a 2mL glass bottle, 200 μl of 88% acetone is added to form a suspension, after shaking for 10 days at room temperature, the supernatant is removed, and the remaining solid is dried to constant weight in a vacuum oven at 40 ℃ to obtain (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-D ] [1,4] oxazepin-9-yl) amino) propanamide form D. A crystalline form D of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-D ] [1,4] oxazepin-9-yl) amino) propanamide having an XRPD pattern as shown in figure 9, a TGA pattern as shown in figure 10 and a DSC pattern as shown in figure 11, as determined by detection analysis.
1.5 Preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form E
30mg of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form A is weighed into a 2mL glass bottle, 200 μl of 1,4 dioxane is added to form a suspension, shaking is carried out at room temperature for 10 days, rapid centrifugation is carried out, the supernatant is removed, and the remaining solid is placed in a vacuum drying oven at 40 ℃ to be dried to constant weight, so that (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form E is obtained. A crystalline form E of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide having an XRPD pattern as shown in figure 12, a TGA pattern as shown in figure 13 and a DSC pattern as shown in figure 14, as determined by assay.
1.6 Preparation of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ F ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form F
The ethanesulfonate salt of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonylthiazolidine-3-yl) -5, 6-dihydrobenzo [ F ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide was weighed into a 2mL glass bottle, 200 μl of water was added to form a suspension, after shaking for 10 days at room temperature, the supernatant was removed, and the remaining solid was dried to constant weight in a vacuum oven at 40 ℃ to give (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonylthiazolidine-3-yl) -5, 6-dihydrobenzo [ F ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form F. A crystalline form F of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ F ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide having an XRPD pattern as shown in figure 15, a TGA pattern as shown in figure 16 and a DSC pattern as shown in figure 17, as determined by assay.
2. Solid stability test
2.1 Experimental purposes:
and (2) examining the physicochemical stability of the (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide under the acceleration condition or the influence factor condition, and providing a basis for crystal form screening and compound storage.
2.2 experimental protocol:
weighing (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f)]Imidazo [1,2-d][1,4]Oxazepin-9-yl) amino) propanamide form A about 2mg, sealed in a 60 ℃ oven, and open at room temperature RH95% (saturated KNO) 3 Aqueous solution) and an illumination box (5000 lx.+ -. 500 lx), the content was measured by HPLC and external standard method for 5 days and 10 days, and the change of the related substances was calculated by chromatographic peak area normalization method.
2.3 experimental results:
the comparison of the liquid phase spectrograms shows that the free alkali crystal form A is added with 2 impurities more than 0 day under the illumination condition of 10, the increase of 0.362% and 0.108% is respectively carried out, and the increase of the impurities is less than 0.05% at 60 ℃ and at room temperature RH of 95% for 10 days and more than 0 day.
2.4 conclusion of experiments
The compound crystal form is unstable under the illumination condition, and light shielding is needed in the later storage process. But is stable under high temperature and high humidity conditions.
3 moisture permeability test
3.1 purpose of experiment
The hygroscopicity of the compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide crystal form A under different relative humidity conditions is examined, and a basis is provided for screening and storing the compound crystal form.
3.2 experimental protocol:
the crystalline form a of the compound (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide is placed in saturated water vapor of different relative humidity, the compound is brought into dynamic equilibrium with the water vapor, and the percentage of the hygroscopic weight gain of the compound after equilibrium is calculated.
3.3 experimental results:
(S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form a has a moisture absorption gain of about 0.188% at 80% RH, no hygroscopicity.
4. Solubility experiments in different Medium
4.1 purpose of experiment
The solubility of the (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide crystal form A in mediums with different pH values, water, artificial Simulated Gastric Fluid (SGF), fasted artificial simulated intestinal fluid (FaSSIF), non-fasted artificial simulated intestinal fluid (FeSSIF) and the like is compared, so that a basis is provided for the evaluation of the patentability.
4.2 experimental protocol:
about 2mg of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide form A is suspended in a different medium for 24 hours, and the thermodynamic solubility of the compound is determined by HPLC, external standard method at 37 ℃.
