WO2018130155A1 - Hdac6选择性抑制剂及其制备方法和应用 - Google Patents

Hdac6选择性抑制剂及其制备方法和应用 Download PDF

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Publication number
WO2018130155A1
WO2018130155A1 PCT/CN2018/072088 CN2018072088W WO2018130155A1 WO 2018130155 A1 WO2018130155 A1 WO 2018130155A1 CN 2018072088 W CN2018072088 W CN 2018072088W WO 2018130155 A1 WO2018130155 A1 WO 2018130155A1
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Prior art keywords
compound
group
pharmaceutically acceptable
acceptable salt
isomer
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PCT/CN2018/072088
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English (en)
French (fr)
Inventor
吴颢
韦昌青
郭强
张贵芬
刘斌
廖勇刚
肖瑶
陈曙辉
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南京明德新药研发股份有限公司
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Priority to BR112019014152-9A priority Critical patent/BR112019014152A2/pt
Priority to RU2019124962A priority patent/RU2772274C2/ru
Priority to US16/476,660 priority patent/US10745389B2/en
Priority to CN201880006463.5A priority patent/CN110382463B/zh
Application filed by 南京明德新药研发股份有限公司 filed Critical 南京明德新药研发股份有限公司
Priority to KR1020197022672A priority patent/KR102549606B1/ko
Priority to SG11201906361YA priority patent/SG11201906361YA/en
Priority to EP18739134.7A priority patent/EP3569592B8/en
Priority to JP2019557669A priority patent/JP7237010B2/ja
Priority to AU2018208160A priority patent/AU2018208160B2/en
Priority to ES18739134T priority patent/ES2927352T3/es
Priority to CA3049834A priority patent/CA3049834A1/en
Priority to MX2019008181A priority patent/MX2019008181A/es
Publication of WO2018130155A1 publication Critical patent/WO2018130155A1/zh
Priority to SA519402090A priority patent/SA519402090B1/ar
Priority to IL267968A priority patent/IL267968B/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D405/00—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom
    • C07D405/14—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing three or more hetero rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D307/00—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom
    • C07D307/02—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings
    • C07D307/04—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings having no double bonds between ring members or between ring members and non-ring members
    • C07D307/10—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings having no double bonds between ring members or between ring members and non-ring members with substituted hydrocarbon radicals attached to ring carbon atoms
    • C07D307/16—Radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/16—Amides, e.g. hydroxamic acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/34—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having five-membered rings with one oxygen as the only ring hetero atom, e.g. isosorbide
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/34—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having five-membered rings with one oxygen as the only ring hetero atom, e.g. isosorbide
    • A61K31/341—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having five-membered rings with one oxygen as the only ring hetero atom, e.g. isosorbide not condensed with another ring, e.g. ranitidine, furosemide, bufetolol, muscarine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/4427—Non condensed pyridines; Hydrogenated derivatives thereof containing further heterocyclic ring systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/4427—Non condensed pyridines; Hydrogenated derivatives thereof containing further heterocyclic ring systems
    • A61K31/443—Non condensed pyridines; Hydrogenated derivatives thereof containing further heterocyclic ring systems containing a five-membered ring with oxygen as a ring hetero atom
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/69—Boron compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0053—Mouth and digestive tract, i.e. intraoral and peroral administration
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D211/00—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings
    • C07D211/04—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D211/80—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having two double bonds between ring members or between ring members and non-ring members
    • C07D211/84—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having two double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms, with at the most one bond to halogen directly attached to ring carbon atoms
    • C07D211/86—Oxygen atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D307/00—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom
    • C07D307/02—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings
    • C07D307/04—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings having no double bonds between ring members or between ring members and non-ring members
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D307/00—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom
    • C07D307/02—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings
    • C07D307/04—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings having no double bonds between ring members or between ring members and non-ring members
    • C07D307/06—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings having no double bonds between ring members or between ring members and non-ring members with only hydrogen atoms or radicals containing only hydrogen and carbon atoms, directly attached to ring carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D309/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only ring hetero atom, not condensed with other rings
    • C07D309/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only ring hetero atom, not condensed with other rings having no double bonds between ring members or between ring members and non-ring members
    • C07D309/04—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only ring hetero atom, not condensed with other rings having no double bonds between ring members or between ring members and non-ring members with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached to ring carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D405/00—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom
    • C07D405/02—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings
    • C07D405/04—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings directly linked by a ring-member-to-ring-member bond

