WO2023116811A1 - SHORT PEPTIDE COMPOUND CONTAINING β-AMINOKETONE AND USE THEREOF - Google Patents

SHORT PEPTIDE COMPOUND CONTAINING β-AMINOKETONE AND USE THEREOF Download PDF

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Publication number
WO2023116811A1
WO2023116811A1 PCT/CN2022/140910 CN2022140910W WO2023116811A1 WO 2023116811 A1 WO2023116811 A1 WO 2023116811A1 CN 2022140910 W CN2022140910 W CN 2022140910W WO 2023116811 A1 WO2023116811 A1 WO 2023116811A1
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compound
pharmaceutically acceptable
acceptable salt
reaction
synthesis
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PCT/CN2022/140910
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French (fr)
Chinese (zh)
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陈曙辉
杨亚讯
张建臣
李鹏
罗志
贺海鹰
王正
黎健
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福建广生中霖生物科技有限公司
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Priority to CN202280016132.6A priority Critical patent/CN116888099A/en
Publication of WO2023116811A1 publication Critical patent/WO2023116811A1/en

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    • 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
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/42—Oxazoles
    • A61K31/423—Oxazoles condensed with carbocyclic rings
    • 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
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/425—Thiazoles
    • A61K31/428—Thiazoles condensed with carbocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A61P31/14—Antivirals for RNA viruses
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D207/00—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom
    • C07D207/02—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D207/18—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having one double bond between ring members or between a ring member and a non-ring member
    • C07D207/22—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having one double bond between ring members or between a ring member and a non-ring member with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D207/24—Oxygen or sulfur atoms
    • C07D207/26—2-Pyrrolidones
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D209/00—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom
    • C07D209/02—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom condensed with one carbocyclic ring
    • C07D209/52—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom condensed with one carbocyclic ring condensed with a ring other than six-membered
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D277/00—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings
    • C07D277/60—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings condensed with carbocyclic rings or ring systems
    • C07D277/62—Benzothiazoles
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D411/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen and sulfur atoms as the only ring hetero atoms
    • C07D411/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen and sulfur atoms as the only ring hetero atoms containing two hetero rings
    • C07D411/06—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen and sulfur atoms as the only ring hetero atoms containing two hetero rings linked by a carbon chain containing only aliphatic carbon atoms

Definitions

  • the invention relates to the technical field of medicinal chemistry, in particular to a short peptide compound containing ⁇ -aminoketone and its application.
  • (3CL pro ) protease belongs to cysteine proteases. Its substrate binding site is highly conserved and has a similar catalytic mechanism. It is a key protease that catalyzes the cleavage of RNA virus precursor protein and plays an important role in virus replication. As a key enzyme for coronavirus replication, 3CL protease (3CL pro ) is an important target for the treatment of diseases caused by various coronaviruses including COVID-19. Using the structural characteristics of 3CL protease to develop drugs for the treatment of new coronavirus infection has great clinical value.
  • the present invention provides a compound represented by formula (II) or a pharmaceutically acceptable salt thereof,
  • Each R 1 is independently selected from halogen, CN, OH, NH 2 , C 1-3 alkyl and C 1-3 alkoxy, and each of the C 1-3 alkyl or C 1-3 alkoxy is independently optionally substituted by 1, 2 or 3 halogens;
  • R 1 and the atoms connected to them form a C 3-6 cycloalkyl group, and the C 3-6 cycloalkyl group is optionally substituted by 1, 2 or 3 R a ;
  • R a is each independently selected from halogen and C 1-3 alkyl
  • n is selected from 0, 1, 2 or 3;
  • R 2 is selected from tert-butyl, C 3-10 cycloalkyl, 3-10 membered heterocycloalkyl and phenyl, each of said C 3-10 cycloalkyl, 3-10 membered heterocycloalkyl and phenyl independently optionally substituted by 1, 2 or 3 R ;
  • R b are each independently selected from halogen and C 1-3 alkyl
  • R 3 is selected from C 1-3 alkyl, C 1-3 alkoxy, -CH 2 R 4 and -CH 2 OR 4 , and the C 1-3 alkyl and C 1-3 alkoxy are each independently optionally substituted by 1, 2 or 3 halogens;
  • R is selected from phenyl and 5-6 membered heteroaryl, each of which is independently optionally substituted by 1, 2 or 3 R;
  • R is selected from halogen and C 1-3 alkyl
  • T1 is selected from O and S;
  • Ring A is selected from
  • a 1 , A 2 , A 3 and A 4 are each independently selected from CH and N.
  • R 1 is selected from F and methyl, and other variables are as defined in the present invention.
  • the above two R 1 and the atoms connected to them form a C 3-6 cycloalkyl group, and other variables are as defined in the present invention.
  • the above two R 1 and the atoms connected to them form a cyclopropyl group, and other variables are as defined in the present invention.
  • R 2 is selected from tert-butyl and adamantyl, and the adamantyl is optionally substituted by 1, 2 or 3 R b , and other variables are as defined in the present invention.
  • R 2 is selected from tert-butyl and adamantyl, and other variables are as defined in the present invention.
  • R 2 is selected from tert-butyl, Other variables are as defined herein.
  • R 3 is selected from -CF 3 , and other variables are as defined in the present invention.
  • the present invention also provides compounds represented by (II-1), (II-2) and (II-3) or pharmaceutically acceptable salts thereof,
  • t is selected from 0, 1 and 2;
  • R 1 , R b , A 1 , A 2 , A 3 , A 4 , T 1 and n are as defined in the present invention.
  • the present invention also provides a compound represented by formula (I) or a pharmaceutically acceptable salt thereof,
  • Each R 1 is independently selected from halogen, CN, OH, NH 2 , C 1-3 alkyl or C 1-3 alkoxy, and the C 1-3 alkyl or C 1-3 alkoxy is optionally each independently substituted by 1, 2 or 3 halogens;
  • R 1 and the atoms connected to them form a C 3-6 cycloalkyl group, and the C 3-6 cycloalkyl group is optionally substituted by 1, 2 or 3 R a ;
  • n is selected from 0, 1, 2 or 3;
  • R a is each independently selected from halogen or C 1-3 alkyl
  • R 2 is selected from tert-butyl, C 3-10 cycloalkyl, 3-10 membered heterocycloalkyl or phenyl, the C 3-10 cycloalkyl, 3-10 membered heterocycloalkyl and phenyl are any optionally replaced by 1, 2 or 3 R b ;
  • Each R b is independently selected from halogen or C 1-3 alkyl
  • R 3 is selected from C 1-3 alkyl, C 1-3 alkoxy, -CH 2 R 4 or -CH 2 OR 4 , and the C 1-3 alkyl and C 1-3 alkoxy are optionally 1, 2 or 3 halogen substitutions;
  • R 4 is selected from phenyl or 5-6 membered heteroaryl, said phenyl and 5-6 membered heteroaryl are optionally substituted independently by 1, 2 or 3 R;
  • R is selected from halogen or C 1-3 alkyl
  • a 1 , A 2 , A 3 and A 4 are each independently selected from CH or N, and 0, 1 or 2 of A 1 , A 2 , A 3 and A 4 are selected from N, and the rest are selected from CH.
  • R 1 is selected from F or methyl, and other variables are as defined in the present invention.
  • the above two R 1 and the atoms connected to them form a C 3-6 cycloalkyl group, and other variables are as defined in the present invention.
  • the above two R 1 and the atoms connected to them form a cyclopropyl group, and other variables are as defined in the present invention.
  • R 2 is selected from tert-butyl, and other variables are as defined in the present invention.
  • R 3 is selected from -CF 3 , and other variables are as defined in the present invention.
  • the present invention also provides the compound represented by (I-1) or a pharmaceutically acceptable salt thereof,
  • R 1 and n are as defined in the present invention.
  • the present invention provides a compound of the following formula or a pharmaceutically acceptable salt thereof:
  • the present invention provides a pharmaceutical composition, which contains a therapeutically effective amount of the above-mentioned compound or a pharmaceutically acceptable salt thereof.
  • the present invention also provides the application of the above-mentioned compound or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating coronavirus infection.
  • the present invention also provides the application of the above-mentioned compound or a pharmaceutically acceptable salt thereof or the above-mentioned composition in the preparation of a medicament for treating coronavirus infection.
  • the aforementioned coronavirus infection is selected from COVID-19.
  • the compound of the present invention has better in vitro anti-new coronavirus Mpro protease activity; better in vitro anti-coronavirus activity at the cellular level, and has no cytotoxicity.
  • pharmaceutically acceptable refers to those compounds, materials, compositions and/or dosage forms, which are suitable for use in contact with human and animal tissues within the scope of sound medical judgment , without undue toxicity, irritation, allergic reaction or other problems or complications, commensurate with a reasonable benefit/risk ratio.
  • pharmaceutically acceptable salt refers to a salt of a compound of the present invention, which is prepared from a compound having a specific substituent found in the present invention and a relatively non-toxic acid or base.
  • base addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of base, either neat solution or in a suitable inert solvent.
  • Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amine or magnesium salts or similar salts.
  • acid addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of the acid, either neat solution or in a suitable inert solvent.
  • Certain specific compounds of the present invention contain basic and acidic functional groups and can thus be converted into either base or acid addition salts.
  • the pharmaceutically acceptable salts of the present invention can be synthesized from the parent compound containing acid groups or bases by conventional chemical methods.
  • such salts are prepared by reacting the free acid or base form of these compounds with a stoichiometric amount of the appropriate base or acid in water or an organic solvent or a mixture of both.
  • the compounds of the invention may exist in particular geometric or stereoisomeric forms.
  • the present invention contemplates all such compounds, including cis and trans isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers isomers, (D)-isomers, (L)-isomers, and their racemic and other mixtures, such as enantiomerically or diastereomerically enriched mixtures, all of which are subject to the present within the scope of the invention.
  • Additional asymmetric carbon atoms may be present in substituents such as alkyl groups. All such isomers, as well as mixtures thereof, are included within the scope of the present invention.
  • enantiomer or “optical isomer” refer to stereoisomers that are mirror images of each other.
  • cis-trans isomers or “geometric isomers” arise from the inability to rotate freely due to the double bond or the single bond of the carbon atoms forming the ring.
  • diastereoisomer refers to stereoisomers whose molecules have two or more chiral centers and which are not mirror images of the molecules.
  • keys with wedge-shaped solid lines and dotted wedge keys Indicates the absolute configuration of a stereocenter, with a straight solid-line bond and straight dashed keys Indicates the relative configuration of the stereocenter, with a wavy line Indicates wedge-shaped solid-line bond or dotted wedge key or with tilde Indicates a straight solid line key or straight dotted key
  • any one or more sites of the group can be linked to other groups through chemical bonds.
  • the chemical bonds that the site is connected with other groups can use straight solid line bonds Straight dotted key or tilde express.
  • the straight-shaped solid-line bond in -OCH3 indicates that it is connected to other groups through the oxygen atom in the group;
  • the straight dotted line bond in indicates that the two ends of the nitrogen atom in the group are connected to other groups;
  • the wavy lines in indicate that the carbon atoms at positions 1 and 2 in the phenyl group are connected to other groups.
  • the terms “enriched in an isomer”, “enriched in an isomer”, “enriched in an enantiomer” or “enantiomerically enriched” refer to one of the isomers or enantiomers
  • the content of the enantiomer is less than 100%, and the content of the isomer or enantiomer is greater than or equal to 60%, or greater than or equal to 70%, or greater than or equal to 80%, or greater than or equal to 90%, or greater than or equal to 95%, or Greater than or equal to 96%, or greater than or equal to 97%, or greater than or equal to 98%, or greater than or equal to 99%, or greater than or equal to 99.5%, or greater than or equal to 99.6%, or greater than or equal to 99.7%, or greater than or equal to 99.8%, or greater than or equal to 99.9%.
  • the terms “isomer excess” or “enantiomeric excess” refer to the difference between the relative percentages of two isomers or two enantiomers. For example, if the content of one isomer or enantiomer is 90% and the other isomer or enantiomer is 10%, then the isomer or enantiomeric excess (ee value) is 80% .
  • Optically active (R)- and (S)-isomers as well as D and L-isomers can be prepared by chiral synthesis or chiral reagents or other conventional techniques. If one enantiomer of a compound of the invention is desired, it can be prepared by asymmetric synthesis or derivatization with chiral auxiliary agents, wherein the resulting diastereomeric mixture is separated and the auxiliary group is cleaved to provide pure desired enantiomer.
  • a diastereoisomeric salt is formed with an appropriate optically active acid or base, and then a diastereomeric salt is formed by a conventional method known in the art. Diastereomeric resolution is performed and the pure enantiomers are recovered. Furthermore, the separation of enantiomers and diastereomers is usually accomplished by the use of chromatography using chiral stationary phases, optionally in combination with chemical derivatization methods (e.g. amines to amino groups formate).
  • the compounds of the present invention may contain unnatural proportions of atomic isotopes at one or more of the atoms that constitute the compounds.
  • compounds may be labeled with radioactive isotopes such as tritium ( 3 H), iodine-125 ( 125 I) or C-14 ( 14 C).
  • radioactive isotopes such as tritium ( 3 H), iodine-125 ( 125 I) or C-14 ( 14 C).
  • heavy hydrogen can be used to replace hydrogen to form deuterated drugs.
  • the bond formed by deuterium and carbon is stronger than the bond formed by ordinary hydrogen and carbon.
  • deuterated drugs can reduce toxic side effects and increase drug stability. , enhance the efficacy, prolong the biological half-life of drugs and other advantages. All changes in isotopic composition of the compounds of the invention, whether radioactive or not, are included within the scope of the invention.
  • substituted means that any one or more hydrogen atoms on a specified atom are replaced by a substituent, which may include deuterium and hydrogen variants, as long as the valence of the specified atom is normal and the substituted compound is stable.
  • Oxygen substitution does not occur on aromatic groups.
  • optionally substituted means that it may or may not be substituted, and unless otherwise specified, the type and number of substituents may be arbitrary on a chemically realizable basis.
  • “Pharmaceutical composition” means a mixture containing one or more compounds described herein, or a physiologically/pharmaceutically acceptable salt or prodrug thereof, and other chemical components, and other components such as a physiologically/pharmaceutically acceptable carrier and excipients.
  • the purpose of the pharmaceutical composition is to promote the administration to the organism, facilitate the absorption of the active ingredient and thus exert biological activity.
  • terapéuticaally effective amount means (i) treating or preventing a particular disease, condition or disorder, (ii) alleviating, ameliorating or eliminating one or more symptoms of a particular disease, condition or disorder, or (iii) preventing or delaying The amount of a compound of the invention for the onset of one or more symptoms of a particular disease, condition or disorder described herein.
  • the amount of a compound of the invention that constitutes a “therapeutically effective amount” will vary depending on the compound, the disease state and its severity, the mode of administration, and the age of the mammal to be treated, but can be routinely determined by one skilled in the art according to its own knowledge and this disclosure.
  • any variable eg, R
  • its definition is independent at each occurrence.
  • said group may optionally be substituted with up to two R, with independent options for each occurrence of R.
  • substituents and/or variations thereof are permissible only if such combinations result in stable compounds.
  • linking group When the number of a linking group is 0, such as -(CRR) 0 -, it means that the linking group is a single bond.
  • a substituent can be bonded to any atom on a ring when the bond of a substituent can cross-link two or more atoms on the ring, e.g., structural unit It means that the substituent R can be substituted at any position on cyclohexyl or cyclohexadiene. When the enumerated substituent does not indicate which atom it is connected to the substituted group, this substituent can be bonded through any atom, for example, pyridyl as a substituent can be connected to any atom on the pyridine ring. The carbon atom is attached to the group being substituted.
  • linking group listed does not indicate its linking direction
  • its linking direction is arbitrary, for example,
  • the linking group L is -MW-, at this time -MW- can be connected to ring A and ring A in the same direction as the reading order from left to right to form It can also be formed by linking loop A and loop A in the opposite direction to the reading order from left to right
  • Combinations of the described linking groups, substituents and/or variations thereof are permissible only if such combinations result in stable compounds.
  • any one or more sites of the group can be linked to other groups through chemical bonds.
  • connection method of the chemical bond is not positioned, and there is an H atom at the connectable site, when the chemical bond is connected, the number of H atoms at the site will decrease correspondingly with the number of chemical bonds connected to become the corresponding valence group.
  • the chemical bonds that the site is connected with other groups can use straight solid line bonds Straight dotted key or tilde express.
  • the straight-shaped solid-line bond in -OCH3 indicates that it is connected to other groups through the oxygen atom in the group;
  • the straight dotted line bond in indicates that the two ends of the nitrogen atom in the group are connected to other groups;
  • the wavy lines in indicate that the 1 and 2 carbon atoms in the phenyl group are connected to other groups.
  • C 1-3 alkyl is used to denote a straight or branched chain saturated hydrocarbon group consisting of 1 to 3 carbon atoms.
  • the C 1-3 alkyl group includes C 1-2 and C 2-3 alkyl groups, etc.; it can be monovalent (such as methyl), divalent (such as methylene) or multivalent (such as methine) .
  • Examples of C 1-3 alkyl include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), and the like.
  • C 1-3 alkoxy denotes those alkyl groups containing 1 to 3 carbon atoms attached to the rest of the molecule through an oxygen atom.
  • the C 1-3 alkoxy group includes C 1-2 , C 2-3 , C 3 and C 2 alkoxy groups and the like.
  • Examples of C 1-3 alkoxy include, but are not limited to, methoxy, ethoxy, propoxy (including n-propoxy and isopropoxy), and the like.
  • the number of atoms in a ring is generally defined as the number of ring members, eg, "5-7 membered ring” means a “ring” with 5-7 atoms arranged around it.
  • C n-n+m or C n -C n+m includes any specific instance of n to n+m carbons, for example C 1-12 includes 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 , also including any range from n to n+m, for example, C 1-12 includes C 1-3 , C 1-6 , C 1-9 , C 3-6 , C 3-9 , C 3-12 , C 6-9 , C 6-12 , and C 9-12 etc.; similarly, n to n +m means that the number of atoms on the ring is from n to n+m, for example, a 3-12-membered ring includes a 3-membered ring, a 4-membered ring, a 5-membered ring, a 6-membered ring, a 7-membered ring, an 8-member
  • C 3-6 cycloalkyl means a saturated cyclic hydrocarbon group composed of 3 to 6 carbon atoms, which includes monocyclic, bicyclic and tricyclic systems, wherein bicyclic and tricyclic systems include Spiral, parallel and bridged rings.
  • the C 3-6 cycloalkyl group includes C 3-5 , C 4-6 , C 4-5 or C 5-6 and the like; it may be monovalent, divalent or multivalent.
  • Examples of C 3-6 cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and the like.
  • C 3-10 cycloalkyl means a saturated cyclic hydrocarbon group composed of 3 to 10 carbon atoms, which includes monocyclic, bicyclic and tricyclic systems, wherein bicyclic and tricyclic systems include Spiral, parallel and bridged rings.
  • the C 3-10 cycloalkyl group includes C 3-8 , C 3-6 , C 3-5 , C 4-10 , C 4-8 , C 4-6 , C 4- 5 , C 5-8 or C 5-6 etc.; it may be monovalent, divalent or multivalent.
  • C 3-10 cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, norbornyl, [2.2.2]bicyclooctane, and the like.
  • the term "3-10 membered heterocycloalkyl" by itself or in combination with other terms denotes a saturated cyclic group consisting of 3 to 10 ring atoms, respectively, whose 1, 2, 3 or 4 ring atoms is a heteroatom independently selected from O, S, N, P, or Se, and the remainder is carbon atoms, wherein the nitrogen atom is optionally quaternized, and the nitrogen, sulfur, and phosphorus heteroatoms can be optionally oxidized (i.e., NO, S (O) p and P(O) p , where p is 1 or 2).
  • the 3-10 membered heterocycloalkyl group includes monocyclic, bicyclic and tricyclic ring systems, wherein bicyclic and tricyclic ring systems include spiro, merged and bridged rings.
  • a heteroatom may occupy the attachment position of the heterocycloalkyl group to the rest of the molecule.
  • the 3-10 membered heterocycloalkyl group includes 3-8 membered, 3-6 membered, 3-5 membered, 4-6 membered, 5-6 membered, 4-membered, 5-membered and 6-membered heterocycloalkyl groups.
  • 3-10 membered heterocycloalkyl groups include, but are not limited to, azetidinyl, oxetanyl, thietanyl, pyrrolidinyl, pyrazolidinyl, imidazolidinyl, tetrahydrothiophenyl ( Including tetrahydrothiophen-2-yl and tetrahydrothiophen-3-yl, etc.), tetrahydrofuranyl (including tetrahydrofuran-2-yl, etc.), tetrahydropyranyl, piperidinyl (including 1-piperidinyl, 2- piperidinyl and 3-piperidinyl, etc.), piperazinyl (including 1-piperazinyl and 2-piperazinyl, etc.), morpholinyl (including 3-morpholinyl and 4-morpholinyl, etc.), Dioxanyl, dithianyl, isoxazolidinyl, isothiazolidin,
  • the terms “5-6-membered heteroaryl ring” and “5-6-membered heteroaryl” in the present invention can be used interchangeably, and the term “5-6-membered heteroaryl” means that there are 5 to 6 ring atoms A monocyclic group with a conjugated ⁇ -electron system, 1, 2, 3 or 4 ring atoms are heteroatoms independently selected from O, S and N, and the rest are carbon atoms. Where the nitrogen atom is optionally quaternized, the nitrogen and sulfur heteroatoms may be optionally oxidized (ie, NO and S(O) p , where p is 1 or 2).