4.3 experimental results: as shown in Table 12
Table 12
Sample name Solubility of Crystal form A (mg/mL)
pH1 7.99
pH2 0.740
pH3 0.173
pH4 0.050
pH5 0.037
pH6 0.021
pH7 0.025
pH8 0.017
Fa 0.036
Fe 0.122
SGF 1.072
water 0.017
5. Polycrystalline screening and stable crystal form confirmation experiment
5.1 Experimental purposes:
through the polycrystalline screening, a relatively stable (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide crystal form was found.
5.2 experimental protocol:
six crystalline forms of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ F ] imidazo [1,2-D ] [1,4] oxazepin-9-yl) amino) propanamide, form a, form B, form C, form D, form E and form F, respectively, were obtained by beating, changing the crystallization solvent, crystallization mode, etc., and the DSC profile of the solid was determined, respectively.
5.3 experimental results:
by comparing the DSC spectra of (S) -2- ((2- ((R) -4- (difluoromethyl) -2-carbonyl thiazolidine-3-yl) -5, 6-dihydrobenzo [ f ] imidazo [1,2-d ] [1,4] oxazepin-9-yl) amino) propanamide of different crystal forms, the crystal form A can be judged to be the most stable crystal form of the six crystal forms in the thermodynamic state.

Claims (16)

1. A crystalline form of a compound having the structure:
the crystal form is selected from one of the following:
the crystal form A, the X-ray powder diffraction pattern has diffraction peaks at the positions of 5.7+/-0.2 degrees, 11.3+/-0.2 degrees, 19.5+/-0.2 degrees, 19.8+/-0.2 degrees, 20.4+/-0.2 degrees, 21.8+/-0.2 degrees and 24.3+/-0.2 degrees of 2 theta;
the X-ray powder diffraction pattern of the crystal form B has diffraction peaks at the positions of 7.6+/-0.2 degrees, 7.9+/-0.2 degrees, 8.3+/-0.2 degrees, 13.1+/-0.2 degrees, 14.2+/-0.2 degrees, 19.5+/-0.2 degrees and 23.6+/-0.2 degrees of 2 theta;
the X-ray powder diffraction pattern of the crystal form C has diffraction peaks at the positions of 7.7+/-0.2 degrees, 13.0+/-0.2 degrees, 15.3+/-0.2 degrees, 17.7+/-0.2 degrees, 19.4+/-0.2 degrees, 19.7+/-0.2 degrees and 22.3+/-0.2 degrees of 2 theta;
the diffraction pattern of the crystal form D, X-ray powder has diffraction peaks at 6.6+/-0.2 degrees, 10.1+/-0.2 degrees, 11.4+/-0.2 degrees, 13.4+/-0.2 degrees, 13.8+/-0.2 degrees, 18.6+/-0.2 degrees, 18.9+/-0.2 degrees and 19.5+/-0.2 degrees of 2 theta;
form E, X-ray powder diffraction pattern having diffraction peaks at 7.7±0.2°, 15.4±0.2°, 17.6±0.2°, 19.4±0.2°, 22.1±0.2°, 23.1±0.2°, 23.9±0.2° and 25.1±0.2° in 2θ;
form F, X-ray powder diffraction pattern having diffraction peaks at 6.5±0.2°, 10.1±0.2°, 13.7±0.2°, 17.3±0.2°, 19.1±0.2°, 24.2±0.2° and 26.2±0.2° in 2θ.
2. The crystalline form of claim 1, wherein,
the X-ray powder diffraction pattern of the crystal form A further has diffraction peaks at 16.9+/-0.2 DEG, 18.5+/-0.2 DEG, 27.4+/-0.2 DEG and 33.9+/-0.2 DEG of 2 theta;
the X-ray powder diffraction pattern of the crystal form B further has diffraction peaks at 14.7+/-0.2 DEG, 15.7+/-0.2 DEG, 21.8+/-0.2 DEG and 24.9+/-0.2 DEG of 2 theta;
the X-ray powder diffraction pattern of form C further has diffraction peaks at 20.4±0.2°, 23.2±0.2°, 24.1±0.2° and 25.3±0.2° in 2θ;
the X-ray powder diffraction pattern of form D further has diffraction peaks at 16.2±0.2°, 20.4±0.2°, 21.9±0.2°, 24.1±0.2° and 26.1±0.2° in 2θ;
the X-ray powder diffraction pattern of form E further has diffraction peaks at 2θ of 12.8±0.2°, 20.5±0.2° and 25.9±0.2°;
the X-ray powder diffraction pattern of form F further has diffraction peaks at 13.4±0.2°, 20.1±0.2°, 25.1±0.2° and 27.7±0.2° in 2θ.