Definitions

  • the present invention relates to a class of compounds which are selective inhibitors of histone deacetylase 6 (HDAC6) and their use in the preparation of a medicament for the treatment of HDAC6 related diseases. Specifically, it relates to a compound of the formula (I) and a pharmaceutically acceptable salt thereof.
  • HDAC6 histone deacetylase 6
  • HDAC inhibitors are widely used in a variety of cancers and can be combined with a variety of drugs to enhance the therapeutic effect of the drug. It is a well-recognized anti-tumor target.
  • HDAC histone deacetylase
  • HAT histone acetyltransferase
  • overexpression of HDAC leads to an increase in deacetylation, thereby increasing the gravitational pull between DNA and histones, making the nucleosomes very tight, which is detrimental to the expression of tumor suppressor genes.
  • the inhibitor (HDACi) can be a new class of anti-tumor drugs by increasing histone acetylation, thereby regulating the expression of apoptosis and differentiation-related proteins, inducing apoptosis and differentiation. Moreover, HDAC is also involved in the regulation of many metabolic diseases, such as Alzheimer's disease, Parkinson's disease, etc. HDACi inhibitors have shown good results in both animal and human trials.
  • HDAC6 is the only deacetylase subtype in the cytoplasm, while the other 17 HDACs are present in the nucleus. HDAC6 does not directly catalyze histones, but uses tubulin and heat shock protein (Hsp90) as substrates through which cell trafficking, adhesion and movement (ie, no gene regulation) are regulated. Therefore, it is believed that it will Gene-related physiological functions produce less effect and thus have fewer side effects. Current clinical trial results have confirmed that HDAC6 selective inhibitors are safe and effective (POC). Clinical studies of the first HDAC6 selective inhibitor, ACY-1215 (Acetylon), have shown that selective HDAC6 inhibitors have better safety and therefore have better commercial prospects.
  • Hsp90 tubulin and heat shock protein
  • Proteasome inhibitors can block the degradation of a large number of regulatory proteins, causing disturbances and overloads in the intracellular signaling system, leading to inhibition of cell growth and ultimately delaying tumor progression. And even stop.
  • HDAC inhibitors are widely used in a variety of cancers, and can be combined with a variety of drugs to enhance the therapeutic effect of the drug, such as the HDAC inhibitor panobinostat combined with the proteasome inhibitor bortezomib (bortezomib) can increase the efficacy of multiple myeloma and Significantly reduce toxicity.
  • the present invention provides a compound of the formula (I), a pharmaceutically acceptable salt and isomers thereof:
  • n is selected from: 0 or 1;
  • T 3 is selected from: C or N;
  • Z 1 , Z 2 , Z 3 are each independently selected from: CH or N;
  • R 1 is selected from C 1 1-3 alkyl, phenyl, 6-membered heteroaryl optionally substituted by 1, 2 or 3;
  • R 2 is selected from the group consisting of: H, F, Cl, Br, I;
  • Ring A is selected from the group consisting of: 4 to 7-membered heterocycloalkyl
  • R is selected from the group consisting of: F, Cl, Br, I;
  • hetero of the 6-membered heteroaryl group and the 4-7-membered heterocycloalkyl group are each independently selected from: -NH-, N, -O-;
  • the number of heteroatoms or heteroatoms is independently selected from 1, 2 or 3.
  • R 1 is selected from the group consisting of: 1, 2 or 3 R substituted: methyl, ethyl, isopropyl, phenyl, pyridyl.
  • R 1 above is selected from: CH 3 , optionally substituted by 1, 2 or 3 R,
  • R 1 is selected from the group consisting of: CH 3 ,
  • the A ring is selected from the group consisting of: butylbutanyl, tetrahydrofuranyl, tetrahydropyranyl, 1,3-dioxolanyl, 1,4-dioxepane, 1 4-dioxaalkyl, 1,4-oxazacycloheptyl, morpholinyl.
  • the A ring is selected from the group consisting of:
  • the structural unit From:
  • the structural unit From:
  • the structural unit From:
  • the structural unit From:
  • the structural unit From:
  • the structural unit From:
  • the structural unit From:
  • the structural unit From:
  • the structural unit From:
  • the structural unit From:
  • the structural unit From:
  • R 1 above is selected from the group consisting of: 1, 2 or 3 R substituted: methyl, ethyl, phenyl, pyridyl, and other variables are as defined herein.
  • R 1 above is selected from: CH 3 , optionally substituted by 1, 2 or 3 R, Other variables are as defined by the present invention.
  • R 1 is selected from the group consisting of: CH 3 ,
  • Other variables are as defined by the present invention.
  • the A ring is selected from the group consisting of: butylbutanyl, tetrahydrofuranyl, tetrahydropyranyl, 1,3-dioxolanyl, 1,4-dioxepane, 1 , 4-dioxanyl, 1,4-oxazepanyl, morpholinyl, other variables are as defined herein.
  • the A ring is selected from the group consisting of: Other variables are as defined by the present invention.
  • the structural unit Selected from: -CH 2 -, -NH-, -C( O)-NH-, Other variables are as defined by the present invention.
  • the above compounds, pharmaceutically acceptable salts and isomers thereof are selected from the group consisting of:
  • a ring, R 1 , R 2 , L 1 and n are as defined in the present invention.
  • the above compounds, pharmaceutically acceptable salts and isomers thereof are selected from the group consisting of:
  • E 1 and E 2 are each independently selected from -O-, -CH 2 - and -CH 2 -CH 2 ;
  • R, R 2 , L 1 and n are as defined in the present invention.
  • the present invention also provides the following compounds and pharmaceutically acceptable salts thereof, selected from the group consisting of:
  • the above compounds, pharmaceutically acceptable salts and isomers thereof are selected from the group consisting of:
  • the present invention also provides a pharmaceutical composition
  • a pharmaceutical composition comprising a therapeutically effective amount of the above compound or a pharmaceutically acceptable salt thereof as an active ingredient together with a pharmaceutically acceptable carrier.
  • the invention also provides the use of a compound, or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the treatment of a HDAC6 related disorder.
  • the invention also provides the use of the above composition for the manufacture of a medicament for the treatment of a HDAC6 related disorder.
  • the above application is characterized in that the medicament is a medicament for treating multiple myeloma.
  • HDAC6 histone deacetylase 6
  • the compound of the present invention has remarkable in vitro activity, and has a remarkable inhibitory effect on HDAC6 enzyme, and has a high selectivity for weak inhibition of HDAC1;
  • co-administration with Ixazomib increased the efficacy of multiple myeloma and significantly reduced toxicity.
  • pharmaceutically acceptable salt refers to a salt of a compound of the invention prepared from a compound having a particular substituent found in the present invention and a relatively non-toxic acid or base.
  • a base addition salt can be obtained by contacting a neutral amount of such a compound with a sufficient amount of a base in a neat solution or a suitable inert solvent.
  • Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic ammonia or magnesium salts or similar salts.
  • an acid addition salt can be obtained by contacting a neutral form of such a compound with a sufficient amount of an acid in a neat solution or a suitable inert solvent.
  • pharmaceutically acceptable acid addition salts include inorganic acid salts including, for example, hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, hydrogencarbonate, phosphoric acid, monohydrogen phosphate, dihydrogen phosphate, sulfuric acid, Hydrogen sulfate, hydroiodic acid, phosphorous acid, etc.; and an organic acid salt, such as acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, Similar acids such as fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid, and me
  • the salt is contacted with a base or acid in a conventional manner, and the parent compound is separated, thereby regenerating the neutral form of the compound.
  • the parent form of the compound differs from the form of its various salts by certain physical properties, such as differences in solubility in polar solvents.
  • a "pharmaceutically acceptable salt” is a derivative of a compound of the invention wherein the parent compound is modified by salt formation with an acid or with a base.
  • pharmaceutically acceptable salts include, but are not limited to, inorganic or organic acid salts of bases such as amines, alkali metal or organic salts of acid groups such as carboxylic acids, and the like.
  • Pharmaceutically acceptable salts include the conventional non-toxic salts or quaternary ammonium salts of the parent compound, for example salts formed from non-toxic inorganic or organic acids.
  • non-toxic salts include, but are not limited to, those derived from inorganic acids and organic acids selected from the group consisting of 2-acetoxybenzoic acid, 2-hydroxyethanesulfonic acid, acetic acid, ascorbic acid, Benzenesulfonic acid, benzoic acid, hydrogencarbonate, carbonic acid, citric acid, edetic acid, ethane disulfonic acid, ethanesulfonic acid, fumaric acid, glucoheptose, gluconic acid, glutamic acid, glycolic acid, Hydrobromic acid, hydrochloric acid, hydroiodide, hydroxyl, hydroxynaphthalene, isethionethane, lactic acid, lactose, dodecylsulfonic acid, maleic acid, malic acid, mandelic acid, methanesulfonic acid, nitric acid, oxalic acid, Pamoic acid, pantothenic acid, phenylacetic acid, phen
  • the pharmaceutically acceptable salts of the present invention can be synthesized from the parent compound containing an acid group or a base by conventional chemical methods.
  • such salts are prepared by reacting these compounds in water or an organic solvent or a mixture of the two via a free acid or base form with a stoichiometric amount of a suitable base or acid.
  • a nonaqueous medium such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile is preferred.
  • the compounds provided herein also exist in the form of prodrugs.
  • Prodrugs of the compounds described herein are readily chemically altered under physiological conditions to convert to the compounds of the invention.
  • prodrugs can be converted to the compounds of the invention by chemical or biochemical methods in an in vivo setting.
  • Certain compounds of the invention may exist in unsolvated or solvated forms, including hydrated forms.
  • the solvated forms are equivalent to the unsolvated forms and are included within the scope of the invention.
  • Certain compounds of the invention may have asymmetric carbon atoms (optical centers) or double bonds. Racemates, diastereomers, geometric isomers and individual isomers are included within the scope of the invention.
  • Wedge and dashed keys unless otherwise stated Represents the absolute configuration of a stereocenter, using wavy lines Indicates a wedge or dashed key or use Indicates the relative configuration of the stereocenter.
  • the compounds described herein contain olefinic double bonds or other centers of geometric asymmetry, they include the E and Z geometric isomers unless otherwise specified. Likewise, all tautomeric forms are included within the scope of the invention.
  • the compounds of the invention may exist in specific geometric or stereoisomeric forms.
  • the present invention contemplates all such compounds, including the cis and trans isomers, the (-)- and (+)-p-enantiomers, the (R)- and (S)-enantiomers, and the diastereomeric a conformation, a (D)-isomer, a (L)-isomer, and a racemic mixture thereof, and other mixtures, such as enantiomerically or diastereomeric enriched mixtures, all of which belong to It is within the scope of the invention.
  • Additional asymmetric carbon atoms may be present in the substituents such as alkyl groups. All such isomers, as well as mixtures thereof, are included within the scope of the invention.
  • optically active (R)- and (S)-isomers as well as the D and L isomers can be prepared by chiral synthesis or chiral reagents or other conventional techniques. If an enantiomer of a compound of the invention is desired, it can be prepared by asymmetric synthesis or by derivatization with a chiral auxiliary wherein the resulting mixture of diastereomers is separated and the auxiliary group cleaved to provide pure The desired enantiomer.
  • a diastereomeric salt is formed with a suitable optically active acid or base, followed by conventional methods well known in the art.
  • the diastereomers are resolved and the pure enantiomer is recovered.
  • the separation of enantiomers and diastereomers is generally accomplished by the use of chromatography using a chiral stationary phase, optionally in combination with chemical derivatization (eg, formation of an amino group from an amine). Formate).
  • the compounds of the present invention may contain unnatural proportions of atomic isotopes on one or more of the atoms that make up the compound.
  • radiolabeled compounds can be used, such as tritium (3 H), iodine -125 (125 I) or C-14 (14 C). Alterations of all isotopic compositions of the compounds of the invention, whether radioactive or not, are included within the scope of the invention.
  • "Optional” or “optionally” means that the subsequently described event or condition may, but is not necessarily, to occur, and that the description includes instances in which the event or condition occurs and instances in which the event or condition does not occur.
  • substituted means that any one or more hydrogen atoms on a particular atom are replaced by a substituent, and may include variants of heavy hydrogen and hydrogen, as long as the valence of the particular atom is normal and the substituted compound is stable. of.
  • Ketone substitution does not occur on the aryl group.
  • optionally substituted means that it may or may not be substituted, and unless otherwise specified, the kind and number of substituents may be arbitrary on the basis of chemically achievable.
  • any variable eg, R
  • its definition in each case is independent.
  • the group may optionally be substituted with at most two R, and each case has an independent option.
  • combinations of substituents and/or variants thereof are permissible only if such combinations result in stable compounds.
  • linking group When the number of one linking group is 0, such as -(CRR) 0 -, it indicates that the linking group is a single bond.
  • one of the variables When one of the variables is selected from a single bond, it means that the two groups to which it is attached are directly linked. For example, when L represents a single bond in A-L-Z, the structure is actually A-Z.
  • substituent When a substituent is vacant, it means that the substituent is absent. For example, when X is vacant in AX, the structure is actually A. When a bond of a substituent can be cross-linked to two atoms on a ring, the substituent can be bonded to any atom on the ring. When the recited substituents do not indicate which atom is attached to a compound included in the chemical structural formula including but not specifically mentioned, such a substituent may be bonded through any atomic phase thereof. Combinations of substituents and/or variants thereof are permissible only if such combinations result in stable compounds. For example, a structural unit or It is indicated that it can be substituted at any position on the cyclohexyl or cyclohexadiene.
  • hetero denotes a hetero atom or a hetero atomic group (ie, a radical containing a hetero atom), including atoms other than carbon (C) and hydrogen (H), and radicals containing such heteroatoms, including, for example, oxygen (O).
  • ring means substituted or unsubstituted cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, cycloalkynyl, heterocycloalkynyl, aryl or heteroaryl. So-called rings include single rings, interlocking rings, spiral rings, parallel rings or bridge rings. The number of atoms on the ring is usually defined as the number of elements of the ring. For example, "5 to 7-membered ring” means 5 to 7 atoms arranged in a circle. Unless otherwise specified, the ring optionally contains from 1 to 3 heteroatoms.
  • 5- to 7-membered ring includes, for example, phenyl, pyridine, and piperidinyl; on the other hand, the term “5- to 7-membered heterocycloalkyl ring” includes pyridyl and piperidinyl, but does not include phenyl.
  • ring also includes ring systems containing at least one ring, each of which "ring” independently conforms to the above definition.
  • heterocycle or “heterocyclyl” means a stable monocyclic, bicyclic or tricyclic ring containing a hetero atom or a heteroatom group which may be saturated, partially unsaturated or unsaturated ( Aromatic) which comprise a carbon atom and 1, 2, 3 or 4 ring heteroatoms independently selected from N, O and S, wherein any of the above heterocycles may be fused to a phenyl ring to form a bicyclic ring.
  • the nitrogen and sulfur heteroatoms can be optionally oxidized (i.e., NO and S(O)p, p is 1 or 2).
  • the nitrogen atom can be substituted or unsubstituted (i.e., N or NR, wherein R is H or other substituents as already defined herein).
  • the heterocyclic ring can be attached to the side groups of any hetero atom or carbon atom to form a stable structure. If the resulting compound is stable, the heterocycles described herein can undergo substitutions at the carbon or nitrogen sites.
  • the nitrogen atom in the heterocycle is optionally quaternized.
  • a preferred embodiment is that when the total number of S and O atoms in the heterocycle exceeds 1, these heteroatoms are not adjacent to each other. Another preferred embodiment is that the total number of S and O atoms in the heterocycle does not exceed one.
  • aromatic heterocyclic group or "heteroaryl” as used herein means a stable 5, 6, or 7 membered monocyclic or bicyclic or aromatic ring of a 7, 8, 9 or 10 membered bicyclic heterocyclic group, It contains carbon atoms and 1, 2, 3 or 4 ring heteroatoms independently selected from N, O and S.
  • the nitrogen atom can be substituted or unsubstituted (i.e., N or NR, wherein R is H or other substituents as already defined herein).
  • the nitrogen and sulfur heteroatoms can be optionally oxidized (i.e., NO and S(O)p, p is 1 or 2).
  • bridged rings are also included in the definition of heterocycles.
  • a bridged ring is formed when one or more atoms (ie, C, O, N, or S) join two non-adjacent carbon or nitrogen atoms.
  • Preferred bridged rings include, but are not limited to, one carbon atom, two carbon atoms, one nitrogen atom, two nitrogen atoms, and one carbon-nitrogen group. It is worth noting that a bridge always converts a single ring into a three ring. In the bridged ring, a substituent on the ring can also be present on the bridge.
  • heterocyclic compounds include, but are not limited to, acridinyl, octanoyl, benzimidazolyl, benzofuranyl, benzofuranylfuranyl, benzindenylphenyl, benzoxazolyl, benzimidin Oxazolinyl, benzothiazolyl, benzotriazolyl, benzotetrazolyl, benzisoxazolyl, benzisothiazolyl, benzimidazolyl, oxazolyl, 4aH-carbazolyl, Porphyrin, chroman, chromene, porphyrin-decahydroquinolinyl, 2H, 6H-1,5,2-dithiazinyl, dihydrofuro[2,3-b] Tetrahydrofuranyl, furyl, furfuryl, imidazolidinyl, imidazolinyl, imidazolyl, 1H-carbazolyl, nonenyl,
  • hydrocarbyl or its subordinate concept (such as alkyl, alkenyl, alkynyl, aryl, etc.), by itself or as part of another substituent, is meant to be straight-chain, branched or cyclic.
  • the hydrocarbon atom group or a combination thereof may be fully saturated (such as an alkyl group), a unit or a polyunsaturated (such as an alkenyl group, an alkynyl group, an aryl group), may be monosubstituted or polysubstituted, and may be monovalent (such as Methyl), divalent (such as methylene) or polyvalent (such as methine), may include divalent or polyvalent radicals with a specified number of carbon atoms (eg, C 1 -C 12 represents 1 to 12 carbons) , C 1-12 is selected from C 1 , C 2 , C 3 , C 4 , C 5 , C 6 , C 7 , C 8 , C 9 , C 10 , C 11 and C 12 ; C 3-12 is selected from C 3 , C 4 , C 5 , C 6 , C 7 , C 8 , C 9 , C 10 , C 11 and C 12 .).
  • C 1-12 is selected from C 1
  • Hydrocarbyl includes, but is not limited to, aliphatic hydrocarbyl groups including chain and cyclic, including but not limited to alkyl, alkenyl, alkynyl groups including, but not limited to, 6-12 members.
  • An aromatic hydrocarbon group such as benzene, naphthalene or the like.
  • hydrocarbyl means a straight or branched chain radical or a combination thereof, which may be fully saturated, unitary or polyunsaturated, and may include divalent and multivalent radicals.
  • saturated hydrocarbon radicals include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl, isobutyl, sec-butyl, isobutyl, cyclohexyl, (cyclohexyl).
  • a homolog or isomer of a methyl group, a cyclopropylmethyl group, and an atomic group such as n-pentyl, n-hexyl, n-heptyl, n-octyl.
  • the unsaturated hydrocarbon group has one or more double or triple bonds, and examples thereof include, but are not limited to, a vinyl group, a 2-propenyl group, a butenyl group, a crotyl group, a 2-isopentenyl group, and a 2-(butadienyl group). , 2,4-pentadienyl, 3-(1,4-pentadienyl), ethynyl, 1- and 3-propynyl, 3-butynyl, and higher homologs and isomers body.
  • heterohydrocarbyl or its subordinate concept (such as heteroalkyl, heteroalkenyl, heteroalkynyl, heteroaryl, etc.), by itself or in combination with another term, means a stable straight chain, branched chain. Or a cyclic hydrocarbon radical or a combination thereof having a number of carbon atoms and at least one heteroatom.
  • heteroalkyl by itself or in conjunction with another term refers to a stable straight chain, branched hydrocarbon radical or combination thereof, having a number of carbon atoms and at least one heteroatom.
  • the heteroatoms are selected from the group consisting of B, O, N, and S, wherein the nitrogen and sulfur atoms are optionally oxidized, and the nitrogen heteroatoms are optionally quaternized.
  • the hetero atom or heteroatom group may be located at any internal position of the heterohydrocarbyl group, including where the hydrocarbyl group is attached to the rest of the molecule, but the terms "alkoxy”, “alkylamino” and “alkylthio” (or thioalkoxy). By customary expression, those alkyl groups which are attached to the remainder of the molecule through an oxygen atom, an amino group or a sulfur atom, respectively.
  • Up to two heteroatoms may be consecutive, for example, -CH 2 -NH-OCH 3.
  • cycloalkyl refers to any heterocyclic alkynyl group, etc., by itself or in combination with other terms, denotes a cyclized “hydrocarbyl group” or “heterohydrocarbyl group”, respectively.
  • a hetero atom may occupy a position at which the hetero ring is attached to the rest of the molecule.
  • cycloalkyl groups include, but are not limited to, cyclopentyl, cyclohexyl, 1-cyclohexenyl, 3-cyclohexenyl, cycloheptyl, and the like.
  • heterocyclic groups include 1-(1,2,5,6-tetrahydropyridyl), 1-piperidinyl, 2-piperidinyl, 3-piperidinyl, 4-morpholinyl, 3-morpholinyl, tetrahydrofuran-2-yl, tetrahydrofuran-3-yl, tetrahydrothiophen-2-yl, tetrahydrothiophen-3-yl, 1-piperazinyl and 2-piperazinyl.
  • alkyl is used to denote a straight or branched saturated hydrocarbon group, which may be monosubstituted (eg, -CH 2 F) or polysubstituted (eg, -CF 3 ), and may be monovalent (eg, Methyl), divalent (such as methylene) or polyvalent (such as methine).
  • alkyl group include methyl (Me), ethyl (Et), propyl (e.g., n-propyl and isopropyl), butyl (e.g., n-butyl, isobutyl, s-butyl). , t-butyl), pentyl (eg, n-pentyl, isopentyl, neopentyl) and the like.
  • alkenyl refers to an alkyl group having one or more carbon-carbon double bonds at any position of the chain, which may be mono- or poly-substituted, and may be monovalent, divalent or multivalent.
  • alkenyl group include a vinyl group, a propenyl group, a butenyl group, a pentenyl group, a hexenyl group, a butadienyl group, a pentadienyl group, a hexadienyl group and the like.
  • alkynyl refers to an alkyl group having one or more carbon-carbon triple bonds at any position of the chain, which may be mono- or poly-substituted, and may be monovalent, divalent or multivalent.
  • alkynyl groups include ethynyl, propynyl, butynyl, pentynyl and the like.
  • a cycloalkyl group includes any stable cyclic or polycyclic hydrocarbon group, any carbon atom which is saturated, may be monosubstituted or polysubstituted, and may be monovalent, divalent or multivalent.
  • Examples of such cycloalkyl groups include, but are not limited to, cyclopropyl, norbornyl, [2.2.2]bicyclooctane, [4.4.0]bicyclononane, and the like.
  • a cycloalkenyl group includes any stable cyclic or polycyclic hydrocarbon group which contains one or more unsaturated carbon-carbon double bonds at any position of the ring, and may be monosubstituted or polysubstituted, It can be one price, two price or multiple price.
  • Examples of such cycloalkenyl groups include, but are not limited to, cyclopentenyl, cyclohexenyl, and the like.
  • a cycloalkynyl group includes any stable cyclic or polycyclic hydrocarbon group which contains one or more carbon-carbon triple bonds at any position of the ring, which may be monosubstituted or polysubstituted, and may be one Price, price or price.
  • halo or “halogen”, by itself or as part of another substituent, denotes a fluorine, chlorine, bromine or iodine atom.
  • haloalkyl is intended to include both monohaloalkyl and polyhaloalkyl.
  • halo(C 1 -C 4 )alkyl is intended to include, but is not limited to, trifluoromethyl, 2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the like. Wait.
  • examples of haloalkyl include, but are not limited to, trifluoromethyl, trichloromethyl, pentafluoroethyl, and pentachloroethyl.
  • alkoxy represents attached through an oxygen bridge
  • C 1-6 alkoxy groups include C 1, C 2, C 3 , C 4, C 5 , and C 6 alkoxy groups.
  • alkoxy groups include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, sec-butoxy, tert-butoxy, n-pentyloxy and S- Pentyloxy.
  • aryl denotes a polyunsaturated, aromatic hydrocarbon substituent which may be monosubstituted or polysubstituted, which may be monovalent, divalent or polyvalent, which may be monocyclic or polycyclic ( For example, 1 to 3 rings; at least one of which is aromatic), they are fused together or covalently linked.
  • heteroaryl refers to an aryl (or ring) containing one to four heteroatoms. In an illustrative example, the heteroatoms are selected from the group consisting of B, N, O, and S, wherein the nitrogen and sulfur atoms are optionally oxidized, and the nitrogen atom is optionally quaternized.
  • a heteroaryl group can be attached to the remainder of the molecule through a heteroatom.
  • aryl or heteroaryl groups include phenyl, naphthyl, biphenyl, pyrrolyl, pyrazolyl, imidazolyl, pyrazinyl, oxazolyl, phenyl-oxazolyl, isomerism Azyl, thiazolyl, furyl, thienyl, pyridyl, pyrimidinyl, benzothiazolyl, indolyl, benzimidazolyl, indolyl, isoquinolyl, quinoxalinyl, quinolinyl, 1 -naphthyl, 2-naphthyl, 4-biphenylyl, 1-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl, 3-pyrazolyl, 2-imidazolyl, 4-imidazolyl, pyrazinyl, 2-oxazolyl
  • aryl groups when used in conjunction with other terms (e.g., aryloxy, arylthio, aralkyl), include aryl and heteroaryl rings as defined above.
  • aralkyl is intended to include those radicals to which an aryl group is attached to an alkyl group (eg, benzyl, phenethyl, pyridylmethyl, and the like), including wherein the carbon atom (eg, methylene) has been, for example, oxygen.
  • alkyl groups substituted by an atom such as phenoxymethyl, 2-pyridyloxymethyl 3-(1-naphthyloxy)propyl and the like.
  • leaving group refers to a functional group or atom which may be substituted by another functional group or atom by a substitution reaction (for example, an affinity substitution reaction).
  • substituent groups include triflate; chlorine, bromine, iodine; sulfonate groups such as mesylate, tosylate, p-bromobenzenesulfonate, p-toluenesulfonic acid Esters and the like; acyloxy groups such as acetoxy, trifluoroacetoxy and the like.
  • protecting group includes, but is not limited to, "amino protecting group", “hydroxy protecting group” or “thiol protecting group”.
  • amino protecting group refers to a protecting group suitable for preventing side reactions at the amino nitrogen position.
  • Representative amino protecting groups include, but are not limited to, formyl; acyl, such as alkanoyl (e.g., acetyl, trichloroacetyl or trifluoroacetyl); alkoxycarbonyl, e.g., tert-butoxycarbonyl (Boc) Arylmethoxycarbonyl, such as benzyloxycarbonyl (Cbz) and 9-fluorenylmethoxycarbonyl (Fmoc); arylmethyl, such as benzyl (Bn), trityl (Tr), 1, 1-di -(4'-methoxyphenyl)methyl; silyl groups such as trimethylsilyl (TMS) and tert-
  • hydroxy protecting group refers to a protecting group suitable for use in preventing hydroxy side reactions.
  • Representative hydroxy protecting groups include, but are not limited to, alkyl groups such as methyl, ethyl and t-butyl groups; acyl groups such as alkanoyl groups (e.g., acetyl); arylmethyl groups such as benzyl (Bn), Oxybenzyl (PMB), 9-fluorenylmethyl (Fm) and diphenylmethyl (diphenylmethyl, DPM); silyl groups such as trimethylsilyl (TMS) and tert-butyl Dimethylsilyl (TBS) and the like.
  • alkyl groups such as methyl, ethyl and t-butyl groups
  • acyl groups such as alkanoyl groups (e.g., acetyl)
  • arylmethyl groups such as benzyl (Bn), Oxybenzyl (PMB), 9-fluoreny
  • the compounds of the present invention can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments set forth below, combinations thereof with other chemical synthetic methods, and those well known to those skilled in the art. Equivalent alternatives, preferred embodiments include, but are not limited to, embodiments of the invention.
  • the solvent used in the present invention is commercially available.
  • the present invention employs the following abbreviations: aq for water; HATU for O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate ; EDC stands for N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride; m-CPBA stands for 3-chloroperoxybenzoic acid; eq stands for equivalent, equivalent; CDI stands for Carbonyldiimidazole; DCM stands for dichloromethane; PE stands for petroleum ether; DIAD stands for diisopropyl azodicarboxylate; DMF stands for N,N-dimethylformamide; DMSO stands for dimethyl sulfoxide; EtOAc stands for acetic acid Esters; EtOH for ethanol; MeOH for methanol; CBz for benzyl
  • Step 1 Compound 1-1 (11.30 g. 80.14 mmol) was added to dry THF (120.00 mL), and then, iPrMgCl (2M, 40.07 mL) was added dropwise at -78 °C, and stirred at 15 ° C for 16 hours under nitrogen atmosphere. After completion of the reaction, saturated NH 4 Cl (50 mL) was added.
  • Step 2 Compound 1-2 (3.70 g, 24.89 mmol) and 2-chloro-4-bromopyridine (4.79 g, 24.89 mmol) were added to dry THF (50.00 mL). BuLi (2.5 M, 9.96 mL) was stirred at 20 ° C for 18 hours under nitrogen. After the completion of the reaction, saturated NH 4 Cl was added, and the residue was extracted and concentrated.
  • Step 3 Compound 1-3 (1.60 g, 7.09 mmol), KOH (795.64 mg, 14.18 mmol) was added to a mixed solvent of dioxane (15.00 mL) / H 2 O (5.00 mL), then Pd was added 2 (dba) 3 (324.62 mg, 354.50 ⁇ mol) and tBuXPhOS (301.07 mg, 709.00 ⁇ mol) were stirred at 115 ° C for 18 hours under nitrogen. After completion of the reaction, it was diluted with water and extracted with ethyl acetate. The residue was purified by silica gel column chromatography to afford Compound 1-4. MS ESI calculated for C 12 H 16 ClNO [M+H] + 208, found 208.
  • Step 4 Compound 1-4 (200.00 mg, 964.92 ⁇ mol) was added to dry DMF (50.00 mL), NaH (60%, 57.90 mg, 1.45 mmol) was added at 0 ° C, stirred at 25 ° C for 0.5 h, then 0 Methyl 4-bromomethylbenzoate (221.04 mg, 964.92 ⁇ mol) was added at ° C and stirred at 25 ° C for 0.5 h. After completion of the reaction, 20 mL of water was added, and the filter residue was dried by filtration and purified by column chromatography to give Compound 1-5. MS ESI calcd for C 21 H 25 NO 4 [M + H] + 356, found 356.
  • Step 5 Compound 1-5 (100 mg, 281.35 ⁇ mol) was added to a mixed solvent of DCM/H 2 O (15.00 mL), 2M NaOH (2 mL) was added at 0 ° C, then 50% NH 2 OH (2 mL) was added Stir at 25 ° C for 0.5 hours.
  • the title compound 1-6 was isolated by preparative HPLC.
  • Step 1 To a compound 2-1 (10.00 g, 51.96 mmol, 1.00 eq) and 5-chloropentan-2-one (6.27 g, 51.96 mmol, 1.00 eq) of tetrahydrofuran (100.00) at -78 ° C under nitrogen atmosphere. n-Butyllithium (2.5 M, 24.94 mL, 1.20 eq) was added dropwise to mL), and then stirred at -78 to 15 ° C for 5 hours under nitrogen atmosphere.
  • Step 2 Compound 2-2 (3.00 g, 15.18 mmol, 1.00 eq) and potassium hydroxide (1.28 g, 22.77 mmol, 1.50 eq) of dioxane (15.00 mL) and water (3.00). To the solution was added Pd 2 (dba) 3 (1.39 g, 1.52 mmol, 0.10 eq) and t-BuXphos (1.29 g, 3.04 mmol, 0.20 eq). It was replaced with nitrogen three times, and then stirred at 110 ° C for 12 hours in a nitrogen atmosphere. The reaction mixture was suction-filtered, poured into water, EtOAc (EtOAc: EtOAc (EtOAc) MS ESI calculated C 10 H 13 NO [M+H] + 180, found 180.
  • EtOAc EtOAc
  • Step 3 Compound 2-3 (230.00 mg, 1.28 mmol, 1.00 eq), methyl 4-(bromomethyl)benzoate (293.97 mg, 1.28 mmol, 1.00 eq), cesium carbonate (627.21 mg, 1.93 mmol, 1.50 eq), potassium iodide (10.65 mg, 64.17 ⁇ mol, 0.05 eq) in tetrahydrofuran (20.00 mL) was replaced with nitrogen three times and then stirred at a temperature of 66 ° C for 12 hours. The reaction mixture was concentrated, and then evaporated, evaporated, evaporated, evaporated MS ESI calcd for C 19 H 21 NO 4 [M + H] + 328, found 328.
  • Step 4 A solution of sodium hydroxide (2M, 2.29 mL, 5.00 eq) was added dropwise to EtOAc (EtOAc (EtOAc) The reaction solution was stirred at 15 ° C for 2.5 hours. The reaction mixture was concentrated, dissolved in water (15 mL), then extracted with dichloromethane:methanol (10:1, 15mL*3), washed with brine, dried over sodium sulfate, 5.
  • Step 1 Add n-butyllithium cyclohexane to a solution of 4-bromo-2-chloro-pyridine (20.00 g, 103.93 mmol, 1.00 eq) in toluene (30.00 mL) at -65 ° C to 75 ° C The solution (2.5 M, 49.89 mL, 1.20 eq) was stirred at -65 ° C to 75 ° C for 1 hour, then to the reaction system was added Compound 3-1 (20.85 g, 103.93 mmol, 17.09 mL, 1.00 eq) of toluene (100.00) The mL) solution was stirred and heated to 25 ° C for 2 hours.
  • Step 2 Compound 3-2 (8.50 g, 30.61 mmol, 1.00 eq), potassium hydroxide (3.44 g, 61.22 mmol, 2.00 eq), tris(dibenzylideneacetone) dipalladium (5.61 g, 6.12 mmol, 0.20 eq), 2-di-tert-butylphosphine-2',4',6'-triisopropylbiphenyl (2.60 g, 6.12 mmol, 0.20 eq) of dioxane (2.00 mL) and water (1.00) The reaction solution was heated to 100 ° C and stirred for 2 hours.
  • Step 3 To a solution of compound 3-3 (7.00 g, 27.00 mmol, 1.00 eq) in N,N-dimethylformamide (70.00 mL), EtOAc (1.08 g, Ment, purity 60%, 1.00 eq), stirred at 0-5 ° C for 10 minutes, then added 4-bromomethylbenzoic acid methyl ester (6.18 g, 27.00 mmol, 1.00 eq) of N, N-di A solution of methylformamide (20.00 mL) was stirred at 10-25 ° C for 2 minutes. A saturated aqueous ammonium chloride solution was added to the reaction mixture, and ethyl acetate was evaporated and evaporated. After concentration, compound 3-4 was obtained and used directly in the next step.
  • Step 4 To a mixed solution of compound 3-4 (11.00 g, 27.00 mmol, 1.00 eq), aqueous hydroxylamine solution (100.00 mL, 50% purity), methanol (20.00 mL) and dichloromethane (10.00 mL) A sodium aqueous solution (2M, 27.00 mL, 2.00 eq), aqueous hydroxyamine (1.00 mL, 50% purity), warmed to 40-50 ° C and stirred for 1 hour. After the reaction mixture was concentrated, the mixture was adjusted to pH 7 with 1M hydrochloric acid, and extracted with dichloromethane.
  • Step 1 To a solution of compound 4-1 (1.50 g, 5.79 mmol) in dimethylformamide (20.00 mL), sodium hydride (347.13 mg, 8.69 mmol, 60% purity), after stirring at 0 ° C for 30 min Further, methyl 4-bromomethylbenzoate (1.99 g, 8.69 mmol) was added, and the mixture was warmed to room temperature and stirred for 2 hr. It was quenched with EtOAc EtOAc (EtOAc) The combined organic phases were washed with EtOAc EtOAc (EtOAc) The crude mixture was obtained afterwards.
  • EtOAc EtOAc
  • the crude mixture was chiralized (chiral column type: OJ (250mm*30mm, 10 ⁇ m); mobile phase: A:CO2B: 0.05% diethylamine/EtOH, EtOH (0.05% diethylamine) and supercritical fluid CO2 From 5% to 40%, flow rate 60 mL/min), compound 4-2 and compound 5-1 were obtained after resolution, and the retention times were 2.226 min and 2.835 min, respectively.
  • Step 2 To a solution of compound 4-2 (700.00 mg, 1.72 mmol) in methanol (5.00 mL), sodium hydroxide (103.20 mg, 2.58 mmol) and hydroxylamine (5.00 mL, 50% aqueous). After concentration, the pH was adjusted to 7 with dilute hydrochloric acid (1M), and purified by reverse-phase preparative HPLC (0.225% FA) to afford compound 4-3.