  • the 5-6 membered heteroaryl can be attached to the rest of the molecule through a heteroatom or a carbon atom.
  • the 5-6 membered heteroaryl includes 5 and 6 membered heteroaryl.
  • Examples of the 5-6 membered heteroaryl groups include, but are not limited to, pyrrolyl (including N-pyrrolyl, 2-pyrrolyl and 3-pyrrolyl, etc.), pyrazolyl (including 2-pyrazolyl and 3-pyrrolyl Azolyl, etc.), imidazolyl (including N-imidazolyl, 2-imidazolyl, 4-imidazolyl and 5-imidazolyl, etc.), oxazolyl (including 2-oxazolyl, 4-oxazolyl and 5- Oxazolyl, etc.), triazolyl (1H-1,2,3-triazolyl, 2H-1,2,3-triazolyl, 1H-1,2,4-triazolyl and 4H-1, 2,4-triazolyl, etc.
  • halogen or halogen by itself or as part of another substituent means a fluorine, chlorine, bromine or iodine atom.
  • the "connected atoms” may be the same atom or different atoms.
  • "two R 1 and the atoms connected to them form a C 3-6 cycloalkyl group", which may form a spiro ring, a bridged ring or a fused ring with ring A.
  • 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 listed below, the embodiments formed by combining them with other chemical synthesis methods, and the methods well known to those skilled in the art Equivalent alternatives, preferred embodiments include but are not limited to the examples of the present invention.
  • the structure of the compounds of the present invention can be confirmed by conventional methods known to those skilled in the art. If the present invention involves the absolute configuration of the compound, the absolute configuration can be confirmed by conventional technical means in the art. For example, in single crystal X-ray diffraction (SXRD), the cultured single crystal is collected with a Bruker D8 venture diffractometer to collect diffraction intensity data, the light source is CuK ⁇ radiation, and the scanning method is: After scanning and collecting relevant data, the absolute configuration can be confirmed by further analyzing the crystal structure by direct method (Shelxs97).
  • SXRD single crystal X-ray diffraction
  • the solvent used in the present invention is commercially available.
  • HATU stands for 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate
  • THF stands for tetrahydrofuran
  • Boc stands for tert-butyloxycarbonyl
  • n-BuLi stands for n-butyl lithium
  • HCl stands for hydrochloric acid
  • ACN stands for acetonitrile
  • MeOH stands for methanol
  • EtOAc or EA stands for ethyl acetate
  • TEA stands for triethylamine
  • DIEA or DIPEA stands for N,N-di Isopropylethylamine
  • DMF stands for N,N-dimethylformamide
  • DMSO stands for dimethyl sulfoxide
  • HPLC stands for high performance liquid chromatography
  • TLC stands for thin layer chromatography
  • PEG stands for polyethylene glycol.
  • reaction solution was poured into 5% citric acid solution for separation, the aqueous phase was extracted with ethyl acetate (20 mL ⁇ 2), the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product.
  • Lithium hydroxide monohydrate (51.25 mg, 1.22 mmol) was added to a solution of starting material 1-9 (0.3 g, 814.19 ⁇ mol) in tetrahydrofuran (2 mL) and water (1 mL), and reacted at 20° C. for 16 hr.
  • Compounds 1-10 were obtained.
  • reaction solution was extracted by adding ethyl acetate (10 mL) and 1M hydrochloric acid (5 mL), and the organic layer was washed with saturated aqueous sodium bicarbonate (10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure.
  • Dissolve compound 2-10 (160 mg, 409.81 ⁇ mol) and 1-5 (160.23 mg, 491.78 ⁇ mol) in acetonitrile (2 mL), cool to 0 ° C, add 1-ethyl-(3-dimethylaminopropyl) Carbodimethimide hydrochloride (79.35mg, 413.91 ⁇ mol) was added dropwise to pyridine (129.66mg, 1.64mmol), and the reaction was stirred at 0°C for 2h.
  • compound 4-2 (8g, 30.97mmol) and dichloromethane (80mL) were added to a dry three-necked flask, and stirring was started; then sodium bicarbonate (2.73g, 32.52mmol, 1.26mL) and Dessert -Martin periodinane (13.79g, 32.52mmol)
  • the reaction solution changed from cloudy to clear, replaced with nitrogen 3 times, stirred for about 20min, the reaction solution turned into a white suspension, and stirred at 15°C for 16hr.
  • the substrate compound 4-5 (0.2 g, 532.74 ⁇ mol) was dissolved in hydrochloric acid/ethyl acetate (4M, 10.00 mL), and stirring was continued for 1 h. Direct concentration gave the hydrochloride salt of compound 4-6.
  • the substrate compound 3-4 (0.35 g, 783.74 ⁇ mol) was dissolved in N,N-dimethylformamide (10 mL), added HATU (447.00 mg, 1.18 mmol), and stirred for 0.5 h.
  • the hydrochloride salt of compound 4-6 (258.92mg) and diisopropylethylamine (405.17mg, 3.13mmol) were added to the reaction system, and stirred at 20°C for 1 hour.
  • the substrate compound 4-9 trifluoroacetate (0.034g, 56.50 ⁇ mol) was dissolved in dichloromethane (1mL), cooled to 0°C, pyridine (31.29mg, 395.52 ⁇ mol) and trifluoroacetic anhydride (47.47mg ,226.01 ⁇ mol), continue stirring for 0.2h.
  • the reaction was washed with saturated sodium bicarbonate solution (10 mL), extracted three times with dichloromethane (10 mL), washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered and concentrated.
  • reaction bottle was replaced with nitrogen, added n-hexane (4000mL), added diethyl zinc solution (1M n-hexane solution, 1.05L), cooled to 0°C, stirred at 0°C for 0.2hr, until the reaction system was clear, added trifluoro Boronium diethyl ether (223.82g, 1.58mol), stirred at 0°C for 0.5hr, added diiodomethane (563.15g, 2.10mol) at 0°C, stirred at 0°C for 0.5hr, added compound 8-1 (50g, 131.41 mmol), raised to 40°C and stirred for 16hr.
  • the compound was dissolved in DMSO, and was diluted with Echo655 according to the 3-fold gradient according to the concentration to be tested, and 10 concentration points were added to the 384-well plate in duplicate wells for each concentration.
  • Dilute Mpro protein and substrate with test buffer 100mM NaCl, 20mM Tris-HCL, 1mM EDTA
  • add Mpro protein to a 384-well test plate and incubate with compound for 30min at room temperature, then add substrate, Mpro protein
  • Inhibition rate% [(Compound-BG Compound )-(ZPE-BG ZPE )]/[(HPE-BG HPE )-(ZPE-BG ZPE )]*100%
  • ZPE no inhibition control, containing 25nM Mpro protein + 25 ⁇ M substrate, no compound
  • test compound well containing 25nM Mpro protein + 25 ⁇ M substrate + compound
  • BG Background control wells containing 25 ⁇ M substrate+compound without Mpro protein
  • the compound of the present invention has better anti-new coronavirus Mpro protease activity in vitro.
  • MRC5 cells and coronavirus HCoV OC43 were purchased from ATCC.
  • MRC5 cells were cultured in MEM (Sigma) medium supplemented with 10% fetal bovine serum (Excell), 1% double antibody (Hyclone), 1% L-glutamine (Gibco) and 1% non-essential amino acids (Gibco).
  • MEM (Sigma) medium supplemented with 5% fetal bovine serum (Excell), 1% double antibody (Hyclone), 1% L-glutamine (Gibco) and 1% non-essential amino acid (Gibco) was used as the experimental medium.
  • Cells were seeded into 96 microwell plates at a certain density (Table 2) and cultured overnight in a 5% CO 2 , 37° C. incubator. On the second day, the compound after doubling dilution (8 concentration points, duplicate wells) was added, 50 ⁇ L per well. Then the diluted virus was added to the cells at 100 TCID 50 per well, 50 ⁇ L per well. Set up cell control (cells, no compound treatment or virus infection), virus control (cells infected with virus, no compound treatment) and culture medium control (only culture medium). The final volume of the experimental culture solution was 200 ⁇ L, and the final concentration of DMSO in the culture solution was 0.5%. The cells were cultured in a 5% CO 2 , 33°C incubator for 5 days. Cell viability was detected using the cell viability assay kit CellTiter Glo (Promega). Cytotoxicity experiments were performed under the same conditions as antiviral experiments, but without virus infection.
  • the antiviral activity and cytotoxicity of the compound are represented by the inhibitory rate (%) and cell viability (%) of the compound on the cytopathic effect caused by the virus at different concentrations, respectively. Calculated as follows:
  • Inhibition rate (%) (test hole reading value-virus control average value)/(cell control average value-virus control average value) ⁇ 100
  • GraphPad Prism was used to conduct nonlinear fitting analysis on the inhibition rate and cell viability of the compound, and calculate the half effective concentration (EC 50 ) and half cytotoxic concentration (CC 50 ) of the compound.
  • the compound of the present invention has good in vitro anti-coronavirus activity at the cellular level, and has no cytotoxicity.
  • C57BL/6J suckling mice were infected with a lethal dose of coronavirus by intranasal instillation, and the suckling mice were treated with vehicle (5% DMSO+40% PEG400+55% water) and the compound of the present invention 2 hours before infection.
  • vehicle 5% DMSO+40% PEG400+55% water
  • the compound of the present invention 2 hours before infection.
  • the body weight, health status and survival of the suckling mice were monitored daily to evaluate the protective effect of the compound of the present invention on the suckling mice at different doses.
  • the compound of the present invention has good anti-coronavirus efficacy in vivo.

Abstract

A short peptide compound containing β-aminoketone and the use thereof, and specifically disclosed is a compound represented by formula (II) or a pharmaceutically acceptable salt thereof.

Description

含β-胺基酮的短肽化合物及其应用Short peptide compound containing β-aminoketone and its application
本申请主张如下优先权This application claims the following priority
申请号：CN2021115809216，申请日：2021年12月22日；Application number: CN2021115809216, application date: December 22, 2021;
申请号：CN2022100451335，申请日：2022年01月14日；Application number: CN2022100451335, application date: January 14, 2022;
申请号：CN2022102216236，申请日：2022年03月08日；Application number: CN2022102216236, application date: March 8, 2022;
申请号：CN2022110234109，申请日：2022年08月24日。Application number: CN2022110234109, application date: August 24, 2022.
技术领域technical field
本发明涉及医药化学技术领域，特别涉及一种含β-胺基酮的短肽化合物及其应用。The invention relates to the technical field of medicinal chemistry, in particular to a short peptide compound containing β-aminoketone and its application.
背景技术Background technique
(3CL pro)蛋白酶属于半胱氨酸蛋白酶。其底物结合位点高度保守且具有相似的催化机制，是催化RNA病毒前体蛋白裂解的关键蛋白酶，对病毒的复制起到重要作用。作为冠状病毒复制的关键酶，3CL蛋白酶(3CL pro)是治疗包括COVID-19在内的多种冠状病毒引起的疾病的重要靶点。利用3CL蛋白酶结构的特点，开发治疗新冠病毒感染的药物具有重大的临床价值。 (3CL pro ) protease belongs to cysteine proteases. Its substrate binding site is highly conserved and has a similar catalytic mechanism. It is a key protease that catalyzes the cleavage of RNA virus precursor protein and plays an important role in virus replication. As a key enzyme for coronavirus replication, 3CL protease (3CL pro ) is an important target for the treatment of diseases caused by various coronaviruses including COVID-19. Using the structural characteristics of 3CL protease to develop drugs for the treatment of new coronavirus infection has great clinical value.
发明内容Contents of the invention
本发明提供式(II)所示化合物或其药学上可接受的盐，The present invention provides a compound represented by formula (II) or a pharmaceutically acceptable salt thereof,
Figure PCTCN2022140910-appb-000001
Figure PCTCN2022140910-appb-000001
其中，in,
R 1各自独立地选自卤素、CN、OH、NH 2、C 1-3烷基和C 1-3烷氧基，所述C 1-3烷基或C 1-3烷氧基各自独立地任选被1、2或3个卤素取代； Each R 1 is independently selected from halogen, CN, OH, NH 2 , C 1-3 alkyl and C 1-3 alkoxy, and each of the C 1-3 alkyl or C 1-3 alkoxy is independently optionally substituted by 1, 2 or 3 halogens;
或者，or,
两个R 1及它们相连的原子形成C 3-6环烷基，所述C 3-6环烷基任选被1、2或3个R a取代； Two R 1 and the atoms connected to them form a C 3-6 cycloalkyl group, and the C 3-6 cycloalkyl group is optionally substituted by 1, 2 or 3 R a ;
R a各自独立地选自卤素和C 1-3烷基； R a is each independently selected from halogen and C 1-3 alkyl;
n选自0、1、2或3；n is selected from 0, 1, 2 or 3;
R 2选自叔丁基、C 3-10环烷基、3-10元杂环烷基和苯基，所述C 3-10环烷基、3-10元杂环烷基和苯基各自独立地任选被1、2或3个R b取代； R 2 is selected from tert-butyl, C 3-10 cycloalkyl, 3-10 membered heterocycloalkyl and phenyl, each of said C 3-10 cycloalkyl, 3-10 membered heterocycloalkyl and phenyl independently optionally substituted by 1, 2 or 3 R ;
R b各自独立地选自卤素和C 1-3烷基； R b are each independently selected from halogen and C 1-3 alkyl;
R 3选自C 1-3烷基、C 1-3烷氧基、-CH 2R 4和-CH 2OR 4，所述C 1-3烷基和C 1-3烷氧基各自独立地任选被1、2或3个卤素取代； R 3 is selected from C 1-3 alkyl, C 1-3 alkoxy, -CH 2 R 4 and -CH 2 OR 4 , and the C 1-3 alkyl and C 1-3 alkoxy are each independently optionally substituted by 1, 2 or 3 halogens;
R 4选自苯基和5-6元杂芳基，所述苯基和5-6元杂芳基各自独立地任选被1、2或3个R取代； R is selected from phenyl and 5-6 membered heteroaryl, each of which is independently optionally substituted by 1, 2 or 3 R;
R选自卤素和C 1-3烷基； R is selected from halogen and C 1-3 alkyl;
T 1选自O和S； T1 is selected from O and S;
环A选自
Figure PCTCN2022140910-appb-000002
Ring A is selected from
Figure PCTCN2022140910-appb-000002
A 1、A 2、A 3和A 4各自独立地选自CH和N。 A 1 , A 2 , A 3 and A 4 are each independently selected from CH and N.
本发明的一些方案中，上述R 1选自F和甲基，其他变量如本发明所定义。 In some solutions of the present invention, the above-mentioned R 1 is selected from F and methyl, and other variables are as defined in the present invention.
本发明的一些方案中，上述两个R 1及它们相连的原子形成C 3-6环烷基，其他变量如本发明所定义。 In some solutions of the present invention, the above two R 1 and the atoms connected to them form a C 3-6 cycloalkyl group, and other variables are as defined in the present invention.
本发明的一些方案中，上述两个R 1及它们相连的原子形成环丙基，其他变量如本发明所定义。 In some solutions of the present invention, the above two R 1 and the atoms connected to them form a cyclopropyl group, and other variables are as defined in the present invention.
本发明的一些方案中，上述结构单元
Figure PCTCN2022140910-appb-000003
选自
Figure PCTCN2022140910-appb-000004
其他变量如本发明所定义。 In some solutions of the present invention, the above structural units
Figure PCTCN2022140910-appb-000003
selected from
Figure PCTCN2022140910-appb-000004
Other variables are as defined herein.
本发明的一些方案中，上述R 2选自叔丁基和金刚烷基，所述金刚烷基任选被1、2或3个R b取代，其他变量如本发明所定义。 In some solutions of the present invention, the above-mentioned R 2 is selected from tert-butyl and adamantyl, and the adamantyl is optionally substituted by 1, 2 or 3 R b , and other variables are as defined in the present invention.
本发明的一些方案中，上述R 2选自叔丁基和金刚烷基，其他变量如本发明所定义。 In some solutions of the present invention, the above-mentioned R 2 is selected from tert-butyl and adamantyl, and other variables are as defined in the present invention.
本发明的一些方案中，上述R 2选自叔丁基、
Figure PCTCN2022140910-appb-000005
其他变量如本发明所定义。 In some schemes of the present invention, above-mentioned R 2 is selected from tert-butyl,
Figure PCTCN2022140910-appb-000005
Other variables are as defined herein.
本发明的一些方案中，上述结构单元
Figure PCTCN2022140910-appb-000006
选自
Figure PCTCN2022140910-appb-000007
Figure PCTCN2022140910-appb-000008
其他变量如本发明所定义。 In some solutions of the present invention, the above structural units
Figure PCTCN2022140910-appb-000006
selected from
Figure PCTCN2022140910-appb-000007
Figure PCTCN2022140910-appb-000008
Other variables are as defined herein.
本发明的一些方案中，上述R 3选自-CF 3，其他变量如本发明所定义。 In some solutions of the present invention, the above-mentioned R 3 is selected from -CF 3 , and other variables are as defined in the present invention.
本发明还提供了(II-1)、(II-2)和(II-3)所示的化合物或其药学上可接受的盐，The present invention also provides compounds represented by (II-1), (II-2) and (II-3) or pharmaceutically acceptable salts thereof,
Figure PCTCN2022140910-appb-000009
Figure PCTCN2022140910-appb-000009
其中，t选自0、1和2；Wherein, t is selected from 0, 1 and 2;
R 1、R b、A 1、A 2、A 3、A 4、T 1和n如本发明所定义。 R 1 , R b , A 1 , A 2 , A 3 , A 4 , T 1 and n are as defined in the present invention.
本发明还提供式(I)所示化合物或其药学上可接受的盐，The present invention also provides a compound represented by formula (I) or a pharmaceutically acceptable salt thereof,
Figure PCTCN2022140910-appb-000010
Figure PCTCN2022140910-appb-000010
其中，in,
R 1各自独立地选自卤素、CN、OH、NH 2、C 1-3烷基或C 1-3烷氧基，所述C 1-3烷基或C 1-3烷氧基任选各自独立地被1、2或3个卤素取代； Each R 1 is independently selected from halogen, CN, OH, NH 2 , C 1-3 alkyl or C 1-3 alkoxy, and the C 1-3 alkyl or C 1-3 alkoxy is optionally each independently substituted by 1, 2 or 3 halogens;
或者，or,
两个R 1及它们相连的原子形成C 3-6环烷基，所述C 3-6环烷基任选被1、2或3个R a取代； Two R 1 and the atoms connected to them form a C 3-6 cycloalkyl group, and the C 3-6 cycloalkyl group is optionally substituted by 1, 2 or 3 R a ;
n选自0、1、2或3；n is selected from 0, 1, 2 or 3;
R a各自独立地选自卤素或C 1-3烷基； R a is each independently selected from halogen or C 1-3 alkyl;
R 2选自叔丁基、C 3-10环烷基、3-10元杂环烷基或苯基，所述C 3-10环烷基、3-10元杂环烷基和苯基任选被1、2或3个R b取代； R 2 is selected from tert-butyl, C 3-10 cycloalkyl, 3-10 membered heterocycloalkyl or phenyl, the C 3-10 cycloalkyl, 3-10 membered heterocycloalkyl and phenyl are any optionally replaced by 1, 2 or 3 R b ;
R b各自独立地选自卤素或C 1-3烷基； Each R b is independently selected from halogen or C 1-3 alkyl;
R 3选自C 1-3烷基、C 1-3烷氧基、-CH 2R 4或-CH 2OR 4，所述C 1-3烷基和C 1-3烷氧基任选被1、2或3个卤素取代； R 3 is selected from C 1-3 alkyl, C 1-3 alkoxy, -CH 2 R 4 or -CH 2 OR 4 , and the C 1-3 alkyl and C 1-3 alkoxy are optionally 1, 2 or 3 halogen substitutions;
R 4选自苯基或5-6元杂芳基，所述苯基和5-6元杂芳基任选独立地被1、2或3个R取代； R 4 is selected from phenyl or 5-6 membered heteroaryl, said phenyl and 5-6 membered heteroaryl are optionally substituted independently by 1, 2 or 3 R;
R选自卤素或C 1-3烷基； R is selected from halogen or C 1-3 alkyl;
A 1、A 2、A 3和A 4各自独立地选自CH或N，且A 1、A 2、A 3和A 4中的0、1或2个选自N，其余选自CH。 A 1 , A 2 , A 3 and A 4 are each independently selected from CH or N, and 0, 1 or 2 of A 1 , A 2 , A 3 and A 4 are selected from N, and the rest are selected from CH.
本发明的一些方案中，上述R 1选自F或甲基，其他变量如本发明所定义。 In some solutions of the present invention, the above-mentioned R 1 is selected from F or methyl, and other variables are as defined in the present invention.
本发明的一些方案中，上述两个R 1及它们相连的原子形成C 3-6环烷基，其他变量如本发明所定义。 In some solutions of the present invention, the above two R 1 and the atoms connected to them form a C 3-6 cycloalkyl group, and other variables are as defined in the present invention.
本发明的一些方案中，上述两个R 1及它们相连的原子形成环丙基，其他变量如本发明所定义。 In some solutions of the present invention, the above two R 1 and the atoms connected to them form a cyclopropyl group, and other variables are as defined in the present invention.
本发明的一些方案中，上述结构单元
Figure PCTCN2022140910-appb-000011
选自
Figure PCTCN2022140910-appb-000012
其他变量如本发明所定义。 In some solutions of the present invention, the above structural units
Figure PCTCN2022140910-appb-000011
selected from
Figure PCTCN2022140910-appb-000012
Other variables are as defined herein.