3. The crystalline form of claim 2, wherein,
the X-ray powder diffraction pattern of form a further has diffraction peaks at 9.8±0.2°, 15.9±0.2°, 18.0±0.2° and 23.7±0.2° in 2θ.
4. The crystalline form of claim 1, wherein,
The X-ray powder diffraction pattern of form A is further substantially as shown in figure 1;
the X-ray powder diffraction pattern of form B is further substantially as shown in figure 4;
the X-ray powder diffraction pattern of form C is further substantially as shown in figure 6;
the X-ray powder diffraction pattern of form D is substantially as shown in FIG. 9;
the X-ray powder diffraction pattern of form E is further substantially as shown in figure 12;
the X-ray powder diffraction pattern of form F is further substantially as shown in figure 15.
5. The crystalline form of claim 1, wherein,
the TGA profile of form a is further substantially as shown in figure 2;
the TGA profile of form C is further substantially as shown in figure 7;
the TGA profile of form D is further substantially shown in figure 10;
the TGA profile of form E is further substantially shown in fig. 13;
the TGA profile of form F is further substantially as shown in figure 16.
6. The crystalline form of claim 1, wherein,
the DSC spectrum of the crystal form A is further basically shown in the figure 3;
the DSC spectrum of the crystal form B is further basically shown in figure 5;
the DSC spectrum of the crystal form C is further basically shown in figure 8;
the DSC spectrum of the crystal form D is further basically shown in figure 11;
The DSC spectrum of the crystal form E is basically further shown in figure 14;
the DSC spectrum of the crystal form F is basically further shown in figure 17.
7. A process for the preparation of crystalline form A, B, C, D or E of a compound according to any one of claims 1 to 6, comprising in particular the steps of:
1) Weighing a proper amount of free alkali or a crystal form thereof, and suspending with a poor solvent;
2) Shaking the suspension obtained above;
3) Centrifuging the suspension, removing supernatant, and drying the rest solid in a vacuum drying oven until the weight is constant to obtain a target product;
the poor solvent is selected from one or more of water, acetone, ethanol, tetrahydrofuran or 1, 4-dioxane.
8. The method of claim 7, wherein the step of determining the position of the probe is performed,
the poor solvent is selected from one or more of water, tetrahydrofuran, acetone or 1, 4-dioxane.
9. The method of claim 7, wherein the step of determining the position of the probe is performed,
the poor solvent is selected from one or more of water, tetrahydrofuran, 88% acetone or 1, 4-dioxane.
10. A process for preparing form F of a compound according to any one of claims 1 to 6, comprising in particular the steps of:
1) Weighing a proper amount of salt of free alkali, and suspending with water;
2) Shaking the suspension obtained above;
3) Centrifuging the suspension rapidly, removing supernatant, and vacuum drying the rest solid to constant weight to obtain target product;
the salt form is selected from ethane sulfonate of free alkali.
11. A pharmaceutical composition comprising a therapeutically effective amount of a crystalline form of a compound according to any one of claims 1-6, and one or more pharmaceutically acceptable excipients.
12. The pharmaceutical composition of claim 11, wherein the excipient is a diluent.
13. The use of a crystalline form of a compound according to any one of claims 1 to 6, and a pharmaceutical composition according to any one of claims 11 to 12, in the manufacture of a PI3K inhibitor medicament.
14. The use of a crystalline form of a compound according to any one of claims 1 to 6, and a pharmaceutical composition according to any one of claims 11 to 12, in the manufacture of a PI3K alpha inhibitor medicament.
15. The use according to claim 13 or 14, wherein the use is in the manufacture of a medicament for the treatment of cancer, bone disease, inflammatory disease, immune disease, neurological disease, metabolic disease, respiratory disease and heart disease; wherein the cancer is selected from the group consisting of breast cancer, pancreatic cancer, non-small cell lung cancer, thyroid cancer, seminoma, melanoma, bladder cancer, liver cancer, kidney cancer, acute myelogenous leukemia, and colorectal cancer.
16. The use according to claim 13 or 14, wherein the use is in the manufacture of a medicament for the treatment of myelodysplastic syndrome.
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