  • Step 1 To a mixture of compound 6-1 (809.15 mg, 7.09 mmol, 697.54 ⁇ L, 1.00 eq) and aluminum trichloride (633.60 mg, 4.75 mmol, 259.67 ⁇ L, 0.67 eq) at 4 ° C Butyryl chloride (500.00 mg, 3.55 mmol, 396.83 ⁇ L, 0.50 eq), and the mixture was stirred at 25 ° C for 3 hr. Filtration and concentration of the filtrate gave Compound 6-2. This liquid was used directly in the next step of the reaction.
  • Step 2 Add a solution of n-butyllithium in cyclohexane (2.5 M) to a solution of 4-bromo-2-chloro-pyridine (527.29 mg, 2.74 mmol, 1.00 eq) in toluene (5.00 mL). , 1.21 mL, 1.10 eq), stirred at -68 ° C for 10 minutes, then a solution of compound 6-2 (600.00 mg, 2.74 mmol, 1.00 eq) in toluene (5.00 mL) was added to the reaction system, and stirred at -68 ° C for 1 hour. Stir at 25 ° C for 14 hours.
  • Step 3 To a solution of compound 6-3 (87.00 mg, 294.21 ⁇ mol, 1.00 eq) in dioxane (2.00 mL) and water (1.00 mL), potassium hydroxide (33.02 mg, 588.42 ⁇ mol, 2.00 eq) , tris(dibenzylideneacetone)dipalladium (26.94 mg, 29.42 ⁇ mol, 0.10 eq), 2-di-tert-butylphosphine-2',4',6'-triisopropylbiphenyl (12.49 mg, 29.42 Molmol, 0.10 eq), and stirred at 80 ° C for 3 hours. Water was added to the reaction mixture, and the mixture was evaporated. The concentrated TLC plate was purified to give the compound 6-4. MS ESI calcd for C 15 H 13 F 2 O 2 N [M + H] + 278, found 278.
  • Step 4 To a solution of compound 6-4 (60.00 mg, 216.40 ⁇ mol, 1.00 eq) in N,N-dimethylformamide (1.00 mL) was added sodium hydride (17.31 mg, 432.80 ⁇ mol, Purity 60%, 2.00 eq), then a solution of methyl 4-bromomethylbenzoate (49.57 mg, 216.40 ⁇ mol, 1.00 eq) in N,N-dimethylformamide (1.00 mL) was added to the reaction system, 25 Stir at ° C for 2 hours. Water was added to the reaction mixture, and the mixture was evaporated. After concentration, purification by TLC plate gave compound 6-5. MS ESI calcd for C 24 H 21 NO 4 F 2 [M + H] + 426, found 426.
  • Step 5 To a solution of compound 6-5 (32.00 mg, 75.22 ⁇ mol, 1.00 eq) in methanol (2.00 mL), aqueous sodium hydroxide (6M, 50.00 ⁇ L, 3.99 eq), aqueous hydroxy ammonia (1.00 mL, 50%) Aqueous solution), stirred at 25 ° C for 17 hours. After the reaction mixture was concentrated, the reverse phase preparative column was purified to give compound 6-6.
  • aqueous sodium hydroxide (6M, 50.00 ⁇ L, 3.99 eq
  • aqueous hydroxy ammonia (1.00 mL, 50%) Aqueous solution
  • Step 2 To a solution of compound 7-2 (70.00 mg, 161.11 mol) in methanol (5.00 mL) was added hydroxylamine (1.00 mL, 50% aqueous solution) and sodium hydroxide NaOH (6.44 mg, 161.11 ⁇ mol), stirred at room temperature 30 minutes. The mixture was adjusted to pH 7 with dilute hydrochloric acid (1M) and purified to afford compound 7-3.
  • Step 1 To a solution of compound 8-1 (70.00 mg, 269.99 ⁇ mol) in dimethylformamide (5.00 mL) was added sodium hydride (21.60 mg, 539.98 ⁇ mol, purity 60%) and stirred at 0 ° C for 30 min. Then, 2-bromomethyl 5-carboxylic acid methylpyrimidine (132.33 mg, 539.98 ⁇ mol) was added, and stirring was continued at 0 ° C for 30 minutes, and the mixture was further stirred for 30 minutes. The mixture was quenched with EtOAc EtOAc (EtOAc)EtOAc.
  • EtOAc EtOAc
  • Step 2 To a solution of compound 8-2 (70.00 mg, 160.72 ⁇ mol,) in methanol (5.00 mL) was added hydroxylamine (1.00 mL, 50% aqueous) and sodium hydroxide NaOH (6.43 mg, 160.72 ⁇ mol) at room temperature Stir for 30 minutes. The mixture was adjusted to pH 7 with dilute hydrochloric acid (1M) and purified to afford compound 8-3.
  • Step 1 To a solution of compound 9-1 (70.00 mg, 269.99 ⁇ mol) in dimethylformamide (5.00 mL) was added sodium hydride (21.60 mg, 539.98 ⁇ mol, purity 60%) and stirred at 0 ° C for 30 min. Then, methyl 2-formate-5-bromomethylpyridine (124.23 mg, 539.98 ⁇ mol) was added, and the mixture was further stirred at 0 ° C for 30 minutes, and the mixture was further stirred for 30 minutes. The mixture was quenched with EtOAc EtOAc (EtOAc)EtOAc. The combined organic phases were washed with water (10 mL*2) and brine (10 mL). MS ESI calcd for C 23 H 21 FN 2 O 4 [M + H] + 409, found 409.
  • Step 2 To a solution of compound 9-2 (50.00 mg, crude) in methanol (5.00 mL), EtOAc (EtOAc, EtOAc, EtOAc minute. The mixture was adjusted to pH 7 with dilute hydrochloric acid (1M) and purified to afford compound 9-3.
  • Step 1 To a mixture of compound 10-1 (4.68 g, 35.46 mmol, 3.66 mL, 2.00 eq) and aluminum trichloride (3.14 g, 23.58 mmol, 1.29 mL, 1.33 eq) was added 4-chlorobutyryl chloride (2.50) g, 17.73 mmol, 1.98 mL, 1.00 eq). After stirring at 30 ° C for 6 hours, the reaction mixture was poured into ice water, stirred, washed with ethyl acetate, washed with brine, dried over anhydrous sodium sulfate . After concentration, it was purified by silica gel column to give Compound 10-2. This liquid was used directly in the next step of the reaction.
  • Step 2 Add a solution of n-butyllithium in cyclohexane (2.5 M) to a solution of 4-bromo-2-chloro-pyridine (488.80 mg, 2.54 mmol, 1.00 eq) in toluene (3.00 mL). 1.12 mL, 1.10 eq), a solution of compound 10-2 (600.00 mg, 2.54 mmol, 1.00 eq) in toluene (1.00 mL) was added to the reaction mixture, and the mixture was warmed to 25 ° C and stirred for 6 hours. After a saturated aqueous solution of ammonium chloride and water were added, and the mixture was evaporated. After concentration, it was purified by silica gel column to give Compound 10-3. MS ESI calcd for C 15 H 11 F 3 ONCI [M+H] + 314, found 314.
  • Step 3 To a solution of compound 10-3 (410.00 mg, 1.31 mmol, 1.00 eq) in dioxane (5.00 mL) and water (1.00 mL), EtOAc (146.67 mg, 2.61 mmol, 2.00 eq) , tris(dibenzylideneacetone)dipalladium (119.96 mg, 131.00 ⁇ mol, 0.10 eq), 2-di-tert-butylphosphine-2',4',6'-triisopropylbiphenyl (55.50 mg, 130.70 Molmol, 0.10 eq), stirred at 80 ° C for 4 hours. A saturated aqueous solution of ammonium chloride and water were added, and the mixture was evaporated. After concentration, the silica gel column was purified to give the compound 10-4.
  • Step 4 Compound 10-4 (100.00 mg, 338.68 ⁇ mol, 1.00 eq), methyl 4-bromomethylbenzoate (155.16 mg, 677.36 ⁇ mol, 2.00 eq), cesium carbonate (220.70 mg, 677.37 ⁇ mol, 2.00 eq)
  • the acetonitrile (2.00 mL) solution was warmed to 80 ° C and stirred for 2 hours. Water was added to the reaction mixture, and the mixture was evaporated. After concentration, purification by preparative thin layer chromatography gave compound 10-5.
  • Step 5 To a solution of compound 10-5 (40.00 mg, 90.21 ⁇ mol, 1.00 eq) in methanol (2.00 mL), sodium hydroxide (3.61 mg, 90.21 ⁇ mol, 1.00 eq), aqueous hydroxy ammonia (1.00 mL, 50%) Purity), stirred at 25 ° C for 17 hours. After the reaction mixture was concentrated, the reverse phase preparative column was purified to give compound 10-6.
  • Step 1 protection of N 2, at -70 °C 4-bromo-2-chloro-3-fluoropyridine (4.00g, 19.01mmol, 1.00eq) in toluene (20.00 mL) was added dropwise n-BuLi (2.5 M, 9.12 mL, 1.20 eq), stirring was continued at this temperature for 30 minutes after completion of the dropwise addition, and then Compound 11-1 (3.81 g, 19.01 mmol, 819.67 ⁇ L, 1.00 eq) was added to the above suspension. The temperature of the reaction system was gradually raised to 40 ° C and stirred for 12 hours.
  • Step 2 N 2 blanket, compound 11-2 (3.86g, 13.05mmol, 1.00eq) , t-BuXPhOS (554.16mg, 1.31mmol, 0.10eq), Pd 2 (dba) 3 (1.20g, 1.31 mmol, 0.10eq) and KOH (1.46g, 26.10mmol, 2.00eq) in dioxane (100 mL) and H 2 O (20mL) was heated to 100 deg.] C for 2 hours.
  • Step 3 N 2 blanket, (60mL) was added to the compound at 0 °C DMF 11-3 (3.20g, 11.54mmol, 1.00eq ) of NaH (553.97mg, 13.85mmol, purity 60%, 1.20eq) and After stirring at this temperature for 20 minutes, methyl 4-bromomethylbenzoate (2.64 g, 11.54 mmol, 1.10 eq) was added to the above suspension and the temperature of the system was raised to 20 ° C and stirring was continued for 1 hour.
  • Step 1 protection of N 2, at -70 °C 4-bromo-2-chloropyridine (515.62mg, 2.68mmol, 1.00eq) in toluene (3.00 mL) was added dropwise n-BuLi (2.5M, 1.29mL After the completion of the dropwise addition, stirring was continued at this temperature for 20 minutes, and then Compound 12-1 (500.00 mg, 2.68 mmol, 1.00 eq) was added to the above suspension. The temperature of the reaction system was gradually raised to 25 ° C and stirred for 6 hours.
  • Step 2 N 2 blanket, compound 12-2 (320.00mg, 1.21mmol, 1.00eq) , t-BuXPhOS (102.76mg, 242.00 ⁇ mol, 0.20eq), Pd 2 (dba) 3 (221.60mg, 242.00 ⁇ mol
  • the reaction mixture was poured into H 2 O (100mL), and extracted with EtOAc (50mL x3), the combined extracts were washed with brine (60 mL) was swished, after dried over anhydrous Na 2 SO 4, filtered, and concentrated under reduced pressure.
  • the obtained solid was purified by preparative TLC (EtOAc /EtOAc) MS ESI calcd for C 14 H 12 FNO 2 [M + H] + 246, found 246.
  • Step 3 N 2 blanket, (5.00mL) was added to the compound NaH DMF 12-3 (290.00mg, 1.18mmol, 1.00eq ) at 0 °C (56.80mg, 1.42mmol, purity 60%, 1.20eq) After stirring at this temperature for 20 minutes, methyl 4-bromomethylbenzoate (297.33 mg, 1.30 mmol, 1.10 eq) was added to the above suspension and the temperature of the system was raised to 20 ° C and stirring was continued for 2 hours.
  • Step 4 sequentially added dropwise at 0 °C to DCM (2.00mL)
  • Compound 12-4 (150.00mg, 381.28 ⁇ mol, 1.00eq) and MeOH (2.00mL) solution of NH 2 OH.H 2 O (150.00 ⁇ L, A 50% aqueous solution) and a NaOH (2M, 150.00 L) aqueous solution were then allowed to continue stirring at 0 °C for 2 hours.
  • Most of the solvent was removed by concentration under reduced pressure, and the residue was purified by preparative HPLC (0.1% NH 4 OH) to afford compound 12-5.
  • Step 1 protection of N 2, at -70 °C 4-bromo-2-chloro-3-fluoropyridine (450.32mg, 2.14mmol, 1.00eq) in toluene (8.00 mL) was added dropwise n-BuLi (2.5 M, 1.03 mL, 1.20 eq). After the completion of the dropwise addition, stirring was continued at this temperature for 20 minutes, and then compound 13-1 (400.00 mg, 2.14 mmol, 1.00 eq) was added to the above suspension. The temperature of the reaction system was gradually raised to 25 ° C and stirred for 6 hours.
  • Step 2 N 2 blanket, compound 13-2 (440.00mg, 1.56mmol, 1.00eq) , t-BuXPhOS (132.49mg, 312.00 ⁇ mol, 0.20eq), Pd 2 (dba) 3 (285.70mg, 312.00 ⁇ mol , (10.00mL) and H 2 O (3.00mL) mixture 0.20eq) and KOH (175.06mg, 3.12mmol, 2.00eq) in dioxane was heated to 100 deg.] C for 2 hours.
  • Step 3 N 2 blanket, (5.00mL) was added to the compound NaH DMF 13-3 (380.00mg, 1.44mmol, 1.00eq ) at 0 °C (69.29mg, 1.73mmol, purity 60%, 1.20eq) After stirring at this temperature for 20 minutes, methyl 4-bromomethylbenzoate (363.74 mg, 1.59 mmol, 1.10 eq) was added to the above suspension and the temperature of the system was raised to 20 ° C and stirring was continued for 2 hours.
  • Step 4 successively added dropwise a solution of compound 13-4 (350.00mg, 850.75 ⁇ mol, 1.00eq) in DCM (3.00mL) and MeOH (3.00mL) at 0 °C NH 2 OH.H 2 O ( 3.00mL, A 50% aqueous solution) and an aqueous solution of NaOH (2M, 3.00 mL) were then stirred and stirred at 0 ° C for one hour. Most of the solvent was removed by concentration under reduced pressure, and the residue was purified by preparative HPLC (0.1%TFA) to afford compound 13-5.
  • Step 1 protection of N 2, at -70 °C solution of 4-bromo-2-chloro-5-fluoropyridine (3.00g, 14.26mmol, 1.00eq) in toluene (80.00mL) was added dropwise n-BuLi (2.5 M, 6.84 mL, 1.20 eq). After completion of the dropwise addition, stirring was continued at this temperature for 30 minutes, and then Compound 14-1 (2.86 g, 14.26 mmol, 2.34 mL, 1.00 eq) was added to the above suspension. The temperature of the reaction system was gradually raised to 40 ° C and stirred for 12 hours.
  • Step 2 N 2 blanket, compound 14-2 (300.00mg, 1.01mmol, 1.00eq) , t-BuXPhOS (85.78mg, 202.00 ⁇ mol, 0.20eq), Pd 2 (dba) 3 (184.98mg, 202.00 ⁇ mol
  • the reaction mixture was poured into H 2 O (100mL), and extracted with EtOAc (50mL x3), the combined extracts were washed with brine (60 mL) was swished, after dried over anhydrous Na 2 SO 4, filtered, and concentrated under reduced pressure.
  • the resulting solid was purified by preparative TLC (EtOAc /EtOAc) MS ESI calcd for C 15 H 13 F 2 NO 2 [M + H] + 278, found 278.
  • Step 3 N 2 blanket, (5.00mL) was added NaH (34.62mg, 865.58 ⁇ mol, purity 60%, 1.20eq) to the compound of DMF 14-3 (200.00mg, 721.32 ⁇ mol, 1.00eq ) at 0 °C And stirred at this temperature for 20 minutes, then methyl 4-bromomethylbenzoate (165.23 mg, 721.32 ⁇ mol, 1.00 eq) was added to the above suspension and the temperature of the system was raised to 20 ° C and stirring was continued for 1 hour. .
  • Step 4 successively added dropwise a solution of DCM (2.00mL) Compound 14-4 (100.00mg, 235.06 ⁇ mol, 1.00eq) and MeOH (4.00mL) at 0 °C NH 2 OH.H 2 O ( 2.00mL, A 50% aqueous solution) and an aqueous solution of NaOH (2.00 mL, 2M) were then stirred and stirred at 0 ° C for one hour. The solvent was concentrated under reduced pressure to remove most of the solvent, and the residue was cooled to 0 ° C, and pH was adjusted to 6-7 with concentrated hydrochloric acid to afford compound 14-5.
  • Step 1 Compound 15-1 (6.20 g, 64.50 mmol, 6.08 mL, 2.00 eq) and aluminum trichloride (4.73 g, 35.48 mmol, 1.10 eq) were added to a 100 ml three-necked flask at 0 °C. Further, 5-chloropentanoyl chloride (5.00 g, 32.25 mmol, 4.17 mL, 1.00 eq) was added, and the mixture was warmed to room temperature and stirred for two hours. The mixture was quenched with EtOAc (EtOAc)EtOAc.
  • EtOAc EtOAc
  • Step 3 To a solution of compound 15-3 (6.00 g, 18.28 mmol, 1.00 eq) in N,N-dimethylformamide (30.00mL), NaH (877.49mg, 21.94mmol) , 60% purity, 1.20 eq), stirred at 0-5 ° C for thirty minutes. Quenched with saturated aqueous ammonium chloride (30 mL), water (100 mL), ethyl acetate (80 mL*3), and the combined organic phase washed with water (100mL*3), saturated brine (50mL*1), sulfuric acid The sodium was dried, filtered and concentrated to give compound 15-4. MS ESI calculated for C 16 H 15 ClFNO [M+H] + 291.
  • Step 4 Under a nitrogen atmosphere, a mixed solution of water (5.00 mL) and 1,4-dioxane (10.00 mL) was added to compound 15-4 (1.00 g, 3.43 mmol, 1.00 eq), potassium hydroxide ( 384.64 mg, 6.86 mmol, 2.00 eq), tris(dibenzylideneacetone)dipalladium (627.74 mg, 686.00 ⁇ mol, 0.20 eq) and 2-di-tert-butylphosphine-2',4',6'-three different Propylbiphenyl (291.10 mg, 686.00 ⁇ mol, 0.20 eq), the mixture was heated to 90 ° C and stirred for three hours.
  • Step 5 Add sodium hydrogen (17.56 mg, 439.08 ⁇ mol) to a solution of compound 15-5 (150.00 mg, 439.08 ⁇ mol, 1.00 eq) in N,N-dimethylformamide (5.00 mL). , 60% purity, 1.00 eq.). After stirring the mixture for ten minutes, methyl 4-bromomethylbenzoate (125.72 mg, 548.85 ⁇ mol, 1.00 eq) was added and the mixture was warmed to room temperature and stirring was continued for 30 minutes. To the system was added saturated ammonium chloride (2 mL), EtOAc (EtOAc) (EtOAc) Purification afforded compound 15-6. MS ESI calcd for C 25 H 24 FNO 4 [M + H] + 421.46, found 422.2.
  • Step 6 To a solution of compound 15-6 (145.00 mg, 292.44 ⁇ mol, 1.00 eq.) in methanol (10.00 mL), EtOAc (EtOAc (EtOAc) After stirring at room temperature for one hour, the mixture was concentrated and subjected to preparative HPLC (0.225% FA) to give Compound 15-7.
  • Step 1 Slowly add n-butyllithium (2.5 M, 88.63 mL, 1.10 eq) to a solution of compound 16-1 (38.76 g, 201.43 mmol, 1.00 eq) in toluene (250.00 mL). After stirring for 30 minutes, p-fluorobenzaldehyde (25.00 g, 201.43 mmol, 21.19 mL, 1.00 eq) was slowly added dropwise to the reaction system, and the system was slowly warmed to room temperature over 30 minutes, and saturated ammonium chloride was added to the system.
  • p-fluorobenzaldehyde 25.00 g, 201.43 mmol, 21.19 mL, 1.00 eq
  • Step 4 To a mixture of compound 16-4 (100.00 mg, 336.09 ⁇ mol, 1.00 eq) in 1,4-dioxane (5.00 mL), tris(dibenzylideneacetone) dipalladium (61.55 mg, 67.22) Molmol, 0.20 eq), potassium hydroxide (40.12 mg, 715.08 ⁇ mol, 2.00 eq) and 2-di-tert-butylphosphine-2',4',6'-triisopropylbiphenyl (28.54 mg, 67.22 ⁇ mol, 0.20 eq), the system was heated to 90 ° C and stirred for 3 hours. The mixture was concentrated, EtOAc (EtOAc)EtOAc. MS ESI calcd for C 14 H 12 FNO 3 [M + H] + 261.25, found 262.2.
  • Step 5 Add sodium hydrogen (22.54 mg, 563.44 ⁇ mol) to a mixture of compound 16-5 (80.00 mg, 281.72 ⁇ mol, 1.00 eq) of N,N-dimethylformamide (10.00 mL). After 60 minutes of stirring, the system was stirred for ten minutes, then methyl 4-bromomethylbenzoate (129.07 mg, 563.44 ⁇ mol, 2.00 eq) was added and stirring was continued for thirty minutes. The mixture was quenched with EtOAc (EtOAc) (EtOAc (EtOAc) 6. MS ESI calcd for C 23 H 20 FNO 5 [M + H] + 409.41, found 410.3.
  • Step 6 To a solution of compound 16-6 (70.00 mg, 141.91 mol, 1.00 eq) in methanol (5.00 mL), EtOAc (1.OmL, 50% purity) and sodium hydroxide (17.03 mg, 425.73 ⁇ mol, 3.00 eq. The mixture was stirred at room temperature for 1 hour. The mixture was concentrated and purified by preparative HPLC (0.225% FA) to afford compound 16-7.
  • Step 1 under N 2, methyl triphenylphosphonium iodide (2.83g, 7.01mmol, 1.10eq) and t-BuOK (1M, 7.01mL, 1.10eq) in THF (20.00mL) was heated at reflux 1 hour. The suspension was then cooled to 0 ° C and compound 17-1 (1.50 g, 6.37 mmol, 1.00 eq) was added and then the obtained mixture was stirred at 20 ° C for 12 hours. The reaction mixture was poured into H 2 O (100mL), and extracted with EtOAc (50mL x3), the combined extracts were washed with brine (50mL) was swished, after dried over anhydrous Na 2 SO 4, filtered, and concentrated under reduced pressure. The obtained solid was isolated and purified by silica gel column chromatography (EtOAc/EtOAc) MS ESI calculated for C 13 H 9 ClFN [M+H] + 234, found 234.
  • Step 2 A solution of compound 17-2 (800.00 mg, 3.42 mmol, 1.00 eq) and OsO 4 (869.47 mg, 3.42 mmol, 177.44 ⁇ L, 1.00 eq) in THF (10.00 mL) and H 2 O (2.00 mL) Stir at 20 ° C for 4 hours. The reaction was then swished with 2M Na 2 SO 3 solution (80 mL), and extracted with EtOAc (30mL x3), the combined extracts were washed with brine (50mL) was swished, after dried over anhydrous Na 2 SO 4, filtered, and reduced Concentration by pressure gives compound 17-3. MS ESI calculated for C 13 H 11 ClFNO 2 [M+H] + 268, found 268.
  • Step 4 Under N 2 protection, Compound 17-4 (180.00mg, 612.83 ⁇ mol, 1.00eq) , t-BuXPhOS (26.02mg, 61.28 ⁇ mol, 0.10eq), Pd 2 (dba) (56.12mg 3, 61.28 A solution of ⁇ mol, 0.10 eq) and KOH (68.77 mg, 1.23 mmol, 2.00 eq) in dioxane (10.00 mL) and H 2 O (2.00 mL) was then warmed to 100 ° C and stirred for 1 hour.
  • Step 5 under N 2, at 0 °C DMF solution of compound 17-5 (90.00mg, 326.95 ⁇ mol, 1.00eq) of (5.00 mL) was added NaH (15.69mg, 392.34 ⁇ mol, purity 60%, 1.20eq solution ) and stirring was continued at this temperature for 20 minutes. Then, methyl 4-(bromomethyl)benzoate (74.89 mg, 326.95 ⁇ mol, 1.00 eq) was added to the above suspension, and the resulting mixture was heated to 15 ° C and stirred for 1 hour.
  • Step 1 To a solution of 1,4-butyrolactone (3.45 g, 40.11 mmol, 3.05 mL, 1.10 eq) in toluene (30.00 mL), EtOAc (1. 5 g, 43.75 mmol, purity 60%) 1.20 eq), a solution of compound 18-1 (5.00 g, 36.46 mmol, 4.39 mL, 1.00 eq) in toluene (10.00 mL) was added at 0 ° C, and the reaction was stirred at 20 ° C for 12 hours. A saturated aqueous ammonium chloride solution and water were added to the mixture and the mixture was evaporated. The organic phase was washed with brine, dried over anhydrous sodium sulfate
  • Step 2 A mixture of compound 18-2 (3.22 g, 16.84 mmol, 1.00 eq), 36% hydrochloric acid (10.20 g, 100.71 mmol, 10.00 mL, 5.98 eq) was taken to 80 ° C and stirred for 3 hours. The reaction mixture was poured into a saturated aqueous solution of potassium carbonate, and the mixture was evaporated. In the next step of the reaction.
  • Step 3 To a solution of 2-chloro-4-iodo-pyridine (730.20 mg, 3.05 mmol, 1.00 eq) in toluene (5.00 mL) was added dropwise n-butyllithium n-hexane (2.5 M, 1.34 mL). 1.10 eq), then a solution of compound 18-3 (560.00 mg, 3.05 mmol, 1.00 eq) in toluene (1.00 mL) was added, and the mixture was gradually warmed to 20 ° C, and the reaction was stirred for 2 hours. A saturated aqueous solution of ammonium chloride and water were added to the mixture, and the mixture was evaporated. MSESI calcd for C 14 H 13 ClN 2 O [M+H]+26.
  • Step 4 To a solution of compound 18-4 (750.00 mg, 2.88 mmol, 1.00 eq) in dioxane (5.00 mL) and water (1.OmL), EtOAc (32.82 mg, 5.75 mmol, 2.00 eq) , tris(dibenzylideneacetone)dipalladium (263.42 mg, 287.66 ⁇ mol, 0.10 eq), 2-di-tert-butylphosphine-2',4',6'-triisopropylbiphenyl (122.15 mg, 287.66 Molmol, 0.10 eq), stirred at 100 ° C for 4 hours.
  • Step 5 To a solution of compound 18-5 (163.00 mg, 672.80 ⁇ mol, 1.00 eq) in N,N-dimethylformamide (2.00 mL) was added sodium hydride (32.29 mg, 807.36 ⁇ mol, 60% purity, 1.20 eq), then a solution of methyl 4-bromomethylbenzoate (154.12 mg, 672.80 ⁇ mol, 1.00 eq) in N,N-dimethylformamide (1.00 mL) was added to the reaction system, 20 Stir at ° C for 2 hours. A saturated aqueous solution of ammonium chloride and water were added to the mixture, and the mixture was evaporated. MS ESI calcd for C 23 H 22 N 2 O 4 [M + H] +391.0, found 391.0.
  • Step 6 To a solution of compound 18-6 (220.00 mg, 563.48 ⁇ mol, 1.00 eq) in methanol (2.00 mL), sodium hydroxide (22.54 mg, 563.48 ⁇ mol, 1.00 eq), aqueous solution of hydroxy ammonia (1.00 mL, 50%) Aqueous solution), stirred at 20 ° C for 6 hours. After concentration of the reaction mixture, purification by reverse phase preparative HPLC gave compound 18-7.
  • Step 1 To a solution of 1,4-butyrolactone (1.00 g, 11.62 mmol, 884.96 ⁇ L, 1.00 eq) in tetrahydrofuran (5.00 mL) was added sodium hydride (1.06 g, 26.38 mmol, purity 60%) , 2.27 eq), stirring at 0 ° C for 30 minutes, then adding a solution of compound 19-1 (2.39 g, 17.43 mmol, 2.06 mL, 1.50 eq) in tetrahydrofuran (2.00 mL) at 0 ° C, stirring at 20 ° C 2 hour. A saturated aqueous ammonium chloride solution, water and ethyl acetate were added to the mixture. The aqueous phase was concentrated to dryness with acetone, filtered, and concentrated to dryness to give compound 19-2 and used directly in the next step.
  • Step 2 A mixture of compound 19-2 (1.50 g, 7.85 mmol, 1.00 eq), 36% hydrochloric acid (15.30 g, 151.11 mmol, 15.00 mL, 19.26 eq) was taken to 80 ° C and the reaction was stirred for 2 hours. The reaction mixture was poured into ice water (0°C), EtOAc (EtOAc)EtOAc.
  • Step 3 To a solution of 2-chloro-4-iodo-pyridine (782.97 mg, 3.27 mmol, 1.00 eq) in toluene (5.00 mL) was added dropwise n-butyllithium n-hexane (2.5 M, 1.44 mL). 1.10 eq), then a solution of compound 19-3 (600.00 mg, 3.27 mmol, 1.00 eq) in toluene (1.00 mL) was added, and the reaction was gradually warmed to 20 ° C, and the reaction was stirred for 12 hours. A saturated aqueous solution of ammonium chloride and water were added to the mixture, and the mixture was evaporated. MS ESI calculated for C 14 H 13 ClN 2 O [M+H]+26.
  • Step 4 To a solution of compound 19-4 (513.00 mg, 1.97 mmol, 1.00 eq) in dioxane (5.00 mL) and water (1.00 mL), EtOAc (221.07 mg, 3.94 mmol, 2.00 eq) , tris(dibenzylideneacetone)dipalladium (180.40 mg, 197.00 ⁇ mol, 0.10 eq) and 2-di-tert-butylphosphine-2',4',6'-triisopropylbiphenyl (83.65 mg, 197.00 Molmol, 0.10 eq), stirred at 100 ° C for 2 hours. Water was added to the reaction mixture, and the mixture was evaporated. After concentration, the silica gel column was purified to give the compound 19-5. MS ESI calculated for C 14 H 14 N 2 O 2 [M+H] + 243, found 243.
  • Step 5 To a solution of 19-5 (240.00 mg, 990.63 ⁇ mol, 1.00 eq) of N,N-dimethylformamide (5.00 mL), EtOAc (47.55 mg, 1. 60%, 1.20 eq), then a solution of methyl 4-bromomethylbenzoate (226.92 mg, 990.63 ⁇ mol, 1.00 eq) in N,N-dimethylformamide (1.00 mL) was added to the reaction system at 20 ° C Stir under 2 hours. Saturated aqueous ammonium chloride solution and water were added to the reaction mixture, and the mixture was evaporated. MS ESI calculated for C 23 H 22 N 2 O 4 [M+H] + 391.1, found 391.1
  • Step 6 To a solution of compound 19-6 (66.00 mg, 169.04 ⁇ mol, 1.00 eq) in methanol (5.00 mL), sodium hydroxide (13.52 mg, 338.08 ⁇ mol, 2.00 eq) and aqueous hydroxy ammonia (1.00 mL, 50%) Purity), stirred at 20 ° C for 12 hours. After the reaction mixture was concentrated, the reverse phase preparative column was purified to give compound 19-7.
  • Step 1 protection of N 2, at -70 °C solution of 1-bromo-4-iodobenzene (15.00g, 53.02mmol, 1.00eq) in THF (150.00mL) was added dropwise n-BuLi (2.5M, 23.33mL , 1.10 eq), after the completion of the dropwise addition, stirring was continued at this temperature for 30 minutes. Then, Compound 20-1 (10.64 g, 53.02 mmol, 8.72 mL, 1.00 eq) was added dropwise to the above yellow suspension. After the dropwise addition was completed, the reaction system was gradually warmed to 20 ° C and stirred for 12 hours.
  • Step 3 Compound 20-3 (100.00 mg, 349.28 ⁇ mol, 1.00 eq), HATU (159.37 mg, 419.14 ⁇ mol, 1.20 eq) and DIEA (67.71 mg, 523.92 ⁇ mol, 91.50 ⁇ L, 1.50 eq) of MeCN (10.00 mL) The solution was stirred at 20 ° C for 30 minutes, then methyl 4-(aminomethyl)benzoate (70.43 mg, 349.28 ⁇ mol, 1.00 eq, HCl salt) was added to the above solution and stirring was continued at 20 ° C for 12 hours.
  • Step 4 (2.00mL) were added dropwise to a solution of DCM (4.00mL) Compound 20-4 (150.00mg, 346.04 ⁇ mol, 1.00eq) and MeOH at 20 °C NH 2 OH.H 2 O ( 3.00mL, 50% aqueous solution) and NaOH (2M, 1.50 mL) in water. After the completion of the dropwise addition, the reaction solution was heated to reflux for 6 hours. It was then isolated and purified directly by preparative HPLC (0.1% TFA) to give compound 20-5.
  • Step 1 Separation of compound 21-1 (1 g, 2.31 mmol) by preparative SFC (chiral separation column type: OJ (250 mm * 30 mm, 5 um), eluent: A: supercritical fluid CO2B: 0.1% NH 3 H 2 O MeOH, increasing phase B from 5% to 40% in 5.5 minutes and maintaining 40% phase B for 3 minutes, finally 5% phase B for 1.5 minutes, flow rate 60 mL/min), yielding two isomers
  • the bodies 21-2 (P1) and 22-1 (P2) have retention times of 6.474 min and 7.256 min, respectively.
  • Step 1 sequentially NH 2 OH.H 2 O was added dropwise at 0 °C solution of compound 22-1 (P2) (450.00mg, 1.04mmol , 1.00eq) in DCM (4.00mL) and MeOH (4.00mL) solution of ( 4.00 mL, 50% aqueous solution) and NaOH (2M, 4.00 mL), after completion of the dropwise addition, stirring was continued at this temperature for 2 hours.
  • Step 1 Compound 23-1 (1.00 g, 3.11 mmol, 1.00 eq), tert-butoxycarbonyl ammonia (546.50 mg, 4.67 mmol, 1.50 eq), Xantphos (179.95 mg, 311.00 smol, 0.10 eq), Pd 2 ( dba) 3 (142.39mg, 155.50 ⁇ mol, 0.05eq) and Cs 2 CO 3 (1.52g, 4.67mmol , 1.50eq) in DMF (15.00mL) was heated to 100 deg.] C was stirred for 12 hours under N 2.
  • Step 2 To a solution of compound 23-2 (250.00 mg, 699.46 s, 1.00 eq) in DCM (5.00 mL) After concentration under reduced pressure, the compound 23-3 was obtained. MS ESI calcd for C 16 H 16 FNO [M+H] + 258.
  • Step 3 Compound 23-3 (250.00 mg, 673.26 ⁇ mol, 1.00 eq, TFA salt), methyl 4-(bromomethyl)benzoate (154.22 mg, 673.26 ⁇ mol, 1.00 eq) and K 2 CO 3 (139.58)
  • a solution of mg, 1.01 mmol, 1.50 eq. in DMF (5.00 mL) was stirred at 25 ° C for 6 h.
  • the reaction was then poured into H 2 O (50mL), and extracted with EtOAc (30mL x2), the combined extracts were washed with brine (30mL) was swished, after dried over anhydrous Na 2 SO 4, filtered, and concentrated under reduced pressure to obtain Compound 23-4 was used directly in the next step.
  • MS ESI calcd for C 25 H 24 FNO 3 [M+H] + 406, found 406.
  • Step 4 (2.00mL) were added dropwise to a solution of DCM (4.00mL) Compound 23-4 (250.00mg, 616.58 ⁇ mol, 1.00eq) and MeOH at 25 °C NH 2 OH.H 2 O ( 3.00mL, A 50% aqueous solution) and an aqueous solution of NaOH (2M, 1.50 mL) were then warmed to 50 ° C and stirred for 2 hours. Most of the solvent was removed by concentration under reduced pressure, and the residue was purified by preparative HPLC (0.1% NH 4 OH) to give Compound 23-5.
  • Step 1 Compound 24-1 (300.00 mg, 934.06 ⁇ mol, 1.00 eq), methyl 4-aminobenzoate (141.19 mg, 934.06 ⁇ mol, 1.00 eq), Xantphos (108.09 mg, 186.81 ⁇ mol, 0.20 eq), Cs A solution of 2 CO 3 (456.50 mg, 1.40 mmol, 1.50 eq) and Pd 2 (dba) 3 (85.53 mg, 93.41 ⁇ mol, 0.10 eq) in DMF (15.00 mL) was heated to 100 ° C under N 2 for 8 hours.
  • Step 2 To a solution of compound 24-2 (250.00 mg, 638.67 ⁇ mol, 1.00 eq) in DCM (2.00 mL) and MeOH (4.00 mL), EtOAc (EtOAc) And an aqueous solution of NaOH (2M, 2.00 mL). After the completion of the dropwise addition, the reaction solution was further stirred at 20 ° C for 12 hours. Most of the solvent was removed and concentrated under reduced pressure, and then DMSO (5mL) a clear solution, the resulting solution was separated by preparative HPLC (0.1% NH 4 OH) to give compound 24-3 with purification.
  • Step 1 Compound 25-1 (800.00 mg, 2.79 mmol, 1.00 eq), HATU (1.59 g, 4.19 mmol, 1.50 eq) and DIEA (722.26 mg, 5.59 mmol, 976.03 ⁇ L, 2.00 eq) of MeCN (5.00 mL) The solution was stirred at 20 ° C for 20 minutes, then methyl 4-aminobenzoate (422.38 mg, 2.79 mmol, 1.00 eq) was added to the above solution and stirring was continued at this temperature for 12 hours.
  • Step 2 were added dropwise a solution of compound 25-2 (1.10g, 2.62mmol, 1.00eq) in DCM (10.00 mL) and MeOH (20.00mL) at 20 °C NH 2 OH.H 2 O ( 11.00mL , 50% aqueous solution) and NaOH (2M, 10.00 mL) aqueous solution, stirring was continued at this temperature for 12 hours after the addition. Most of the solvent was removed by concentration under reduced pressure and then purified with EtOAc (5 mL).
  • Step 1 protection of N 2, at -70 °C (50.00mL) was added dropwise to the toluene compound 26-1 (5.00g, 25.98mmol, 1.00eq) of n-BuLi (2.5M, 12.47mL, 1.20eq) After the completion of the dropwise addition, stirring was continued at this temperature for 30 minutes. 4-Chloro-1-(4-fluorophenyl)butyl-1-one (5.21 g, 25.98 mmol, 4.27 mL, 1.00 eq) was then added to the above yellow suspension. The reaction system was warmed to 15 ° C and stirring was continued for 12 hours.
  • Step 2 Compound 26-2 (1.00 g, 3.60 mmol, 1.00 eq), DPPP (148.51 mg, 360.00 ⁇ mol, 0.10 eq), Pd 2 (dba) 3 (164.86 mg, 180.00 ⁇ mol, 0.05 eq) and TEA ( A mixture of 546.54 mg, 5.40 mmol, 748.68 [mu]L, 1.50 eq.
  • Step 3 A (1.00g, 3.32mmol, 1.00eq) and NaOH (265.60mg, 6.64mmol, 2.00eq) in MeOH (10.00mL) and compound 26-3 H 2 O (10.00mL) and the mixture was heated to reflux Stir for 2 hours. The MeOH was then concentrated under reduced pressure. EtOAc (EtOAc) (EtOAc) MS ESI calculated for C 16 H 14 FNO 3 [M+H] + 288, found 288.
  • Step 4 A solution of 26-4 (900.00 mg, 3.13 mmol, 1.00 eq), HATU (1.79 g, 4.70 mmol, 1.50 eq) and DIEA (1.21 g, 9.40 mmol, 3.00 eq) in MeCN (15.00 mL) Stir at °C for 30 minutes. Methyl 4-(aminomethyl)benzoate (631.71 mg, 3.13 mmol, 1.00 eq, HCl salt) was then added to the above solution and stirring was continued at 20 ° C for 3 hours. It was concentrated to dryness under reduced pressure.
  • Step 5 (4.00mL) were added dropwise to a solution of 26-5 (200.00mg, 460.34 ⁇ mol, 1.00eq) in DCM (2.00mL) and MeOH NH 2 OH.H 2 O (47.04mg , 460.34 at 0 °C A solution of ⁇ mol, 2.00 mL, 50% aqueous solution) and NaOH (2M, 230.17 ⁇ L) was added and stirred at this temperature for 3 hours. Most of the solvent was removed by concentration under reduced pressure and then purified with EtOAc (5 mL).
  • the multi-plate reader was used to detect the change of the fluorescence signal intensity in the 384-well microplate to determine the degree of deacetylation of the acetylated substrate.
  • the IC50 value of the compound inhibitor was used to evaluate the deacetylation of histones. Inhibition of the enzyme (HDAC).
  • the compound was diluted to 2 mM in DMSO and added to a 384 compound plate, 3 fold dilutions, 10 gradients, double duplicate wells using a Bravo fully automated liquid workstation; Echo liquid handlers were transferred from 384 compound plates. 0.15 ⁇ L of the compound was placed in a 384 assay plate.
  • 1.2Assay buffer Configuration A configuration with a final concentration of 20mM Hepes, 137mM NaCl, 2.7mM KCl , 1mM MgCl 2, 0.05% BSA, 0.5mM TCEP in 1x assay buffer.
  • the rapidly dissolving histone deacetylase was added to the 1x assay buffer and mixed well.
  • the 1.5x enzyme mixture was added to a 20 ⁇ L to 384-well assay plate, mixed by centrifugation, placed in a 23 ° C incubator, and the enzyme was incubated with the compound for 20 minutes.
  • the 3x substrate solution was configured with 1 x assay buffer and mixed well. 10 ⁇ L was added to a 384-well experimental plate, mixed by centrifugation, and placed in a 23 ° C incubator for 90 minutes to carry out a reaction.
  • the stop mixture of trypsin and the positive compound SAHA (Vorinostat, vorinostat) was configured with 1 x assay buffer and mixed well. The reaction was stopped by adding 30 ⁇ L to a 384-well assay plate, centrifuged, and placed in a 23 ° C incubator for 60 minutes.
  • the 384-well assay plate was placed on an Envision multi-plate reader and the fluorescence value was read.
  • the compounds of the present invention have a significant inhibitory effect on HDAC6, and have a high selectivity for the weak inhibition of HDAC1.
  • the Beagle's pharmacopoeetic characteristics after oral administration of the compound were tested in a standard protocol.
  • the candidate compound was formulated into a uniform suspension and administered to a beagle dog for single oral administration.
  • Oral vehicle is a certain proportion of polyethylene glycol-polyoxyethylene castor oil aqueous solution. Collect whole blood samples within 24 hours, centrifuge at 3000g for 10 minutes, separate the supernatant to obtain plasma samples, take 30 ⁇ L of plasma samples, add 300 ⁇ L of internal standard acetonitrile solution to precipitate protein, vortex centrifuge to take 30 ⁇ L of supernatant and add equal volume of water. The supernatant was sampled by vortexing, and the plasma concentration was quantitatively analyzed by LC-MS/MS analysis, and the pharmacokinetic parameters such as peak concentration, peak time, half-life, and area under the curve of the drug were calculated.
  • the compounds of the present invention have a good single or partial index of canine pharmacokinetics.
  • HDAC inhibitors are widely used in a variety of cancers and can be combined with a variety of drugs to enhance the therapeutic effect of the drug. It is a well-recognized anti-tumor target. HDAC inhibitors are widely used in a variety of cancers, and can be combined with a variety of drugs to enhance the therapeutic effect of the drug, such as the combination of panobinostat and the proteasome inhibitor bortezomib (bortezomib) can increase the efficacy of multiple myeloma and significantly reduce toxicity .
  • Human myeloma MM.1S cells were cultured in vitro in a single layer, cultured in RPMI-1640 medium (supplier: Gibco; Cat. No. 22400-089) supplemented with 10% fetal bovine serum, and cultured at 37 ° C in 5% CO 2 incubator. Passage was routinely digested with trypsin-EDTA twice a week. When the cell saturation is 80%-90%, the cells are collected, counted, and inoculated.
  • the drug should be thoroughly mixed before administration to the animal.
  • the experimental indicator is to investigate whether tumor growth is inhibited, delayed or cured.
  • Tumor diameters were measured with vernier calipers three times a week.
  • TGI (%) ((1 - (the average tumor volume at the end of a treatment group - the average tumor volume at the beginning of the treatment group)) / (the average tumor at the end of the treatment of the solvent control group) Volume-solvent control group average tumor volume at the start of treatment)) x 100%.
  • T weight and C weight indicate the tumor weights of the drug-administered group and the vehicle control group, respectively.
  • Statistical analysis included mean and standard error (SEM) of tumor volume at each time point for each group.
  • the treatment group showed the best therapeutic effect on the 21st day after the administration at the end of the trial, and therefore statistical analysis was performed based on this data to evaluate the difference between the groups.
  • T-test was used for comparison between the two groups, and one-way ANOVA was used for comparison between three groups or groups. If there was a significant difference in F values, the Games-Howell method was used for the test. If there is no significant difference in F values, the Dunnet (2-sided) method is used for analysis. All data analysis was performed with SPSS 17.0. A significant difference was considered at p ⁇ 0.05.
  • the tumor volume was 218 mm 3 .
  • mice showed good tolerance and no significant weight loss.