本发明的一些方案中，上述R 2选自叔丁基，其他变量如本发明所定义。 In some solutions of the present invention, the above-mentioned R 2 is selected from tert-butyl, and other variables are as defined in the present invention.
本发明的一些方案中，上述R 3选自-CF 3，其他变量如本发明所定义。 In some solutions of the present invention, the above-mentioned R 3 is selected from -CF 3 , and other variables are as defined in the present invention.
本发明还提供了(I-1)所示的化合物或其药学上可接受的盐，The present invention also provides the compound represented by (I-1) or a pharmaceutically acceptable salt thereof,
Figure PCTCN2022140910-appb-000013
Figure PCTCN2022140910-appb-000013
其中，R 1和n如本发明所定义。 Wherein, R 1 and n are as defined in the present invention.
本发明还有一些方案是由上述各变量任意组合而来。Some schemes of the present invention are formed by any combination of the above-mentioned variables.
本发明提供下式化合物或其药学上可接受的盐：The present invention provides a compound of the following formula or a pharmaceutically acceptable salt thereof:
Figure PCTCN2022140910-appb-000014
Figure PCTCN2022140910-appb-000014
Figure PCTCN2022140910-appb-000015
Figure PCTCN2022140910-appb-000015
本发明提供了一种药物组合物，其含有治疗有效量的上述的化合物或其药学上可接受的盐。The present invention provides a pharmaceutical composition, which contains a therapeutically effective amount of the above-mentioned compound or a pharmaceutically acceptable salt thereof.
本发明还提供了上述的化合物或其药学上可接受的盐在制备治疗冠状病毒感染的药物中的应用。The present invention also provides the application of the above-mentioned compound or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating coronavirus infection.
本发明还提供了上述的化合物或其药学上可接受的盐或上述的组合物在制备治疗冠状病毒感染的药物中的应用。The present invention also provides the application of the above-mentioned compound or a pharmaceutically acceptable salt thereof or the above-mentioned composition in the preparation of a medicament for treating coronavirus infection.
本发明的一些方案中，上述的冠状病毒感染选自COVID-19。In some aspects of the present invention, the aforementioned coronavirus infection is selected from COVID-19.
技术效果technical effect
本发明化合物具有较好的体外抗新型冠状病毒Mpro蛋白酶活性；较好的细胞水平的体外抗冠状病毒活性，且无细胞毒性。The compound of the present invention has better in vitro anti-new coronavirus Mpro protease activity; better in vitro anti-coronavirus activity at the cellular level, and has no cytotoxicity.
定义和说明Definition and Description
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。Unless otherwise stated, the following terms and phrases used herein are intended to have the following meanings. A specific term or phrase should not be considered indeterminate or unclear if it is not specifically defined, but should be understood according to its ordinary meaning. When a trade name appears herein, it is intended to refer to its corresponding trade name or its active ingredient.
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。The term "pharmaceutically acceptable" as used herein refers to those compounds, materials, compositions and/or dosage forms, which are suitable for use in contact with human and animal tissues within the scope of sound medical judgment , without undue toxicity, irritation, allergic reaction or other problems or complications, commensurate with a reasonable benefit/risk ratio.
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、有机胺或镁盐或类似的盐。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。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。The term "pharmaceutically acceptable salt" refers to a salt of a compound of the present invention, which is prepared from a compound having a specific substituent found in the present invention and a relatively non-toxic acid or base. When compounds of the present invention contain relatively acidic functional groups, base addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of base, either neat solution or in a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amine or magnesium salts or similar salts. When compounds of the present invention contain relatively basic functionalities, acid addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of the acid, either neat solution or in a suitable inert solvent. Certain specific compounds of the present invention contain basic and acidic functional groups and can thus be converted into either base or acid addition salts.
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。The pharmaceutically acceptable salts of the present invention can be synthesized from the parent compound containing acid groups or bases by conventional chemical methods. In general, such salts are prepared by reacting the free acid or base form of these compounds with a stoichiometric amount of the appropriate base or acid in water or an organic solvent or a mixture of both.
除非另有说明，术语“异构体”意在包括几何异构体、顺反异构体、立体异构体、对映异构体、旋光异构体、非对映异构体和互变异构体。Unless otherwise stated, the term "isomer" is intended to include geometric isomers, cis-trans isomers, stereoisomers, enantiomers, optical isomers, diastereoisomers and interconversions isomer.
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。The compounds of the invention may exist in particular geometric or stereoisomeric forms. The present invention contemplates all such compounds, including cis and trans isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers isomers, (D)-isomers, (L)-isomers, and their racemic and other mixtures, such as enantiomerically or diastereomerically enriched mixtures, all of which are subject to the present within the scope of the invention. Additional asymmetric carbon atoms may be present in substituents such as alkyl groups. All such isomers, as well as mixtures thereof, are included within the scope of the present invention.
除非另有说明，术语“对映异构体”或者“旋光异构体”是指互为镜像关系的立体异构体。Unless otherwise stated, the terms "enantiomer" or "optical isomer" refer to stereoisomers that are mirror images of each other.
除非另有说明，术语“顺反异构体”或者“几何异构体”系由因双键或者成环碳原子单键不能自由旋转而引起。Unless otherwise stated, the terms "cis-trans isomers" or "geometric isomers" arise from the inability to rotate freely due to the double bond or the single bond of the carbon atoms forming the ring.
除非另有说明，术语“非对映异构体”是指分子具有两个或多个手性中心，并且分子间为非镜像的关系的立体异构体。Unless otherwise indicated, the term "diastereoisomer" refers to stereoisomers whose molecules have two or more chiral centers and which are not mirror images of the molecules.
除非另有说明，“(+)”表示右旋，“(-)”表示左旋，“(±)”表示外消旋。Unless otherwise specified, "(+)" means dextrorotation, "(-)" means levorotation, and "(±)" means racemization.
除非另有说明，用楔形实线键
Figure PCTCN2022140910-appb-000016
和楔形虚线键
Figure PCTCN2022140910-appb-000017
表示一个立体中心的绝对构型，用直形实线键
Figure PCTCN2022140910-appb-000018
和直形虚线键
Figure PCTCN2022140910-appb-000019
表示立体中心的相对构型，用波浪线
Figure PCTCN2022140910-appb-000020
表示楔形实线键
Figure PCTCN2022140910-appb-000021
或楔形虚线键
Figure PCTCN2022140910-appb-000022
或用波浪线
Figure PCTCN2022140910-appb-000023
表示直形实线键
Figure PCTCN2022140910-appb-000024
或直形虚线键
Figure PCTCN2022140910-appb-000025
Unless otherwise noted, keys with wedge-shaped solid lines
Figure PCTCN2022140910-appb-000016
and dotted wedge keys
Figure PCTCN2022140910-appb-000017
Indicates the absolute configuration of a stereocenter, with a straight solid-line bond
Figure PCTCN2022140910-appb-000018
and straight dashed keys
Figure PCTCN2022140910-appb-000019
Indicates the relative configuration of the stereocenter, with a wavy line
Figure PCTCN2022140910-appb-000020
Indicates wedge-shaped solid-line bond
Figure PCTCN2022140910-appb-000021
or dotted wedge key
Figure PCTCN2022140910-appb-000022
or with tilde
Figure PCTCN2022140910-appb-000023
Indicates a straight solid line key
Figure PCTCN2022140910-appb-000024
or straight dotted key
Figure PCTCN2022140910-appb-000025
除非另有规定，当某一基团具有一个或多个可连接位点时，该基团的任意一个或多个位点可以通过化学键与其他基团相连。所述位点与其他基团连接的化学键可以用直形实线键
Figure PCTCN2022140910-appb-000026
直形虚线键
Figure PCTCN2022140910-appb-000027
或波浪线
Figure PCTCN2022140910-appb-000028
表示。例如-OCH 3中的直形实线键表示通过该基团中的氧原子与其他基团相连；
Figure PCTCN2022140910-appb-000029
中的直形虚线键表示通过该基团中的氮原子的两端与其他基团相连；
Figure PCTCN2022140910-appb-000030
中的波浪线表示通过该苯基集团中的1和2位的碳原子与其他基团相连。 Unless otherwise specified, when a group has one or more linkable sites, any one or more sites of the group can be linked to other groups through chemical bonds. The chemical bonds that the site is connected with other groups can use straight solid line bonds
Figure PCTCN2022140910-appb-000026
Straight dotted key
Figure PCTCN2022140910-appb-000027
or tilde
Figure PCTCN2022140910-appb-000028
express. For example, the straight-shaped solid-line bond in -OCH3 indicates that it is connected to other groups through the oxygen atom in the group;
Figure PCTCN2022140910-appb-000029
The straight dotted line bond in indicates that the two ends of the nitrogen atom in the group are connected to other groups;
Figure PCTCN2022140910-appb-000030
The wavy lines in indicate that the carbon atoms at positions 1 and 2 in the phenyl group are connected to other groups.
除非另有说明，术语“富含一种异构体”、“异构体富集”、“富含一种对映体”或者“对映体富集”指其中一种异构体或对映体的含量小于100％，并且，该异构体或对映体的含量大于等于60％，或者大于等于70％，或者大于等于80％，或者大于等于90％，或者大于等于95％，或者大于等于96％，或者大于等于97％，或者大于等于98％，或者大于等于99％，或者大于等于99.5％，或者大于等于99.6％，或者大于等于99.7％，或者大于等于99.8％，或者大于等于99.9％。Unless otherwise stated, the terms "enriched in an isomer", "enriched in an isomer", "enriched in an enantiomer" or "enantiomerically enriched" refer to one of the isomers or enantiomers The content of the enantiomer is less than 100%, and the content of the isomer or enantiomer is greater than or equal to 60%, or greater than or equal to 70%, or greater than or equal to 80%, or greater than or equal to 90%, or greater than or equal to 95%, or Greater than or equal to 96%, or greater than or equal to 97%, or greater than or equal to 98%, or greater than or equal to 99%, or greater than or equal to 99.5%, or greater than or equal to 99.6%, or greater than or equal to 99.7%, or greater than or equal to 99.8%, or greater than or equal to 99.9%.
除非另有说明，术语“异构体过量”或“对映体过量”指两种异构体或两种对映体相对百分数之间的差值。例如，其中一种异构体或对映体的含量为90％，另一种异构体或对映体的含量为10％，则异构体或对映体过量(ee值)为80％。Unless otherwise stated, the terms "isomer excess" or "enantiomeric excess" refer to the difference between the relative percentages of two isomers or two enantiomers. For example, if the content of one isomer or enantiomer is 90% and the other isomer or enantiomer is 10%, then the isomer or enantiomeric excess (ee value) is 80% .
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)。Optically active (R)- and (S)-isomers as well as D and L-isomers can be prepared by chiral synthesis or chiral reagents or other conventional techniques. If one enantiomer of a compound of the invention is desired, it can be prepared by asymmetric synthesis or derivatization with chiral auxiliary agents, wherein the resulting diastereomeric mixture is separated and the auxiliary group is cleaved to provide pure desired enantiomer. Alternatively, when the molecule contains a basic functional group (such as an amino group) or an acidic functional group (such as a carboxyl group), a diastereoisomeric salt is formed with an appropriate optically active acid or base, and then a diastereomeric salt is formed by a conventional method known in the art. Diastereomeric resolution is performed and the pure enantiomers are recovered. Furthermore, the separation of enantiomers and diastereomers is usually accomplished by the use of chromatography using chiral stationary phases, optionally in combination with chemical derivatization methods (e.g. amines to amino groups formate).
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)。又例如，可用重氢取代氢形成氘代药物，氘与碳构成的键比普通氢与碳构成的键更坚固，相比于未氘化药物，氘代药物有降低毒副作用、增加药物稳定性、增强疗效、延长药物生物半衰期等优势。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。 The compounds of the present invention may contain unnatural proportions of atomic isotopes at one or more of the atoms that constitute the compounds. For example, compounds may be labeled with radioactive isotopes such as tritium ( 3 H), iodine-125 ( 125 I) or C-14 ( 14 C). For another example, heavy hydrogen can be used to replace hydrogen to form deuterated drugs. The bond formed by deuterium and carbon is stronger than the bond formed by ordinary hydrogen and carbon. Compared with non-deuterated drugs, deuterated drugs can reduce toxic side effects and increase drug stability. , enhance the efficacy, prolong the biological half-life of drugs and other advantages. All changes in isotopic composition of the compounds of the invention, whether radioactive or not, are included within the scope of the invention.
术语“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。The term "optional" or "optionally" means that the subsequently described event or circumstance can but need not occur, and that the description includes instances where said event or circumstance occurs and instances where said event or circumstance does not occur .
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，取代基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。当取代基为氧(即＝O)时，意味着两个氢原子被取代。氧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。The term "substituted" means that any one or more hydrogen atoms on a specified atom are replaced by a substituent, which may include deuterium and hydrogen variants, as long as the valence of the specified atom is normal and the substituted compound is stable. When a substituent is oxygen (ie =0), it means that two hydrogen atoms are replaced. Oxygen substitution does not occur on aromatic groups. The term "optionally substituted" means that it may or may not be substituted, and unless otherwise specified, the type and number of substituents may be arbitrary on a chemically realizable basis.
“药物组合物”表示含有一种或多种本文所述化合物或其生理学上/可药用的盐或前体药物与其他化学组分的混合物，以及其他组分例如生理学/可药用的载体和赋形剂。药物组合物的目的是促进对生物体的给药，利于活性成分的吸收进而发挥生物活性。"Pharmaceutical composition" means a mixture containing one or more compounds described herein, or a physiologically/pharmaceutically acceptable salt or prodrug thereof, and other chemical components, and other components such as a physiologically/pharmaceutically acceptable carrier and excipients. The purpose of the pharmaceutical composition is to promote the administration to the organism, facilitate the absorption of the active ingredient and thus exert biological activity.
术语“治疗有效量”意指(i)治疗或预防特定疾病、病况或障碍，(ii)减轻、改善或消除特定疾病、病况或障碍的一种或多种症状，或(iii)预防或延迟本文中所述的特定疾病、病况或障碍的一种或多种症状发作的本发明化合物的用量。构成“治疗有效量”的本发明化合物的量取决于该化合物、疾病状态及其严重性、给药方式以及待被治疗的哺乳动物的年龄而改变，但可例行性地由本领域技术人员根据其自身的知识及本公开内容而确定。The term "therapeutically effective amount" means (i) treating or preventing a particular disease, condition or disorder, (ii) alleviating, ameliorating or eliminating one or more symptoms of a particular disease, condition or disorder, or (iii) preventing or delaying The amount of a compound of the invention for the onset of one or more symptoms of a particular disease, condition or disorder described herein. The amount of a compound of the invention that constitutes a "therapeutically effective amount" will vary depending on the compound, the disease state and its severity, the mode of administration, and the age of the mammal to be treated, but can be routinely determined by one skilled in the art according to its own knowledge and this disclosure.
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。When any variable (eg, R) occurs more than once in the composition or structure of a compound, its definition is independent at each occurrence. Thus, for example, if a group is substituted with 0-2 R, said group may optionally be substituted with up to two R, with independent options for each occurrence of R. Also, combinations of substituents and/or variations thereof are permissible only if such combinations result in stable compounds.
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。 When the number of a linking group is 0, such as -(CRR) 0 -, it means that the linking group is a single bond.
当一个取代基数量为0时，表示该取代基是不存在的，比如-A-(R) 0表示该结构实际上是-A。 When the number of a substituent is 0, it means that the substituent does not exist, such as -A-(R) 0 means that the structure is actually -A.
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。When a substituent is vacant, it means that the substituent does not exist. For example, when X in A-X is vacant, it means that the structure is actually A.
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。When one of the variables is selected from a single bond, it means that the two groups connected are directly connected. For example, when L in A-L-Z represents a single bond, it means that the structure is actually A-Z.
当一个取代基的键可以交叉连接到一个环上的两个以上原子时，这种取代基可以与这个环上的任意原子相键合，例如，结构单元
Figure PCTCN2022140910-appb-000031
表示其取代基R可在环己基或者环己二烯上的任意一个位置发生取代。当所列举的取代基中没有指明其通过哪一个原子连接到被取代的基团上时，这种取代基可以通过 其任何原子相键合，例如，吡啶基作为取代基可以通过吡啶环上任意一个碳原子连接到被取代的基团上。 A substituent can be bonded to any atom on a ring when the bond of a substituent can cross-link two or more atoms on the ring, e.g., structural unit
Figure PCTCN2022140910-appb-000031
It means that the substituent R can be substituted at any position on cyclohexyl or cyclohexadiene. When the enumerated substituent does not indicate which atom it is connected to the substituted group, this substituent can be bonded through any atom, for example, pyridyl as a substituent can be connected to any atom on the pyridine ring. The carbon atom is attached to the group being substituted.
当所列举的连接基团没有指明其连接方向，其连接方向是任意的，例如，
Figure PCTCN2022140910-appb-000032
中连接基团L为-M-W-，此时-M-W-既可以按与从左往右的读取顺序相同的方向连接环A和环A构成
Figure PCTCN2022140910-appb-000033
也可以按照与从左往右的读取顺序相反的方向连接环A和环A构成
Figure PCTCN2022140910-appb-000034
所述连接基团、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。 When the linking group listed does not indicate its linking direction, its linking direction is arbitrary, for example,
Figure PCTCN2022140910-appb-000032
The linking group L is -MW-, at this time -MW- can be connected to ring A and ring A in the same direction as the reading order from left to right to form
Figure PCTCN2022140910-appb-000033
It can also be formed by linking loop A and loop A in the opposite direction to the reading order from left to right
Figure PCTCN2022140910-appb-000034
Combinations of the described linking groups, substituents and/or variations thereof are permissible only if such combinations result in stable compounds.
除非另有规定，当某一基团具有一个或多个可连接位点时，该基团的任意一个或多个位点可以通过化学键与其他基团相连。当该化学键的连接方式是不定位的，且可连接位点存在H原子时，则连接化学键时，该位点的H原子的个数会随所连接化学键的个数而对应减少变成相应价数的基团。所述位点与其他基团连接的化学键可以用直形实线键
Figure PCTCN2022140910-appb-000035
直形虚线键
Figure PCTCN2022140910-appb-000036
或波浪线
Figure PCTCN2022140910-appb-000037
表示。例如-OCH 3中的直形实线键表示通过该基团中的氧原子与其他基团相连；
Figure PCTCN2022140910-appb-000038
中的直形虚线键表示通过该基团中的氮原子的两端与其他基团相连；
Figure PCTCN2022140910-appb-000039
中的波浪线表示通过该苯基基团中的1和2位碳原子与其他基团相连。 Unless otherwise specified, when a group has one or more linkable sites, any one or more sites of the group can be linked to other groups through chemical bonds. When the connection method of the chemical bond is not positioned, and there is an H atom at the connectable site, when the chemical bond is connected, the number of H atoms at the site will decrease correspondingly with the number of chemical bonds connected to become the corresponding valence group. The chemical bonds that the site is connected with other groups can use straight solid line bonds
Figure PCTCN2022140910-appb-000035
Straight dotted key
Figure PCTCN2022140910-appb-000036
or tilde
Figure PCTCN2022140910-appb-000037
express. For example, the straight-shaped solid-line bond in -OCH3 indicates that it is connected to other groups through the oxygen atom in the group;
Figure PCTCN2022140910-appb-000038
The straight dotted line bond in indicates that the two ends of the nitrogen atom in the group are connected to other groups;
Figure PCTCN2022140910-appb-000039
The wavy lines in indicate that the 1 and 2 carbon atoms in the phenyl group are connected to other groups.
除非另有规定，术语“C 1-3烷基”用于表示直链或支链的由1至3个碳原子组成的饱和碳氢基团。所述C 1-3烷基包括C 1-2和C 2-3烷基等；其可以是一价(如甲基)、二价(如亚甲基)或者多价(如次甲基)。C 1-3烷基的实例包括但不限于甲基(Me)、乙基(Et)、丙基(包括n-丙基和异丙基)等。 Unless otherwise specified, the term "C 1-3 alkyl" is used to denote a straight or branched chain saturated hydrocarbon group consisting of 1 to 3 carbon atoms. The C 1-3 alkyl group includes C 1-2 and C 2-3 alkyl groups, etc.; it can be monovalent (such as methyl), divalent (such as methylene) or multivalent (such as methine) . Examples of C 1-3 alkyl include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), and the like.
除非另有规定，术语“C 1-3烷氧基”表示通过一个氧原子连接到分子的其余部分的那些包含1至3个碳原子的烷基基团。所述C 1-3烷氧基包括C 1-2、C 2-3、C 3和C 2烷氧基等。C 1-3烷氧基的实例包括但不限于甲氧基、乙氧基、丙氧基(包括正丙氧基和异丙氧基)等。 Unless otherwise specified, the term "C 1-3 alkoxy" denotes those alkyl groups containing 1 to 3 carbon atoms attached to the rest of the molecule through an oxygen atom. The C 1-3 alkoxy group includes C 1-2 , C 2-3 , C 3 and C 2 alkoxy groups and the like. Examples of C 1-3 alkoxy include, but are not limited to, methoxy, ethoxy, propoxy (including n-propoxy and isopropoxy), and the like.
除非另有规定，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个原子的“环”。Unless otherwise specified, the number of atoms in a ring is generally defined as the number of ring members, eg, "5-7 membered ring" means a "ring" with 5-7 atoms arranged around it.