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Abstract

一类作为组蛋白去乙酰化酶6（HDAC6）选择性抑制剂的化合物，及其在制备治疗HDAC6相关疾病的药物中的应用。具体公开了式（I）所示化合物及其药学上可接受的盐。

Description

HDAC6选择性抑制剂及其制备方法和应用
相关申请的交叉引用
本申请主张2017.01.10提交的中国专利申请CN201710017287.2的优先权，其内容在此并入本申请。
技术领域
本发明涉及了一类作为组蛋白去乙酰化酶6(HDAC6)选择性抑制剂的化合物，及其在制备治疗HDAC6相关疾病的药物的应用。具体涉及式(Ⅰ)所示化合物及其药学上可接受的盐。
背景技术
WHO专家预测，2020年全球人口将达80亿，癌症发病将达到2000万人，死亡将达到1200万人。癌症将成为新世纪人类的第一杀手，对人类生存构成最严重的威胁。处于工业化进程中的中国，居美国之后成为世界第二癌症高发国，其癌症发病率及死亡率呈现明显上升趋势。城市癌症占全死因的第一位，农村癌症占全死因的第二位。随着我国癌症发病率和死亡率的快速升高，全国每年用于癌症的医疗费用已超过1,500亿元人民币。
HDAC抑制剂广泛应用于多种癌症，并能够与多种药物联合而增强该药物的治疗效果，是一个被充分证实的抗肿瘤靶点。在细胞核内组蛋白去乙酰化酶(histone deacetylase,HDAC)和组蛋白乙酰化转移酶(histone acetyl trans-ferase,HAT)共同调控基因的转录。在癌细胞中,HDAC的过度表达导致去乙酰化作用的增强，从而增加DNA与组蛋白之间的引力,使核小体变得十分紧密,不利于肿瘤抑制基因的表达。抑制剂(HDACi)则可通过提高组蛋白乙酰化，从而调控细胞凋亡及分化相关蛋白的表达，诱导细胞凋亡及分化，成为一类新的抗肿瘤药物。不仅如此，HDAC还参与了很多代谢性疾病的调控，如阿尔兹海默症(Alzheimer),帕金森症(Parkinson)等疾病，HDACi抑制剂在动物和人体试验中都展示了良好的效果。
在总共有18种去乙酰化酶亚型中，HDAC6是唯一在细胞质中的去乙酰化酶亚型，而其他17种HDAC都是存在于细胞核中。HDAC6不直接催化组蛋白，而是以微管蛋白(tubulin)和热休克蛋白(Hsp90)作为底物,通过他们调节细胞贩运，粘附和运动(即不基因调控).因此，相信它将对基因相关的生理功能产生更少的作用，从而副作用更小。目前的临床实验结果已经证实HDAC6选择性抑制剂安全有效(POC)。第一个HDAC6选择性抑制剂ACY-1215(Acetylon)的临床研究证明，选择性HDAC6抑制剂有更好的安全性，因此有更好的商业前景。
Figure PCTCN2018072088-appb-000001
蛋白酶体抑制剂(Drug Design,Development and Therapy 2016:10 217–226)可以阻断大量调节蛋白的降解，引起细胞内信号系统的紊乱和超负荷，导致细胞生长的抑制，最终使肿瘤进展过程延缓，甚至停止。HDAC抑制剂广泛应用于多种癌症，并能够与多种药物联合而增强该药物的治疗效果，如HDAC抑制剂panobinostat与蛋白酶体抑制剂硼替佐米(bortezomib)合用能增加多发性骨髓瘤疗效并明显降低毒性。
发明内容
本发明提供了式(Ⅰ)所示化合物、药学上可接受的盐及其异构体：
Figure PCTCN2018072088-appb-000002
Figure PCTCN2018072088-appb-000003
选自：单键或双键；
n选自：0或1；
T 1、T 2分别独立地选自：CH、CH 2、-C(＝O)-、N；
T 3选自：C或N；
Z 1、Z 2、Z 3分别独立地选自：CH或N；
L 1选自：单键、-NH-、-C(＝O)-NH-；
R 1选自任选被1、2或3个R取代的：C 1-3烷基、苯基、6元杂芳基；
R 2选自：H、F、Cl、Br、I；
A环选自：4～7元杂环烷基；
R选自：F、Cl、Br、I；
所述6元杂芳基、4～7元杂环烷基之“杂”分别独立地选自：-NH-、N、-O-；
以上任何一种情况下，杂原子或杂原子团的数目分别独立地选自1、2或3。
本发明的一些方案中，上述R 1选自任选被1、2或3个R取代的：甲基、乙基、异丙基、苯基、吡啶基。
本发明的一些方案中，上述R 1选自任选被1、2或3个R取代的：CH 3、
Figure PCTCN2018072088-appb-000004
Figure PCTCN2018072088-appb-000005
本发明的一些方案中，上述R 1选自：CH 3、
Figure PCTCN2018072088-appb-000006
Figure PCTCN2018072088-appb-000007
本发明的一些方案中，上述A环选自：恶丁环基、四氢呋喃基、四氢吡喃基、1,3-二氧戊环基、1,4-二氧杂环庚烷基、1,4-二噁烷基、1,4-氧氮杂环庚烷基、吗啉基。
本发明的一些方案中，上述A环选自：
Figure PCTCN2018072088-appb-000008
Figure PCTCN2018072088-appb-000009
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000010
选自：
Figure PCTCN2018072088-appb-000011
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000012
选自：
Figure PCTCN2018072088-appb-000013
Figure PCTCN2018072088-appb-000014
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000015
选自：
Figure PCTCN2018072088-appb-000016
Figure PCTCN2018072088-appb-000017
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000018
选自：
Figure PCTCN2018072088-appb-000019
Figure PCTCN2018072088-appb-000020
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000021
选自：
Figure PCTCN2018072088-appb-000022
Figure PCTCN2018072088-appb-000023
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000024
选自：
Figure PCTCN2018072088-appb-000025
Figure PCTCN2018072088-appb-000026
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000027
选自：
Figure PCTCN2018072088-appb-000028
Figure PCTCN2018072088-appb-000029
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000030
选自：
Figure PCTCN2018072088-appb-000031
Figure PCTCN2018072088-appb-000032
Figure PCTCN2018072088-appb-000033
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000034
选自：
Figure PCTCN2018072088-appb-000035
Figure PCTCN2018072088-appb-000036
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000037
选自：-CH 2-、-NH-、-C(＝O)-NH-、
Figure PCTCN2018072088-appb-000038
Figure PCTCN2018072088-appb-000039
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000040
选自：
Figure PCTCN2018072088-appb-000041
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000042
选自：
Figure PCTCN2018072088-appb-000043
Figure PCTCN2018072088-appb-000044
本发明的一些方案中，上述R 1选自任选被1、2或3个R取代的：甲基、乙基、苯基、吡啶基，其他变量如本发明所定义。
本发明的一些方案中，上述R 1选自任选被1、2或3个R取代的：CH 3、
Figure PCTCN2018072088-appb-000045
Figure PCTCN2018072088-appb-000046
其他变量如本发明所定义。
本发明的一些方案中，上述R 1选自：CH 3、
Figure PCTCN2018072088-appb-000047
Figure PCTCN2018072088-appb-000048
其他变量如本发明所定义。
本发明的一些方案中，上述A环选自：恶丁环基、四氢呋喃基、四氢吡喃基、1,3-二氧戊环基、1,4-二氧杂环庚烷基、1,4-二噁烷基、1,4-氧氮杂环庚烷基、吗啉基，其他变量如本发明所定义。
本发明的一些方案中，上述A环选自：
Figure PCTCN2018072088-appb-000049
Figure PCTCN2018072088-appb-000050
其他变量如本发明所定义。
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000051
选自：
Figure PCTCN2018072088-appb-000052
其他变量如本发明所定义。
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000053
选自：
Figure PCTCN2018072088-appb-000054
Figure PCTCN2018072088-appb-000055
其他变量如本发明所定义。
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000056
选自：
Figure PCTCN2018072088-appb-000057
Figure PCTCN2018072088-appb-000058
其他变量如本发明所定义。
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000059
选自：
Figure PCTCN2018072088-appb-000060
Figure PCTCN2018072088-appb-000061
其他变量如本发明所定义。
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000062
选自：
Figure PCTCN2018072088-appb-000063
Figure PCTCN2018072088-appb-000064
Figure PCTCN2018072088-appb-000065
其他变量如本发明所定义。
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000066
选自：
Figure PCTCN2018072088-appb-000067
Figure PCTCN2018072088-appb-000068
Figure PCTCN2018072088-appb-000069
其他变量如本发明所定义。
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000070
选自：
Figure PCTCN2018072088-appb-000071
Figure PCTCN2018072088-appb-000072
Figure PCTCN2018072088-appb-000073
其他变量如本发 明所定义。
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000074
选自：
Figure PCTCN2018072088-appb-000075
Figure PCTCN2018072088-appb-000076
Figure PCTCN2018072088-appb-000077
其他变量如本发明所定义。
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000078
选自：
Figure PCTCN2018072088-appb-000079
Figure PCTCN2018072088-appb-000080
Figure PCTCN2018072088-appb-000081
Figure PCTCN2018072088-appb-000082
其他变量如本发明所定义。
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000083
选自：-CH 2-、-NH-、-C(＝O)-NH-、
Figure PCTCN2018072088-appb-000084
Figure PCTCN2018072088-appb-000085
其他变量如本发明所定义。
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000086
选自：
Figure PCTCN2018072088-appb-000087
其他变量如本发明所定义。
本发明的一些方案中，上述结构单元
Figure PCTCN2018072088-appb-000088
选自：
Figure PCTCN2018072088-appb-000089
Figure PCTCN2018072088-appb-000090
其他变量如本发明所定义。
本发明还有一些方案是由上述各变量任意组合而来。
本发明的一些方案中，上述的化合物、药学上可接受的盐及其异构体，其选自：
Figure PCTCN2018072088-appb-000091
Figure PCTCN2018072088-appb-000092
其中，A环、R 1、R 2、L 1和n如本发明所定义。
本发明的一些方案中，上述的化合物、药学上可接受的盐及其异构体，其选自：
Figure PCTCN2018072088-appb-000093
其中，
E 1、E 2分别独立地选自-O-、-CH 2-和-CH 2-CH2-；
R、R 2、L 1和n如本发明所定义。
本发明还提供了下列化合物及其药学上可接受的盐，选自：
Figure PCTCN2018072088-appb-000094
本发明的一些方案中，上述的化合物、药学上可接受的盐及其异构体，其选自：
Figure PCTCN2018072088-appb-000095
本发明还提供了一种药物组合物，包括治疗有效量的上述化合物或其药学上可接受的盐作为活性成分以及药学上可接受的载体。
本发明还提供了述的化合物或其药学上可接受的盐在制备治疗HDAC6相关病症的药物上的应用。
本发明还提供了上述组合物在制备治疗HDAC6相关病症的药物上的应用。
本发明的一些方案中，上述的应用，其特征在于，所述药物是治疗多发性骨髓瘤的药物。
技术效果
作为新型的组蛋白去乙酰化酶6(HDAC6)选择性抑制剂，本发明的化合物具有显著的体外活性，对HDAC6酶的抑制作用显著，而同时对HDAC1弱抑制，具有选择性高的特点；另外，与Ixazomib联合给药增加多发性骨髓瘤疗效并明显降低毒性。
定义和说明
除非另有说明，本文所用的下列术语和短语旨在具有下列含义。一个特定的术语或短语在没有特别定义的情况下不应该被认为是不确定的或不清楚的，而应该按照普通的含义去理解。当本文中出现商品名时，意在指代其对应的商品或其活性成分。这里所采用的术语“药学上可接受的”，是针对那些化合物、材料、组合物和/或剂型而言，它们在可靠的医学判断的范围之内，适用于与人类和动物的组织接触使用，而没有过多的毒性、刺激性、过敏性反应或其它问题或并发症，与合理的利益/风险比相称。
术语“药学上可接受的盐”是指本发明化合物的盐，由本发明发现的具有特定取代基的化合物与相对无毒的酸或碱制备。当本发明的化合物中含有相对酸性的功能团时，可以通过在纯的溶液或合适的惰性溶剂中用足够量的碱与这类化合物的中性形式接触的方式获得碱加成盐。药学上可接受的碱加成盐包括钠、钾、钙、铵、有机氨或镁盐或类似的盐。当本发明的化合物中含有相对碱性的官能团时，可以通过在纯的溶液或合适的惰性溶剂中用足够量的酸与这类化合物的中性形式接触的方式获得酸加成盐。药学上可接受的酸加成盐的实例包括无机酸盐，所述无机酸包括例如盐酸、氢溴酸、硝酸、碳酸，碳酸氢根，磷酸、磷酸一氢根、磷酸二氢根、硫酸、硫酸氢根、氢碘酸、亚磷酸等；以及有机酸盐，所述 有机酸包括如乙酸、丙酸、异丁酸、马来酸、丙二酸、苯甲酸、琥珀酸、辛二酸、反丁烯二酸、乳酸、扁桃酸、邻苯二甲酸、苯磺酸、对甲苯磺酸、柠檬酸、酒石酸和甲磺酸等类似的酸；还包括氨基酸(如精氨酸等)的盐，以及如葡糖醛酸等有机酸的盐(参见Berge et al.,"Pharmaceutical Salts",Journal of Pharmaceutical Science 66:1-19(1977))。本发明的某些特定的化合物含有碱性和酸性的官能团，从而可以被转换成任一碱或酸加成盐。
优选地，以常规方式使盐与碱或酸接触，再分离母体化合物，由此再生化合物的中性形式。化合物的母体形式与其各种盐的形式的不同之处在于某些物理性质，例如在极性溶剂中的溶解度不同。
本文所用的“药学上可接受的盐”属于本发明化合物的衍生物，其中，通过与酸成盐或与碱成盐的方式修饰所述母体化合物。药学上可接受的盐的实例包括但不限于：碱基比如胺的无机酸或有机酸盐、酸根比如羧酸的碱金属或有机盐等等。药学上可接受的盐包括常规的无毒性的盐或母体化合物的季铵盐，例如无毒的无机酸或有机酸所形成的盐。常规的无毒性的盐包括但不限于那些衍生自无机酸和有机酸的盐，所述的无机酸或有机酸选自2-乙酰氧基苯甲酸、2-羟基乙磺酸、乙酸、抗坏血酸、苯磺酸、苯甲酸、碳酸氢根、碳酸、柠檬酸、依地酸、乙烷二磺酸、乙烷磺酸、富马酸、葡庚糖、葡糖酸、谷氨酸、乙醇酸、氢溴酸、盐酸、氢碘酸盐、羟基、羟萘、羟乙磺酸、乳酸、乳糖、十二烷基磺酸、马来酸、苹果酸、扁桃酸、甲烷磺酸、硝酸、草酸、双羟萘酸、泛酸、苯乙酸、磷酸、多聚半乳糖醛、丙酸、水杨酸、硬脂酸、亚乙酸、琥珀酸、氨基磺酸、对氨基苯磺酸、硫酸、单宁、酒石酸和对甲苯磺酸。
本发明的药学上可接受的盐可由含有酸根或碱基的母体化合物通过常规化学方法合成。一般情况下，这样的盐的制备方法是：在水或有机溶剂或两者的混合物中，经由游离酸或碱形式的这些化合物与化学计量的适当的碱或酸反应来制备。一般地，优选醚、乙酸乙酯、乙醇、异丙醇或乙腈等非水介质。
除了盐的形式，本发明所提供的化合物还存在前药形式。本文所描述的化合物的前药容易地在生理条件下发生化学变化从而转化成本发明的化合物。此外，前体药物可以在体内环境中通过化学或生化方法被转换到本发明的化合物。
本发明的某些化合物可以以非溶剂化形式或者溶剂化形式存在，包括水合物形式。一般而言，溶剂化形式与非溶剂化的形式相当，都包含在本发明的范围之内。
本发明的某些化合物可以具有不对称碳原子(光学中心)或双键。外消旋体、非对映异构体、几何异构体和单个的异构体都包括在本发明的范围之内。
除非另有说明，用楔形键和虚线键
Figure PCTCN2018072088-appb-000096
表示一个立体中心的绝对构型，用波浪线
Figure PCTCN2018072088-appb-000097
表示楔形键或虚线键
Figure PCTCN2018072088-appb-000098
或
Figure PCTCN2018072088-appb-000099
用
Figure PCTCN2018072088-appb-000100
表示立体中心的相对构型。当本文所述化合物含有烯属双键或其它几何不对称中心，除非另有规定，它们包括E、Z几何异构体。同样地，所有的互变异构形式均包括在本发明的范围之内。
本发明的化合物可以存在特定的几何或立体异构体形式。本发明设想所有的这类化合物，包括顺式和反式异构体、(-)-和(+)-对对映体、(R)-和(S)-对映体、非对映异构体、(D)-异构体、(L)-异构体，及其外消旋混合物和其他混合物，例如对映异构体或非对映体富集的混合物，所有这些混合物都属于本发明的范围之内。烷基等取代基中可存在另外的不对称碳原子。所有这些异构体以及它们的混合物，均包括在本发明的范围之内。
可以通过的手性合成或手性试剂或者其他常规技术制备光学活性的(R)-和(S)-异构体以及D和L异构体。如果想得到本发明某化合物的一种对映体，可以通过不对称合成或者具有手性助剂的衍生作用来制备，其中将所得非对映体混合物分离，并且辅助基团裂开以提供纯的所需对映异构体。或者，当分子中含有碱性官能团(如氨基)或酸性官能团(如羧基)时，与适当的光学活性的酸或碱形成非对映异构体的盐，然后通过本领域所公知的常规方法进行非对映异构体拆分，然后回收得到纯的对映体。此外，对映异构体和非对映异构体的分离通常是通过使用色谱法完成的，所述色谱法采用手性固定相，并任选地与化学衍生法相结合(例如由胺生成氨基甲酸盐)。
本发明的化合物可以在一个或多个构成该化合物的原子上包含非天然比例的原子同位素。例如，可用放射性同位素标记化合物，比如氚( 3H)，碘-125( 125I)或C-14( 14C)。本发明的化合物的所有同位素组成的变换，无论放射性与否，都包括在本发明的范围之内。“任选”或“任选地”指的是随后描述的事件或状况可能但不是必需出现的，并且该描述包括其中所述事件或状况发生的情况以及所述事件或状况不发生的情况。
术语“被取代的”是指特定原子上的任意一个或多个氢原子被取代基取代，可以包括重氢和氢的变体，只要特定原子的价态是正常的并且取代后的化合物是稳定的。当取代基为酮基(即＝O)时，意味着两个氢原子被取代。酮取代不会发生在芳香基上。术语“任选被取代的”是指可以被取代，也可以不被取代，除非另有规定，取代基的种类和数目在化学上可以实现的基础上可以是任意的。
当任何变量(例如R)在化合物的组成或结构中出现一次以上时，其在每一种情况下的定义都是独立的。因此，例如，如果一个基团被0-2个R所取代，则所述基团可以任选地至多被两个R所取代，并且每种情况下的R都有独立的选项。此外，取代基和/或其变体的组合只有在这样的组合会产生稳定的化合物的情况下才是被允许的。
当一个连接基团的数量为0时，比如-(CRR) 0-，表示该连接基团为单键。
当其中一个变量选自单键时，表示其连接的两个基团直接相连，比如A-L-Z中L代表单键时表示该结构实际上是A-Z。
当一个取代基为空缺时，表示该取代基是不存在的，比如A-X中X为空缺时表示该结构实际上是A。当一个取代基的键可以交叉连接到一个环上的两个原子时，这种取代基可以与这个环上的任意原子相键合。当所列举的取代基中没有指明其通过哪一个原子连接到化学结构通式中包括但未具体提及的 化合物时，这种取代基可以通过其任何原子相键合。取代基和/或其变体的组合只有在这样的组合会产生稳定的化合物的情况下才是被允许的。例如，结构单元
Figure PCTCN2018072088-appb-000101
或
Figure PCTCN2018072088-appb-000102
表示其可在环己基或者环己二烯上的任意一个位置发生取代。
除非另有规定，术语“杂”表示杂原子或杂原子团(即含有杂原子的原子团)，包括碳(C)和氢(H)以外的原子以及含有这些杂原子的原子团，例如包括氧(O)、氮(N)、硫(S)、硅(Si)、锗(Ge)、铝(Al)、硼(B)、-O-、-S-、＝O、＝S、-C(＝O)O-、-C(＝O)-、-C(＝S)-、-S(＝O)、-S(＝O) 2-，以及任选被取代的-C(＝O)N(H)-、-N(H)-、-C(＝NH)-、-S(＝O) 2N(H)-或-S(＝O)N(H)-。
除非另有规定，“环”表示被取代或未被取代的环烷基、杂环烷基、环烯基、杂环烯基、环炔基、杂环炔基、芳基或杂芳基。所谓的环包括单环、联环、螺环、并环或桥环。环上原子的数目通常被定义为环的元数，例如，“5～7元环”是指环绕排列5～7个原子。除非另有规定，该环任选地包含1～3个杂原子。因此，“5～7元环”包括例如苯基、吡啶和哌啶基；另一方面，术语“5～7元杂环烷基环”包括吡啶基和哌啶基，但不包括苯基。术语“环”还包括含有至少一个环的环系，其中的每一个“环”均独立地符合上述定义。
除非另有规定，术语“杂环”或“杂环基”意指稳定的含杂原子或杂原子团的单环、双环或三环，它们可以是饱和的、部分不饱和的或不饱和的(芳族的)，它们包含碳原子和1、2、3或4个独立地选自N、O和S的环杂原子，其中上述任意杂环可以稠合到一个苯环上形成双环。氮和硫杂原子可任选被氧化(即NO和S(O)p，p是1或2)。氮原子可以是被取代的或未取代的(即N或NR，其中R是H或本文已经定义过的其他取代基)。该杂环可以附着到任何杂原子或碳原子的侧基上从而形成稳定的结构。如果产生的化合物是稳定的，本文所述的杂环可以发生碳位或氮位上的取代。杂环中的氮原子任选地被季铵化。一个优选方案是，当杂环中S及O原子的总数超过1时，这些杂原子彼此不相邻。另一个优选方案是，杂环中S及O原子的总数不超过1。如本文所用，术语“芳族杂环基团”或“杂芳基”意指稳定的5、6、7元单环或双环或7、8、9或10元双环杂环基的芳香环，它包含碳原子和1、2、3或4个独立地选自N、O和S的环杂原子。氮原子可以是被取代的或未取代的(即N或NR，其中R是H或本文已经定义过的其他取代基)。氮和硫杂原子可任选被氧化(即NO和S(O)p，p是1或2)。值得注意的是，芳香杂环上S和O原子的总数不超过1。桥环也包含在杂环的定义中。当一个或多个原子(即C、O、N或S)连接两个不相邻的碳原子或氮原子时形成桥环。优选的桥环包括但不限于：一个碳原子、两个碳原子、一个氮原子、两个氮原子和一个碳-氮基。值得注意的是，一个桥总是将单环转换成三环。桥环中，环上的取代基也可以出现在桥上。
杂环化合物的实例包括但不限于：吖啶基、吖辛因基、苯并咪唑基、苯并呋喃基、苯并巯基呋喃基、苯并巯基苯基、苯并恶唑基、苯并恶唑啉基、苯并噻唑基、苯并三唑基、苯并四唑基、苯并异恶唑基、 苯并异噻唑基、苯并咪唑啉基、咔唑基、4aH-咔唑基、咔啉基、苯并二氢吡喃基、色烯、噌啉基十氢喹啉基、2H,6H-1,5,2-二噻嗪基、二氢呋喃并[2,3-b]四氢呋喃基、呋喃基、呋咱基、咪唑烷基、咪唑啉基、咪唑基、1H-吲唑基、吲哚烯基、二氢吲哚基、中氮茚基、吲哚基、3H-吲哚基、异苯并呋喃基、异吲哚基、异二氢吲哚基、异喹啉基、异噻唑基、异恶唑基、亚甲二氧基苯基、吗啉基、萘啶基，八氢异喹啉基、恶二唑基、1,2,3-恶二唑基、1,2,4-恶二唑基、1,2,5-恶二唑基、1,3,4-恶二唑基、恶唑烷基、恶唑基、羟吲哚基、嘧啶基、菲啶基、菲咯啉基、吩嗪、吩噻嗪、苯并黄嘌呤基、酚恶嗪基、酞嗪基、哌嗪基、哌啶基、哌啶酮基、4-哌啶酮基、胡椒基、蝶啶基、嘌呤基、吡喃基、吡嗪基、吡唑烷基、吡唑啉基、吡唑基、哒嗪基、吡啶并恶唑、吡啶并咪唑、吡啶并噻唑、吡啶基、吡咯烷基、吡咯啉基、2H-吡咯基、吡咯基、喹唑啉基、喹啉基、4H-喹嗪基、喹喔啉基、奎宁环基、四氢呋喃基、四氢异喹啉基、四氢喹啉基、四唑基，6H-1,2,5-噻二嗪基、1,2,3-噻二唑基、1,2,4-噻二唑基、1,2,5-噻二唑基、1,3,4-噻二唑基、噻蒽基、噻唑基、异噻唑基噻吩基、噻吩并恶唑基、噻吩并噻唑基、噻吩并咪唑基、噻吩基、三嗪基、1,2,3-三唑基、1,2,4-三唑基、1,2,5-三唑基、1,3,4-三唑基和呫吨基。还包括稠环和螺环化合物。
除非另有规定，术语“烃基”或者其下位概念(比如烷基、烯基、炔基、芳基等等)本身或者作为另一取代基的一部分表示直链的、支链的或环状的烃原子团或其组合，可以是完全饱和的(如烷基)、单元或多元不饱和的(如烯基、炔基、芳基)，可以是单取代或多取代的，可以是一价(如甲基)、二价(如亚甲基)或者多价(如次甲基)，可以包括二价或多价原子团，具有指定数量的碳原子(如C 1-C 12表示1至12个碳，C 1-12选自C 1、C 2、C 3、C 4、C 5、C 6、C 7、C 8、C 9、C 10、C 11和C 12；C 3-12选自C 3、C 4、C 5、C 6、C 7、C 8、C 9、C 10、C 11和C 12。)。“烃基”包括但不限于脂肪烃基和芳香烃基，所述脂肪烃基包括链状和环状，具体包括但不限于烷基、烯基、炔基，所述芳香烃基包括但不限于6-12元的芳香烃基，例如苯、萘等。在一些实施例中，术语“烃基”表示直链的或支链的原子团或它们的组合，可以是完全饱和的、单元或多元不饱和的，可以包括二价和多价原子团。饱和烃原子团的实例包括但不限于甲基、乙基、正丙基、异丙基、正丁基、叔丁基、异丁基、仲丁基、异丁基、环己基、(环己基)甲基、环丙基甲基，以及正戊基、正己基、正庚基、正辛基等原子团的同系物或异构体。不饱和烃基具有一个或多个双键或三键，其实例包括但不限于乙烯基、2-丙烯基、丁烯基、巴豆基、2-异戊烯基、2-(丁二烯基)、2，4-戊二烯基、3-(1，4-戊二烯基)、乙炔基、1-和3-丙炔基，3-丁炔基，以及更高级的同系物和异构体。
除非另有规定，术语“杂烃基”或者其下位概念(比如杂烷基、杂烯基、杂炔基、杂芳基等等)本身或者与另一术语联合表示稳定的直链的、支链的或环状的烃原子团或其组合，有一定数目的碳原子和至少一个杂原子组成。在一些实施例中，术语“杂烷基”本身或者与另一术语联合表示稳定的直链的、支链的烃原子团或其组合物，有一定数目的碳原子和至少一个杂原子组成。在一个典型实施例中，杂原子选自B、O、N和S，其中氮和硫原子任选地被氧化，氮杂原子任选地被季铵化。杂原子或杂原子团可以 位于杂烃基的任何内部位置，包括该烃基附着于分子其余部分的位置，但术语“烷氧基”、“烷氨基”和“烷硫基”(或硫代烷氧基)属于惯用表达，是指分别通过一个氧原子、氨基或硫原子连接到分子的其余部分的那些烷基基团。实例包括但不限于-CH 2-CH 2-O-CH 3、-CH 2-CH 2-NH-CH 3、-CH 2-CH 2-N(CH 3)-CH 3、-CH 2-S-CH 2-CH 3、-CH 2-CH 2、-S(O)-CH 3、-CH 2-CH 2-S(O) 2-CH 3、-CH＝CH-O-CH 3、-CH 2-CH＝N-OCH 3和–CH＝CH-N(CH 3)-CH 3。至多两个杂原子可以是连续的，例如-CH 2-NH-OCH 3。
除非另有规定，术语“环烃基”、“杂环烃基”或者其下位概念(比如芳基、杂芳基、环烷基、杂环烷基、环烯基、杂环烯基、环炔基、杂环炔基等等)本身或与其他术语联合分别表示环化的“烃基”、“杂烃基”。此外，就杂烃基或杂环烃基(比如杂烷基、杂环烷基)而言，杂原子可以占据该杂环附着于分子其余部分的位置。环烃基的实例包括但不限于环戊基、环己基、1-环己烯基、3-环己烯基、环庚基等。杂环基的非限制性实例包括1-(1,2,5,6-四氢吡啶基)、1-哌啶基、2-哌啶基，3-哌啶基、4-吗啉基、3-吗啉基、四氢呋喃-2-基、四氢呋喃吲哚-3-基、四氢噻吩-2-基、四氢噻吩-3-基，1-哌嗪基和2-哌嗪基。
除非另有规定，术语“烷基”用于表示直链或支链的饱和烃基,可以是单取代(如-CH 2F)或多取代的(如-CF 3)，可以是一价(如甲基)、二价(如亚甲基)或者多价(如次甲基)。烷基的例子包括甲基(Me)，乙基(Et)，丙基(如，n-丙基和异丙基)，丁基(如，n-丁基，异丁基，s-丁基，t-丁基)，戊基(如，n-戊基，异戊基，新戊基)等。
除非另有规定，“烯基”指在链的任何位点上具有一个或多个碳碳双键的烷基,可以是单取代或多取代的，可以是一价、二价或者多价。烯基的例子包括乙烯基，丙烯基，丁烯基，戊烯基，己烯基，丁间二烯基，戊间二烯基，己间二烯基等。
除非另有规定，“炔基”指在链的任何位点上具有一个或多个碳碳三键的烷基，可以是单取代或多取代的，可以是一价、二价或者多价。炔基的例子包括乙炔基，丙炔基，丁炔基，戊炔基等。