除非另有规定，C n-n+m或C n-C n+m包括n至n+m个碳的任何一种具体情况，例如C 1-12包括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，也包括n至n+m中的任何一个范围，例如C 1-12包括C 1-3、C 1-6、C 1-9、C 3-6、C 3-9、C 3-12、C 6-9、C 6-12、和C 9-12等；同理，n元至n+m元表示环上原子数为n至n+m个，例如3-12元环包括3元环、4元环、5元环、6元环、7元环、8元环、9元环、10元环、11元环、和12元环，也包括n至n+m中的任何一个范围，例如3-12元环包括3-6元环、3-9元环、5-6元环、5-7元环、6-7元环、6-8元环、和6-10 元环等。 Unless otherwise specified, C n-n+m or C n -C n+m includes any specific instance of n to n+m carbons, for example C 1-12 includes 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 , also including any range from n to n+m, for example, C 1-12 includes C 1-3 , C 1-6 , C 1-9 , C 3-6 , C 3-9 , C 3-12 , C 6-9 , C 6-12 , and C 9-12 etc.; similarly, n to n +m means that the number of atoms on the ring is from n to n+m, for example, a 3-12-membered ring includes a 3-membered ring, a 4-membered ring, a 5-membered ring, a 6-membered ring, a 7-membered ring, an 8-membered ring, and a 9-membered ring , 10-membered rings, 11-membered rings, and 12-membered rings, also including any range from n to n+m, for example, 3-12-membered rings include 3-6-membered rings, 3-9-membered rings, 5-6-membered rings ring, 5-7-membered ring, 6-7-membered ring, 6-8-membered ring, and 6-10-membered ring, etc.
除非另有规定，“C 3-6环烷基”表示由3至6个碳原子组成的饱和环状碳氢基团，其包括单环、双环和三环体系，其中双环和三环体系包括螺环、并环和桥环。所述C 3-6环烷基包括C 3-5、C 4-6、C 4-5或C 5-6等；其可以是一价、二价或者多价。C 3-6环烷基的实例包括，但不限于，环丙基、环丁基、环戊基、环己基等。 Unless otherwise specified, "C 3-6 cycloalkyl" means a saturated cyclic hydrocarbon group composed of 3 to 6 carbon atoms, which includes monocyclic, bicyclic and tricyclic systems, wherein bicyclic and tricyclic systems include Spiral, parallel and bridged rings. The C 3-6 cycloalkyl group includes C 3-5 , C 4-6 , C 4-5 or C 5-6 and the like; it may be monovalent, divalent or multivalent. Examples of C 3-6 cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and the like.
除非另有规定，“C 3-10环烷基”表示由3至10个碳原子组成的饱和环状碳氢基团，其包括单环、双环和三环体系，其中双环和三环体系包括螺环、并环和桥环。所述C 3-10环烷基包括C 3-8、C 3-6、C 3-5、C 4-10、C 4-8、C 4-6、C 4- 5、C 5-8或C 5-6等；其可以是一价、二价或者多价。C 3-10环烷基的实例包括，但不限于，环丙基、环丁基、环戊基、环己基、环庚基、降冰片烷基、[2.2.2]二环辛烷等。 Unless otherwise specified, "C 3-10 cycloalkyl" means a saturated cyclic hydrocarbon group composed of 3 to 10 carbon atoms, which includes monocyclic, bicyclic and tricyclic systems, wherein bicyclic and tricyclic systems include Spiral, parallel and bridged rings. The C 3-10 cycloalkyl group includes C 3-8 , C 3-6 , C 3-5 , C 4-10 , C 4-8 , C 4-6 , C 4- 5 , C 5-8 or C 5-6 etc.; it may be monovalent, divalent or multivalent. Examples of C 3-10 cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, norbornyl, [2.2.2]bicyclooctane, and the like.
除非另有规定，术语“3-10元杂环烷基”本身或者与其他术语联合分别表示由3至10个环原子组成的饱和环状基团，其1、2、3或4个环原子为独立选自O、S、N、P或Se的杂原子，其余为碳原子，其中氮原子任选地被季铵化，氮、硫和磷杂原子可任选被氧化(即NO、S(O) p和P(O) p，p是1或2)。其包括单环、双环和三环体系，其中双环和三环体系包括螺环、并环和桥环。此外，就该“3-10元杂环烷基”而言，杂原子可以占据杂环烷基与分子其余部分的连接位置。所述3-10元杂环烷基包括3-8元、3-6元、3-5元、4-6元、5-6元、4元、5元和6元杂环烷基等。3-10元杂环烷基的实例包括但不限于氮杂环丁基、氧杂环丁基、硫杂环丁基、吡咯烷基、吡唑烷基、咪唑烷基、四氢噻吩基(包括四氢噻吩-2-基和四氢噻吩-3-基等)、四氢呋喃基(包括四氢呋喃-2-基等)、四氢吡喃基、哌啶基(包括1-哌啶基、2-哌啶基和3-哌啶基等)、哌嗪基(包括1-哌嗪基和2-哌嗪基等)、吗啉基(包括3-吗啉基和4-吗啉基等)、二噁烷基、二噻烷基、异噁唑烷基、异噻唑烷基、1,2-噁嗪基、1,2-噻嗪基、六氢哒嗪基、高哌嗪基、高哌啶基或二氧杂环庚烷基等。 Unless otherwise specified, the term "3-10 membered heterocycloalkyl" by itself or in combination with other terms denotes a saturated cyclic group consisting of 3 to 10 ring atoms, respectively, whose 1, 2, 3 or 4 ring atoms is a heteroatom independently selected from O, S, N, P, or Se, and the remainder is carbon atoms, wherein the nitrogen atom is optionally quaternized, and the nitrogen, sulfur, and phosphorus heteroatoms can be optionally oxidized (i.e., NO, S (O) p and P(O) p , where p is 1 or 2). It includes monocyclic, bicyclic and tricyclic ring systems, wherein bicyclic and tricyclic ring systems include spiro, merged and bridged rings. In addition, as for the "3-10 membered heterocycloalkyl group", a heteroatom may occupy the attachment position of the heterocycloalkyl group to the rest of the molecule. The 3-10 membered heterocycloalkyl group includes 3-8 membered, 3-6 membered, 3-5 membered, 4-6 membered, 5-6 membered, 4-membered, 5-membered and 6-membered heterocycloalkyl groups. Examples of 3-10 membered heterocycloalkyl groups include, but are not limited to, azetidinyl, oxetanyl, thietanyl, pyrrolidinyl, pyrazolidinyl, imidazolidinyl, tetrahydrothiophenyl ( Including tetrahydrothiophen-2-yl and tetrahydrothiophen-3-yl, etc.), tetrahydrofuranyl (including tetrahydrofuran-2-yl, etc.), tetrahydropyranyl, piperidinyl (including 1-piperidinyl, 2- piperidinyl and 3-piperidinyl, etc.), piperazinyl (including 1-piperazinyl and 2-piperazinyl, etc.), morpholinyl (including 3-morpholinyl and 4-morpholinyl, etc.), Dioxanyl, dithianyl, isoxazolidinyl, isothiazolidinyl, 1,2-oxazinyl, 1,2-thiazinyl, hexahydropyridazinyl, homopiperazinyl, homopiperidinyl Pyridyl or dioxepanyl, etc.
除非另有规定，本发明术语“5-6元杂芳环”和“5-6元杂芳基”可以互换使用，术语“5-6元杂芳基”表示由5至6个环原子组成的具有共轭π电子体系的单环基团，其1、2、3或4个环原子为独立选自O、S和N的杂原子，其余为碳原子。其中氮原子任选地被季铵化，氮和硫杂原子可任选被氧化(即NO和S(O) p，p是1或2)。5-6元杂芳基可通过杂原子或碳原子连接到分子的其余部分。所述5-6元杂芳基包括5元和6元杂芳基。所述5-6元杂芳基的实例包括但不限于吡咯基(包括N-吡咯基、2-吡咯基和3-吡咯基等)、吡唑基(包括2-吡唑基和3-吡唑基等)、咪唑基(包括N-咪唑基、2-咪唑基、4-咪唑基和5-咪唑基等)、噁唑基(包括2-噁唑基、4-噁唑基和5-噁唑基等)、***基(1H-1,2,3-***基、2H-1,2,3-***基、1H-1,2,4-***基和4H-1,2,4-***基等)、四唑基、异噁唑基(3-异噁唑基、4-异噁唑基和5-异噁唑基等)、噻唑基(包括2-噻唑基、4-噻唑基和5-噻唑基等)、呋喃基(包括2-呋喃基和3-呋喃基等)、噻吩基(包括2-噻吩基和3-噻吩基等)、吡啶基(包括2-吡啶基、3-吡啶基和4-吡啶基等)、吡嗪基或嘧啶基(包括2-嘧啶基和4-嘧啶基等)。 Unless otherwise specified, the terms "5-6-membered heteroaryl ring" and "5-6-membered heteroaryl" in the present invention can be used interchangeably, and the term "5-6-membered heteroaryl" means that there are 5 to 6 ring atoms A monocyclic group with a conjugated π-electron system, 1, 2, 3 or 4 ring atoms are heteroatoms independently selected from O, S and N, and the rest are carbon atoms. Where the nitrogen atom is optionally quaternized, the nitrogen and sulfur heteroatoms may be optionally oxidized (ie, NO and S(O) p , where p is 1 or 2). The 5-6 membered heteroaryl can be attached to the rest of the molecule through a heteroatom or a carbon atom. The 5-6 membered heteroaryl includes 5 and 6 membered heteroaryl. Examples of the 5-6 membered heteroaryl groups include, but are not limited to, pyrrolyl (including N-pyrrolyl, 2-pyrrolyl and 3-pyrrolyl, etc.), pyrazolyl (including 2-pyrazolyl and 3-pyrrolyl Azolyl, etc.), imidazolyl (including N-imidazolyl, 2-imidazolyl, 4-imidazolyl and 5-imidazolyl, etc.), oxazolyl (including 2-oxazolyl, 4-oxazolyl and 5- Oxazolyl, etc.), triazolyl (1H-1,2,3-triazolyl, 2H-1,2,3-triazolyl, 1H-1,2,4-triazolyl and 4H-1, 2,4-triazolyl, etc.), tetrazolyl, isoxazolyl (3-isoxazolyl, 4-isoxazolyl and 5-isoxazolyl, etc.), thiazolyl (including 2-thiazolyl , 4-thiazolyl and 5-thiazolyl, etc.), furyl (including 2-furyl and 3-furyl, etc.), thienyl (including 2-thienyl and 3-thienyl, etc.), pyridyl (including 2 -pyridyl, 3-pyridyl and 4-pyridyl, etc.), pyrazinyl or pyrimidyl (including 2-pyrimidyl and 4-pyrimidyl, etc.).
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。Unless otherwise specified, the term "halogen" or "halogen" by itself or as part of another substituent means a fluorine, chlorine, bromine or iodine atom.
除非另有说明，当两个取代基及它们相连的原子成环时，所述“相连的原子”可以是同一原子，也可以是不 同原子。例如，本发明中“两个R 1及它们相连的原子形成C 3-6环烷基”，其可以与环A形成螺环、桥环或并环。 Unless otherwise stated, when two substituents and their connected atoms form a ring, the "connected atoms" may be the same atom or different atoms. For example, in the present invention, "two R 1 and the atoms connected to them form a C 3-6 cycloalkyl group", which may form a spiro ring, a bridged ring or a fused ring with ring A.
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。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 listed below, the embodiments formed by combining them with other chemical synthesis methods, and the methods well known to those skilled in the art Equivalent alternatives, preferred embodiments include but are not limited to the examples of the present invention.
本发明的化合物可以通过本领域技术人员所熟知的常规方法来确认结构，如果本发明涉及化合物的绝对构型，则该绝对构型可以通过本领域常规技术手段予以确证。例如单晶X射线衍射法(SXRD)，把培养出的单晶用Bruker D8 venture衍射仪收集衍射强度数据，光源为CuKα辐射，扫描方式：
Figure PCTCN2022140910-appb-000040
扫描，收集相关数据后，进一步采用直接法(Shelxs97)解析晶体结构，便可以确证绝对构型。 The structure of the compounds of the present invention can be confirmed by conventional methods known to those skilled in the art. If the present invention involves the absolute configuration of the compound, the absolute configuration can be confirmed by conventional technical means in the art. For example, in single crystal X-ray diffraction (SXRD), the cultured single crystal is collected with a Bruker D8 venture diffractometer to collect diffraction intensity data, the light source is CuKα radiation, and the scanning method is:
Figure PCTCN2022140910-appb-000040
After scanning and collecting relevant data, the absolute configuration can be confirmed by further analyzing the crystal structure by direct method (Shelxs97).
本发明所使用的溶剂可经市售获得。The solvent used in the present invention is commercially available.
本发明采用下述缩略词：HATU代表2-(7-氮杂苯并三氮唑)-N,N,N',N'-四甲基脲六氟磷酸酯；THF代表四氢呋喃；Boc代表叔丁基氧羰基；n-BuLi代表正丁基锂；HCl代表盐酸；ACN代表乙腈；MeOH代表甲醇；EtOAc或EA代表乙酸乙酯；TEA代表三乙胺；DIEA或DIPEA代表N,N-二异丙基乙胺；DMF代表N,N-二甲基甲酰胺；DMSO代表二甲基亚砜；HPLC代表高效液相色谱；TLC代表薄层色谱；PEG代表聚乙二醇。The following abbreviations are used in the present invention: HATU stands for 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate; THF stands for tetrahydrofuran; Boc stands for tert-butyloxycarbonyl; n-BuLi stands for n-butyl lithium; HCl stands for hydrochloric acid; ACN stands for acetonitrile; MeOH stands for methanol; EtOAc or EA stands for ethyl acetate; TEA stands for triethylamine; DIEA or DIPEA stands for N,N-di Isopropylethylamine; DMF stands for N,N-dimethylformamide; DMSO stands for dimethyl sulfoxide; HPLC stands for high performance liquid chromatography; TLC stands for thin layer chromatography; PEG stands for polyethylene glycol.
化合物依据本领域常规命名原则或者使用
Figure PCTCN2022140910-appb-000041
软件命名，市售化合物采用供应商目录名称。 Compounds are named according to the conventional naming principles in this field or using
Figure PCTCN2022140910-appb-000041
The software is named, and the commercially available compounds adopt the supplier catalog name.
具体实施方式Detailed ways
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。The present invention will be described in detail through examples below, but it does not imply any unfavorable limitation to the present invention. The present invention has been described in detail herein, and its specific embodiments are also disclosed. For those skilled in the art, various changes and improvements can be made to the specific embodiments of the present invention without departing from the spirit and scope of the present invention. will be obvious.
实施例1Example 1
Figure PCTCN2022140910-appb-000042
Figure PCTCN2022140910-appb-000042
合成路线：synthetic route:
Figure PCTCN2022140910-appb-000043
Figure PCTCN2022140910-appb-000043
步骤1：化合物1-2的合成Step 1: Synthesis of Compound 1-2
将化合物1-1(5g，34.93mmol)溶于甲醇(50mL)，加入氢氧化钠水溶液(4M，43.66mL)，在20℃搅拌4h。向反应体系加入柠檬酸(50mL)，饱和食盐水(40mL)，乙酸乙酯(50mL)萃取，分得有机相，无水硫酸钠干燥，旋干得到化合物1-2。 1H NMR(400MHz,DMSO-d6)δ＝–2.56-12.41(m,1H),–7.67-7.59(m,1H),–7.26-7.20(m,1H),–3.98-3.88(m,1H),–3.34-3.30(m,1H),–3.19-3.08(m,2H),–2.36-2.08(m,2H),–1.69-1.50(m,2H),–1.41-1.32(m,9H). Compound 1-1 (5 g, 34.93 mmol) was dissolved in methanol (50 mL), and aqueous sodium hydroxide solution (4M, 43.66 mL) was added, and stirred at 20° C. for 4 h. Citric acid (50 mL), saturated brine (40 mL) and ethyl acetate (50 mL) were added to the reaction system for extraction, and the organic phase was separated, dried over anhydrous sodium sulfate, and spin-dried to obtain compound 1-2. 1 H NMR (400MHz, DMSO-d6) δ=–2.56-12.41(m,1H),–7.67-7.59(m,1H),–7.26-7.20(m,1H),–3.98-3.88(m,1H ),–3.34-3.30(m,1H),–3.19-3.08(m,2H),–2.36-2.08(m,2H),–1.69-1.50(m,2H),–1.41-1.32(m,9H ).
步骤2：化合物1-3的合成Step 2: Synthesis of compounds 1-3
将化合物1-2(3.5g,12.95mmol)溶于N,N-二甲基甲酰胺(40mL)中，依次加入1-(3-二甲基氨基丙基)-3-乙基碳二亚胺(2.11g,11.01mmol)，1-羟基苯并三氮唑(1.63g,12.04mmol)，N,O-二甲基羟胺盐酸盐(1.26g,12.95mmol)，降温至0℃，缓慢滴加三乙胺(1.22g,12.04mmol,1.68mL)，恢复至20℃，反应16hr。用饱和柠檬酸(50mL)调至pH在3到5之间，加入乙酸乙酯(80mL)，乙酸乙酯经饱和碳酸钠溶液(60mL)洗，饱和食盐水(50mL)洗，有机层干燥减压浓缩，得到粗品。经硅胶柱层析(二氯甲烷:甲醇＝1:0～20:1)纯化，得到化合物1-3。 1H NMR(400MHz,DMSO-d 6)δ＝–7.98-7.92(m,2H),–7.22-7.13(m,1H),–4.48-4.37(m,1H),–3.75-3.68(m,2H),–3.33-3.30(m,1H),–3.19-3.12(m,2H),–3.12-3.08(m,1H),–2.75-2.71(m,5H),–2.52-2.47(m,1H),–1.45-1.30(m,9H)。 Compound 1-2 (3.5g, 12.95mmol) was dissolved in N,N-dimethylformamide (40mL), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiethylene was added successively Amine (2.11g, 11.01mmol), 1-hydroxybenzotriazole (1.63g, 12.04mmol), N, O-dimethylhydroxylamine hydrochloride (1.26g, 12.95mmol), cooled to 0°C, slowly Triethylamine (1.22g, 12.04mmol, 1.68mL) was added dropwise, returned to 20°C, and reacted for 16hr. Use saturated citric acid (50mL) to adjust the pH between 3 and 5, add ethyl acetate (80mL), wash ethyl acetate with saturated sodium carbonate solution (60mL) and saturated brine (50mL), and dry the organic layer to reduce Concentrate under reduced pressure to obtain a crude product. Purify by silica gel column chromatography (dichloromethane:methanol=1:0～20:1) to obtain compound 1-3. 1 H NMR (400MHz, DMSO-d 6 )δ=–7.98-7.92(m,2H),–7.22-7.13(m,1H),–4.48-4.37(m,1H),–3.75-3.68(m, 2H),–3.33-3.30(m,1H),–3.19-3.12(m,2H),–3.12-3.08(m,1H),–2.75-2.71(m,5H),–2.52-2.47(m, 1H),–1.45-1.30(m,9H).
步骤3：化合物1-4的合成Step 3: Synthesis of Compounds 1-4
将2-溴苯并噻唑(3.39g,15.85mmol,)溶于THF(30mL)中，氮气置换三次，降温至-78℃，缓慢滴加正丁基锂的四氢呋喃溶液(2.5M,5.07mL)，搅拌30min，之后缓慢加入化合物1-3(0.5g,1.59mmol)的四氢呋喃(5mL)溶液，反应1hr。将饱和氯化铵溶液(40mL)加入反应液中，搅拌10min，加入乙酸乙酯(50mL)反复萃取三次，合并有机相，饱和食盐水(50mL)洗，有机层经无水硫酸钠干燥，减压浓缩得到粗品。经硅胶柱层析(二氯甲烷:甲醇＝1:0～20:1)纯化，得到化合物1-4。Dissolve 2-bromobenzothiazole (3.39g, 15.85mmol,) in THF (30mL), replace with nitrogen three times, cool down to -78°C, slowly add n-butyllithium tetrahydrofuran solution (2.5M, 5.07mL) dropwise , stirred for 30min, then slowly added a solution of compound 1-3 (0.5g, 1.59mmol) in tetrahydrofuran (5mL) and reacted for 1hr. Add saturated ammonium chloride solution (40mL) to the reaction solution, stir for 10min, add ethyl acetate (50mL) and extract repeatedly three times, combine the organic phases, wash with saturated brine (50mL), and dry the organic layer over anhydrous sodium sulfate, reduce Concentrate under reduced pressure to obtain the crude product. Purify by silica gel column chromatography (dichloromethane:methanol=1:0～20:1) to obtain compound 1-4.
步骤4：化合物1-5的合成Step 4: Synthesis of Compounds 1-5
将化合物1-4(30mg,77.03μmol)溶于盐酸/乙酸乙酯溶液(5mL)中，20℃下搅拌1hr，反应液直接减压浓缩，得到化合物1-5。Compound 1-4 (30 mg, 77.03 μmol) was dissolved in hydrochloric acid/ethyl acetate solution (5 mL), stirred at 20°C for 1 hr, and the reaction solution was directly concentrated under reduced pressure to obtain compound 1-5.
MS–ESI m/z:[M+H] +＝290.1. MS-ESI m/z: [M+H] + = 290.1.
步骤5：1-7的合成Step 5: Synthesis of 1-7
在0℃，向原料1-6(400.00mg,1.66mmol)的甲醇(2mL)溶液中加入甲苯(4mL)，三甲基硅重氮甲烷(2M,1.66mL)，在20℃反应16hr。将反应液减压浓缩得到粗品。粗品用硅胶柱层析法分离(石油醚/乙酸乙酯＝10:1).得到产物1-7。 1H NMR(400MHz,CDCl 3)δ＝–4.39-4.20(m,1H),–3.87-3.68(m,4H),–2.74-2.62(m,1H),–1.98-1.87(m,1H),–1.83-1.61(m,3H),–1.56-1.49(m,1H),–1.48-1.36(m,9H),–1.30-1.21(m,1H). To a methanol (2 mL) solution of starting material 1-6 (400.00 mg, 1.66 mmol) was added toluene (4 mL), trimethylsilyldiazomethane (2M, 1.66 mL) at 0°C, and reacted at 20°C for 16 hr. The reaction solution was concentrated under reduced pressure to obtain a crude product. The crude product was separated by silica gel column chromatography (petroleum ether/ethyl acetate=10:1). The product 1-7 was obtained. 1 H NMR (400MHz, CDCl 3 )δ=–4.39-4.20(m,1H),–3.87-3.68(m,4H),–2.74-2.62(m,1H),–1.98-1.87(m,1H) ,–1.83-1.61(m,3H),–1.56-1.49(m,1H),–1.48-1.36(m,9H),–1.30-1.21(m,1H).
步骤6：1-8的合成Step 6: Synthesis of 1-8
向原料1-7(0.28g,1.10mmol)的反应瓶中加入盐酸/乙酸乙酯(4M,5mL)，在20℃反应2hr。将反应液减压浓缩得到粗品。得到化合物1-8。 1H NMR(400MHz,CD 3OD)δ＝–4.21-4.07(m,2H),–3.94-3.79(m,3H),–3.03-2.91(m,1H),–2.04-2.03(m,1H),–2.05-2.00(m,1H),–1.93-1.80(m,3H),1.73(s,2H). Add hydrochloric acid/ethyl acetate (4M, 5 mL) to the reaction flask of raw material 1-7 (0.28 g, 1.10 mmol), and react at 20° C. for 2 hr. The reaction solution was concentrated under reduced pressure to obtain a crude product. Compounds 1-8 were obtained. 1 H NMR (400MHz, CD 3 OD) δ=–4.21-4.07(m,2H),–3.94-3.79(m,3H),–3.03-2.91(m,1H),–2.04-2.03(m,1H) ),–2.05-2.00(m,1H),–1.93-1.80(m,3H),1.73(s,2H).
步骤7：1-9的合成Step 7: Synthesis of 1-9
在0℃，向原料1-8(305.30mg,1.32mmol)的N,N-二甲基甲酰胺(2mL)溶液中加入N-Boc-L-叔亮氨酸(0.21g,1.10mmol)，N,N-二异丙基乙胺(426.49mg,3.30mmol,574.79μL)，HATU(627.38mg,1.65mmol)，在20℃反应16hr。将反应液倒入5％柠檬酸溶液中分液，水相用乙酸乙酯萃取(20mL×2)，有机相用无水硫酸钠干燥，过滤，减压浓缩得到粗品。粗品用硅胶柱层析法分离(石油醚/乙酸乙酯＝3:1)，得到化合物1-9。 1H NMR(400MHz,CDCl 3)δ＝5.21(br d,J＝9.6Hz,1H),–4.48-4.37(m,1H),–4.30-4.21(m,1H),–4.01-3.95(m,1H),–3.68-3.60(m,3H),–2.73-2.61(m,1H),–1.97-1.87(m,1H),–1.79-1.56(m,4H),–1.39-1.33(m,10H),0.97(s,9H). To a solution of starting material 1-8 (305.30 mg, 1.32 mmol) in N,N-dimethylformamide (2 mL) was added N-Boc-L-tert-leucine (0.21 g, 1.10 mmol) at 0°C, N,N-Diisopropylethylamine (426.49mg, 3.30mmol, 574.79μL), HATU (627.38mg, 1.65mmol), reacted at 20°C for 16hr. The reaction solution was poured into 5% citric acid solution for separation, the aqueous phase was extracted with ethyl acetate (20 mL×2), the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was separated by silica gel column chromatography (petroleum ether/ethyl acetate=3:1) to obtain compound 1-9. 1 H NMR (400MHz, CDCl 3 ) δ=5.21(br d,J=9.6Hz,1H),–4.48-4.37(m,1H),–4.30-4.21(m,1H),–4.01-3.95(m ,1H),–3.68-3.60(m,3H),–2.73-2.61(m,1H),–1.97-1.87(m,1H),–1.79-1.56(m,4H),–1.39-1.33(m ,10H),0.97(s,9H).
步骤8：1-10的合成Step 8: Synthesis of 1-10
向原料1-9(0.3g,814.19μmol)的四氢呋喃(2mL)和水(1mL)溶液中加入一水合氢氧化锂(51.25mg,1.22mmol)，在20℃反应16hr。向反应液中加入20mL的5％柠檬酸水溶液，加入20mL乙酸乙酯分液，有机相用无水硫酸钠干燥，过滤，减压浓缩得到粗品。得到化合物1-10。 1H NMR(400MHz,CDCl 3)δ＝5.24(d,J＝9.9Hz,1H),–4.51-4.45(m,1H),–4.41-4.33(m,1H),–4.18-4.15(m,1H),–3.06-2.99(m,1H),–1.99-1.89(m,1H),–1.85-1.75 (m,3H),–1.59-1.49(m,2H),–1.46-1.42(m,9H),–1.05-1.01(m,9H). Lithium hydroxide monohydrate (51.25 mg, 1.22 mmol) was added to a solution of starting material 1-9 (0.3 g, 814.19 μmol) in tetrahydrofuran (2 mL) and water (1 mL), and reacted at 20° C. for 16 hr. Add 20 mL of 5% citric acid aqueous solution to the reaction solution, add 20 mL of ethyl acetate to separate the layers, dry the organic phase with anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain a crude product. Compounds 1-10 were obtained. 1 H NMR (400MHz, CDCl 3 )δ=5.24(d,J=9.9Hz,1H),–4.51-4.45(m,1H),–4.41-4.33(m,1H),–4.18-4.15(m, 1H),–3.06-2.99(m,1H),–1.99-1.89(m,1H),–1.85-1.75(m,3H),–1.59-1.49(m,2H),–1.46-1.42(m, 9H),–1.05-1.01(m,9H).
步骤9：化合物1-11的合成Step 9: Synthesis of Compounds 1-11
将化合物1-10(0.5g,1.41mmol)溶于二氯甲烷(5mL)，加入三氟乙酸(1.5mL)，在20℃反应1h。反应直接浓缩得到化合物1-11。Compound 1-10 (0.5g, 1.41mmol) was dissolved in dichloromethane (5mL), added trifluoroacetic acid (1.5mL), and reacted at 20°C for 1h. The reaction was directly concentrated to obtain compound 1-11.
MS–ESI m/z:[M+H] +＝255.3。 MS-ESI m/z: [M+H] + = 255.3.
步骤10：化合物1-12的合成Step 10: Synthesis of Compounds 1-12
将化合物1-11(0.45g,1.55mmol)溶于甲醇(10mL)，将三氟乙酸甲酯(1.98g,15.48mmol)加入到反应体系，然后将三乙胺(939.57mg,9.29mmol)加入到反应体系。反应在38℃搅拌12h。粗品反应液直接减压浓缩。粗品用二氯甲烷(30mL)溶解，并用1N的盐酸溶液(10mL)洗涤，有机相并用饱和食盐水(20mL)洗涤，过滤，浓缩得到化合物1-12。Compound 1-11 (0.45g, 1.55mmol) was dissolved in methanol (10mL), methyl trifluoroacetate (1.98g, 15.48mmol) was added to the reaction system, and then triethylamine (939.57mg, 9.29mmol) was added to the reaction system. The reaction was stirred at 38 °C for 12 h. The crude reaction solution was directly concentrated under reduced pressure. The crude product was dissolved in dichloromethane (30 mL), washed with 1N hydrochloric acid solution (10 mL), and the organic phase was washed with saturated brine (20 mL), filtered, and concentrated to obtain compound 1-12.
MS–ESI m/z:[M+H] +＝351.1。 MS-ESI m/z: [M+H] + = 351.1.
步骤11：化合物1的合成Step 11: Synthesis of Compound 1
将化合物1-12(25.1mg,77.04μmol)和化合物1-5(29.69mg,84.74μmol)溶于乙腈(5mL)，将反应体系降温至0℃，加入1-乙基-(3-二甲基氨基丙基)碳酰二甲亚胺盐酸盐(16.25mg,84.74μmol)，随后滴加吡啶(24.37mg,308.15μmol)，反应在0℃下进行2hr。反应液加入乙酸乙酯(10mL)和1M盐酸(5mL)进行萃取，有机层用饱和碳酸氢钠水溶液(10mL)洗涤，无水硫酸钠干燥，过滤，减压浓缩。通过制备TLC(乙酸乙酯，Rf＝0.5)纯化得化合物1。Dissolve compound 1-12 (25.1 mg, 77.04 μmol) and compound 1-5 (29.69 mg, 84.74 μmol) in acetonitrile (5 mL), cool the reaction system to 0 ° C, add 1-ethyl-(3-dimethyl Aminopropyl) carbodicarboximide hydrochloride (16.25 mg, 84.74 μmol), followed by dropwise addition of pyridine (24.37 mg, 308.15 μmol), and the reaction was carried out at 0° C. for 2 hr. The reaction solution was extracted by adding ethyl acetate (10 mL) and 1M hydrochloric acid (5 mL), and the organic layer was washed with saturated aqueous sodium bicarbonate (10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. Compound 1 was purified by preparative TLC (ethyl acetate, Rf=0.5).
MS–ESI m/z:[M+H] +＝621.67。 1H NMR(400MHz,CDCl 3)δppm–8.16-8.21(m,1H),–7.95-8.03(m,1H),–7.82-7.87(m,1H),–7.52-7.62(m,2H),–7.00-7.12(m,2H),–5.77-5.91(m,1H),–5.41-5.51(m,1H),4.67(s,1H),–4.41-4.54(m,1H),4.03(s,1H),–3.31-3.44(m,2H),–2.87-2.96(m,1H),–2.54-2.73(m,2H),–1.74-2.32(m,7H),–1.23-1.35(m,1H),1.10(s,8H). MS-ESI m/z: [M+H] + = 621.67. 1 H NMR (400MHz, CDCl 3 )δppm–8.16-8.21(m,1H),–7.95-8.03(m,1H),–7.82-7.87(m,1H),–7.52-7.62(m,2H), –7.00-7.12(m,2H),–5.77-5.91(m,1H),–5.41-5.51(m,1H),4.67(s,1H),–4.41-4.54(m,1H),4.03(s ,1H),–3.31-3.44(m,2H),–2.87-2.96(m,1H),–2.54-2.73(m,2H),–1.74-2.32(m,7H),–1.23-1.35(m ,1H),1.10(s,8H).
实施例2Example 2
Figure PCTCN2022140910-appb-000044
Figure PCTCN2022140910-appb-000044
Figure PCTCN2022140910-appb-000045
Figure PCTCN2022140910-appb-000045
步骤1：化合物2-2的合成Step 1: Synthesis of compound 2-2
将化合物2-1(5g,54.32mmol)溶于甲醇(50mL)，在70℃回流48h。反应体系减压浓缩得到目标产物粗品。粗品纯度较高，直接用于下一步反应，得到化合物2-2。 1H NMR(400MHz,CDCl 3)δ＝4.81(s,1H),3.77(s,3H),3.43(s,3H)。 Compound 2-1 (5 g, 54.32 mmol) was dissolved in methanol (50 mL), and refluxed at 70° C. for 48 h. The reaction system was concentrated under reduced pressure to obtain the crude product of the target product. The crude product was of high purity and was directly used in the next reaction to obtain compound 2-2. 1 H NMR (400 MHz, CDCl 3 ) δ = 4.81 (s, 1H), 3.77 (s, 3H), 3.43 (s, 3H).
步骤2：化合物2-3的合成Step 2: Synthesis of Compound 2-3
将化合物2-2溶于甲苯(3mL)，降温至0℃，缓慢滴加化合物(R)-(+)苯乙胺(1.5g,12.38mmol,1.60mL)，在20℃搅拌1h。向反应体系加入乙酸乙酯(60mL)和饱和食盐水(30mL)萃取分得有机相，无水硫酸钠干燥，旋干得到粗品。经硅胶柱层析(石油醚:乙酸乙酯＝1:0～5:1)纯化，得到目标化合物2-3。 1H NMR(400MHz,CDCl 3)δ＝–7.95-7.56(m,1H),–7.31-7.17(m,5H),–4.71-4.40(m,1H),–3.95-3.71(m,3H),–1.67-1.51(m,3H)。 Compound 2-2 was dissolved in toluene (3 mL), cooled to 0 °C, compound (R)-(+) phenethylamine (1.5 g, 12.38 mmol, 1.60 mL) was slowly added dropwise, and stirred at 20 °C for 1 h. Ethyl acetate (60 mL) and saturated brine (30 mL) were added to the reaction system for extraction to obtain an organic phase, which was dried over anhydrous sodium sulfate and spin-dried to obtain a crude product. Purification by silica gel column chromatography (petroleum ether:ethyl acetate=1:0～5:1) gave the target compound 2-3. 1 H NMR (400MHz, CDCl 3 )δ=–7.95-7.56(m,1H),–7.31-7.17(m,5H),–4.71-4.40(m,1H),–3.95-3.71(m,3H) ,–1.67-1.51 (m,3H).
步骤3：化合物2-4的合成Step 3: Synthesis of compounds 2-4
将化合物2-3(0.5g,2.61mmol)溶于2,2,2-三氟乙醇(5mL)，加入三氟乙酸(313.04mg,2.75mmol,203.28μL)降温至-10℃，搅拌1h，控制温度-10℃，缓慢滴加环戊二烯(207.40mg,3.14mmol)，继续搅拌0.5h。反应体系减压浓缩，加入甲基叔丁基醚(60mL)和饱和碳酸氢钠溶液(30mL×2)搅拌10min，萃取分得有机相，无水硫酸钠干燥，减压浓缩。经硅胶柱层析(石油醚:乙酸乙酯＝1:0～5:1)纯化，得到化合物2-4，构型经二维NMR确认。 1H NMR(400 MHz,CDCl 3)δ＝–7.34-7.18(m,5H),–6.59-6.41(m,1H),6.31(dd,J＝1.6,5.6Hz,1H),4.35(br d,J＝1.3Hz,1H),3.39(s,3H),–3.18-3.03(m,1H),2.95(br s,1H),–2.33-2.22(m,1H),2.14(br d,J＝8.4Hz,1H),–1.54-1.41(m,4H)。 Compound 2-3 (0.5g, 2.61mmol) was dissolved in 2,2,2-trifluoroethanol (5mL), added trifluoroacetic acid (313.04mg, 2.75mmol, 203.28μL) and cooled to -10°C, stirred for 1h, Control the temperature at -10°C, slowly add cyclopentadiene (207.40mg, 3.14mmol) dropwise, and continue stirring for 0.5h. The reaction system was concentrated under reduced pressure, methyl tert-butyl ether (60 mL) and saturated sodium bicarbonate solution (30 mL×2) were added and stirred for 10 min, the organic phase was obtained by extraction, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. Purification by silica gel column chromatography (petroleum ether: ethyl acetate = 1:0-5:1) gave compound 2-4, whose configuration was confirmed by two-dimensional NMR. 1 H NMR (400 MHz, CDCl 3 )δ=–7.34-7.18(m,5H),–6.59-6.41(m,1H),6.31(dd,J=1.6,5.6Hz,1H),4.35(br d ,J＝1.3Hz,1H),3.39(s,3H),–3.18-3.03(m,1H),2.95(br s,1H),–2.33-2.22(m,1H),2.14(br d,J =8.4Hz, 1H), -1.54-1.41(m, 4H).
MS–ESI m/z:[M+H] +＝258.2。 MS-ESI m/z: [M+H] + = 258.2.
步骤4：化合物2-5的合成Step 4: Synthesis of Compound 2-5
将化合物2-4(100.00mg,388.61μmol)溶于四氢呋喃(1.25mL)，降温至-70℃，缓慢滴加硼烷四氢呋喃络合物(1M,427.47μL)，缓慢升温至20℃搅拌1h。降温至0℃，加入10％氢氧化钠水溶液(0.55mL)溶液和30％的双氧水(220.28mg,1.94mmol,186.68μL)溶液，缓慢升温至20℃，搅拌1h。向反应体系加入硫代硫酸钠饱和溶液(10mL)搅拌10min淬灭反应，随后加入饱和食盐水(20mL)和乙酸乙酯(60mL×2)萃取，分得有机相。取少量样品溶液，用3％柠檬酸调节pH小于8，再淀粉碘化钾试纸检测呈阴性后，无水硫酸钠干燥，30℃减压浓缩。经硅胶柱层析(石油醚:乙酸乙酯＝1:0～5:1)纯化得到化合物2-5。 1H NMR(400MHz,CDCl 3)δ＝–7.30-7.13(m,5H),3.93(br d,J＝6.5Hz,1H),3.78(br s,1H),–3.70-3.54(m,1H),–3.39-3.32(m,1H),–3.31-3.24(m,3H),–2.49-2.40(m,1H),2.26(s,1H),–2.09-2.00(m,1H),1.72(br d,J＝10.1Hz,1H),1.46(br d,J＝6.5Hz,1H),–1.41-1.33(m,3H)。 Compound 2-4 (100.00mg, 388.61μmol) was dissolved in tetrahydrofuran (1.25mL), cooled to -70°C, slowly added borane tetrahydrofuran complex (1M, 427.47μL) dropwise, slowly heated to 20°C and stirred for 1h. Cool down to 0°C, add 10% aqueous sodium hydroxide solution (0.55mL) and 30% hydrogen peroxide (220.28mg, 1.94mmol, 186.68μL) solution, slowly raise the temperature to 20°C, and stir for 1h. Saturated sodium thiosulfate solution (10 mL) was added to the reaction system and stirred for 10 min to quench the reaction, then saturated brine (20 mL) and ethyl acetate (60 mL×2) were added for extraction, and the organic phase was separated. Take a small amount of sample solution, adjust the pH to less than 8 with 3% citric acid, and then test negative with starch potassium iodide test paper, dry it over anhydrous sodium sulfate, and concentrate under reduced pressure at 30°C. Compound 2-5 was obtained after purification by silica gel column chromatography (petroleum ether:ethyl acetate=1:0～5:1). 1 H NMR (400MHz, CDCl 3 ) δ=–7.30-7.13(m,5H), 3.93(br d,J=6.5Hz,1H), 3.78(br s,1H),–3.70-3.54(m,1H ),–3.39-3.32(m,1H),–3.31-3.24(m,3H),–2.49-2.40(m,1H),2.26(s,1H),–2.09-2.00(m,1H),1.72 (br d, J = 10.1Hz, 1H), 1.46 (br d, J = 6.5Hz, 1H), -1.41-1.33 (m, 3H).
MS–ESI m/z:[M+H] +＝276.1。 MS-ESI m/z: [M+H] + = 276.1.
步骤5：化合物2-6的盐酸盐的合成Step 5: Synthesis of the hydrochloride salt of compound 2-6
将化合物2-5(3g,10.90mmol)溶于乙醇(80mL)，加入浓盐酸(1.19g,32.69mmol)和湿钯碳(15g,10.68mmol,5％)。反应在20℃搅拌16h。将反应液用硅藻土过滤，然后直接旋干得到粗品化合物2-6的盐酸盐。Compound 2-5 (3 g, 10.90 mmol) was dissolved in ethanol (80 mL), and concentrated hydrochloric acid (1.19 g, 32.69 mmol) and wet palladium on carbon (15 g, 10.68 mmol, 5%) were added. The reaction was stirred at 20 °C for 16 h. The reaction solution was filtered with celite, and then directly spin-dried to obtain the crude compound 2-6 hydrochloride.
MS–ESI m/z:[M+H] +＝172.0。 MS-ESI m/z: [M+H] + = 172.0.
步骤6：化合物2-8的合成Step 6: Synthesis of compounds 2-8
将化合物2-7(1.87g,10.90mmol)溶于N,N-二甲基甲酰胺(20mL)，加入HATU(4.78g,12.58mmol)和二异丙基乙胺(4.34g,33.55mmol)，在搅拌30min后，加入化合物2-6的盐酸盐(190mg,1.12mmol)。反应在20℃搅拌16h。向反应液加入水(15mL)，乙酸乙酯(60mL)萃取两次，合并有机相，将有机相经5％柠檬酸(30mL)洗涤两次，食盐水(20mL)洗涤四次，无水硫酸钠干燥，过滤，旋干。经柱层析(石油醚：乙酸乙酯＝3:1)纯化得到化合物2-8。 1H NMR(400MHz,CDCl 3)δ＝–5.28-5.16(m,1H),4.50(br s,1H),4.28(d,J＝9.8Hz,1H),3.92(s,1H),3.74(s,3H),2.81(s,1H),2.67(s,1H),2.17(br dd,J＝6.1,12.7Hz,1H),–1.99-1.93(m,1H),–1.90-1.84(m,1H),1.59(br d,J＝13.3Hz,2H),1.43(s,9H),1.04(s,9H)。 Compound 2-7 (1.87g, 10.90mmol) was dissolved in N,N-dimethylformamide (20mL), and HATU (4.78g, 12.58mmol) and diisopropylethylamine (4.34g, 33.55mmol) were added , after stirring for 30 min, the hydrochloride salt of compound 2-6 (190 mg, 1.12 mmol) was added. The reaction was stirred at 20 °C for 16 h. Add water (15mL) to the reaction solution, extract twice with ethyl acetate (60mL), combine the organic phases, wash the organic phase twice with 5% citric acid (30mL), wash four times with brine (20mL), and wash with anhydrous sulfuric acid Sodium-dried, filtered, and spin-dried. Compound 2-8 was obtained after purification by column chromatography (petroleum ether: ethyl acetate = 3:1). 1 H NMR (400MHz, CDCl 3 ) δ=–5.28-5.16 (m, 1H), 4.50 (br s, 1H), 4.28 (d, J=9.8Hz, 1H), 3.92 (s, 1H), 3.74 ( s,3H),2.81(s,1H),2.67(s,1H),2.17(br dd,J＝6.1,12.7Hz,1H),–1.99-1.93(m,1H),–1.90-1.84(m , 1H), 1.59 (br d, J=13.3Hz, 2H), 1.43 (s, 9H), 1.04 (s, 9H).