除非另有规定，环烷基包括任何稳定的环状或多环烃基，任何碳原子都是饱和的，可以是单取代或多取代的，可以是一价、二价或者多价。这些环烷基的实例包括，但不限于，环丙基、降冰片烷基、[2.2.2]二环辛烷、[4.4.0]二环癸烷等。
除非另有规定，环烯基包括任何稳定的环状或多环烃基，该烃基在环的任何位点含有一个或多个不饱和的碳-碳双键，可以是单取代或多取代的，可以是一价、二价或者多价。这些环烯基的实例包括，但不限于，环戊烯基、环己烯基等。
除非另有规定，环炔基包括任何稳定的环状或多环烃基，该烃基在环的任何位点含有一个或多个碳-碳三键，可以是单取代或多取代的，可以是一价、二价或者多价。
除非另有规定，术语“卤代素”或“卤素”本身或作为另一取代基的一部分表示氟、氯、溴或碘原子。此外，术语“卤代烷基”意在包括单卤代烷基和多卤代烷基。例如，术语“卤代(C 1-C 4)烷基”意在包括但不仅限于三氟甲基、2，2，2-三氟乙基、4-氯丁基和3-溴丙基等等。除非另有规定，卤代烷基的实例包 括但不仅限于：三氟甲基、三氯甲基、五氟乙基，和五氯乙基。
“烷氧基”代表通过氧桥连接的具有特定数目碳原子的上述烷基，除非另有规定，C 1-6烷氧基包括C 1、C 2、C 3、C 4、C 5和C 6的烷氧基。烷氧基的例子包括但不限于：甲氧基、乙氧基、正丙氧基、异丙氧基、正丁氧基、仲丁氧基、叔丁氧基、正戊氧基和S-戊氧基。
除非另有规定，术语“芳基”表示多不饱和的芳族烃取代基，可以是单取代或多取代的，可以是一价、二价或者多价，它可以是单环或多环(比如1至3个环；其中至少一个环是芳族的)，它们稠合在一起或共价连接。术语“杂芳基”是指含有一至四个杂原子的芳基(或环)。在一个示范性实例中，杂原子选自B、N、O和S，其中氮和硫原子任选地被氧化，氮原子任选地被季铵化。杂芳基可通过杂原子连接到分子的其余部分。芳基或杂芳基的非限制性实施例包括苯基、萘基、联苯基、吡咯基、吡唑基、咪唑基、吡嗪基、恶唑基、苯基-恶唑基、异恶唑基、噻唑基、呋喃基、噻吩基、吡啶基、嘧啶基、苯并噻唑基、嘌呤基、苯并咪唑基、吲哚基、异喹啉基、喹喔啉基、喹啉基、1-萘基、2-萘基、4-联苯基、1-吡咯基、2-吡咯基、3-吡咯基、3-吡唑基、2-咪唑基、4-咪唑基、吡嗪基、2-恶唑基、4-恶唑基、2-苯基-4-恶唑基、5-恶唑基、3-异恶唑基、4-异恶唑基、5-异恶唑基、2-噻唑基、4-噻唑基、5-噻唑基、2-呋喃基、3-呋喃基、2-噻吩基、3-噻吩基、2-吡啶基、3-吡啶基、4-吡啶基、2-嘧啶基、4-嘧啶基、5-苯并噻唑基、嘌呤基、2-苯并咪唑基、5-吲哚基、1-异喹啉基、5-异喹啉基、2-喹喔啉基、5-喹喔啉基、3-喹啉基和6-喹啉基。上述任意一个芳基和杂芳基环系的取代基选自下文所述的可接受的取代基。
除非另有规定，芳基在与其他术语联合使用时(例如芳氧基、芳硫基、芳烷基)包括如上定义的芳基和杂芳基环。因此，术语“芳烷基”意在包括芳基附着于烷基的那些原子团(例如苄基、苯乙基、吡啶基甲基等)，包括其中碳原子(如亚甲基)已经被例如氧原子代替的那些烷基，例如苯氧基甲基、2-吡啶氧甲基3-(1-萘氧基)丙基等。
术语“离去基团”是指可以被另一种官能团或原子通过取代反应(例如亲和取代反应)所取代的官能团或原子。例如，代表性的离去基团包括三氟甲磺酸酯；氯、溴、碘；磺酸酯基，如甲磺酸酯、甲苯磺酸酯、对溴苯磺酸酯、对甲苯磺酸酯等；酰氧基，如乙酰氧基、三氟乙酰氧基等等。
术语“保护基”包括但不限于“氨基保护基”、“羟基保护基”或“巯基保护基”。术语“氨基保护基”是指适合用于阻止氨基氮位上副反应的保护基团。代表性的氨基保护基包括但不限于：甲酰基；酰基，例如链烷酰基(如乙酰基、三氯乙酰基或三氟乙酰基)；烷氧基羰基，如叔丁氧基羰基(Boc)；芳基甲氧羰基，如苄氧羰基(Cbz)和9-芴甲氧羰基(Fmoc)；芳基甲基，如苄基(Bn)、三苯甲基(Tr)、1,1-二-(4'-甲氧基苯基)甲基；甲硅烷基，如三甲基甲硅烷基(TMS)和叔丁基二甲基甲硅烷基(TBS)等等。术语“羟基保护基”是指适合用于阻止羟基副反应的保护基。代表性羟基保护基包括但不限于：烷基，如甲基、乙基和叔丁基；酰基，例如链烷酰基(如乙酰基)；芳基甲基，如苄基(Bn)，对甲氧基苄基(PMB)、9-芴基甲基(Fm)和二苯基甲基(二苯甲基，DPM)；甲硅烷基，如三甲基甲硅烷基(TMS)和叔丁基二甲 基甲硅烷基(TBS)等等。
本发明的化合物可以通过本领域技术人员所熟知的多种合成方法来制备，包括下面列举的具体实施方式、其与其他化学合成方法的结合所形成的实施方式以及本领域技术上人员所熟知的等同替换方式，优选的实施方式包括但不限于本发明的实施例。
本发明所使用的溶剂可经市售获得。本发明采用下述缩略词：aq代表水；HATU代表O-(7-氮杂苯并三唑-1-基)-N,N,N',N'-四甲基脲六氟磷酸盐；EDC代表N-(3-二甲基氨基丙基)-N'-乙基碳二亚胺盐酸盐；m-CPBA代表3-氯过氧苯甲酸；eq代表当量、等量；CDI代表羰基二咪唑；DCM代表二氯甲烷；PE代表石油醚；DIAD代表偶氮二羧酸二异丙酯；DMF代表N,N-二甲基甲酰胺；DMSO代表二甲亚砜；EtOAc代表乙酸乙酯；EtOH代表乙醇；MeOH代表甲醇；CBz代表苄氧羰基，是一种胺保护基团；BOC代表叔丁基羰基是一种胺保护基团；HOAc代表乙酸；NaCNBH 3代表氰基硼氢化钠；r.t.代表室温；O/N代表过夜；THF代表四氢呋喃；Boc 2O代表二-叔丁基二碳酸酯；TFA代表三氟乙酸；DIPEA代表二异丙基乙基胺；SOCl 2代表氯化亚砜；CS 2代表二硫化碳；TsOH代表对甲苯磺酸；NFSI代表N-氟-N-(苯磺酰基)苯磺酰胺；NCS代表1-氯吡咯烷-2,5-二酮；n-Bu 4NF代表氟化四丁基铵；iPrOH代表2-丙醇；mp代表熔点；LDA代表二异丙基胺基锂；EA代表乙酸乙酯；DPPF代表1,1'-双二苯基磷二茂铁；Et代表乙基；Me代表甲基；DCM代表二氯甲烷；TMSCHN 2代表三甲基硅烷化重氮甲烷；DCE代表二氯乙烷；BSA代表牛血清蛋白；TCEP代表三(2-羧乙基)膦；BH 3-Me 2S代表硼烷甲基硫醚；Pd(OAc) 2代表醋酸钯；DPPP代表1,3-双(二苯基膦)丙烷；TEA代表三乙基胺；TMSCl代表三甲基氯硅烷；EDCI代表1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐；Et 3N代表三乙胺；MeI代表碘甲烷；KHMDS代表六甲基二硅基胺基钾；n-BuLi代表正丁基锂；Pd 2(dba) 3代表三(二亚苄基丙酮)二钯；t-BuXPhOS代表2-二-叔丁膦基-2',4',6'-三异丙基联；DIEA代表N,N-二异丙基乙胺。
化合物经手工或者
Figure PCTCN2018072088-appb-000103
软件命名，市售化合物采用供应商目录名称。
具体实施方式
下面通过实施例对本发明进行详细描述，但并不意味着对本发明任何不利限制。本文已经详细地描述了本发明，其中也公开了其具体实施例方式，对本领域的技术人员而言，在不脱离本发明精神和范围的情况下针对本发明具体实施方式进行各种变化和改进将是显而易见的。
实施例1
Figure PCTCN2018072088-appb-000104
步骤1:将化合物1-1(11.30g.80.14mmol)加入到无水THF(120.00mL)中，-78℃下滴加iPrMgCl(2M,40.07mL)，氮气保护下于15℃搅拌16小时。反应完毕后加入饱和NH 4Cl(50mL)，萃取浓缩后将残余物经硅胶柱层析纯化得化合物1-2。 1HNMR(400MHz,CDCl 3)δ3.57(t,J＝6.0Hz,2H),2.59～2.66(m,3H),2.02～2.07(m,2H),1.11(d,J＝6.8Hz,6H).
步骤2:将化合物1-2(3.70g,24.89mmol)和2-氯-4-溴吡啶(4.79g,24.89mmol)加入到无水THF(50.00mL)中，-78℃下滴加n-BuLi(2.5M,9.96mL)，氮气保护下于20℃搅拌18小时。反应完毕后加入饱和NH 4Cl，萃取浓缩后将残余物硅胶柱层析纯化得化合物1-3。 1HNMR(400MHz,CDCl 3)δ8.31(d,J＝6.8Hz,1H),7.33(d,J＝1.6Hz,1H),7.15～7.19(m,1H),3.92～3.98(m,1H),3.75～3.81(m,1H),2.08～2.17(m,2H),1.89～2.01(m,2H),1.60～1.72(m,2H),0.78(d,J＝6.8Hz,3H),0.72(d,J＝6.8Hz,3H).
步骤3:将化合物1-3(1.60g,7.09mmol)，KOH(795.64mg,14.18mmol)加入到二氧六环(15.00mL)/H 2O(5.00mL)的混合溶剂中，然后加入Pd 2(dba) 3(324.62mg,354.50μmol)和tBuXPhOS(301.07mg,709.00μmol)，氮气保护下，115℃搅拌18小时。反应完毕后用水稀释并用乙酸乙酯萃取，萃取液用盐水和无水硫酸钠干燥后浓缩。将残余物通过硅胶柱层析纯化，得到化合物1-4。MS ESI计算值C 12H 16ClNO[M+H] +208,实测值208.
步骤4:将化合物1-4(200.00mg,964.92μmol)加入到无水DMF(50.00mL)中，0℃下加NaH(60％, 57.90mg,1.45mmol)，25℃搅拌0.5小时，然后0℃下加4-溴甲基苯甲酸甲酯(221.04mg,964.92μmol)，25℃搅拌0.5小时。反应完毕后加入20mL水，过滤将滤渣干燥后柱层析纯化得化合物1-5。MS ESI计算值C 21H 25NO 4[M+H] +356,实测值356.
步骤5:将化合物1-5(100mg,281.35μmol)加入到DCM/H 2O(15.00mL)的混合溶剂中，0℃下加入2M NaOH(2mL)，然后加入50％NH 2OH(2mL)，25℃搅拌0.5小时。用制备HPLC分离得到目标化合物1-6。 1H NMR(400MHz,MeOD)δ7.74(d,J＝8.0Hz,2H),7.68(d,J＝6.8Hz,1H),7.40(d,J＝8.0Hz,2H),6.61(d,J＝0.8Hz,1H),6.46(dd,J＝6.8Hz&0.8Hz,1H),5.25(s,2H),3.91～3.97(m,1H),3.79～3.84(m,1H),2.10～2.19(m,3H),2.01～2.05(m,1H),1.79～1.94(m,1H),0.95(d,J＝7.2Hz,3H),0.83(d,J＝7.2Hz,3H).MS ESI计算值C 20H 24N 2O 4[M+H] +357,实测值357.
实施例2
Figure PCTCN2018072088-appb-000105
步骤1：在-78℃，氮气氛围下，向化合物2-1(10.00g,51.96mmol,1.00eq)和5-氯戊-2-酮(6.27g,51.96mmol,1.00eq)的四氢呋喃(100.00mL)中滴加正丁基锂(2.5M,24.94mL,1.20eq)，然后再氮气环境中-78～15℃搅拌5小时。将该反应液浓缩，分散于水(100mL)和乙酸乙酯(100mL)，萃取合并有机相，经饱和食盐水(100mL*3)洗涤，Na 2SO 4干燥，浓缩后，用硅胶柱纯化得到化合物2-2。MS ESI计算值C 10H 12ClNO[M+H] +198,实测值198.
步骤2：在氮气氛围下，向化合物2-2(3.00g,15.18mmol,1.00eq)和氢氧化钾(1.28g,22.77mmol,1.50eq)的二氧六环(15.00mL)和水(3.00mL)溶液中加入Pd 2(dba) 3(1.39g,1.52mmol,0.10eq)和t-BuXphos(1.29 g,3.04mmol,0.20eq)。用氮气置换3次，然后再氮气环境中110℃下搅拌12个小时。将反应液抽滤，倒入水中，用二氯甲烷：甲醇(10:1,20mL*3)萃取，经饱和食盐水洗涤，硫酸钠干燥，浓缩，用硅胶柱纯化得到化合物化合物2-3。MS ESI计算值C 10H 13NO[M+H] +180,,实测值180.
步骤3：将化合物2-3(230.00mg,1.28mmol,1.00eq)，4-(溴甲基)苯甲酸甲酯(293.97mg,1.28mmol,1.00eq)，碳酸铯(627.21mg,1.93mmol,1.50eq)，碘化钾(10.65mg,64.17μmol,0.05eq)的四氢呋喃(20.00mL)用氮气置换3次，然后再氮气环境66℃下搅拌12小时。将该反应液浓缩，然后溶解在水，用乙酸乙酯萃取，经饱和食盐水洗涤，硫酸钠干燥，浓缩得化合物2-4。MS ESI计算值C 19H 21NO 4[M+H] +328,实测值328.
步骤4：在氮气环境，0℃下，向化合物2-4(300.00mg,916.39μmol,1.00eq)的甲醇(15.00mL)滴加氢氧化钠(2M,2.29mL,5.00eq)水溶液。该反应液在15℃下搅拌2.5小时。将反应液浓缩，溶解在水中(15mL)，然后用二氯甲烷：甲醇(10:1,15mL*3)萃取，经饱和食盐水洗涤，硫酸钠干燥，浓缩，制备HPLC分离纯化得化合物2-5. 1H NMR(400MHz,CDCl 3)δ7.40(m,2H),7.19(d,J＝7.2,1H),6.87(m,2H),6.72(s,1H),6.27(d,J＝5.6Hz,2H),4.01(m,2H),3.86(d,J＝6.0,2H),2.46(m,2H),2.16(s,2H),1.96(m,2H),1.65(m,4H),1.32(s,4H).MS ESI计算值C 18H 20N 2O 4[M+H] +329,实测值329.
实施例3
Figure PCTCN2018072088-appb-000106
步骤1：在-65℃到75℃下，向4-溴-2-氯-吡啶(20.00g,103.93mmol,1.00eq)的甲苯(30.00mL)溶液中滴加正丁基锂的环己烷溶液(2.5M,49.89mL,1.20eq)，-65℃到75℃下搅拌1小时，然后向反应体系中加入化合物3-1(20.85g,103.93mmol,17.09mL,1.00eq)的甲苯(100.00mL)溶液，升温到25℃下搅拌2小时。向反应液加入饱和氯化铵水溶液，用乙酸乙酯萃取，然后无水硫酸钠干燥，过滤，浓缩。浓缩后，用硅胶柱纯化得到化合物3-2。 1H NMR(400MHz,CDCl 3)δ8.29(d,J＝5.3Hz,1H),7.43-7.37(m,3H),7.24(dd,J＝1.6,5.2Hz,1H),7.06-7.00(m,2H),4.13-4.00(m,2H),2.64-2.56(m,1H),2.46(m,1H),2.04-1.91(m,2H)。MS ESI计算值C 15H 13ClFNO[M+H]+278,实测值278.
步骤2：将化合物3-2(8.50g,30.61mmol,1.00eq)，氢氧化钾(3.44g,61.22mmol,2.00eq)，三(二亚苄基丙酮)二钯(5.61g,6.12mmol,0.20eq)，2-二叔丁基膦-2’,4’,6’-三异丙基联苯(2.60g,6.12mmol,0.20eq)的二氧六环(2.00mL)和水(1.00mL)反应液升温到100℃，搅拌2小时。向反应液滴加1M稀盐酸调节pH到7，加入水，用乙酸乙酯萃取，无水硫酸钠干燥，过滤，浓缩得到粗品。打浆纯化得到化合物3-3。 1H NMR(400MHz,CDCl 3)δ12.84(brs,1H),7.40(dd,J＝5.3,8.7Hz,2H),7.25(d,J＝6.8Hz,1H),7.01(t,J＝8.8Hz,2H),6.69(s,1H),6.29(dd,J＝1.4,6.8Hz,1H),4.12-3.98(m,2H),2.48(t,J＝7.2Hz,2H),1.98(m,2H),1.25(dd,J＝6.9,9.5Hz,2H)。MS ESI计算值C 15H 14FNO 2[M+H] +260,实测值260.
步骤3：在0-5℃下，向化合物3-3(7.00g,27.00mmol,1.00eq)的N,N-二甲基甲酰胺(70.00mL)的溶液中加入氢化钠(1.08g,27.00mmol,纯度60％,1.00eq)，0-5℃下搅拌10分钟，然后向反应体系中加入4-溴甲基苯甲酸甲酯(6.18g,27.00mmol,1.00eq)的N,N-二甲基甲酰胺(20.00mL)溶液，10-25℃下搅拌2分钟。向反应液中加入饱和氯化铵水溶液，乙酸乙酯萃取无水硫酸钠干燥，浓缩。浓缩后得到化合物3-4并直接用于下步反应。
步骤4：向化合物3-4(11.00g,27.00mmol,1.00eq)，羟氨水溶液(100.00mL,50％纯度)，甲醇(20.00mL)和二氯甲烷(10.00mL)混合溶液中加入氢氧化钠水溶液(2M,27.00mL,2.00eq)、羟氨水溶液(1.00mL,50％纯度)，升温到40-50℃，搅拌1小时。反应液浓缩后，用1M盐酸调节pH到7,二氯甲烷萃取，浓缩后反相制备HPLC纯化得到化合物3-5。 1H NMR(400MHz,DMSO-d 6)δ11.17(s,1H),9.04(br s,1H),7.74-7.64(m,3H),7.55-7.45(m,2H),7.32(d,J＝8.3Hz,2H),7.14(t,J＝8.9Hz,2H),6.50(d,J＝1.9Hz,1H),6.25(dd,J＝2.0,7.2Hz,1H),6.29-6.22(m,1H),5.05(s,2H),3.98-3.83(m,2H),2.48-2.41(m,2H),1.85(m,2H)。MS ESI计算值C 23H 21FN 2O 4[M+H] +409,实测值409.
实施例4
Figure PCTCN2018072088-appb-000107
步骤1：向化合物4-1(1.50g,5.79mmol)的二甲基甲酰胺(20.00mL)溶液中加入氢化钠(347.13mg,8.69mmol,60％纯度)，在0℃下搅拌30分钟后，再加入4-溴甲基苯甲酸甲酯(1.99g,8.69mmol)，升至室温，并搅拌2小时。在0℃下用饱和的氯化铵水溶液(15mL)淬灭并用水(20mL)稀释，乙酸乙酯EtOAc(20mL*4)萃取。合并的有机相用水(20mL*2)和饱和的食盐水(20mL*1)洗，干燥，过滤，浓缩，经硅胶柱层析纯化(PE/EtOAc＝3/1,1:1to 0/1)后得到粗品混合物。粗品混合物采用手性拆分(手性柱型号:OJ(250mm*30mm,10μm)；流动相:A:CO2B:0.05％二乙胺/EtOH，EtOH(0.05％二乙胺)与超临界流体CO2从5％到40％,流速60mL/min)，拆分后得到化合物4-2和化合物5-1，保留时间分别为2.226min，2.835min。化合物4-2的 1H NMR(400MHz,DMSO-d 6)δ＝7.95-7.88(m,2H),7.71(d,J＝7.2Hz,1H),7.54-7.46(m,2H),7.38(d,J＝8.4Hz,2H),7.19-7.09(m,2H),6.51(d,J＝2.0Hz,1H),6.27(dd,J＝2.0,7.2Hz,1H),5.09(s,2H),3.98-3.85(m,2H),3.83(s,3H),2.49-2.41(m,2H),1.91-1.75(m,2H).MS ESI计算值C 24H 22FNO 4[M+H] +407.43,实测值408.1.
.步骤2：向化合物4-2(700.00mg,1.72mmol)的甲醇(5.00mL)溶液中加入氢氧化钠(103.20mg,2.58mmol)和羟胺(5.00mL，50％水溶液)，室温搅拌30分钟.浓缩后用稀盐酸(1M)将pH调至7，反相制备HPLC(0.225％FA)分离纯化后得到化合物4-3。 1H NMR(400MHz,DMSO-d6)δ＝11.49-10.50(br s,1H),9.04(br s,1H),7.69(m,3H),7.49(m,2H),7.33(m,2H),7.14(m,2H),6.50(m,1H),6.25(m,1H),5.05(s,2H),3.90(m,2H),2.45(m,2H),1.85(m,2H)。MS ESI计算值C 23H 21N 2O 4F[M+H] +408.42,实测值 408.9。
实施例5
Figure PCTCN2018072088-appb-000108
步骤1：向化合物5-1(900.00mg,2.21mmol)的甲醇(5.00mL)溶液中加入氢氧化钠(132.60mg,3.32mmol)和羟胺(5.00mL，50％水溶液),室温搅拌30分钟。浓缩，用稀盐酸(1M)调pH至7，用乙酸乙酯(50mL*4)萃取，合并有机相并浓缩，反相制备HPLC((0.225％FA)纯化后得到化合物5-2。 1H NMR(400MHz,DMSO-d 6)δ12.07-10.09(br s,1H),9.08(br s,1H),7.68(m,3H),7.56-7.44(m,2H),7.32(m,2H),7.14(m,2H),6.50(m,1H),6.25(m,1H),5.01(s,2H),4.09-3.74(m,2H),2.45(m,2H),1.94-1.76(m,2H).MS ESI计算值C23H21FN2O4[M+H] +408.42,实测值408.8.
实施例6
Figure PCTCN2018072088-appb-000109
Figure PCTCN2018072088-appb-000110
步骤1：在5℃下，向化合物6-1(809.15mg,7.09mmol,697.54μL,1.00eq)和三氯化铝(633.60mg,4.75mmol,259.67μL,0.67eq)混合物中加入4-氯丁酰氯(500.00mg,3.55mmol,396.83μL,0.50eq)，25℃下搅拌3小时后，将反应液加入到冰水中，搅拌，用乙酸乙酯提取，饱和食盐水洗涤，无水硫酸钠干燥，过滤，将滤液浓缩，得化合物6-2。该液体直接用于下步反应。
步骤2：在-68℃下，向4-溴-2-氯-吡啶(527.29mg,2.74mmol,1.00eq)的甲苯(5.00mL)溶液中加入正丁基锂的环己烷溶液(2.5M,1.21mL,1.10eq)，-68℃下搅拌10分钟，然后向反应体系中加入化合物6-2(600.00mg,2.74mmol,1.00eq)的甲苯(5.00mL)溶液，-68℃搅拌1小时，25℃下搅拌14小时。向反应液依加入饱和氯化铵水溶液和水，用乙酸乙酯萃取，然后饱和食盐水洗涤，无水硫酸钠干燥，浓缩。浓缩后，用TLC板纯化得到化合物化合物6-3。MS ESI计算值C 15H 12F 2ONCl[M+H] +296,实测值296.
步骤3：向化合物6-3(87.00mg,294.21μmol,1.00eq)的二氧六环(2.00mL)和水(1.00mL)溶液中，加入氢氧化钾(33.02mg,588.42μmol,2.00eq)、三(二亚苄基丙酮)二钯(26.94mg,29.42μmol,0.10eq)、2-二叔丁基膦-2’,4’,6’-三异丙基联苯(12.49mg,29.42μmol,0.10eq)，80℃下搅拌3小时。向反应液中加入水，用乙酸乙酯萃取，饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩。浓缩后TLC板纯化得到化合物6-4。MS ESI计算值C 15H 13F 2O 2N[M+H] +278,实测值278.
步骤4：在0℃下，向化合物6-4(60.00mg,216.40μmol,1.00eq)的N,N-二甲基甲酰胺(1.00mL)的溶液中加入氢化钠(17.31mg,432.80μmol,纯度60％,2.00eq)，然后向反应体系中加入4-溴甲基苯甲酸甲酯(49.57mg,216.40μmol,1.00eq)的N,N-二甲基甲酰胺(1.00mL)溶液，25℃下搅拌2小时。向反应 液中加入水，乙酸乙酯萃取，饱和食盐水洗涤，无水硫酸钠干燥，浓缩。浓缩后，用TLC板纯化得到化合物6-5。MS ESI计算值C 24H 21NO 4F 2[M+H] +426,实测值426.
步骤5：向化合物6-5(32.00mg,75.22μmol,1.00eq)的甲醇(2.00mL)溶液中加入氢氧化钠水溶液(6M,50.00μL,3.99eq)、羟氨水溶液(1.00mL,50％水溶液)，25℃下搅拌17小时。反应液浓缩后，反相制备柱纯化得到化合物6-6。 1H NMR(400MHz,MeOD)δppm 1.89-2.08(m,2H)2.48-2.74(m,2H)3.93-4.11(m,2H)5.20(s,2H)6.46(dd,J＝7.09,1.82Hz,1H)6.66(s,1H)6.88-7.03(m,2H)7.37(d,J＝8.16Hz,2H)7.63(d,J＝7.15Hz,1H)7.67-7.72(m,2H)7.72-7.75(m,1H)。MS ESI计算值C 23H 20N 2O 4F 2[M+H] +427,实测值427.
实施例7
Figure PCTCN2018072088-appb-000111
步骤1：向化合物7-1(70.00mg,269.99μmol)的二甲基甲酰胺(5.00mL)的溶液中加入氢化钠(21.60mg,539.98μmol,60％纯度)，在0℃下搅拌30分钟，再向混合物中加入6-溴甲基-3-甲酸甲酯吡啶(124.23mg,539.98μmol)，在0℃下搅拌30分钟。升至室温，继续搅拌30分钟。混合物用饱和的氯化铵水溶液(1mL)，加水(15mL)，乙酸乙酯(10mL*4)萃取。合并有机相，用水(10mL*3)和饱和的食盐水(10mL)洗涤，干燥，过滤，浓缩。板分离(PE/EA＝1/1)得到化合物7-2。MS ESI计算值C 23H 21FN 2O 4[M+H] +409,实测值409。
步骤2：向化合物7-2(70.00mg,161.11μmol)的甲醇(5.00mL)溶液中加入羟胺(1.00mL，50％的水溶液)和氢氧化钠NaOH(6.44mg,161.11μmol)，室温下搅拌30分钟。混合物用稀盐酸(1M)调pH至7，纯化后得到化合物7-3。 1H NMR(400MHz,DMSO-d 6)δ8.77(d,J＝1.8Hz,1H),8.05(dd,J＝2.4,8.2 Hz,1H),7.69(d,J＝7.2Hz,1H),7.57-7.48(m,2H),7.30(d,J＝8.2Hz,1H),7.21-7.11(m,2H),6.48(d,J＝1.8Hz,1H),6.28(dd,J＝2.0,7.2Hz,1H),5.14(s,2H),4.01-3.83(m,2H),2.49-2.44(m,2H),1.92-1.79(m,2H).MS ESI计算值C 22H 20FN 3O 4[M+H] +410,实测值410.
实施例8
Figure PCTCN2018072088-appb-000112
步骤1：向化合物8-1(70.00mg,269.99μmol)的二甲基甲酰胺(5.00mL)溶液中加入氢化钠(21.60mg,539.98μmol,纯度60％)，在0℃下搅拌30分钟，然后加入2-溴甲基5-甲酸甲酯嘧啶(132.33mg,539.98μmol)，继续在0℃下搅拌30分钟，升至室温后继续搅拌30分钟。混合物用饱和的氯化铵(1mL)淬灭，加水(15mL)，乙酸乙酯萃取(10mL*4)。合并后的有机相用水(5mL*3)和饱和的食盐水(10mL)洗涤，干燥，过滤浓缩，制备薄层层析(PE/EA＝1/1)分离纯化后得到化合物8-2。MS ESI计算值C22H20FN3O4[M+H] +424,实测值424.
步骤2：向化合物8-2(70.00mg,160.72μmol,)的甲醇(5.00mL)溶液中加入羟胺(1.00mL，50％的水溶液)和氢氧化钠NaOH(6.43mg,160.72μmol)，室温下搅拌30分钟。混合物用稀盐酸(1M)调pH至7，纯化后得到化合物8-3。 1H NMR(400MHz,DMSO-d6)δ8.97(s,2H),7.69(d,J＝7.2Hz,1H),7.59-7.51(m,2H),7.20-7.13(m,2H),6.47(d,J＝2.0Hz,1H),6.31(dd,J＝2.0,7.2Hz,1H),5.26(s,2H),4.00-3.87(m,2H),2.53(m,2H),1.93-1.82(m,2H).MS ESI计算值C 21H 19FN 4O 4[M+H] +411,实测值411.
实施例9
Figure PCTCN2018072088-appb-000113
步骤1：向化合物9-1(70.00mg,269.99μmol)的二甲基甲酰胺(5.00mL)溶液中加入氢化钠(21.60mg,539.98μmol,纯度60％)，在0℃下搅拌30分钟，然后加入2-甲酸甲酯-5-溴甲基吡啶(124.23mg,539.98μmol)，继续在0℃下搅拌30分钟，升至室温后继续搅拌30分钟。混合物用饱和的氯化铵(1mL*1)淬灭，加水(15mL)，乙酸乙酯萃取(10mL*3)。合并后的有机相用水(10mL*2)和饱和的食盐水(10mL)洗涤，干燥，过滤浓缩，制备TLC(PE/EA＝1/1)分离纯化后得到化合物9-2。MS ESI计算值C 23H 21FN 2O 4[M+H] +409,实测值409.
步骤2：向化合物9-2(50.00mg,粗品)的甲醇(5.00mL)溶液中加入羟胺(1.00mL，50％的水溶液)和氢氧化钠NaOH(3.06mg,76.39μmol)，室温下搅拌30分钟。混合物用稀盐酸(1M)调pH至7，纯化后得到化合物9-3。 1H NMR(400MHz,DMSO-d 6)δ8.53(s,1H),7.97-7.87(m,1H),7.86-7.72(m,2H),7.50(m,2H),7.14(m,2H),6.51(m,1H),6.28(m,1H),5.10(s,2H),4.01-3.81(m,2H),2.48-2.42(m,2H),1.92-1.76(m,2H).MS ESI计算值C 22H 20FN 3O 4[M+H] +410,实测值410.
实施例10
Figure PCTCN2018072088-appb-000114
Figure PCTCN2018072088-appb-000115
步骤1：向化合物10-1(4.68g,35.46mmol,3.66mL,2.00eq)和三氯化铝(3.14g,23.58mmol,1.29mL,1.33eq)混合物中滴加4-氯丁酰氯(2.50g,17.73mmol,1.98mL,1.00eq)，30℃下搅6小时后，将反应液加入到冰水中，搅拌，用乙酸乙酯提取，饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩。浓缩后，用硅胶柱纯化得化合物10-2。该液体直接用于下步反应。
步骤2：在-68℃下，向4-溴-2-氯-吡啶(488.80mg,2.54mmol,1.00eq)的甲苯(3.00mL)溶液中加入正丁基锂的环己烷溶液(2.5M,1.12mL,1.10eq)，然后向反应体系中加入化合物10-2(600.00mg,2.54mmol,1.00eq)的甲苯(1.00mL)溶液，升温到25℃搅拌6小时。向反应液加入饱和氯化铵水溶液和水，用乙酸乙酯萃取，然后饱和食盐水洗涤，无水硫酸钠干燥，浓缩。浓缩后，用硅胶柱纯化得到化合物10-3。MS ESI计算值C 15H 11F 3ONCl[M+H] +314,实测值314.
步骤3：向化合物10-3(410.00mg,1.31mmol,1.00eq)的二氧六环(5.00mL)和水(1.00mL)溶液中，加入氢氧化钾(146.67mg,2.61mmol,2.00eq)，三(二亚苄基丙酮)二钯(119.96mg,131.00μmol,0.10eq)，2-二叔丁基膦-2’,4’,6’-三异丙基联苯(55.50mg,130.70μmol,0.10eq)，80℃下搅拌4小时。向反应液中加入饱和氯化铵水溶液，水，用乙酸乙酯萃取，饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩。浓缩后硅胶柱纯化得到化合物10-4。
步骤4：化合物10-4(100.00mg,338.68μmol,1.00eq)，4-溴甲基苯甲酸甲酯(155.16mg,677.36μmol,2.00eq)，碳酸铯(220.70mg,677.37μmol,2.00eq)的乙腈(2.00mL)溶液升温到80℃，搅拌2小时。向反应液中加入水，乙酸乙酯萃取，饱和食盐水洗涤，无水硫酸钠干燥，浓缩。浓缩后，用制备薄层层析纯化得到化合物10-5。
步骤5：向化合物10-5(40.00mg,90.21μmol,1.00eq)的甲醇(2.00mL)溶液中加入氢氧化钠(3.61mg,90.21μmol,1.00eq)、羟氨水溶液(1.00mL,50％纯度)，25℃下搅拌17小时。反应液浓缩后，反相制备柱纯化得到化合物10-6。 1H NMR(400MHz,MEOD)δppm 1.75-2.11(m,2H)2.53-2.84(m,2H)3.90-4.20(m,2H)5.41(s,2H)6.79-6.88(m,2H)6.92(s,1H)6.97(dd,J＝5.40,1.25Hz,1H)7.54(d,J＝8.28Hz,2H)7.76(d,J＝8.28Hz,2H)8.05(d,J＝5.40Hz,1H)，MS ESI计算值C23H19N2O4F3[M+H] +445,实测值445.
实施例11
Figure PCTCN2018072088-appb-000116
步骤1：在N 2保护，-70℃下向4-溴-2-氯-3-氟吡啶(4.00g,19.01mmol,1.00eq)的甲苯(20.00mL)溶液中滴加n-BuLi(2.5M,9.12mL,1.20eq)，滴加完成后继续在该温度下搅拌30分钟,然后将化合物11-1(3.81g,19.01mmol,819.67μL,1.00eq)加入到上述悬浊液中。将反应体系温度逐渐升至40℃搅拌12小时。将反应液倾倒入H 2O(80mL),并用EtOAc(40mL x3)萃取，合并的萃取液用盐水(50mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经硅胶柱层析(PE/EtOAc＝10/1)即得化合物11-2。MSESI计算值C 15H 12ClF 2NO[M+H] +296,实测值296.
.步骤2：在N 2保护，将化合物11-2(3.86g,13.05mmol,1.00eq),t-BuXPhOS(554.16mg,1.31mmol,0.10eq),Pd 2(dba) 3(1.20g,1.31mmol,0.10eq)和KOH(1.46g,26.10mmol,2.00eq)的二氧六环(100mL)及H 2O(20mL)混合物加热至100℃搅拌2小时。将反应液倾倒入H 2O(80mL),并用EtOAc/MeOH(10/1,30mL x3)萃取，合并的萃取液用盐水(40mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经PE(20mL)打浆，过滤，干燥即得化合物11-3。MS ESI计算值C 15H 13F 2NO 2[M+H] +278,实测值278.
步骤3：在N 2保护，0℃下向化合物11-3(3.20g,11.54mmol,1.00eq)的DMF(60mL)溶液中加入NaH(553.97mg,13.85mmol,纯度60％,1.20eq)并在该温度下搅拌20分钟,然后将4-溴甲基苯甲酸甲酯(2.64g,11.54mmol,1.10eq)加入到上述悬浊液中并将体系温度升至20℃继续搅拌1小时。将反应液倾倒入H 2O(250mL),并用EtOAc/MeOH(10/1,50mL x3)萃取，合并的萃取液用盐水(100mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经硅胶柱层析(PE/EtOAc＝3/1)即得化合物11-4。MS ESI计算值C 24H 21F 2NO 4[M+H] +426,实测值426.
步骤4：在0℃下向化合物11-4(2.00g,4.70mmol,1.00eq)的MeOH(20.0mL)及DCM(10.0mL)溶液中依次滴加NH 2OH.H 2O(20.0mL，50％水溶液)和NaOH(2M,20.0mL)水溶液，然后将反应继续在0℃搅拌1小时。减压浓缩除去大部分溶剂，剩余溶液冷却至0℃并用浓盐酸调节pH＝7-8，后处理即得化合物11-5。1H NMR(400MHz,DMSO-d 6)δ11.18(s,1H),9.90-9.90(m,1H),9.04(brs,1H),7.98(d,J＝6.78Hz,1H),7.70(d,J＝8.03Hz,2H),7.44(dd,J＝8.41,5.65Hz,2H),7.37(d,J＝8.03Hz,2H),7.15(t,J＝8.91Hz,2H),6.60(d,J＝7.53Hz,1H),4.90-5.15(m,2H),3.80-4.05(m,2H),2.60-2.75(m,1H),2.38-2.44(m,1H),1.93(q,J＝7.09Hz,2H).MS ESI计算值C 23H 20F 2N 2O 4[M+H]+427,实测值427.
实施例12
Figure PCTCN2018072088-appb-000117
Figure PCTCN2018072088-appb-000118
步骤1：在N 2保护，-70℃下向4-溴-2-氯吡啶(515.62mg,2.68mmol,1.00eq)的甲苯(3.00mL)溶液中滴加n-BuLi(2.5M,1.29mL,1.20eq)，滴加完成后在该温度下继续搅拌20分钟,然后将化合物12-1(500.00mg,2.68mmol,1.00eq)加入到上述悬浊液中。将反应体系温度逐渐升至25℃搅拌6小时。减压浓缩除去甲苯，将剩余固体溶解在DMF(3.00mL)中，在25℃下加入NaH(160.76mg,4.02mmol,纯度60％,1.50eq)然后将体系加热至60℃搅拌2小时。将反应液倾倒入H 2O(80mL),并用EtOAc(30mL x3)萃取，合并的萃取液用盐水(50mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经硅胶柱层析(PE/EtOAc＝10/1)即得化合物12-2。MS ESI计算值C 14H 11ClFNO[M+H] +264,实测值264.
步骤2：在N 2保护，将化合物12-2(320.00mg,1.21mmol,1.00eq),t-BuXPhOS(102.76mg,242.00μmol,0.20eq),Pd 2(dba) 3(221.60mg,242.00μmol,0.20eq)和KOH(135.79mg,2.42mmol,2.00eq)的二氧六环(10.00mL)及H 2O(3.00mL)混合物加热至100℃搅拌2小时。将反应液倾倒入H 2O(100mL),并用EtOAc(50mL x3)萃取，合并的萃取液用盐水(60mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经制备TLC(PE/EtOAc＝3/1)即得化合物12-3。MS ESI计算值C 14H 12FNO 2[M+H] +246,实测值246.
步骤3：在N 2保护，0℃下向化合物12-3(290.00mg,1.18mmol,1.00eq)的DMF(5.00mL)溶液中加入NaH(56.80mg,1.42mmol,纯度60％,1.20eq)并在该温度下搅拌20分钟,然后将4-溴甲基苯甲酸甲酯(297.33mg,1.30mmol,1.10eq)加入到上述悬浊液中并将体系温度升至20℃继续搅拌2小时。将反应液倾倒入H 2O(80mL)，并用EtOAc/MeOH(10/1,30mL x3)萃取，合并的萃取液用盐水(50mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经制备TLC(PE/EtOAc＝1/1)分离纯化即得化合物12- 4。MS ESI计算值C 23H 20FNO 4[M+H] +394,实测值394.
步骤4：在0℃下向化合物12-4(150.00mg,381.28μmol,1.00eq)的DCM(2.00mL)及MeOH(2.00mL)溶液中依次滴加NH 2OH.H 2O(150.00μL,50％水溶液)和NaOH(2M,150.00μL)水溶液，然后将反应继续在0℃搅拌2小时。减压浓缩除去大部分溶剂，剩余溶液经制备HPLC(0.1％NH 4OH)分离纯化即得化合物12-5。 1H NMR(400MHz,DMSO-d 6)δ10.99(brs,1H),8.67-9.79(brs,1H),7.77(d,J＝7.28Hz,1H),7.69(d,J＝8.03Hz,2H),7.46(dd,J＝8.66,5.40Hz,2H),7.34(d,J＝8.03Hz,2H),7.20(t,J＝8.91Hz,2H),6.52(d,J＝1.51Hz,1H),6.18(dd,J＝7.03,1.76Hz,1H),5.08(s,2H),4.39-4.58(m,2H),3.01-3.19(m,2H).MS ESI计算值C 22H 19FN 2O 4[M+H]+395,实测值395.
实施例13
Figure PCTCN2018072088-appb-000119
步骤1：在N 2保护，-70℃下向4-溴-2-氯-3-氟吡啶(450.32mg,2.14mmol,1.00eq)的甲苯(8.00mL)溶液中滴加n-BuLi(2.5M,1.03mL,1.20eq)，滴加完成后在该温度下继续搅拌20分钟,然后将化合物13-1(400.00mg,2.14mmol,1.00eq)加入到上述悬浊液中。将反应体系温度逐渐升至25℃搅拌6小时。减压浓缩除去甲苯，将剩余固体溶解在DMF(8.00mL)中，在25℃下加入NaH(128.40mg,3.21mmol, 纯度60％,1.50eq)然后将体系加热至60℃搅拌2小时。将反应液倾倒入H 2O(80mL),并用EtOAc(30mL x3)萃取，合并的萃取液用盐水(50mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经硅胶柱层析(PE/EtOAc＝10/1)即化合物13-2。MS ESI计算值C 14H 10ClF 2NO[M+H] +282,实测值282.
步骤2：在N 2保护，将化合物13-2(440.00mg,1.56mmol,1.00eq),t-BuXPhOS(132.49mg,312.00μmol,0.20eq)，Pd 2(dba) 3(285.70mg,312.00μmol,0.20eq)和KOH(175.06mg,3.12mmol,2.00eq)的二氧六环(10.00mL)及H 2O(3.00mL)的混合物加热至100℃搅拌2小时。将反应液倾倒入H 2O(100mL),并用EtOAc(50mL x3)萃取，合并的萃取液用盐水(60mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经制备TLC(PE/EtOAc＝3/1)即得化合物13-3。MS ESI计算值C 14H 11F 2NO 2[M+H] +264,实测值264.
步骤3：在N 2保护，0℃下向化合物13-3(380.00mg,1.44mmol,1.00eq)的DMF(5.00mL)溶液中加入NaH(69.29mg,1.73mmol,纯度60％,1.20eq)并在该温度下搅拌20分钟，然后将4-溴甲基苯甲酸甲酯(363.74mg,1.59mmol,1.10eq)加入到上述悬浊液中并将体系温度升至20℃继续搅拌2小时。将反应液倾倒入H 2O(80mL)，并用EtOAc/MeOH(10/1,30mL x3)萃取，合并的萃取液用盐水(50mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经制备TLC(PE/EtOAc＝1/1)分离纯化即得化合物13-4。MS ESI计算值C 23H 19F 2NO 4[M+H] +412,实测值412.
步骤4：在0℃下向化合物13-4(350.00mg,850.75μmol,1.00eq)的DCM(3.00mL)及MeOH(3.00mL)溶液中依次滴加NH 2OH.H 2O(3.00mL,50％水溶液)和NaOH(2M,3.00mL)水溶液，然后将反应继续在0℃搅拌1小时。减压浓缩除去大部分溶剂，剩余溶液经制备HPLC(0.1％TFA)分离纯化即得化合物13-5。 1H NMR(400MHz,DMSO-d 6)δ11.19(brs,1H),8.06(d,J＝6.52Hz,1H),7.71(d,J＝8.28Hz,2H),7.38-7.50(m,5H),7.22(t,J＝8.91Hz,2H),6.65(d,J＝7.28Hz,1H),4.98-5.08(m,2H),4.49-4.59(m,2H),3.25-3.34(m,1H),3.05-3.15(m,1H).MS ESI计算值C 22H 18F 2N 2O 4[M+H]+413,实测值413.
实施例14
Figure PCTCN2018072088-appb-000120
Figure PCTCN2018072088-appb-000121
步骤1：在N 2保护，-70℃下向4-溴-2-氯-5-氟吡啶(3.00g,14.26mmol,1.00eq)的甲苯(80.00mL)溶液中滴加n-BuLi(2.5M,6.84mL,1.20eq)，滴加完成后在该温度下继续搅拌30分钟,然后将化合物14-1(2.86g,14.26mmol,2.34mL,1.00eq)加入到上述悬浊液中。将反应体系温度逐渐升至40℃搅拌12小时。将反应液倾倒入H 2O(300mL),并用EtOAc(100mL x3)萃取，合并的萃取液用盐水(100mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经硅胶柱层析(PE/EtOAc＝10/1)即得化合物14-2。MS ESI计算值C 15H 12ClF 2NO[M+H] +296,实测值296.
步骤2：在N 2保护，将化合物14-2(300.00mg,1.01mmol,1.00eq),t-BuXPhOS(85.78mg,202.00μmol,0.20eq)，Pd 2(dba) 3(184.98mg,202.00μmol,0.20eq)和KOH(113.34mg,2.02mmol,2.00eq)的二氧六环(10.00mL)及H 2O(3.00mL)的混合物加热至100℃搅拌2小时。将反应液倾倒入H 2O(100mL),并用EtOAc(50mL x3)萃取，合并的萃取液用盐水(60mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经制备TLC(PE/EtOAc＝3/1)即得化合物14-3。MS ESI计算值C 15H 13F 2NO 2[M+H] +278,实测值278.
步骤3：在N 2保护，0℃下向化合物14-3(200.00mg,721.32μmol,1.00eq)的DMF(5.00mL)溶液中加入NaH(34.62mg,865.58μmol,纯度60％,1.20eq),并在该温度下搅拌20分钟，然后将4-溴甲基苯甲酸甲酯(165.23mg,721.32μmol,1.00eq)加入到上述悬浊液中并将体系温度升至20℃继续搅拌1小时。将反应液倾倒入H 2O(50mL)，并用EtOAc/MeOH(10/1,20mL x3)萃取，合并的萃取液用盐水(30mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经制备TLC(PE/EtOAc＝1/1)分离纯化即得化合物14-4。MS ESI计算值C 24H 21F 2NO 4[M+H] +426,实测值426.
步骤4：在0℃下向化合物14-4(100.00mg,235.06μmol,1.00eq)的DCM(2.00mL)及MeOH(4.00mL)溶液中依次滴加NH 2OH.H 2O(2.00mL,50％水溶液)和NaOH(2.00mL,2M)水溶液，然后将反应继续在0℃搅拌1小时。减压浓缩除去大部分溶剂，剩余溶液冷却至0℃，用浓盐酸调pH＝6-7，后处理即得化合物14-5。 1H NMR(400MHz,DMSO-d 6)δ11.17(brs,1H),9.05(brs,1H),8.61-9.43(m,1H),7.64-7.73(m,3H),7.46(dd,J＝8.5,5.5Hz,2H),7.34(d,J＝8.0Hz,2H),7.16(t,J＝8.9Hz,2H),6.48(t,J＝6.9Hz,1H),5.07-5.18(m,2H),3.90(t,J＝7.2Hz,2H),2.64-2.70(m,1H),2.41-2.47(m,1H),1.82-1.97(m,2H).MSESI计算值C 23H 20F 2N 2O 4[M+H]+427,实测值427.
实施例15
Figure PCTCN2018072088-appb-000122
步骤1：在0℃下，将化合物15-1(6.20g,64.50mmol,6.08mL,2.00eq)和三氯化铝(4.73g,35.48mmol,1.10eq)加入至100毫升的三口瓶中，再加入5-氯戊酰氯(5.00g,32.25mmol,4.17mL,1.00eq)，混合物升至室温，搅拌两个小时。混合物用冰水(20mL)淬灭，加水(100mL)，乙酸乙酯(100mL*2)萃取，合并有机相，饱和食盐水洗涤，硫酸钠干燥，过滤，浓缩后得到化合物15-2。 1H NMR(400MHz,CDCl 3)δ8.08-7.91(m,2H),7.13(m,2H),3.63-3.55(m,2H),2.99(m,2H),1.98-1.82(m,4H).
步骤2：-78℃的氮气氛围下，向4-溴-2-氯吡啶(5.83g,30.28mmol,1.00eq)的甲苯(100.00mL)溶液中缓慢滴加入叔丁基锂(2.5M,13.32mL,1.10eq)，搅拌十分钟后缓慢滴加入化合物15-2(6.50g,30.28 mmol,1.00eq)，继续搅拌三十分钟。饱和的氯化铵(20mL)淬灭，加水(100mL)，乙酸乙酯(100mL*3)萃取，合并后的有机相用饱和的食盐水(100mL)洗涤，硫酸钠干燥，过滤，浓缩，柱分离(硅胶柱,石油醚/乙酸乙酯＝10/1)后得到化合物15-3。MS ESI计算值C 16H 16Cl 2FNO[M+H] +328.2,实测值328.2.
步骤3：在0℃氮气氛围下，向化合物15-3(6.00g,18.28mmol,1.00eq)的N,N-二甲基甲酰胺(30.00mL)的溶液中加入NaH(877.49mg,21.94mmol,60％纯度,1.20eq)，0-5℃下搅拌三十分钟。用饱和的氯化铵(30mL)淬灭，加水(100mL)，乙酸乙酯(80mL*3)萃取，合并后的有机相用水(100mL*3)、饱和食盐水(50mL*1)洗涤，硫酸钠干燥，过滤，浓缩后得到化合物15-4。MS ESI计算值C 16H 15ClFNO[M+H] +291.75,实测值292.0.
步骤4：在氮气保护下，水(5.00mL)和1,4-二氧六环(10.00mL)的混合溶液中加入化合物15-4(1.00g,3.43mmol,1.00eq)，氢氧化钾(384.64mg,6.86mmol,2.00eq)，三(二亚苄基丙酮)二钯(627.74mg,686.00μmol,0.20eq)和2-二叔丁基膦-2’,4’,6’-三异丙基联苯(291.10mg,686.00μmol,0.20eq)，混合物加热至90℃，搅拌三小时。混合物浓缩，加水(50mL)，乙酸乙酯(20mL*3)萃取，合并后的有机相用饱和食盐水(50mL)洗涤，硫酸钠干燥，过滤，浓缩，柱纯化(硅胶柱,石油醚/乙酸乙酯＝10/1至2/1、二氯甲烷/甲醇＝20/1)后得到化合物15-5。MS ESI计算值C 16H 16FNO 2[M+H] +273.30,实测值274.1.
步骤5：在0℃氮气保护下，向化合物15-5(150.00mg,439.08μmol,1.00eq)的N,N-二甲基甲酰胺(5.00mL)溶液中加入钠氢(17.56mg,439.08μmol,60％纯度,1.00eq)，混合物搅拌十分钟后加入4-溴甲基苯甲酸甲酯(125.72mg,548.85μmol,1.00eq)，混合物升至室温，继续搅拌三十分钟。向体系中加入饱和的氯化铵(2mL)，加水(15mL)，用乙酸乙酯(10mL*3)萃取，合并后的有机相用饱和食盐水(15mL)洗涤，硫酸钠干燥，过滤浓缩后纯化得到化合物15-6。MS ESI计算值C 25H 24FNO 4[M+H] +421.46,实测值422.2。
步骤6：向化合物15-6(145.00mg,292.44μmol,1.00eq)的甲醇(10.00mL)溶液中加入羟胺水溶液(2.00mL，50％水溶液)和氢氧化钠(35.09mg,877.32μmol,3.00eq)，室温下搅拌一个小时混合物浓缩后制备HPLC(0.225％FA)分离后得化合物15-7。 1H NMR(400MHz,DMSO-d 6)δ＝7.73-7.62(m,3H),7.46(m,2H),7.32(d,J＝8.2Hz,2H),7.16(t,J＝8.8Hz,2H),6.47(d,J＝1.6Hz,1H),6.20(dd,J＝1.6,7.2Hz,1H),5.04(s,2H),3.71-3.53(m,2H),2.34-2.07(m,2H),1.65-1.46(m,4H).MS ESI计算值C 24H 23FN 2O 4[M+H] +422.45,实测值423.4.
实施例16
Figure PCTCN2018072088-appb-000123
步骤1：在-78℃氮气保护下，向化合物16-1(38.76g,201.43mmol,1.00eq)的甲苯(250.00mL)溶液中缓慢滴加入正丁基锂(2.5M,88.63mL,1.10eq)，搅拌三十分钟后向反应体系中缓慢滴加对氟苯甲醛(25.00g,201.43mmol,21.19mL,1.00eq)，在30分钟内体系缓慢升至室温，向体系中加入饱和氯化铵(100mL)溶液，再加入水(200mL)，用乙酸乙酯(150mL*3)萃取，合并有机相，用饱和食盐水(150mL)洗涤，硫酸钠干燥，过滤，浓缩，柱(硅胶柱,石油醚：乙酸乙酯＝5:1)分离纯化后得到化合物16-2。 1HNMR(400MHz,CDCl 3)δ8.30(d,J＝5.2Hz,1H),7.43-7.37(m,1H),7.35-7.29(m,2H),7.20(dd,J＝0.8,5.2Hz,1H),7.11-7.03(m,2H),5.78(d,J＝2.6Hz,1H),2.67(d,J＝3.4Hz,1H).MS ESI计算值C 12H 9ClFNO[M+H] +237.66,实测值238.2.
步骤2：在0℃下，向化合物16-2(23.00g,96.78mmol,1.00eq)的二氯甲烷(230.00mL)中加入戴斯-马丁氧化剂(45.15g,106.46mmol,32.96mL,1.10eq)，体系升温至25℃，搅拌两小时。将混合体系过滤，过滤后的滤液浓缩，经硅胶柱(石油醚：乙酸乙酯＝5:1)层析纯化后得到化合物16-3。MS ESI计算值C 12H 7ClFNO[M+H] +235.64,实测值236.2..
步骤3：向化合物16-3(3.00g,12.73mmol,1.00eq)的甲苯(100.00mL)溶液中加入对甲苯磺酸(7.27g,38.19mmol,3.00eq)和乙二醇(1.58g,25.46mmol,1.42mL,2.00eq)，反应体系加热至110℃，搅拌5小 时。混合物浓缩后柱纯化(石油醚：乙酸乙酯＝10:1至5:1)后得到化合物16-4。MS ESI计算值C 14H 11ClFNO 2[M+H] +279.69,实测值280.0.
步骤4：向化合物16-4(100.00mg,336.09μmol,1.00eq)的1,4-二氧六环(5.00mL)混合物中加入，三(二亚苄基丙酮)二钯(61.55mg,67.22μmol,0.20eq)，氢氧化钾(40.12mg,715.08μmol,2.00eq)和2-二叔丁基膦-2’,4’,6’-三异丙基联苯(28.54mg,67.22μmol,0.20eq)，体系加热至90℃搅拌3小时。混合物浓缩，加水(10mL)，用乙酸乙酯(10mL*3)萃取，合并有机相，用饱和食盐水(10mL)洗涤，硫酸钠干燥，过滤，浓缩，纯化后得到化合物16-5。MS ESI计算值C 14H 12FNO 3[M+H] +261.25,实测值262.2.
步骤5：在0℃氮气保护下，向化合物16-5(80.00mg,281.72μmol,1.00eq)的N,N-二甲基甲酰胺(10.00mL)混合物中加入钠氢(22.54mg,563.44μmol,60％纯度,2.00eq)，体系搅拌十分钟后，加入4-溴甲基苯甲酸甲酯(129.07mg,563.44μmol,2.00eq)，继续搅拌三十分钟。混合物用水(5mL)淬灭，乙酸乙酯萃取(5mL*3)，合并后的有机相用水(10mL)，饱和食盐水(10mL)洗涤，用硫酸钠干燥，过滤，浓缩后纯化得到化合物16-6。MS ESI计算值C 23H 20FNO 5[M+H] +409.41,实测值410.3.
步骤6：向化合物16-6(70.00mg,141.91μmol,1.00eq)的甲醇(5.00mL)溶液中加入羟胺水溶液(1.00mL，50％纯度)和氢氧化钠(17.03mg,425.73μmol,3.00eq)，混合物在室温下搅拌1小时。混合物浓缩后经制备HPLC(0.225％FA)分离纯化后得到化合物16-7。 1H NMR(400MHz,DMSO-d 6)δ＝7.77(d,J＝7.2Hz,1H),7.68(d,J＝8.2Hz,2H),7.53-7.45(m,2H),7.33(d,J＝8.2Hz,2H),7.19(t,J＝8.8Hz,2H),6.48(d,J＝1.8Hz,1H),6.20(dd,J＝1.8,7.2Hz,1H),5.08(s,2H),3.99(m,4H).MS ESI计算值C 22H 19FN 2O 5[M+H] +410.40,实测值411.1.
实施例17
Figure PCTCN2018072088-appb-000124
Figure PCTCN2018072088-appb-000125
步骤1：在N 2保护下，将碘化甲基三苯基磷(2.83g,7.01mmol,1.10eq)和t-BuOK(1M,7.01mL,1.10eq)的THF(20.00mL)溶液加热回流1小时。然后将该悬浊液冷却至0℃并加入化合物17-1(1.50g,6.37mmol,1.00eq)然后将所得反应液在20℃下搅拌12小时。将反应液倾倒入H 2O(100mL),并用EtOAc(50mL x3)萃取,合并的萃取相用盐水(50mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经硅胶柱层析(PE/EtOAc＝20/1)分离纯化即得化合物17-2。MS ESI计算值C 13H 9ClFN[M+H]+234,实测值234.
步骤2：将化合物17-2(800.00mg,3.42mmol,1.00eq)和OsO 4(869.47mg,3.42mmol,177.44μL,1.00eq)的THF(10.00mL)及H 2O(2.00mL)溶液在20℃搅拌4小时。然后将反应液用2M Na 2SO 3水溶液(80mL)荡洗，并用EtOAc(30mL x3)萃取,合并的萃取相用盐水(50mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩即得化合物17-3。MS ESI计算值C 13H 11ClFNO 2[M+H]+268,实测值268.
步骤3：将化合物17-3(800.00mg,2.99mmol,1.00eq),1,2-二溴乙烷(561.45mg,2.99mmol,225.48μL,1.00eq)和t-BuOK(503.03mg,4.49mmol,1.50eq)的DMSO(10.00mL)加热至70℃搅拌6小时。将反应液倾倒入H 2O(100mL),并用EtOAc/MeOH(10/1,50mL x3)萃取,合并的萃取相用盐水(80mL)荡洗,后经无水Na2SO4干燥，过滤，减压浓缩。所得固体经制备TLC(PE/EtOAc＝2/1)分离纯化即得化合物17-4。MS ESI计算值C 15H 13ClFNO 2[M+H]+294,实测值294.
步骤4：在N 2保护下，将化合物17-4(180.00mg,612.83μmol,1.00eq),t-BuXPhOS(26.02mg,61.28μmol,0.10eq),Pd 2(dba) 3(56.12mg,61.28μmol,0.10eq)和KOH(68.77mg,1.23mmol,2.00eq)的二氧六环(10.00mL)及H 2O(2.00mL)溶液加热至100℃搅拌1小时。将反应液倾倒入H 2O(80mL),并用2M HCl调节pH＝6，EtOAc(30mL x3)萃取,合并的萃取相用盐水(50mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经制备TLC(PE/EtOAc＝3/1)分离纯化即得化合物17-5。MS ESI计算值C 15H 14FNO 3[M+H]+276,实测值276.
步骤5：在N 2保护下，于0℃向化合物17-5(90.00mg,326.95μmol,1.00eq)的DMF(5.00mL)溶液中加入NaH(15.69mg,392.34μmol,纯度60％,1.20eq)并在该温度下继续搅拌20分钟。然后将4-(溴甲基)苯甲酸甲酯(74.89mg,326.95μmol,1.00eq)加入到上述悬浊液中,所得反应液升温至15℃搅拌1小时。将反应液倾倒入H 2O(50mL),并用EtOAc(20mL x3)萃取,合并的萃取相用盐水(50mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得固体经制备TLC(PE/EtOAc＝1/2)分离纯化即得化合物17-6。MS ESI计算值C 24H 22FNO 5[M+H]+424,实测值424.
步骤6：在0℃下向化合物17-6(100.00mg,236.17μmol,1.00eq)的DCM(1.00mL)及MeOH(2.00mL)溶液中依次滴加入NH 2OH.H 2O(1.00mL,50％水溶液)和NaOH水溶液(2M,1.00mL)。滴加完成后将反应液在0℃搅拌6小时。减压浓缩除去大部分溶剂。剩余溶液冷却至0℃并用2M HCl调pH＝8,过滤并用H 2O(5mL x3)反复润洗,最后真空干燥即得化合物17-7。 1H NMR(400MHz,DMSO-d 6)δ11.17(brs,1H),9.04(brs,1H),7.74(d,J＝7.28Hz,1H),7.69(d,J＝8.28Hz,2H),7.46(dd,J＝9.03,5.52Hz,2H),7.33(d,J＝8.28Hz,2H),7.19(t,J＝8.91Hz,2H),6.57(d,J＝1.76Hz,1H),6.13(dd,J＝7.15,1.88Hz,1H),5.07(s,2H),4.11-4.24(m,1H),3.99-4.08(m,1H),3.54-3.71(m,4H).MS ESI计算值C 23H 21FN 2O 5[M+H]+425,实测值425.
实施例18
Figure PCTCN2018072088-appb-000126
Figure PCTCN2018072088-appb-000127
步骤1：在0℃下，向1,4-丁内酯(3.45g,40.11mmol,3.05mL,1.10eq)的甲苯(30.00mL)溶液中加入氢化钠(1.75g,43.75mmol,纯度60％,1.20eq)，0℃下加入化合物18-1(5.00g,36.46mmol,4.39mL,1.00eq)的甲苯(10.00mL)溶液，在20℃下搅拌反应12小时。向反应液加入饱和氯化铵水溶液和水，用乙酸乙酯萃取。有机相用饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩得到化合物化合物18-2并直接用于下步反应。
步骤2：将装有化合物18-2(3.22g,16.84mmol,1.00eq),36％盐酸(10.20g,100.71mmol,10.00mL,5.98eq)混合物升高到80℃，搅拌反应3小时。反应液倒入到0℃饱和的碳酸钾水溶液中，加水，用乙酸乙酯萃取，有机相饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩，硅胶柱纯化得到化合物18-3并直接用于下步反应。
步骤3：在-68℃下，向2-氯-4-碘-吡啶(730.20mg,3.05mmol,1.00eq)甲苯(5.00mL)溶液滴加正丁基锂正己烷溶液(2.5M,1.34mL,1.10eq)，然后加入化合物18-3(560.00mg,3.05mmol,1.00eq)的甲苯(1.00mL)溶液，反应逐渐升温到20℃，搅拌反应2小时。向反应液加入饱和氯化铵水溶液，水，用乙酸乙酯萃取，有机相用饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩，硅胶柱纯化得到化合物18-4。MSESI计算值C 14H 13ClN 2O[M+H]+261.0,实测值261.0.