MS–ESI m/z:[M+H] +＝385.2。 MS-ESI m/z: [M+H] + = 385.2.
步骤7：化合物2-9的合成Step 7: Synthesis of compounds 2-9
将化合物2-8(500mg,1.30mmol)溶解于乙腈(7.5mL)中，加入2-碘酰基苯甲酸(976.31mg,3.49mmol)，60℃搅拌16h。反应液直接用硅藻土过滤，旋干。无纯化得到化合物2-9。Compound 2-8 (500mg, 1.30mmol) was dissolved in acetonitrile (7.5mL), 2-iodoxybenzoic acid (976.31mg, 3.49mmol) was added, and stirred at 60°C for 16h. The reaction solution was directly filtered through celite and spin-dried. Compound 2-9 was obtained without purification.
MS–ESI m/z:[M-55] +＝327.1。 MS-ESI m/z: [M-55] + = 327.1.
步骤8：化合物2-10的合成Step 8: Synthesis of Compound 2-10
将化合物2-9(480mg,1.26mmol)溶解于二氯甲烷(4.8mL)，降温至0℃，加入二乙胺基三氟化硫(1.01g,6.28mmol)，20℃搅拌16h。0℃将反应液缓慢加入饱和碳酸氢钠溶液(20mL)中，并用二氯甲烷(20mL)萃取2次，合并有机相并用无水硫酸钠干燥，过滤，浓缩。无纯化得到化合物2-10。Compound 2-9 (480mg, 1.26mmol) was dissolved in dichloromethane (4.8mL), cooled to 0°C, diethylaminosulfur trifluoride (1.01g, 6.28mmol) was added, and stirred at 20°C for 16h. The reaction solution was slowly added to saturated sodium bicarbonate solution (20 mL) at 0° C., and extracted twice with dichloromethane (20 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated. Compound 2-10 was obtained without purification.
MS–ESI m/z:[M-100] +＝304.0。 MS-ESI m/z: [M-100] + = 304.0.
步骤9：化合物2-11的合成Step 9: Synthesis of Compound 2-11
将化合物2-10(475mg,1.17mmol)溶解于四氢呋喃(5.5mL)，水(1.84mL)和甲醇(1.84mL)中，加入一水合氢氧化锂(147.84mg,3.52mmol)，20℃搅拌16h。向反应液加入水(20mL)和5％柠檬酸(25mL)，用乙酸乙酯(40mL)萃取两次，合并有机相，经饱和食盐水(40mL)洗涤，再用无水硫酸钠干燥，过滤，旋干。无纯化得到化合物2-11。 1H NMR(400MHz,CDCl 3)δ＝5.18(br d,J＝9.8Hz,1H),–4.65-4.53(m,2H),4.31(d,J＝9.9Hz,1H),3.33(br d,J＝7.3Hz,1H),–2.55-2.41(m,1H),–2.33-2.07(m,3H),1.45(s,9H),1.03(s,9H)。 Compound 2-10 (475mg, 1.17mmol) was dissolved in tetrahydrofuran (5.5mL), water (1.84mL) and methanol (1.84mL), added lithium hydroxide monohydrate (147.84mg, 3.52mmol), stirred at 20°C for 16h . Add water (20mL) and 5% citric acid (25mL) to the reaction solution, extract twice with ethyl acetate (40mL), combine the organic phases, wash with saturated brine (40mL), dry over anhydrous sodium sulfate, and filter , spin dry. Compound 2-11 was obtained without purification. 1 H NMR (400MHz, CDCl 3 ) δ=5.18(br d, J=9.8Hz, 1H), –4.65-4.53(m, 2H), 4.31(d, J=9.9Hz, 1H), 3.33(br d , J=7.3Hz, 1H), -2.55-2.41(m, 1H), -2.33-2.07(m, 3H), 1.45(s, 9H), 1.03(s, 9H).
MS–ESI m/z:[M-55] +＝335.1。 MS-ESI m/z: [M-55] + = 335.1.
步骤10：化合物2-12的合成Step 10: Synthesis of Compound 2-12
将化合物2-10(160mg,409.81μmol)和1-5(160.23mg,491.78μmol)溶解于乙腈(2mL)，降温至0℃，加入1-乙基-(3-二甲基氨基丙基)碳酰二甲亚胺盐酸盐(79.35mg,413.91μmol)，然后将吡啶(129.66mg,1.64mmol)滴加进去，反应0℃搅拌2h。向反应液加入水(15mL)，乙酸乙酯(30mL)萃取两次，合并有机相，将有机相经5％柠檬酸(20mL)洗涤两次，食盐水(10mL)洗涤一次，无水硫酸钠干燥，过滤，旋干。经柱层析(二氯甲烷：甲醇＝20:1)纯化得到化合物2-12。 1H NMR(400MHz,CDCl 3)δ＝8.19(s,1H),–8.02-7.97(m,1H),–7.61-7.55(m,2H),5.83(q,J＝7.8Hz,1H),5.55(s,1H),5.26(br d,J＝10.4Hz,1H),–4.58-4.51(m,2H),4.31(d,J＝10.1Hz,1H),–3.40-3.33(m,2H),3.20(br d,J＝6.6Hz,1H),–2.67-2.53(m,2H),–2.36-2.16(m,6H),–2.13-2.07(m,1H),1.94(br d,J＝10.0Hz,1H),1.45(s,9H),1.05(s,9H)。 Dissolve compound 2-10 (160 mg, 409.81 μmol) and 1-5 (160.23 mg, 491.78 μmol) in acetonitrile (2 mL), cool to 0 ° C, add 1-ethyl-(3-dimethylaminopropyl) Carbodimethimide hydrochloride (79.35mg, 413.91μmol) was added dropwise to pyridine (129.66mg, 1.64mmol), and the reaction was stirred at 0°C for 2h. Add water (15mL) to the reaction solution, extract twice with ethyl acetate (30mL), combine the organic phases, wash the organic phase twice with 5% citric acid (20mL), wash once with brine (10mL), anhydrous sodium sulfate Dry, filter and spin dry. Compound 2-12 was obtained after purification by column chromatography (dichloromethane:methanol=20:1). 1 H NMR (400MHz, CDCl 3 )δ=8.19(s,1H),–8.02-7.97(m,1H),–7.61-7.55(m,2H),5.83(q,J=7.8Hz,1H), 5.55(s,1H),5.26(br d,J=10.4Hz,1H),–4.58-4.51(m,2H),4.31(d,J=10.1Hz,1H),–3.40-3.33(m,2H ),3.20(br d,J＝6.6Hz,1H),–2.67-2.53(m,2H),–2.36-2.16(m,6H),–2.13-2.07(m,1H),1.94(br d, J=10.0Hz, 1H), 1.45(s, 9H), 1.05(s, 9H).
MS–ESI m/z:[M+H] +＝662.3。 MS-ESI m/z: [M+H] + = 662.3.
步骤11：化合物2-13的三氟乙酸盐的合成Step 11: Synthesis of Trifluoroacetate Salt of Compound 2-13
将化合物2-12(210mg,317.34μmol)溶于二氯甲烷(2.1mL)和三氟乙酸(0.7mL)中，反应在20℃下搅拌2h。反应液油泵旋干，二氯甲烷重复旋蒸直至形状为淡黄色泡沫。得到化合物2-13的三氟乙酸盐。Compound 2-12 (210 mg, 317.34 μmol) was dissolved in dichloromethane (2.1 mL) and trifluoroacetic acid (0.7 mL), and the reaction was stirred at 20° C. for 2 h. The reaction solution was spin-dried with an oil pump, and dichloromethane was repeatedly rotary-evaporated until the shape was light yellow foam. The trifluoroacetate salt of compound 2-13 was obtained.
MS–ESI m/z:[M+H] +＝562.2。 MS-ESI m/z: [M+H] + = 562.2.
步骤12：化合物2的合成Step 12: Synthesis of compound 2
将化合物2-13的三氟乙酸盐(170mg,302.68μmol)溶于甲醇(3.4mL)中，然后加入三乙胺(245.03mg,2.42mmol)和三氟乙酸甲酯(387.59mg,3.03mmol)。反应升温至室温38℃，搅拌16h。将反应液直接旋干，加入水(10mL)和乙酸乙酯(10mL)溶解，加入5％柠檬酸(10mL)，调节溶液为酸性，分液，再用乙酸乙酯(10mL)萃取两次, 合并有机相并用饱和食盐水(10mL)洗涤两次，无水硫酸钠干燥，过滤，浓缩。通过制备TLC(乙酸乙酯，R f＝0.5)纯化得化合物2。 1H NMR(400MHz,CDCl 3)δ＝–8.22-8.11(m,1H),–8.01-7.95(m,1H),–7.61-7.52(m,2H),4.68(d,J＝9.2Hz,1H),–4.61-4.53(m,2H),–3.44-3.35(m,2H),3.20(br d,J＝5.7Hz,1H),–2.81-2.73(m,1H),–2.33-1.88(m,9H),1.07(s,9H)。 The trifluoroacetate (170 mg, 302.68 μmol) of compound 2-13 was dissolved in methanol (3.4 mL), then triethylamine (245.03 mg, 2.42 mmol) and methyl trifluoroacetate (387.59 mg, 3.03 mmol) were added ). The reaction was warmed to room temperature 38°C and stirred for 16h. Spin the reaction solution to dryness directly, add water (10mL) and ethyl acetate (10mL) to dissolve, add 5% citric acid (10mL) to adjust the solution to acidity, separate the layers, and then extract twice with ethyl acetate (10mL). The combined organic phases were washed twice with saturated brine (10 mL), dried over anhydrous sodium sulfate, filtered and concentrated. Compound 2 was purified by preparative TLC (ethyl acetate, R f =0.5). 1 H NMR (400MHz, CDCl 3 )δ=–8.22-8.11(m,1H),–8.01-7.95(m,1H),–7.61-7.52(m,2H),4.68(d,J=9.2Hz, 1H),–4.61-4.53(m,2H),–3.44-3.35(m,2H),3.20(br d,J＝5.7Hz,1H),–2.81-2.73(m,1H),–2.33-1.88 (m,9H), 1.07(s,9H).
MS–ESI m/z:[M+H] +＝658.3。 MS-ESI m/z: [M+H] + = 658.3.
实施例3Example 3
Figure PCTCN2022140910-appb-000046
Figure PCTCN2022140910-appb-000046
合成路线：synthetic route:
Figure PCTCN2022140910-appb-000047
Figure PCTCN2022140910-appb-000047
步骤1：化合物3-3的合成Step 1: Synthesis of compound 3-3
将化合物3-1(4.62g,21.01mmol)溶解于N,N-二甲基甲酰胺(50mL)中，加入HATU(9.22g,24.24mmol)和N,N-二异丙基乙胺(8.35g,64.64mmol,11.26mL)，搅拌0.5小时，加入化合物3-2(5g,16.16mmol)，20℃搅拌1小时。反应加入水(150mL)，乙酸乙酯(30mL×2)萃取，合并有机相，经5％柠檬酸(10mL)和食盐水(20mL×4)洗涤，无水硫酸钠干燥，过滤，旋干。经柱层析(石油醚:乙酸乙酯＝5:1)纯化得到化合物3-3。Compound 3-1 (4.62g, 21.01mmol) was dissolved in N,N-dimethylformamide (50mL), and HATU (9.22g, 24.24mmol) and N,N-diisopropylethylamine (8.35 g, 64.64mmol, 11.26mL), stirred for 0.5 hours, added compound 3-2 (5g, 16.16mmol), stirred at 20°C for 1 hour. Water (150 mL) was added to the reaction, extracted with ethyl acetate (30 mL×2), the organic phases were combined, washed with 5% citric acid (10 mL) and brine (20 mL×4), dried over anhydrous sodium sulfate, filtered, and spin-dried. Compound 3-3 was obtained after purification by column chromatography (petroleum ether:ethyl acetate=5:1).
MS–ESI m/z:[M+H] +＝475.6. MS-ESI m/z: [M+H] + = 475.6.
步骤2：化合物3-4的合成Step 2: Synthesis of Compound 3-4
在干燥的单口瓶中加入化合物3-3(6g,12.64mmol)和溶剂四氢呋喃(90mL)，甲醇(30mL)，水(30mL)，加入一水合氢氧化锂(1.59g,37.92mmol)，反应在25℃下反应12小时。取锥形瓶，缓慢滴加1M盐酸，调节pH至7， 加入20mL水和20mL二氯甲烷，萃取三次，分离出有机相，用20mL饱和食盐水洗涤，无水硫酸钠干燥，过滤减压浓缩得到残余物。得到化合物3-4。Add compound 3-3 (6g, 12.64mmol) and solvent tetrahydrofuran (90mL), methanol (30mL), water (30mL) in a dry single-necked bottle, add lithium hydroxide monohydrate (1.59g, 37.92mmol), and react in The reaction was carried out at 25°C for 12 hours. Take a conical flask, slowly add 1M hydrochloric acid dropwise, adjust the pH to 7, add 20mL water and 20mL dichloromethane, extract three times, separate the organic phase, wash with 20mL saturated brine, dry over anhydrous sodium sulfate, filter and concentrate under reduced pressure to get the residue. Compound 3-4 was obtained.
MS–ESI m/z:[M+H] +＝447.5. MS-ESI m/z: [M+H] + = 447.5.
步骤3：化合物3-5的合成Step 3: Synthesis of Compound 3-5
将化合物3-4(0.35g,783.74μmol)和化合物1-5(0.2g,691.20μmol)，溶于乙腈(10mL)，降温至0℃，加入1-(3-二甲氨基丙基)-3-乙醛盐酸盐(150.24mg,783.74μmol)，滴加吡啶(225.43mg,2.85mmol,230.03μL)，反应在0℃进行2小时。反应加入水(15mL)，乙酸乙酯(30mL×2)萃取，合并有机相，经5％柠檬酸(10mL)和食盐水(20mL×4)洗涤，无水硫酸钠干燥，过滤，旋干。经柱层析(二氯甲烷:甲醇＝5:1)纯化得到化合物3-5。Dissolve compound 3-4 (0.35g, 783.74μmol) and compound 1-5 (0.2g, 691.20μmol) in acetonitrile (10mL), cool to 0°C, add 1-(3-dimethylaminopropyl)- 3-Acetaldehyde hydrochloride (150.24mg, 783.74μmol) was added dropwise with pyridine (225.43mg, 2.85mmol, 230.03μL), and the reaction was carried out at 0°C for 2 hours. Water (15 mL) was added to the reaction, extracted with ethyl acetate (30 mL×2), the organic phases were combined, washed with 5% citric acid (10 mL) and brine (20 mL×4), dried over anhydrous sodium sulfate, filtered and spin-dried. Compound 3-5 was obtained after purification by column chromatography (dichloromethane:methanol=5:1).
MS–ESI m/z:[M+H] +＝718.9. MS-ESI m/z: [M+H] + = 718.9.
步骤4：化合物3-6三氟乙酸盐的合成Step 4: Synthesis of Compound 3-6 Trifluoroacetate Salt
将化合物3-5(0.3g,417.88μmol)溶于二氯甲烷(10mL)和三氟乙酸(2.5mL)，15℃搅拌1小时。反应液减压浓缩得到化合物3-6三氟乙酸盐。反应未经纯化直接投入下一步反应。Compound 3-5 (0.3 g, 417.88 μmol) was dissolved in dichloromethane (10 mL) and trifluoroacetic acid (2.5 mL), and stirred at 15° C. for 1 hour. The reaction solution was concentrated under reduced pressure to obtain compound 3-6 trifluoroacetate. The reaction was directly put into the next reaction without purification.
MS–ESI m/z:[M+H] +＝618.8. MS-ESI m/z: [M+H] + = 618.8.
步骤5：化合物3的合成Step 5: Synthesis of compound 3
将化合物3-6三氟乙酸盐(0.17g,275.17μmol)溶于二氯甲烷(1mL)，降温至0℃，加入吡啶(152.36mg,1.93mmol,155.47μL)和三氟乙酸酐(231.18mg,1.10mmol,153.10μL)，0℃反应0.5小时。反应加入水(15mL)，乙酸乙酯(30mL×2)萃取，合并有机相，经5％柠檬酸(10mL)和食盐水(20mL×4)洗涤，无水硫酸钠干燥，过滤，旋干。将粗产品根据制备HPLC(柱型:Waters Xbridge Prep OBD C18 150*40mm*10μm；流动相:[水(NH 4HCO 3)-乙腈]；乙腈％:50％-80％,8min)分离，馏分减压浓缩得到化合物3。 Dissolve compound 3-6 trifluoroacetic acid salt (0.17g, 275.17μmol) in dichloromethane (1mL), cool to 0°C, add pyridine (152.36mg, 1.93mmol, 155.47μL) and trifluoroacetic anhydride (231.18 mg, 1.10mmol, 153.10μL), react at 0℃ for 0.5 hours. Water (15 mL) was added to the reaction, extracted with ethyl acetate (30 mL×2), the organic phases were combined, washed with 5% citric acid (10 mL) and brine (20 mL×4), dried over anhydrous sodium sulfate, filtered and spin-dried. The crude product was separated according to preparative HPLC (column type: Waters Xbridge Prep OBD C18 150*40mm*10 μm; mobile phase: [water (NH 4 HCO 3 )-acetonitrile]; acetonitrile %: 50%-80%, 8min), and fractions Concentration under reduced pressure gave compound 3.
MS–ESI m/z:[M+H] +＝714.8. 1H NMR(400MHz,CDCl 3)δ＝8.25-8.14(m,1H),8.05-7.94(m,1H),7.63-7.51(m,2H),5.87-5.76(m,1H),5.76-5.75(m,1H),5.71-5.59(m,1H),4.63-4.45(m,1H),4.41-4.19(m,1H),3.99-3.75(m,1H),3.65-3.32(m,3H),2.72-1.95(m,10H),1.75-1.37(m,16H). MS-ESI m/z: [M+H] + = 714.8. 1 H NMR (400MHz, CDCl 3 ) δ = 8.25-8.14 (m, 1H), 8.05-7.94 (m, 1H), 7.63-7.51 (m ,2H),5.87-5.76(m,1H),5.76-5.75(m,1H),5.71-5.59(m,1H),4.63-4.45(m,1H),4.41-4.19(m,1H),3.99 -3.75(m,1H),3.65-3.32(m,3H),2.72-1.95(m,10H),1.75-1.37(m,16H).
实施例4Example 4
Figure PCTCN2022140910-appb-000048
Figure PCTCN2022140910-appb-000048
合成路线：synthetic route:
Figure PCTCN2022140910-appb-000049
Figure PCTCN2022140910-appb-000049
步骤1：化合物4-2的合成Step 1: Synthesis of Compound 4-2
在预先干燥过的三口烧瓶中加入原料化合物1-1(10g,34.93mmol)加入溶剂甲醇(100mL)氮气抽换气三次。将该反应器置于冰浴中，体系温度(内)控制在-10～-5℃，缓慢滴加硼氢化钠(5.39g,142.47mmol)，加毕，混合物在15℃继续搅拌12小时。反应液中加入水(100mL)搅拌10分钟，乙酸乙酯(200mL×4)萃取，有机层经无水硫酸钠干燥后，过滤，有机相减压浓缩得到化合物4-2。Add raw material compound 1-1 (10 g, 34.93 mmol) into a pre-dried three-necked flask, add solvent methanol (100 mL) and pump nitrogen three times. The reactor was placed in an ice bath, the system temperature (inside) was controlled at -10~-5°C, sodium borohydride (5.39 g, 142.47 mmol) was slowly added dropwise, and the mixture was stirred at 15°C for 12 hours. Water (100 mL) was added to the reaction solution and stirred for 10 minutes, extracted with ethyl acetate (200 mL×4), the organic layer was dried over anhydrous sodium sulfate, filtered, and the organic phase was concentrated under reduced pressure to obtain compound 4-2.
MS–ESI m/z:[M+H] +＝259.2. MS-ESI m/z: [M+H] + = 259.2.
步骤2：化合物4-3的合成Step 2: Synthesis of compound 4-3
15℃下，在干燥的三口瓶中加入化合物4-2(8g,30.97mmol)和二氯甲烷(80mL)，开启搅拌；随后加入碳酸氢钠(2.73g,32.52mmol,1.26mL)和戴斯-马丁过碘烷(13.79g,32.52mmol)反应液由混浊变澄清，氮气置换3次，搅拌约20min反应液变为白色悬浊液，15℃搅拌16hr。反应液加入硫代硫酸钠(100mL)淬灭，二氯甲烷(100mL×3)萃取，合并有机相，有机相经过饱和的碳酸氢钠溶液(200mL)洗涤。有机层经无水硫酸钠干燥后，过滤，减压浓缩。粗产品通过自动过柱机分离(梯度淋洗：二氯甲烷：甲醇＝100：0-90:10)得到化合物4-3。At 15°C, compound 4-2 (8g, 30.97mmol) and dichloromethane (80mL) were added to a dry three-necked flask, and stirring was started; then sodium bicarbonate (2.73g, 32.52mmol, 1.26mL) and Dessert -Martin periodinane (13.79g, 32.52mmol) The reaction solution changed from cloudy to clear, replaced with nitrogen 3 times, stirred for about 20min, the reaction solution turned into a white suspension, and stirred at 15°C for 16hr. The reaction solution was quenched by adding sodium thiosulfate (100 mL), extracted with dichloromethane (100 mL×3), and the combined organic phases were washed with saturated sodium bicarbonate solution (200 mL). The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The crude product was separated by an automatic column machine (gradient elution: dichloromethane: methanol = 100:0-90:10) to obtain compound 4-3.