步骤4：向化合物18-4(750.00mg,2.88mmol,1.00eq)的二氧六环(5.00mL)和水(1.00mL)溶液中，加入氢氧化钾(322.82mg,5.75mmol,2.00eq)、三(二亚苄基丙酮)二钯(263.42mg,287.66μmol,0.10eq)、2-二叔丁基膦-2’,4’,6’-三异丙基联苯(122.15mg,287.66μmol,0.10eq)，100℃下搅拌4小时。向反应液中加入饱和氯化铵，水，用乙酸乙酯萃取，饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩。浓缩后硅胶柱纯化得到化合物18-5。MS ESI计算值C 14H 14N 2O 2[M+H]+243,实测值243.
步骤5：在0℃下，向化合物18-5(163.00mg,672.80μmol,1.00eq)的N,N-二甲基甲酰胺(2.00mL)的溶液中加入氢化钠(32.29mg,807.36μmol,60％纯度,1.20eq)，然后向反应体系中加入4-溴甲基苯甲酸甲酯(154.12mg,672.80μmol,1.00eq)的N,N-二甲基甲酰胺(1.00mL)溶液，20℃下搅拌2小时。向反应液中加入饱和氯化铵水溶液和水，乙酸乙酯萃取，饱和食盐水洗涤，无水硫酸钠干燥，浓缩得到化合物18-6。MS ESI计算值C 23H 22N 2O 4[M+H]+391.0,实测值391.0.
步骤6：向化合物18-6(220.00mg,563.48μmol,1.00eq)的甲醇(2.00mL)溶液中加入氢氧化钠(22.54mg,563.48μmol,1.00eq)，羟氨水溶液(1.00mL,50％水溶液)，20℃下搅拌6小时。反应液浓缩后，反相制备HPLC纯化得到化合物18-7。 1H NMR(400MHz,METHANOL-d4)δ＝8.53(dd,J＝0.8,4.8Hz,1H),7.82-7.74(m,1H),7.70(d,J＝8.2Hz,2H),7.67(d,J＝7.9Hz,1H),7.60(d,J＝7.1Hz,1H),7.36(d,J＝8.3Hz,2H),7.27(ddd,J＝1.1,4.9,7.5Hz,1H),6.79(d,J＝1.8Hz,1H),6.60(dd,J＝2.0,7.1Hz,1H),5.18(s,2H),4.17-3.96(m,2H),3.04-2.91(m,1H),2.38(td,J＝7.6,12.6Hz,1H),2.06-1.80(m,2H).MS ESI计算值C 22H 21N 3O 4[M+H]+392,实测值392.
实施例19
Figure PCTCN2018072088-appb-000128
步骤1：在0℃下，向1,4-丁内酯(1.00g,11.62mmol,884.96μL,1.00eq)的四氢呋喃(5.00mL)溶液中加入氢化钠(1.06g,26.38mmol,纯度60％,2.27eq)，在0℃下搅拌30分钟，然后0℃下加入化合物 19-1(2.39g,17.43mmol,2.06mL,1.50eq)的四氢呋喃(2.00mL)溶液，在20℃下搅拌反应2小时。向反应液加入饱和氯化铵水溶液，水，乙酸乙酯。水相浓缩干加入丙酮，搅拌过滤，母液浓缩干得化合物19-2并直接用于下步反应。
步骤2：将装有化合物19-2(1.50g,7.85mmol,1.00eq)，36％盐酸(15.30g,151.11mmol,15.00mL,19.26eq)混合物升高到80℃，搅拌反应2小时。反应液倒入到0℃冰水中，用乙酸乙酯萃取，有机相饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩，硅胶柱纯化得到化合物19-3并直接用于下步反应。
步骤3：在-68℃下，向2-氯-4-碘-吡啶(782.97mg,3.27mmol,1.00eq)甲苯(5.00mL)溶液滴加正丁基锂正己烷溶液(2.5M,1.44mL,1.10eq)，然后加入化合物19-3(600.00mg,3.27mmol,1.00eq)的甲苯(1.00mL)溶液，反应逐渐升温到20℃，搅拌反应12小时。向反应液加入饱和氯化铵水溶液，水，用乙酸乙酯萃取，有机相用饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩，硅胶柱纯化得到化合物19-4。MS ESI计算值C 14H 13ClN 2O[M+H]+261.0,实测值261.0.
步骤4：向化合物19-4(513.00mg,1.97mmol,1.00eq)的二氧六环(5.00mL)和水(1.00mL)溶液中，加入氢氧化钾(221.07mg,3.94mmol,2.00eq)，三(二亚苄基丙酮)二钯(180.40mg,197.00μmol,0.10eq)和2-二叔丁基膦-2’,4’,6’-三异丙基联苯(83.65mg,197.00μmol,0.10eq)，100℃下搅拌2小时。向反应液中加入水，用乙酸乙酯萃取，饱和食盐水洗涤，无水硫酸钠干燥，过滤，浓缩。浓缩后硅胶柱纯化得到化合物19-5。MS ESI计算值C 14H 14N 2O 2[M+H]+243,实测值243.
步骤5：在0℃下，向19-5(240.00mg,990.63μmol,1.00eq)的N,N-二甲基甲酰胺(5.00mL)的溶液中加入氢化钠(47.55mg,1.19mmol,纯度60％,1.20eq)，然后向反应体系中加入4-溴甲基苯甲酸甲酯(226.92mg,990.63μmol,1.00eq)的N,N-二甲基甲酰胺(1.00mL)溶液，20℃下搅拌2小时。向反应液中加入饱和氯化铵水溶液和水，乙酸乙酯萃取，饱和食盐水洗涤，无水硫酸钠干燥，浓缩后层析板纯化得到化合物19-6。MS ESI计算值C 23H 22N 2O 4[M+H]+391.1,实测值391.1
步骤6：向化合物19-6(66.00mg,169.04μmol,1.00eq)的甲醇(5.00mL)溶液中加入氢氧化钠(13.52mg,338.08μmol,2.00eq)和羟氨水溶液(1.00mL,50％纯度)，20℃下搅拌12小时。反应液浓缩后，反相制备柱纯化得到化合物19-7。 1H NMR(400MHz,MeOD)δ8.51(d,J＝6.1Hz,2H),8.45(brs,1H),8.47-8.41(m,1H),7.71(d,J＝8.3Hz,2H),7.65(d,J＝7.2Hz,1H),7.62-7.56(m,2H),7.37(d,J＝8.3Hz,2H),6.76(d,J＝1.9Hz,1H),6.49(dd,J＝1.9,7.2Hz,1H),5.20(s,2H),4.13-3.99(m,2H),2.58(t,J＝7.2Hz,2H),2.09- 1.88(m,2H).MS ESI计算值C 22H 21N 3O 4[M+H]+392,实测值392.
实施例20
Figure PCTCN2018072088-appb-000129
步骤1：在N 2保护，-70℃下向1-溴-4-碘苯(15.00g,53.02mmol,1.00eq)的THF(150.00mL)溶液中滴加n-BuLi(2.5M,23.33mL,1.10eq)，滴加完成后，继续在该温度下搅拌30分钟。然后将化合物20-1(10.64g,53.02mmol,8.72mL,1.00eq)滴加入上述黄色悬浊液中，滴加完成后，将反应体系逐渐升温至20℃搅拌12小时。反应完成后，加水(300mL)淬灭，并用EtOAc(100mL x3)萃取，合并的萃取液用盐水(100mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩即得化合物20-2。 1H NMR(400MHz,CDCl 3)δ7.25-7.50(m,6H),6.90-7.00(m,2H),4.03(t,J＝6.8Hz,2H),2.40-2.55(m,2H),1.85-2.00(m,2H).
.步骤2：在N 2保护，-70℃下向化合物20-2(1.00g,3.11mmol,1.00eq)的THF(15.00mL)溶液中滴加n-BuLi(2.5M,1.24mL,1.00eq),滴加完成后，继续在该温度下搅拌30分钟。然后将干冰(约5g)加入上述反应体系并逐渐升温至20℃搅拌2小时。反应完成后，将反应液倾倒入H 2O(30mL),用2M KOH调pH＝9,用EtOAc(10mL x2)萃取。然后水相用2M HCl调pH＝5,用EtOAc(20mL x3)萃取。将该萃取液用盐水(30mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩即得化合物20-3。 1H NMR(400MHz,DMSO-d 6)δ7.87(d,J＝8.0Hz,2H),7.56(d,J＝7.6Hz,2H),7.40-7.50(m,2H),7.12(t,J＝8.8Hz,2H),3.90-4.00(m,2H),2.51-2.60(m,2H),1.75-1.90(m,2H).
步骤3：将化合物20-3(100.00mg,349.28μmol,1.00eq),HATU(159.37mg,419.14μmol,1.20eq)和DIEA(67.71mg,523.92μmol,91.50μL,1.50eq)的MeCN(10.00mL)溶液在20℃搅拌30分钟,然后将4-(氨甲基)苯甲酸甲酯(70.43mg,349.28μmol,1.00eq,HCl盐)加入到上述溶液中并在20℃继续搅拌12小时。反应完成后，浓缩反应液，残留物用H 2O(3mL)打浆,过滤后，真空干燥即得化合物20-4，并直接用于下步反应。MS ESI计算值C 26H 24FNO 4[M+H] +434,实测值434.
步骤4：在20℃下向化合物20-4(150.00mg,346.04μmol,1.00eq)的DCM(4.00mL)和MeOH(2.00mL)溶液中依次滴加NH 2OH.H 2O(3.00mL,50％水溶液)和NaOH(2M,1.50mL)水溶液。滴加完成后，将反应液加热回流6小时。然后直接经制备HPLC(0.1％TFA)分离纯化即得化合物20-5。 1H NMR(400MHz,DMSO-d 6)δ11.15(brs,1H),8.95-9.05(m,2H),7.80(d,J＝8.0Hz,2H),7.67(d,J＝8.0Hz,2H),7.52(d,J＝8.4Hz,1H),7.40-7.50(m,2H),7.33(d,J＝8.4Hz,2H),7.02-7.10(m,2H),4.67(d,J＝5.6Hz,2H),3.92(t,J＝6.8Hz,2H),2.51-2.60(m,2H),1.85-1.95(m,2H).MS ESI计算值C 25H 23FN 2O 4[M+H]+435,实测值435.
实施例21
Figure PCTCN2018072088-appb-000130
步骤1：将化合物21-1(1g,2.31mmol)经制备SFC分离(手性分离柱型号：OJ(250mm*30mm,5um)， 洗脱剂：A:超临界流体CO2B:0.1％NH 3H 2O MeOH，在5.5分钟内将B相从5％提高到到40％并保持40％B相洗脱3分钟,最后5％B相保持1.5分钟,流速60mL/min)，得到两个异构体21-2(P1)和22-1(P2)，保留时间分别为6.474min，7.256min。
.步骤2：在0℃下向化合物21-2(P1)(450.00mg,1.04mmol,1.00eq)的DCM(4.00mL)和MeOH(4.00mL)溶液中依次滴加NH 2OH.H 2O(4.00mL,50％水溶液)和NaOH(2M,4.00mL)，滴加完成后，继续在该温度下搅拌2小时。减压浓缩除去大部分溶剂，剩余溶剂冷却至0℃并用8M HCl调节pH＝7-8,将析出固体过滤，滤饼用H 2O(5mL x2)润洗,然后减压干燥即得化合物21-3。 1H NMR(400MHz,MeOD)δ7.79(d,J＝8.4Hz,2H),7.71(d,J＝8.4Hz,2H),7.54(d,J＝8.4Hz,2H),7.40-7.05(m,4H),6.95-7.05(m,2H),4.60(d,J＝4.0Hz,2H),4.02(t,J＝7.2Hz,2H),2.50-2.60(m,2H),1.85-2.00(m,2H).MS ESI计算值C 25H 23FN 2O 4[M+H] +435,实测值435.
实施例22
Figure PCTCN2018072088-appb-000131
步骤1：在0℃下向化合物22-1(P2)(450.00mg,1.04mmol,1.00eq)的DCM(4.00mL)和MeOH(4.00mL)溶液中依次滴加NH 2OH.H 2O(4.00mL,50％水溶液)和NaOH(2M,4.00mL)，滴加完成后，继续在该温度下搅拌2小时。减压浓缩除去大部分溶剂，剩余溶剂冷却至0℃并用8M HCl调节pH＝7-8,后处理即得化合物22-2(P2)。 1H NMR(400MHz,MeOD)δ7.79(d,J＝8.4Hz,2H),7.71(d,J＝8.4Hz,2H),7.54(d,J＝8.4Hz,2H),7.40-7.05(m,4H),6.95-7.05(m,2H),4.60(s,2H),4.02(t,J＝7.2Hz,2H),2.50-2.60(m,2H),1.85-2.00(m,2H).MS ESI计算值C 25H 23FN 2O 4[M+H] +435,实测值435.
实施例23
Figure PCTCN2018072088-appb-000132
步骤1：将化合物23-1(1.00g,3.11mmol,1.00eq)，叔丁氧羰基氨(546.50mg,4.67mmol,1.50eq),Xantphos(179.95mg,311.00μmol,0.10eq),Pd 2(dba) 3(142.39mg,155.50μmol,0.05eq)和Cs 2CO 3(1.52g,4.67mmol,1.50eq)的DMF(15.00mL)溶液在N 2保护下加热至100℃搅拌12小时。将反应液倾倒入H 2O(100mL),并用EtOAc(50mL x3)萃取,合并的萃取液用盐水(80mL)荡洗,后经无水Na 2SO4干燥，过滤，减压浓缩。所得产物经硅胶柱层析(PE/EtOAc＝9/1)即得化合物23-2。MS ESI计算值C 21H 24FNO 3[M+H]+358,实测值358.
.步骤2：在25℃下向化合物23-2(250.00mg,699.46μmol,1.00eq)的DCM(5.00mL)溶液中加入TFA(2.00mL)并搅拌30分钟。后减压浓缩即得化合物23-3。MS ESI计算值C 16H 16FNO[M+H] +258,实测值258.
步骤3：将化合物23-3(250.00mg,673.26μmol,1.00eq,TFA盐),4-(溴甲基)苯甲酸甲酯(154.22mg,673.26 μmol,1.00eq)和K 2CO 3(139.58mg,1.01mmol,1.50eq)的DMF(5.00mL)溶液在25℃下搅拌6小时。然后将反应液倾倒入H 2O(50mL),并用EtOAc(30mL x2)萃取,合并的萃取液用盐水(30mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩即得化合物23-4并直接用于下步反应。MS ESI计算值C 25H 24FNO 3[M+H] +406,实测值406.
步骤4：在25℃下向化合物23-4(250.00mg,616.58μmol,1.00eq)的DCM(4.00mL)和MeOH(2.00mL)溶液中依次滴加NH 2OH.H 2O(3.00mL,50％水溶液)和NaOH(2M,1.50mL)水溶液，然后将反应体系升温至50℃搅拌2小时。减压浓缩除去大部分溶剂，剩余溶液经制备HPLC(0.1％NH 4OH)分离纯化即得化合物23-5。 1H NMR(400MHz,DMSO-d 6)δ11.04(brs,1H),9.03(brs,1H),7.67(d,J＝8.0Hz,2H),7.35-7.45(m,4H),6.95-7.05(m,4H),6.45(d,J＝8.4Hz,2H),6.28(t,J＝5.6Hz,1H),4.25(d,J＝6.0Hz,2H),3.70-3.90(m,2H),2.20-2.45(m,4H),1.65-1.90(m,2H).MS ESI计算值C 25H 24FNO 3[M+H]+406,实测值406.
实施例24
Figure PCTCN2018072088-appb-000133
步骤1：将化合物24-1(300.00mg,934.06μmol,1.00eq),4-氨基苯甲酸甲酯(141.19mg,934.06μmol,1.00eq),Xantphos(108.09mg,186.81μmol,0.20eq),Cs 2CO 3(456.50mg,1.40mmol,1.50eq)和Pd 2(dba) 3(85.53mg,93.41μmol,0.10eq)的DMF(15.00mL)溶液在N 2保护下加热至100℃搅拌8小时。将反应液倾倒入H 2O(150mL),并用EtOAc(80mL x2)萃取,合并的萃取液用盐水(50mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得产物经硅胶柱层析分离纯化(PE/EtOAc＝5/1)即得化合物24-2。MS ESI计算 值C 24H 22FNO 3[M+H]+392,实测值392.
.步骤2：在25℃下向化合物24-2(250.00mg,638.67μmol,1.00eq)的DCM(2.00mL)和MeOH(4.00mL)溶液中依次滴加NH2OH.H2O(2.00mL,50％水溶液)和NaOH(2M,2.00mL)水溶液。滴加完成后，将反应液在20℃继续搅拌12小时。减压浓缩除去大部分溶剂，然后用DMSO(5mL)溶清，所得溶液经制备HPLC(0.1％NH 4OH)分离纯化即得化合物24-3。
1H NMR(400MHz,DMSO-d6)δ10.88(brs,1H),8.85(brs,1H),8.50(brs,1H),6.90-7.80(m,12H),3.80-4.00(m,2H),2.30-2.45(m,2H),1.80-1.90(m,2H).MS ESI计算值C 23H 21FN 2O 3[M+H]+393,实测值393.
实施例25
Figure PCTCN2018072088-appb-000134
步骤1：将化合物25-1(800.00mg,2.79mmol,1.00eq),HATU(1.59g,4.19mmol,1.50eq)和DIEA(722.26mg,5.59mmol,976.03μL,2.00eq)的MeCN(5.00mL)溶液在20℃搅拌20分钟,然后将4-氨基苯甲酸甲酯(422.38mg,2.79mmol,1.00eq)加入到上述溶液中并在该温度下继续搅拌12小时。减压浓缩除去溶剂后，剩余固体用EtOAc(80mL)溶解,并用2M NaOH水溶液(30mL x2),盐水(50mL)依次荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩干。所得产物经硅胶柱层析(PE/EtOAc＝10/1)即得化合物25-2。MSESI计算值C 25H 22FNO 4[M+H]+420,实测值420.
.步骤2：在20℃下向化合物25-2(1.10g,2.62mmol,1.00eq)的DCM(10.00mL)和MeOH(20.00mL)溶液中依次滴加NH 2OH.H 2O(11.00mL,50％水溶液)和NaOH(2M,10.00mL)水溶液，加完后继续在该温度下搅拌12小时。减压浓缩除去大部分溶剂，然后用DMSO(5mL)溶清，所得溶液经制备HPLC(0.1％HCl)分离纯化即得化合物25-3。
1H NMR(400MHz,DMSO-d 6)δ11.14(brs,1H),10.39(brs,1H),7.86(d,J＝8.4Hz,2H),7.82(d,J＝8.4Hz,2H),7.40(d,J＝8.8Hz,2H),7.61(d,J＝8.8Hz,2H),7.45-7.55(m,2H),7.13(t,J＝8.8Hz,2H),3.96(t,J＝7.6Hz,2H),2.58(t,J＝6.8Hz,2H),1.80-1.95(m,2H).MS ESI计算值C 24H 21FN 2O 4[M+H]+421,实测值421.
实施例26
Figure PCTCN2018072088-appb-000135
步骤1：在N 2保护，-70℃下向化合物26-1(5.00g,25.98mmol,1.00eq)的甲苯(50.00mL)溶液中滴加n-BuLi(2.5M,12.47mL,1.20eq)，滴加完成后继续在该温度下搅拌30分钟。然后将4-氯-1-(4-氟苯基)丁基-1-酮(5.21g,25.98mmol,4.27mL,1.00eq)加入到上述黄色悬浊液中。并将反应体系升温至15℃继续搅拌12小时。将反应液倾倒入H 2O(100mL),并用EtOAc(50mL x3)萃取,合并的萃取相用盐水(80mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩。所得产物经硅胶柱层析(PE/EtOAc＝10/1)分离纯化即得化合物26-2。MS ESI计算值C 15H 13ClFNO[M+H]+278,实测值278.
步骤2：将化合物26-2(1.00g,3.60mmol,1.00eq),DPPP(148.51mg,360.00μmol,0.10eq),Pd 2(dba) 3(164.86mg,180.00μmol,0.05eq)和TEA(546.54mg,5.40mmol,748.68μL,1.50eq)的DMSO(10.00mL)及MeOH(3.00mL)溶液的混合物加热至80℃在CO(50psi)气氛中搅拌12小时。将反应液倾倒入H 2O(100mL),并用EtOAc(50mL x3)萃取,合并的萃取相用盐水(80mL)荡洗,后经无水Na 2SO 4干燥，过滤，减压浓缩即得化合物26-3。MS ESI计算值C 17H 16FNO 3[M+H]+302,实测值302.
步骤3：将化合物26-3(1.00g,3.32mmol,1.00eq)和NaOH(265.60mg,6.64mmol,2.00eq)的MeOH(10.00mL)及H 2O(10.00mL)混合物溶液加热至回流并搅拌2小时。然后减压浓缩除去MeOH，所得水相用EtOAc(10mL x2)萃取,然后将水相用浓盐酸调pH＝6.最后减压浓缩干即得化合物26-4。MS ESI计算值C 16H 14FNO 3[M+H]+288,实测值288.
步骤4：将26-4(900.00mg,3.13mmol,1.00eq)，HATU(1.79g,4.70mmol,1.50eq)和DIEA(1.21g,9.40mmol,3.00eq)的MeCN(15.00mL)溶液在20℃搅拌30分钟。然后将4-(氨甲基)苯甲酸甲酯(631.71mg,3.13mmol,1.00eq,HCl盐)加入到上述溶液中并继续在20℃搅拌3小时。减压浓缩干。并用水H 2O(80mL)稀释，然后用EtOAc(30mL x3)萃取，合并的萃取相用盐水(30mL)荡洗，后经无水Na 2SO 4干燥，过滤，减压浓缩。后经硅胶柱层析(PE/EtOAc＝5/1to 3/1)分离纯化即得26-5。MS ESI计算值C 25H 23FN 2O 4[M+H]+435,实测值435.
步骤5：在0℃下向26-5(200.00mg,460.34μmol,1.00eq)的DCM(2.00mL)和MeOH(4.00mL)溶液中依次滴加NH 2OH.H 2O(47.04mg,460.34μmol,2.00mL,50％水溶液)和NaOH(2M,230.17μL)水溶液，加完后继续在该温度下搅拌3小时。减压浓缩除去大部分溶剂，然后用DMSO(5mL)溶清，所得溶液经制备HPLC(0.05％HCl)分离纯化即得26-6。 1H NMR(400MHz,DMSO-d 6)δ11.15(brs,1H),10.22(brs,1H),9.35(t,J＝6.4Hz,1H),8.72(d,J＝1.5Hz,1H),8.01-8.07(m,1H),7.94-8.00(m,1H),7.68(d,J＝8.3Hz,2H),7.49-7.56(m,2H),7.34(d,J＝8.3Hz,2H),7.15(t,J＝8.9Hz,2H),4.51(d,J＝6.3Hz,2H),3.98(t,J＝7.2Hz,2H),2.62(t,J＝7.2Hz,2H),1.89(qd,J＝7.2,5.1Hz,2H).MS ESI计算值C 24H 22FN 3O 4[M+H]+436,实测值436.
实验例1：体外评价
通过
Figure PCTCN2018072088-appb-000136
多功能酶标仪检测384微孔板内荧光信号强度的变化，从而测定乙酰化底物的去乙酰化程度，以化合物抑制剂的半抑制浓度IC50值为指标，来评价化合物对组蛋白去乙酰化酶(HDAC) 的抑制作用。
1.实验步骤和方法：
1.1化合物稀释加样：
用DMSO将化合物稀释成2mM加到384化合物板中，利用Bravo全自动液体工作站进行3倍稀释，10个梯度，双复孔；Echo超声波液体工作站设备(Echo liquid handler)再从384化合物板中转移0.15μL化合物到384实验板中。
1.2Assay buffer配置：配置含有终浓度为20mM Hepes，137mM NaCl，2.7mM KCl，1mM MgCl 2，0.05％BSA，0.5mM TCEP的1x assay buffer。
1.3 1.5x酶混合液的配置：
快速溶解组蛋白去乙酰化酶加入到1x assay buffer中，混合均匀。将1.5x的酶混合液加入20μL到384孔实验板中，离心混合，放置23℃培养箱，酶与化合物孵育20分钟。
1.4 3x底物混合液的配置：
用1x assay buffer配置3x底物溶液，混合均匀。加入10μL到384孔实验板中，离心混合，放置23℃培养箱孵育90分钟，进行反应。
1.5准备终止混合液：
用1x assay buffer配置胰蛋白酶与阳性化合物SAHA(Vorinostat，伏立诺他)的终止混合液，混合均匀。加入30μL到384孔实验板中终止反应，离心混合，放置23℃培养箱孵育60分钟。
1.6读数：
终止反应60分钟后，将384孔实验板放置在Envision多功能酶标仪上，读取荧光值。
1.7分析数据:用XLfit5软件来分析数据，计算化合物的IC50值。
表1.本发明化合物体外筛选试验结果
Figure PCTCN2018072088-appb-000137
Figure PCTCN2018072088-appb-000138
注：A≤10nM；10nM＜B≤50nM；50nM＜C≤200nM；D＞200nM；
结论：本发明化合物对HDAC6的抑制作用显著，而对HDAC 1弱抑制，具有选择性高的特点。
实验例2：化合物药代动力学评价
实验目的：测试化合物在公比格犬体内药代动力学
实验材料：
比格犬(公,6-15公斤,6月-4年龄，Marshall Bioresources,北京，中国)
实验操作：
以标准方案测试化合物口服给药后的比格犬药代特征，实验中候选化合物配成均一的混悬液，给予比格犬单次口服给药。口服溶媒为一定比例的聚乙二醇-聚氧乙烯蓖麻油水溶液。收集24小时内的全血样品，3000g离心10分钟，分离上清得血浆样品，取30μL血浆样品加入300μL含内标的乙腈溶液沉淀蛋白，涡旋离心取30μL上清液加入等倍体积的水再涡旋离心取上清进样，以LC-MS/MS分析方法定量分 析血药浓度，并计算药代参数，如达峰浓度，达峰时间，半衰期，药时曲线下面积等。
实验结果如表2所示：
表2.药代动力学测试结果
Figure PCTCN2018072088-appb-000139
结论：本发明化合物具有很好的犬药代动力学单项或部分指标。
实验例3：联合Ixazomib给药对人骨髓瘤MM.1S细胞皮下异种移植肿瘤CB-17SCID模型的体内药效学研究
1.实验目的：
HDAC抑制剂广泛应用于多种癌症，并能够与多种药物联合而增强该药物的治疗效果，是一个被充分证实的抗肿瘤靶点。HDAC抑制剂广泛应用于多种癌症，并能够与多种药物联合而增强该药物的治疗效果，如panobinostat与蛋白酶体抑制剂硼替佐米(bortezomib)合用能增加多发性骨髓瘤疗效并明显降低毒性。
本实验的目的是研究本发明化合物与蛋白酶体抑制剂Ixazomib联合给药对人骨髓瘤MM.1S细胞皮下异种移植瘤在CB-17SCID模型体内药效进行评估。
2.实验方法与步骤
2.1细胞培养
人骨髓瘤MM.1S细胞体外单层培养，培养条件为RPMI-1640培养基(供应商：Gibco；货号:22400-089)中加10％胎牛血清，37℃ 5％CO 2孵箱培养。一周两次用胰酶-EDTA进行常规消化处理传代。当细胞饱和度为80％-90％时，收取细胞，计数，接种。
2.2肿瘤细胞接种
将0.2mL 5×10 6个MM.1S细胞皮下接种于每只小鼠(北京维通利华生物科技股份有限公司,雌性,6－8周龄)的右后背(PBS:Matrigel＝1:1)。肿瘤平均体积达到100-150mm 3时开始分组给药。
2.3受试物的配制
表3.受试物配制方法
Figure PCTCN2018072088-appb-000140
Figure PCTCN2018072088-appb-000141
注：在给动物给药前需要轻轻将药物充分混匀。
2.4实验动物日常观察
每天监测动物的健康状况及死亡情况，例行检查包括观察肿瘤生长和药物治疗对动物日常行为表现的影响如行为活动，摄食摄水量(仅目测)，体重变化(每隔一天测量一次)，外观体征或其它不正常情况。基于各组动物数量记录了组内动物死亡数和副作用。
2.5肿瘤测量和实验指标
实验指标是考察肿瘤生长是否被抑制、延缓或治愈。每周三次用游标卡尺测量肿瘤直径。肿瘤体积的计算公式为：V＝0.5a×b 2，a和b分别表示肿瘤的长径和短径。
化合物的抑瘤疗效用TGI(％)或相对肿瘤增殖率T/C(％)评价。TGI(％)，反映肿瘤生长抑制率。TGI(％)的计算：TGI(％)＝((1-(某处理组给药结束时平均瘤体积－该处理组开始给药时平均瘤体积))/(溶剂对照组治疗结束时平均瘤体积－溶剂对照组开始治疗时平均瘤体积))×100％。
相对肿瘤增殖率T/C(％)：计算公式如下：T/C％＝T RTV/C RTV×100％(T RTV：治疗组RTV；C RTV：阴性对照组RTV)。根据肿瘤测量的结果计算出相对肿瘤体积(relative tumor volume，RTV)，计算公式为RTV＝V t/V 0，其中V 0是分组给药时(即d 0)测量所得平均肿瘤体积，V t为某一次测量时的平均肿瘤体积，T RTV与C RTV取同一天数据。
在实验结束后将检测肿瘤重量，并计算T/C weight百分比，T weight和C weight分别表示给药组和溶媒对照组的瘤重。
3.统计分析
统计分析，包括每个组的每个时间点的肿瘤体积的平均值和标准误(SEM)。治疗组在试验结束时给药后第21天表现出最好的治疗效果，因此基于此数据进行统计学分析评估组间差异。两组间比较用T-test进行分析，三组或多组间比较用one-way ANOVA进行分析，如果F值有显著性差异，应用Games-Howell法进行检验。如果F值无显著性差异，应用Dunnet(2-sided)法进行分析。用SPSS 17.0进行所有数 据分析。p<0.05认为有显著性差异。
4.实验结果及讨论
在本实验中，我们评价了实施例26化合物与Ixazomib联合给药在人骨髓瘤MM.1S细胞异种移植瘤模型中的体内药效。该组在不同时间点的瘤体积数据表明。开始给药后第21天，溶剂对照组荷瘤鼠的瘤体积达到2611mm 3，受试物Ixazomib(4mg/kg)单独给药组与溶剂对照组相比具有显著的抑瘤作用(T/C＝34.97％,TGI＝68.65％,p＝0.030)，瘤体积为916mm 3。实施例26化合物(30mg/kg)与Ixazomib(4mg/kg)联合给药组与溶剂对照组相比具有显著的抑瘤作用(T/C＝8.34％,TGI＝96.88％,p＝0.001)，瘤体积为218mm 3。
综上所述，在人骨髓瘤MM.1S细胞异种移植瘤模型中实施例26化合物(30mg/kg)与Ixazomib(4mg/kg)联合给药组与溶剂对照组比较抑瘤效果最好，小鼠显示出良好的耐受性，且无明显体重下降。