步骤3：化合物4-5的合成Step 3: Synthesis of compounds 4-5
在预先干燥过的三口烧瓶中加入化合物4-4(111mg,931.83μmol)，加入溶剂四氢呋喃(1mL)氮气置换三次，将该反应器置于冰浴中。体系温度控制在0-5℃，缓慢滴加异丙基镁氯化锂络合物的四氢呋喃溶液(2M,465.91μL)，加毕，混合物在0℃继续搅拌0.5小时。随后控制在0-5℃，将化合物4-3(0.3g,1.17mmol)和四氢呋喃(3mL)的混合溶剂加入上述体系，加毕，缓慢升温至15℃，继续搅拌2hr。加入5mL饱和氯化铵水溶液淬灭反应，用乙酸 乙酯10mL×3萃取，合并有机相无水硫酸钠干燥，减压浓缩得到粗产品。粗产品通过自动过柱机分离(梯度淋洗：二氯甲烷：甲醇＝100:0-95:5)得到化合物4-5。Compound 4-4 (111 mg, 931.83 μmol) was added to a pre-dried three-necked flask, the solvent tetrahydrofuran (1 mL) was added to replace nitrogen three times, and the reactor was placed in an ice bath. The temperature of the system was controlled at 0-5°C, and a tetrahydrofuran solution (2M, 465.91 μL) of isopropylmagnesium lithium chloride complex was slowly added dropwise. After the addition was complete, the mixture was stirred at 0°C for 0.5 hours. Then, controlled at 0-5°C, a mixed solvent of compound 4-3 (0.3g, 1.17mmol) and tetrahydrofuran (3mL) was added to the above system. After the addition was completed, the temperature was slowly raised to 15°C, and stirring was continued for 2hr. Add 5 mL of saturated ammonium chloride aqueous solution to quench the reaction, extract with ethyl acetate 10 mL × 3, combine the organic phases to dry over anhydrous sodium sulfate, and concentrate under reduced pressure to obtain a crude product. The crude product was separated by an automatic column machine (gradient elution: dichloromethane: methanol = 100:0-95:5) to obtain compound 4-5.
MS–ESI m/z:[M+1] +＝376.1. MS-ESI m/z: [M+1] + = 376.1.
步骤4：化合物4-6的盐酸盐合成Step 4: Synthesis of hydrochloride salt of compound 4-6
将底物化合物4-5(0.2g,532.74μmol)溶于盐酸/乙酸乙酯(4M,10.00mL)中，继续搅拌1h。直接浓缩得到化合物4-6的盐酸盐。The substrate compound 4-5 (0.2 g, 532.74 μmol) was dissolved in hydrochloric acid/ethyl acetate (4M, 10.00 mL), and stirring was continued for 1 h. Direct concentration gave the hydrochloride salt of compound 4-6.
步骤5：化合物4-7的合成Step 5: Synthesis of compounds 4-7
将底物化合物3-4(0.35g,783.74μmol)溶解于N,N-二甲基甲酰胺(10mL)中，加入HATU(447.00mg,1.18mmol)，搅拌0.5h。将化合物4-6的盐酸盐(258.92mg)和二异丙基乙胺(405.17mg,3.13mmol)分别加入到反应体系,20℃搅拌1小时。反应液中加入水(150mL)，乙酸乙酯(30mL)萃取2次，合并有机相，经5％柠檬酸(10mL)和食盐水(20mL×4)洗涤，无水硫酸钠干燥，过滤，旋干。由TLC(石油醚:乙酸乙酯＝3:1,Rf＝0.3)，经柱层析(石油醚:乙酸乙酯＝5:1)纯化得到化合物4-7。The substrate compound 3-4 (0.35 g, 783.74 μmol) was dissolved in N,N-dimethylformamide (10 mL), added HATU (447.00 mg, 1.18 mmol), and stirred for 0.5 h. The hydrochloride salt of compound 4-6 (258.92mg) and diisopropylethylamine (405.17mg, 3.13mmol) were added to the reaction system, and stirred at 20°C for 1 hour. Add water (150mL) to the reaction solution, extract twice with ethyl acetate (30mL), combine the organic phases, wash with 5% citric acid (10mL) and brine (20mL×4), dry over anhydrous sodium sulfate, filter, and spin dry . Compound 4-7 was obtained by TLC (petroleum ether: ethyl acetate = 3:1, Rf = 0.3) and column chromatography (petroleum ether: ethyl acetate = 5:1).
MS–ESI m/z:[M+H] +＝704.4. MS-ESI m/z: [M+H] + = 704.4.
步骤6：化合物4-8的合成Step 6: Synthesis of compounds 4-8
将化合物4-7(0.2g,284.15μmol)溶于二氯甲烷(2mL)，将戴斯-马丁过碘烷(361.55mg,852.44μmol)加入到反应体系，在15℃搅拌1h。用硫代硫酸钠饱和溶液(10mL)和碳酸氢钠饱和溶液(10mL)洗涤，二氯甲烷萃取(10mL)3次，合并有机相并用饱和食盐水洗涤(30mL)，无水硫酸钠干燥，过滤，浓缩得到化合物4-8。Compound 4-7 (0.2 g, 284.15 μmol) was dissolved in dichloromethane (2 mL), Dess-Martin periodinane (361.55 mg, 852.44 μmol) was added to the reaction system, and stirred at 15° C. for 1 h. Wash with saturated sodium thiosulfate solution (10 mL) and saturated sodium bicarbonate solution (10 mL), extract with dichloromethane (10 mL) 3 times, combine organic phases and wash with saturated brine (30 mL), dry over anhydrous sodium sulfate, filter , concentrated to obtain compound 4-8.
MS–ESI m/z:[M+H] +＝702.3. MS-ESI m/z: [M+H] + = 702.3.
步骤7：化合物4-9三氟乙酸盐的合成Step 7: Synthesis of Compound 4-9 Trifluoroacetate Salt
将化合物4-8(0.09g,128.23μmol)溶于二氯甲烷(2mL)，将三氟乙酸(462.00mg,4.05mmol,300.00μL)加入到反应体系。反应在15℃下继续搅拌2h。反应直接浓缩得到化合物4-9三氟乙酸盐。Compound 4-8 (0.09 g, 128.23 μmol) was dissolved in dichloromethane (2 mL), and trifluoroacetic acid (462.00 mg, 4.05 mmol, 300.00 μL) was added to the reaction system. The reaction was stirred for an additional 2 h at 15 °C. The reaction was directly concentrated to obtain the trifluoroacetate salt of compound 4-9.
步骤8：化合物4的合成Step 8: Synthesis of compound 4
将底物化合物4-9三氟乙酸盐(0.034g,56.50μmol)溶于二氯甲烷(1mL)，降温到0℃，加入吡啶(31.29mg,395.52μmol)和三氟乙酸酐(47.47mg,226.01μmol)，继续搅拌0.2h。反应用饱和碳酸氢钠溶液(10mL)洗涤，二氯甲烷(10mL)萃取3次，饱和食盐水(20mL)洗涤，无水硫酸钠干燥，过滤，浓缩。制备HPLC分离[柱型:Waters Xbridge BEH C18 100*30mm*10μm；流动相:[H 2O(NH 4HCO 3)-乙腈]；乙腈％:55％-75％,8min]得到化合物4。 The substrate compound 4-9 trifluoroacetate (0.034g, 56.50μmol) was dissolved in dichloromethane (1mL), cooled to 0°C, pyridine (31.29mg, 395.52μmol) and trifluoroacetic anhydride (47.47mg ,226.01μmol), continue stirring for 0.2h. The reaction was washed with saturated sodium bicarbonate solution (10 mL), extracted three times with dichloromethane (10 mL), washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered and concentrated. Preparative HPLC separation [column type: Waters Xbridge BEH C18 100*30mm*10μm; mobile phase: [H 2 O(NH 4 HCO 3 )-acetonitrile]; acetonitrile%: 55%-75%, 8min] to obtain compound 4.
MS–ESI m/z:[M+H] +＝698.3. MS-ESI m/z: [M+H] + = 698.3.
实施例5Example 5
Figure PCTCN2022140910-appb-000050
Figure PCTCN2022140910-appb-000050
合成路线：synthetic route:
Figure PCTCN2022140910-appb-000051
Figure PCTCN2022140910-appb-000051
步骤1：化合物5-4的合成Step 1: Synthesis of compound 5-4
将噻唑[4,5-c]吡啶(2.16g,15.86mmol)溶于THF(15mL)中，氮气保护，冷却-78℃，滴加n-BuLi(2.5M THF溶液,6.35mL)，搅拌反应30分钟后，将化合物5-3(1g,3.17mmol)的THF(5mL)溶液滴加到反应液中，-78℃搅拌反应2小时。缓慢升温至0℃搅拌反应0.5hr。反应完毕，依次加入饱和氯化铵水溶液(20mL)和水(20mL)淬灭，乙酸乙酯(80mL*3)萃取，无水硫酸钠干燥，过滤，减压浓缩。残余物经柱层析(DCM:MeOH＝1:0至20:1)纯化后经高效液相制备分离(中性：柱型:Xtimate C18 150*40mm*5μm；流动相:[H 2O(NH 4OH v/v)-ACN]；ACN％:21％-51％,20min)得到化合物5-4。 Dissolve thiazo[4,5-c]pyridine (2.16g, 15.86mmol) in THF (15mL), under nitrogen protection, cool at -78°C, add n-BuLi (2.5M THF solution, 6.35mL) dropwise, and stir the reaction After 30 minutes, a solution of compound 5-3 (1 g, 3.17 mmol) in THF (5 mL) was added dropwise to the reaction liquid, and the reaction was stirred at -78°C for 2 hours. Slowly raise the temperature to 0°C and stir the reaction for 0.5hr. After the reaction was complete, it was quenched by adding saturated aqueous ammonium chloride (20 mL) and water (20 mL) successively, extracted with ethyl acetate (80 mL*3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by column chromatography (DCM:MeOH=1:0 to 20:1) and separated by HPLC preparation (neutral: column type: Xtimate C18 150*40mm*5μm; mobile phase: [H 2 O( NH 4 OH v/v)-ACN]; ACN%: 21%-51%, 20 min) to obtain compound 5-4.
MS–ESI m/z:[M+H] +＝391.1. MS-ESI m/z: [M+H] + = 391.1.
步骤2：化合物5-5的盐酸盐的合成Step 2: Synthesis of the hydrochloride salt of compound 5-5
将化合物5-4(100mg,256.11μmol)溶于EtOAc(2mL)中，20℃滴加HCl/EtOAc(4M,6mL)，滴加完毕后，20℃搅拌反应2hr。反应完毕后将反应液减压浓缩，加入乙酸乙酯(20mL)稀释。无需进一步纯化。减压浓缩得到化合物5-5的盐酸盐。Compound 5-4 (100mg, 256.11μmol) was dissolved in EtOAc (2mL), and HCl/EtOAc (4M, 6mL) was added dropwise at 20°C. After the addition was complete, the reaction was stirred at 20°C for 2hr. After the reaction was complete, the reaction solution was concentrated under reduced pressure, and ethyl acetate (20 mL) was added to dilute. No further purification was required. Concentration under reduced pressure gave the hydrochloride salt of compound 5-5.
MS–ESI m/z:[M+H] +＝290.9. MS-ESI m/z: [M+H] + = 290.9.
步骤3：化合物5-6的合成Step 3: Synthesis of Compound 5-6
将化合物5-5的盐酸盐(70mg,214.20μmol)和化合物3-4(96mg,214.97μmol)，HATU(126.00mg,331.38μmol)，TEA(32.72mg,323.30μmol,45μL)加入到DMF(1mL)中，25℃搅拌反应12小时。反应完毕后，依次加水(20mL)和饱和碳酸氢钠水溶液(10mL)稀释淬灭，用乙酸乙酯(50mL*3)萃取，有机相合并用无水硫酸钠干燥，过滤，减压浓缩得到化合物5-6。残余物无需进一步纯化，直接用于下一步。The hydrochloride of compound 5-5 (70mg, 214.20μmol) and compound 3-4 (96mg, 214.97μmol), HATU (126.00mg, 331.38μmol), TEA (32.72mg, 323.30μmol, 45μL) were added to DMF ( 1 mL), stirred at 25°C for 12 hours. After the reaction is complete, add water (20mL) and saturated aqueous sodium bicarbonate solution (10mL) successively to dilute and quench, extract with ethyl acetate (50mL*3), combine organic phases with anhydrous sodium sulfate, dry, filter, and concentrate under reduced pressure to obtain compound 5-6. The residue was used in the next step without further purification.
MS–ESI m/z:[M+H] +＝719.1. MS-ESI m/z: [M+H] + = 719.1.
步骤4：化合物5-7盐酸盐的合成Step 4: Synthesis of Compound 5-7 Hydrochloride
将粗品化合物5-6(180mg,250.38μmol)溶于EtOAc(2mL)中，冷却至0℃，滴加HCl/EtOAc(4M,62.60μL)，滴加完毕，20℃反应2hr.反应完毕后，将反应液减压浓缩，得到化合物5-7盐酸盐。MS–ESI m/z:619.1[M+H] +. Crude compound 5-6 (180mg, 250.38μmol) was dissolved in EtOAc (2mL), cooled to 0°C, and HCl/EtOAc (4M, 62.60μL) was added dropwise, and the reaction was completed at 20°C for 2hr. After the reaction was completed, The reaction solution was concentrated under reduced pressure to obtain compound 5-7 hydrochloride. MS–ESI m/z:619.1[M+H] + .
步骤5：化合物5的合成Step 5: Synthesis of compound 5
将粗品化合物5-7盐酸盐(200mg,323.21μmol)溶于MeOH(10mL)中，加入TEA(163.57mg,1.62mmol,225μL)和三氟乙酸甲酯(414.02mg,3.23mmol,326μL)，38℃搅拌反应12hr，反应完毕后将反应液减压浓缩。残余物经高效液相制备分离(柱型:Welch Xtimate C18 100*40mm*3μm；流动相:[H 2O(HCl)-ACN]；ACN％:30％-70％,9.5min)，得到化合物5。 The crude compound 5-7 hydrochloride (200 mg, 323.21 μmol) was dissolved in MeOH (10 mL), TEA (163.57 mg, 1.62 mmol, 225 μL) and methyl trifluoroacetate (414.02 mg, 3.23 mmol, 326 μL) were added, The reaction was stirred at 38° C. for 12 hours, and the reaction solution was concentrated under reduced pressure after the reaction was completed. The residue was separated by high performance liquid phase preparation (column type: Welch Xtimate C18 100*40mm*3μm; mobile phase: [H 2 O(HCl)-ACN]; ACN%: 30%-70%, 9.5min), to obtain the compound 5.
MS–ESI m/z:[M+H] +＝715.1. MS-ESI m/z: [M+H] + = 715.1.
1H NMR(400MHz,CD 3OD)δppm 9.80-9.53(m,1H),8.98-8.65(m,2H),5.85-5.49(m,1H),4.74-4.47(m,1H),4.46-3.58(m,4H),3.44-3.35(m,1H),2.87-2.04(m,6H),2.03-1.24(m,22H). 1 H NMR (400MHz, CD 3 OD) δppm 9.80-9.53(m,1H),8.98-8.65(m,2H),5.85-5.49(m,1H),4.74-4.47(m,1H),4.46-3.58 (m,4H),3.44-3.35(m,1H),2.87-2.04(m,6H),2.03-1.24(m,22H).
实施例6Example 6
Figure PCTCN2022140910-appb-000052
Figure PCTCN2022140910-appb-000052
合成路线：synthetic route:
Figure PCTCN2022140910-appb-000053
Figure PCTCN2022140910-appb-000053
步骤1：化合物6-2的合成Step 1: Synthesis of compound 6-2
将2-溴噻唑[5,4-b]吡啶(340.98mg,1.59mmol)溶于THF(5mL)中，氮气保护，冷却-78℃，滴加n-BuLi(2.5M THF溶液,634.18μL)，搅拌反应30分钟后，将化合物1-3(100mg,317.09μmol)的THF(1mL)溶液滴加到反应液中，-78℃搅拌反应2小时。反应完毕后，加水(20mL)淬灭(內温低于-50℃)，乙酸乙酯(30mL*3)萃取，有机相合并，减压浓缩。残余物经薄层色谱制备板(EtOAc)纯化。得到化合物6-2。Dissolve 2-bromothiazolo[5,4-b]pyridine (340.98mg, 1.59mmol) in THF (5mL), under nitrogen protection, cool to -78°C, add dropwise n-BuLi (2.5M THF solution, 634.18μL) , after stirring and reacting for 30 minutes, THF (1 mL) solution of compound 1-3 (100 mg, 317.09 μmol) was added dropwise to the reaction liquid, and stirred and reacted at -78°C for 2 hours. After the reaction was completed, water (20 mL) was added to quench (the internal temperature was lower than -50°C), extracted with ethyl acetate (30 mL*3), the organic phases were combined, and concentrated under reduced pressure. The residue was purified by thin layer chromatography preparative plate (EtOAc). Compound 6-2 was obtained.
MS–ESI m/z:[M+H] +＝391.0. MS-ESI m/z: [M+H] + = 391.0.
1H NMR(400MHz,CDCl 3)δppm 8.69(dd,J＝1.5,4.5Hz,1H),8.36(dd,J＝1.3,8.3Hz,1H),7.48(dd,J＝4.8,8.3Hz,1H),5.93-5.70(m,2H),5.59-5.43(m,1H),3.38-3.27(m,2H),2.64-2.49(m,2H),2.18-1.90(m,3H),1.37(s,9H).步骤2：化合物6-3的盐酸盐的合成 1 H NMR (400MHz, CDCl 3 ) δppm 8.69(dd, J=1.5,4.5Hz,1H),8.36(dd,J=1.3,8.3Hz,1H),7.48(dd,J=4.8,8.3Hz,1H ),5.93-5.70(m,2H),5.59-5.43(m,1H),3.38-3.27(m,2H),2.64-2.49(m,2H),2.18-1.90(m,3H),1.37(s , 9H). Step 2: Synthesis of the hydrochloride salt of compound 6-3
将化合物6-2(0.1g,256.11μmol)溶于EtOAc(1mL)，加入HCl/EtOAc(4M,64.03μL)在20℃下搅拌3hr。反应结束将反应液浓缩除去溶剂，得到化合物6-3的盐酸盐，直接用于下一步。Compound 6-2 (0.1 g, 256.11 μmol) was dissolved in EtOAc (1 mL), added with HCl/EtOAc (4M, 64.03 μL) and stirred at 20° C. for 3 hr. After the reaction, the reaction solution was concentrated to remove the solvent to obtain the hydrochloride of compound 6-3, which was directly used in the next step.
MS-ESI m/z:[M+H] +＝291.0. MS-ESI m/z: [M+H] + = 291.0.
步骤3：化合物6-4的合成Step 3: Synthesis of compound 6-4
将化合物6-3的盐酸盐(0.09g,309.98μmol)，化合物3-4(166.12mg,371.98μmol)溶于DMF(3mL)，加入HATU(235.73mg,619.96μmol)和DIEA(200.31mg,1.55mmol,269.97μL)，在20℃下搅拌1hr。反应结束后向反应液中加入EA(20mL)和水(20mL)，分液，将水相用EA(20mL)萃取两次，合并有机相用饱和NaHCO 3(20mL)和饱和NaCl(20mL)进行洗涤，有机相浓缩旋干。经柱层析分离(DCM：MeOH＝1:0至20:1)得到化合物6-4。 The hydrochloride of compound 6-3 (0.09g, 309.98μmol), compound 3-4 (166.12mg, 371.98μmol) was dissolved in DMF (3mL), added HATU (235.73mg, 619.96μmol) and DIEA (200.31mg, 1.55mmol, 269.97μL), stirred at 20°C for 1hr. After the reaction, add EA (20mL) and water (20mL) to the reaction solution, separate the layers, extract the aqueous phase with EA (20mL) twice, and combine the organic phases with saturated NaHCO 3 (20mL) and saturated NaCl (20mL). After washing, the organic phase was concentrated and spin-dried. Compound 6-4 was obtained by separation by column chromatography (DCM:MeOH=1:0 to 20:1).
ESI m/z:[M+H] +＝719.3. ESI m/z: [M+H] + ＝719.3.
步骤4：化合物6-5的盐酸盐的合成Step 4: Synthesis of the hydrochloride salt of compound 6-5
将化合物6-4(0.12g,166.92μmol)溶于EtOAc(1.3mL)，加入HCl/EtOAc(4M,1.23mL)，在20℃下搅拌3hr。反应结束将反应液浓缩除去溶剂，得到化合物6-5的盐酸盐无需纯化，直接用于下一步。Compound 6-4 (0.12 g, 166.92 μmol) was dissolved in EtOAc (1.3 mL), HCl/EtOAc (4M, 1.23 mL) was added, and stirred at 20° C. for 3 hr. After the reaction, the reaction solution was concentrated to remove the solvent to obtain the hydrochloride of compound 6-5, which was directly used in the next step without further purification.