Claims (25)

  1. 式(Ⅰ)所示化合物、药学上可接受的盐及其异构体：
    Figure PCTCN2018072088-appb-100001
    Figure PCTCN2018072088-appb-100002
    选自：单键或双键；
    n选自：0或1；
    T 1、T 2分别独立地选自：CH、CH 2、-C(＝O)-、N；
    T 3选自：C或N；
    Z 1、Z 2、Z 3分别独立地选自：CH或N；
    L 1选自：单键、-NH-、-C(＝O)-NH-；
    R 1选自任选被1、2或3个R取代的：C 1-3烷基、苯基、6元杂芳基；
    R 2选自：H、F、Cl、Br、I；
    A环选自：4～7元杂环烷基；
    R选自：F、Cl、Br、I；
    所述6元杂芳基、4～7元杂环烷基之“杂”分别独立地选自：-NH-、N、-O-；
    以上任何一种情况下，杂原子或杂原子团的数目分别独立地选自1、2或3。
  2. 根据权利要求1所述的化合物、药学上可接受的盐及其异构体，其中，R 1选自任选被1、2或3个R取代的：甲基、乙基、异丙基、苯基、吡啶基。
  3. 根据权利要求2所述的化合物、药学上可接受的盐及其异构体，其中，R 1选自任选被1、2或3个R取代的：CH 3、
    Figure PCTCN2018072088-appb-100003
  4. 根据权利要求3所述的化合物、药学上可接受的盐及其异构体，其中，R 1选自：CH 3、
    Figure PCTCN2018072088-appb-100004
    Figure PCTCN2018072088-appb-100005
  5. 根据权利要求1～4任意一项所述的化合物、药学上可接受的盐及其异构体，其中，A环选自：恶丁环基、四氢呋喃基、四氢吡喃基、1,3-二氧戊环基、1,4-二氧杂环庚烷基、1,4-二噁烷基、1,4-氧氮杂环庚烷基、吗啉基。
  6. 根据权利要求5所述的化合物、药学上可接受的盐及其异构体，其中，A环选自：
    Figure PCTCN2018072088-appb-100006
    Figure PCTCN2018072088-appb-100007
  7. 根据权利要求1～4任意一项所述的化合物、药学上可接受的盐及其异构体，其中，结构单元
    Figure PCTCN2018072088-appb-100008
    选自：
    Figure PCTCN2018072088-appb-100009
  8. 根据权利要求1～4任意一项所述的化合物、药学上可接受的盐及其异构体，其中，结构单元
    Figure PCTCN2018072088-appb-100010
    选自：
    Figure PCTCN2018072088-appb-100011
  9. 根据权利要求8所述的化合物、药学上可接受的盐及其异构体，其中，结构单元
    Figure PCTCN2018072088-appb-100012
    选自：
    Figure PCTCN2018072088-appb-100013
  10. 根据权利要求1～4任意一项所述的化合物、药学上可接受的盐及其异构体，其中，结构单元
    Figure PCTCN2018072088-appb-100014
    选自：
    Figure PCTCN2018072088-appb-100015
    Figure PCTCN2018072088-appb-100016
  11. 根据权利要求10所述的化合物、药学上可接受的盐及其异构体，其中，结构单元
    Figure PCTCN2018072088-appb-100017
    选自：
    Figure PCTCN2018072088-appb-100018
    Figure PCTCN2018072088-appb-100019
  12. 根据权利要求11所述的化合物、药学上可接受的盐及其异构体，其中，结构单元
    Figure PCTCN2018072088-appb-100020
    选自：
    Figure PCTCN2018072088-appb-100021
    Figure PCTCN2018072088-appb-100022
    Figure PCTCN2018072088-appb-100023
  13. 根据权利要求12所述的化合物、药学上可接受的盐及其异构体，其中，结构单元
    Figure PCTCN2018072088-appb-100024
    选自：
    Figure PCTCN2018072088-appb-100025
    Figure PCTCN2018072088-appb-100026
  14. 根据权利要求1或13所述的化合物、药学上可接受的盐及其异构体，其中，结构单元
    Figure PCTCN2018072088-appb-100027
    选自：
    Figure PCTCN2018072088-appb-100028
    Figure PCTCN2018072088-appb-100029
  15. 根据权利要求14所述的化合物、药学上可接受的盐及其异构体，其中，结构单元
    Figure PCTCN2018072088-appb-100030
    选自：
    Figure PCTCN2018072088-appb-100031
    Figure PCTCN2018072088-appb-100032
    Figure PCTCN2018072088-appb-100033
  16. 根据权利要求1～4任意一项所述的化合物、药学上可接受的盐及其异构体，其中，结构单元
    Figure PCTCN2018072088-appb-100034
    选自：-CH 2-、-NH-、-C(＝O)-NH-、
    Figure PCTCN2018072088-appb-100035
  17. 根据权利要求1～4任意一项所述的化合物、药学上可接受的盐及其异构体，其中，结构单元
    Figure PCTCN2018072088-appb-100036
    选自：
    Figure PCTCN2018072088-appb-100037
  18. 根据权利要求1～4任意一项所述的化合物、药学上可接受的盐及其异构体，其中，结构单元
    Figure PCTCN2018072088-appb-100038
    选自：
    Figure PCTCN2018072088-appb-100039
    Figure PCTCN2018072088-appb-100040
  19. 根据权利要求1～4任意一项所述的化合物、药学上可接受的盐及其异构体，其选自：
    Figure PCTCN2018072088-appb-100041
    Figure PCTCN2018072088-appb-100042
    其中，
    A环、R、R 2、L 1和n如权利要求1所定义；
    R 1如权利要求1～4任意一项所定义。
  20. 根据权利要求19所述的化合物、药学上可接受的盐及其异构体，其选自：
    Figure PCTCN2018072088-appb-100043
    其中，
    E 1、E 2分别独立地选自-O-、-CH 2-和-CH 2-CH 2-；
    R、R 2、L 1和n如权利要求1所定义。
  21. 下列化合物及其药学上可接受的盐，选自：
    Figure PCTCN2018072088-appb-100044
    Figure PCTCN2018072088-appb-100045
  22. 根据权利要求21所述的化合物、药学上可接受的盐及其异构体，其选自：
    Figure PCTCN2018072088-appb-100046
    Figure PCTCN2018072088-appb-100047
  23. 一种药物组合物，包括治疗有效量的根据权利要求1～22任意一项所述的化合物或其药学上可接受的盐作为活性成分以及药学上可接受的载体。
  24. 根据权利要求1～22任意一项所述的化合物或其药学上可接受的盐或根据权利要求23所述的组合物在制备治疗HDAC6相关病症的药物上的应用。
  25. 根据权利要求24所述的应用，其特征在于，所述药物是治疗多发性骨髓瘤的药物。
PCT/CN2018/072088 2017-01-10 2018-01-10 Hdac6选择性抑制剂及其制备方法和应用 WO2018130155A1 (zh)

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2020007329A1 (zh) * 2018-07-04 2020-01-09 基石药业（苏州）有限公司 Hdac6选择性抑制剂的晶型及其应用
WO2021110121A1 (zh) * 2019-12-05 2021-06-10 基石药业（苏州）有限公司 Hdac6选择性抑制剂及其组合疗法

Families Citing this family (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
AU2022262655A1 (en) 2021-04-23 2023-11-02 Tenaya Therapeutics, Inc. Hdac6 inhibitors for use in the treatment of dilated cardiomyopathy
EP4333841A1 (en) 2021-05-04 2024-03-13 Tenaya Therapeutics, Inc. 2-fluoroalkyl-1,3,4-oxadiazol-5-yl-thiazol, hdac6 inhibitors for use in the treatment of metabolic disease and hfpef

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101641338A (zh) * 2006-09-11 2010-02-03 柯瑞斯公司 作为抗增殖制剂的多功能小分子
WO2011106632A1 (en) * 2010-02-26 2011-09-01 Millennium Pharmaceuticals, Inc. Substituted hydroxamic acids and uses thereof
CN103429574A (zh) * 2010-11-16 2013-12-04 阿塞蒂隆制药公司 作为蛋白质去乙酰化酶抑制剂的嘧啶羟基酰胺化合物和其使用方法
WO2015007870A1 (en) * 2013-07-18 2015-01-22 Institut De Recherche Pour Le Developpement (I.R.D.) New compounds for the treatment and/or prevention of parasitic diseases and method of production of thereof
WO2015100363A1 (en) * 2013-12-23 2015-07-02 The Trustees Of Columbia University In The City Of New York Selective hdac6 inhibitors

Family Cites Families (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
AU2003215112A1 (en) * 2002-02-07 2003-09-02 Axys Pharmaceuticals Novel bicyclic hydroxamates as inhibitors of histone deacetylase
US8088771B2 (en) * 2008-07-28 2012-01-03 Gilead Sciences, Inc. Cycloalkylidene and heterocycloalkylidene inhibitor compounds
US8404738B2 (en) * 2011-01-21 2013-03-26 Hoffmann-La Roche Inc. 4-amino-N-hydroxy-benzamides for the treatment of cancer
US9464073B2 (en) * 2014-02-26 2016-10-11 Acetylon Pharmaceuticals, Inc. Pyrimidine hydroxy amide compounds as HDAC6 selective inhibitors

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101641338A (zh) * 2006-09-11 2010-02-03 柯瑞斯公司 作为抗增殖制剂的多功能小分子
WO2011106632A1 (en) * 2010-02-26 2011-09-01 Millennium Pharmaceuticals, Inc. Substituted hydroxamic acids and uses thereof
CN103429574A (zh) * 2010-11-16 2013-12-04 阿塞蒂隆制药公司 作为蛋白质去乙酰化酶抑制剂的嘧啶羟基酰胺化合物和其使用方法
WO2015007870A1 (en) * 2013-07-18 2015-01-22 Institut De Recherche Pour Le Developpement (I.R.D.) New compounds for the treatment and/or prevention of parasitic diseases and method of production of thereof
WO2015100363A1 (en) * 2013-12-23 2015-07-02 The Trustees Of Columbia University In The City Of New York Selective hdac6 inhibitors

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
BERGE ET AL.: "Pharmaceutical Salts", JOURNAL OF PHARMACEUTICAL SCIENCE, vol. 66, 1977, pages 1 - 19, XP002675560, doi:10.1002/jps.2600660104
DRUG DESIGN, DEVELOPMENT AND THERAPY, vol. 10, 2016, pages 217 - 226
See also references of EP3569592A4

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2020007329A1 (zh) * 2018-07-04 2020-01-09 基石药业（苏州）有限公司 Hdac6选择性抑制剂的晶型及其应用
US11345682B2 (en) 2018-07-04 2022-05-31 Cstone Pharmaceuticals (Suzhou) Co., Ltd. Polymorph of HDAC6-selective inhibitor and application thereof
WO2021110121A1 (zh) * 2019-12-05 2021-06-10 基石药业（苏州）有限公司 Hdac6选择性抑制剂及其组合疗法

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