MS-ESI m/z:[M+H] +＝619.3. MS-ESI m/z: [M+H] + = 619.3.
步骤5：化合物6的合成Step 5: Synthesis of compound 6
将化合物6-5(0.1g,161.61μmol)溶于MeOH(5mL)，加入TEA(81.76mg,808.03μmol,112.47μL)，三氟乙酸甲酯(206.94mg,1.62mmol,162.94μL)在38℃下搅拌16hr。之后将其用乙腈溶解。反应液经prep-HPLC(柱型:Phenomenex C18 80*40mm*3μm；流动相:[H 2O(NH 3H 2O)-ACN]；ACN％:50％-80％,8min)分离纯化得到化合物6。MS-ESI m/z:715.2[M+H] +. 1H NMR(400MHz,CDCl 3)δppm 8.69-8.83(m,1H)8.41-8.53(m,1H)7.51-7.64(m,1H)5.56-5.92(m,2H)4.45-4.62(m,1H)4.20-4.43(m,1H)3.75-4.04(m,1H)3.28-3.69(m,3H)3.07-3.19(m,2H)2.70-2.99(m,3H)2.48-2.68(m,1H)2.17-2.28(m,1H)1.96-2.09(m,5H)1.69-1.72(m,1H)1.67(br d,J＝16.31Hz,16H)1.40-1.49(m,3H). Compound 6-5 (0.1g, 161.61μmol) was dissolved in MeOH (5mL), TEA (81.76mg, 808.03μmol, 112.47μL), methyl trifluoroacetate (206.94mg, 1.62mmol, 162.94μL) were added at 38°C Stir for 16 hr. It was then dissolved with acetonitrile. The reaction solution was separated and purified by prep-HPLC (column type: Phenomenex C18 80*40mm*3μm; mobile phase: [H 2 O(NH 3 H 2 O)-ACN]; ACN%: 50%-80%, 8min) to obtain Compound 6. MS-ESI m/z:715.2[M+H] + . 1 H NMR(400MHz, CDCl 3 )δppm 8.69-8.83(m,1H)8.41-8.53(m,1H)7.51-7.64(m,1H)5.56 -5.92(m,2H)4.45-4.62(m,1H)4.20-4.43(m,1H)3.75-4.04(m,1H)3.28-3.69(m,3H)3.07-3.19(m,2H)2.70-2.99 (m,3H)2.48-2.68(m,1H)2.17-2.28(m,1H)1.96-2.09(m,5H)1.69-1.72(m,1H)1.67(br d,J＝16.31Hz,16H)1.40 -1.49(m,3H).
实施例7Example 7
Figure PCTCN2022140910-appb-000054
Figure PCTCN2022140910-appb-000054
合成路线：synthetic route:
Figure PCTCN2022140910-appb-000055
Figure PCTCN2022140910-appb-000055
步骤1：化合物7-3的合成Step 1: Synthesis of compound 7-3
选取50mL的三口烧瓶，将7-2(100mg,271.40μmol)和1-5(78.53mg,271.40μmol)溶于DMF(10mL)中，然后加入HATU(206.39mg,542.79μmol)和DIEA(105.23mg,814.19μmol,141.82μL)后，在20℃下搅拌12小时。反应完毕后，向所得反应液中加入30ml的水并用30ml的乙酸乙酯萃取两次，保留有机相，经饱和的食盐水洗涤，无水硫酸钠干燥，减压蒸馏浓缩得到产物粗品7-3。Choose a 50mL three-necked flask, dissolve 7-2 (100mg, 271.40μmol) and 1-5 (78.53mg, 271.40μmol) in DMF (10mL), then add HATU (206.39mg, 542.79μmol) and DIEA (105.23mg , 814.19 μmol, 141.82 μL), stirred at 20°C for 12 hours. After the reaction was completed, 30 ml of water was added to the resulting reaction solution and extracted twice with 30 ml of ethyl acetate, the organic phase was retained, washed with saturated brine, dried over anhydrous sodium sulfate, concentrated by distillation under reduced pressure to obtain the crude product 7-3 .
MS–ESI m/z:[M+Na] +＝662.1。 MS-ESI m/z: [M+Na] + = 662.1.
步骤2：化合物7-4盐酸盐的合成Step 2: Synthesis of Compound 7-4 Hydrochloride
选取50mL的三口烧瓶，将7-3(158mg,246.95μmol)溶于HCl/EtOAc(4M,5mL)中，在20℃下搅拌12小时。反应完毕后，减压蒸馏浓缩得到产物7-4盐酸盐。A 50 mL three-neck flask was selected, 7-3 (158 mg, 246.95 μmol) was dissolved in HCl/EtOAc (4M, 5 mL), and stirred at 20° C. for 12 hours. After the reaction was completed, the product was concentrated by distillation under reduced pressure to obtain the product 7-4 hydrochloride.
MS–ESI m/z:[M+H] +＝540.0。 MS-ESI m/z: [M+H] + = 540.0.
步骤3：化合物7的合成Step 3: Synthesis of compound 7
选取50mL的三口烧瓶将7-4盐酸盐(160mg,296.47μmol)溶于MeOH(10mL)中，然后加入三氟乙酸甲酯(379.63mg,2.96mmol,298.92μL)和TEA(180.00mg,1.78mmol,247.59μL)后，在20℃下搅拌12小时。反应完毕后，将反应液旋干得到粗品,经高效液相分离(柱型:Xtimate C18 150*40mm*5μm；流动相:[水(HCl)-ACN]；ACN％:40％-70％,10min)得到化合物7。Select a 50mL three-necked flask and dissolve 7-4 hydrochloride (160mg, 296.47μmol) in MeOH (10mL), then add methyl trifluoroacetate (379.63mg, 2.96mmol, 298.92μL) and TEA (180.00mg, 1.78 mmol, 247.59 μL), stirred at 20°C for 12 hours. After the reaction was completed, the reaction solution was spin-dried to obtain the crude product, which was separated by high performance liquid phase (column type: Xtimate C18 150*40mm*5 μm; mobile phase: [water (HCl)-ACN]; ACN%: 40%-70%, 10 min) to obtain compound 7.
MS–ESI m/z:[M+H] +＝636.1. MS-ESI m/z: [M+H] + = 636.1.
1H NMR(400MHz,CD 3OD)δppm 8.24-8.18(m,1H)8.16-8.09(m,1H)7.71-7.56(m,2H)5.78(dd,J＝11.8,3.3Hz,1H)4.72(s,1H)4.51(br s,1H)4.39(s,1H)3.41-3.33(m,2H)2.90-2.78(m,1H)2.59-2.47(m,2H)2.30-2.14(m,3H)2.14–1.91(m,4H)1.54(br d,J＝9.8Hz,1H)1.12(d,J＝7.0Hz,3H)1.08(s,9H)。 1 H NMR (400MHz, CD 3 OD) δppm 8.24-8.18 (m, 1H) 8.16-8.09 (m, 1H) 7.71-7.56 (m, 2H) 5.78 (dd, J = 11.8, 3.3Hz, 1H) 4.72 ( s,1H)4.51(br s,1H)4.39(s,1H)3.41-3.33(m,2H)2.90-2.78(m,1H)2.59-2.47(m,2H)2.30-2.14(m,3H)2.14 -1.91 (m, 4H) 1.54 (br d, J = 9.8Hz, 1H) 1.12 (d, J = 7.0Hz, 3H) 1.08 (s, 9H).
实施例8Example 8
Figure PCTCN2022140910-appb-000056
Figure PCTCN2022140910-appb-000056
步骤1：化合物8-2的合成Step 1: Synthesis of compound 8-2
选取100mL的三口烧瓶，将8-1(2.2g,5.78mmol)溶解在30mL的甲醇中，然后加入湿钯碳(0.5g,10％钯含量)，在氢气氛围下于25℃搅拌16h。待原料反应完成后，将所得反应液经硅藻土过滤得到反应液，减压浓缩得到粗产物8-2。In a 100 mL three-neck flask, 8-1 (2.2 g, 5.78 mmol) was dissolved in 30 mL of methanol, then wet palladium on carbon (0.5 g, 10% palladium content) was added, and stirred at 25° C. for 16 h under a hydrogen atmosphere. After the reaction of the raw materials was completed, the obtained reaction liquid was filtered through diatomaceous earth to obtain the reaction liquid, and concentrated under reduced pressure to obtain the crude product 8-2.
MS–ESI m/z:[M+Na] +＝405.1 MS-ESI m/z: [M+Na] + = 405.1
步骤2：化合物8-3盐酸盐的合成Step 2: Synthesis of Compound 8-3 Hydrochloride
选取100mL的三口烧瓶，将8-2(0.5g,1.31mmol)溶解在20mL的乙酸乙酯中，然后加入HCl/EtOAc(4M,3.26mL)，于25℃搅拌2h。待原料反应完成后，将所得反应液减压浓缩得到粗产物8-3盐酸盐。A 100 mL three-necked flask was selected, and 8-2 (0.5 g, 1.31 mmol) was dissolved in 20 mL of ethyl acetate, then HCl/EtOAc (4M, 3.26 mL) was added, and stirred at 25° C. for 2 h. After the reaction of the raw materials was completed, the resulting reaction solution was concentrated under reduced pressure to obtain the crude product 8-3 hydrochloride.
MS–ESI m/z:[M+H] +＝282.9。 MS-ESI m/z: [M+H] + = 282.9.
步骤3：化合物8-4的合成Step 3: Synthesis of Compound 8-4
选取50mL的三口烧瓶，将8-3盐酸盐(471mg,1.67mmol)溶于二氯甲烷(5mL)中，然后加入三氟乙酸甲酯(2.14g,16.68mmol,1.68mL)和TEA(1.01g,10.01mmol,1.39mL)后，在38℃下搅拌12小时。反应完毕后，将反应液旋干得到粗品8-4。Choose a 50mL three-necked flask, dissolve 8-3 hydrochloride (471mg, 1.67mmol) in dichloromethane (5mL), then add methyl trifluoroacetate (2.14g, 16.68mmol, 1.68mL) and TEA (1.01 g, 10.01mmol, 1.39mL), stirred at 38°C for 12 hours. After the reaction was completed, the reaction solution was spin-dried to obtain the crude product 8-4.
MS–ESI m/z:[M+H] +＝379.1。 MS-ESI m/z: [M+H] + = 379.1.
步骤4：化合物8-5的合成Step 4: Synthesis of compound 8-5
选取50mL的三口烧瓶，将8-4(0.5g,1.32mmol)溶于甲醇(10mL)中，然后加入一水合氢氧化锂(110.90mg,2.64mmol)和水(5mL)后，在25℃下搅拌12小时。反应完毕后，向所得反应液加入1M的盐酸调节pH 4～5，然后加入20mL的水并用20mL的乙酸乙酯萃取三次，保留有机相，加入饱和食盐水洗涤，无水硫酸钠干燥，减压蒸馏得到粗品8-5。Select a 50mL three-necked flask, dissolve 8-4 (0.5g, 1.32mmol) in methanol (10mL), then add lithium hydroxide monohydrate (110.90mg, 2.64mmol) and water (5mL), and then Stir for 12 hours. After the reaction was completed, 1M hydrochloric acid was added to the resulting reaction solution to adjust the pH to 4-5, then 20 mL of water was added and extracted three times with 20 mL of ethyl acetate, the organic phase was retained, washed with saturated brine, dried over anhydrous sodium sulfate, and decompressed. Distillation afforded crude 8-5.
MS–ESI m/z:[M+H] +＝365.0。 MS-ESI m/z: [M+H] + = 365.0.
步骤5：化合物8的合成Step 5: Synthesis of Compound 8
选取50mL的三口烧瓶，将8-5(232mg,318.37μmol,50％纯度)和6-3(208.09mg,318.37μmol,50％纯度)溶于DMF(10mL)中，然后加入HATU(242.11mg,636.73μmol)和DIEA(123.44mg,955.10μmol,166.36μL)后，在30℃下搅拌2小时。反应完毕后，向所得反应液中加入30mL的水并用30mL的乙酸乙酯萃取两次，保留有机相，经饱和的食盐水洗涤，无水硫酸钠干燥，减压蒸馏浓缩得到产物粗品，经高效液相制备分离(柱型:Phenomenex C18 80*40mm*3μm；流动相:[水(NH 3H 2O+NH 4HCO 3)-ACN]；ACN％:45％-75％,8min)得到产物8。 Choose a 50mL three-necked flask, dissolve 8-5 (232mg, 318.37μmol, 50% purity) and 6-3 (208.09mg, 318.37μmol, 50% purity) in DMF (10mL), then add HATU (242.11mg, 636.73 μmol) and DIEA (123.44 mg, 955.10 μmol, 166.36 μL), stirred at 30° C. for 2 hours. After the reaction was completed, 30 mL of water was added to the resulting reaction solution and extracted twice with 30 mL of ethyl acetate, the organic phase was retained, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated by distillation under reduced pressure to obtain the crude product. Liquid phase preparation and separation (column type: Phenomenex C18 80*40mm*3μm; mobile phase: [water (NH 3 H 2 O+NH 4 HCO 3 )-ACN]; ACN%: 45%-75%, 8min) to obtain the product 8.
MS–ESI m/z:[M+H] +＝637.1。 MS-ESI m/z: [M+H] + = 637.1.
1H NMR(400MHz,DMSO-d 6)δppm 8.83(br d,J＝6.3Hz,1H)7.75(dd,J＝8.3,4.6Hz,1H)7.67(s,1H)7.17(br s,3H)5.74-5.39(m,1H)4.58(s,1H)4.41(s,1H)4.22(s,1H)3.20(br t,J＝9.1Hz,1H)3.15-3.01(m,1H)2.66(br s,1H)2.33(s,3H)2.13–1.97(m,2H)1.94-1.71(m,3H)1.37(br d,J＝9.0Hz,1H)1.23(s,1H)0.97-1.02(s,9H)0.93-0.96(m,3H)。 1 H NMR (400MHz,DMSO-d 6 )δppm 8.83(br d,J=6.3Hz,1H)7.75(dd,J=8.3,4.6Hz,1H)7.67(s,1H)7.17(br s,3H) 5.74-5.39(m,1H)4.58(s,1H)4.41(s,1H)4.22(s,1H)3.20(br t,J＝9.1Hz,1H)3.15-3.01(m,1H)2.66(br s ,1H)2.33(s,3H)2.13–1.97(m,2H)1.94-1.71(m,3H)1.37(br d,J＝9.0Hz,1H)1.23(s,1H)0.97-1.02(s,9H )0.93-0.96 (m,3H).
实施例9Example 9
Figure PCTCN2022140910-appb-000057
Figure PCTCN2022140910-appb-000057
合成路线：synthetic route:
Figure PCTCN2022140910-appb-000058
Figure PCTCN2022140910-appb-000058
步骤1：化合物9-4-1的合成Step 1: Synthesis of Compound 9-4-1
反应瓶置换氮气，加入正己烷(4000mL)，加入二乙基锌溶液(1M正己烷溶液,1.05L)，降温至0℃，0℃搅拌0.2hr，至反应体系澄清，在0℃加入三氟化硼***(223.82g,1.58mol)，0℃搅拌0.5hr，在0℃加入二碘甲烷(563.15g,2.10mol)，0℃搅拌0.5hr，在0℃加入化合物8-1(50g,131.41mmol)，升至40℃搅拌16hr。过滤反应体系，滤饼用正己烷(1500mL)洗涤，将滤饼溶解在乙酸乙酯(5L)中，加入水(10L)，缓慢加入925g碳酸钠，再加入1.85kg乙二胺四乙酸二钠盐，搅拌15min，静置30min，分液，水相用乙酸乙酯(2L)萃取一次，合并有机相，有机相用饱和氯化钠溶液(3L)洗四次，无水硫酸钠干燥，过滤，减压浓缩。未经纯化直接用于下一步反应。得到化合物9-4-1。The reaction bottle was replaced with nitrogen, added n-hexane (4000mL), added diethyl zinc solution (1M n-hexane solution, 1.05L), cooled to 0°C, stirred at 0°C for 0.2hr, until the reaction system was clear, added trifluoro Boronium diethyl ether (223.82g, 1.58mol), stirred at 0°C for 0.5hr, added diiodomethane (563.15g, 2.10mol) at 0°C, stirred at 0°C for 0.5hr, added compound 8-1 (50g, 131.41 mmol), raised to 40°C and stirred for 16hr. Filter the reaction system, wash the filter cake with n-hexane (1500mL), dissolve the filter cake in ethyl acetate (5L), add water (10L), slowly add 925g of sodium carbonate, then add 1.85kg of disodium edetate salt, stirred for 15 min, stood still for 30 min, separated, the aqueous phase was extracted once with ethyl acetate (2L), the organic phase was combined, and the organic phase was washed four times with saturated sodium chloride solution (3L), dried over anhydrous sodium sulfate, filtered , concentrated under reduced pressure. It was directly used in the next reaction without purification. Compound 9-4-1 was obtained.
MS–ESI m/z:[M+H] +＝295.1。 MS-ESI m/z: [M+H] + = 295.1.
步骤2：化合物9-4-2的合成Step 2: Synthesis of Compound 9-4-2
将化合物9-4-1(26g,78.59mmol)溶于四氢呋喃(260mL)和水(85mL)中，加入碳酸氢钠(13.20g,157.17mmol)和二碳酸二叔丁酯(25.73g,117.88mmol)，20℃下搅拌16hr。向反应体系中加入乙酸乙酯(200mL)萃取，有机相用饱和氯化钠溶液(150mL)洗涤，无水硫酸钠干燥，过滤，减压浓缩。粗产品经石油醚(5V)打浆30min，过滤，滤饼旋干。得到化合物9-4-2。Compound 9-4-1 (26g, 78.59mmol) was dissolved in tetrahydrofuran (260mL) and water (85mL), sodium bicarbonate (13.20g, 157.17mmol) and di-tert-butyl dicarbonate (25.73g, 117.88mmol) were added ), stirred at 20°C for 16hr. Ethyl acetate (200 mL) was added to the reaction system for extraction, and the organic phase was washed with saturated sodium chloride solution (150 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was beaten with petroleum ether (5V) for 30 minutes, filtered, and the filter cake was spin-dried. Compound 9-4-2 was obtained.
MS–ESI m/z:[M+H] +＝395.2。 MS-ESI m/z: [M+H] + = 395.2.
步骤3：化合物9-4-3的合成Step 3: Synthesis of compound 9-4-3
将化合物9-4-2(23g,58.30mmol)溶于四氢呋喃(140mL)，加入水(50mL)，甲醇(50mL)，一水合氢氧化锂(7.34g,174.90mmol)，15℃搅拌16hr。反应液中加入乙酸乙酯(200mL)，水(50mL)，分离取水相，加入5％柠檬 酸(100mL)，乙酸乙酯(200mL×2)萃取，饱和氯化钠溶液(300mL)洗有机相，有机层经无水硫酸钠干燥后，过滤，减压浓缩得到化合物9-4-3。Compound 9-4-2 (23g, 58.30mmol) was dissolved in tetrahydrofuran (140mL), water (50mL), methanol (50mL), lithium hydroxide monohydrate (7.34g, 174.90mmol) were added, and stirred at 15°C for 16hr. Add ethyl acetate (200mL) and water (50mL) to the reaction solution, separate the aqueous phase, add 5% citric acid (100mL), extract with ethyl acetate (200mL×2), wash the organic phase with saturated sodium chloride solution (300mL) , the organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain compound 9-4-3.
MS–ESI m/z:[M+H] +＝381.1。 MS-ESI m/z: [M+H] + = 381.1.
步骤4：化合物9-4-4的盐酸盐的合成Step 4: Synthesis of the hydrochloride salt of compound 9-4-4
将化合物9-4-3(10g,26.28mmol)溶于二氯甲烷(38mL)中，加入氯化氢/二氧六环(4M,32.85mL)，20℃反应16hr。减压浓缩。得到化合物9-4-4的盐酸盐。Compound 9-4-3 (10 g, 26.28 mmol) was dissolved in dichloromethane (38 mL), hydrogen chloride/dioxane (4M, 32.85 mL) was added, and reacted at 20° C. for 16 hr. Concentrate under reduced pressure. The hydrochloride salt of compound 9-4-4 was obtained.
MS–ESI m/z:[M+H] +＝281.2。 MS-ESI m/z: [M+H] + = 281.2.
步骤5：化合物9-4的合成Step 5: Synthesis of compound 9-4
将化合物9-4-4的盐酸盐(8.3g,26.20mmol)溶于甲醇(83mL)，加入三乙胺(7.95g,78.59mmol,10.94mL)和三氟乙酸甲酯(13.42g,104.79mmol)，40℃搅拌16hr。旋干，用乙酸乙酯(10mL)和水(10mL)溶解，分液后，乙酸乙酯(10mL×2)萃取水相，合并有机相，经5％柠檬酸(20mL)和饱和食盐水(10mL)洗涤，无水硫酸钠干燥，过滤，旋干。乙酸乙酯：石油醚＝1:5(5V)打浆，过滤，旋干。得到化合物9-4。The hydrochloride (8.3g, 26.20mmol) of compound 9-4-4 was dissolved in methanol (83mL), triethylamine (7.95g, 78.59mmol, 10.94mL) and methyl trifluoroacetate (13.42g, 104.79 mmol), stirred at 40°C for 16hr. Rotate to dryness, dissolve with ethyl acetate (10mL) and water (10mL), separate the layers, extract the aqueous phase with ethyl acetate (10mL×2), combine the organic phases, wash with 5% citric acid (20mL) and saturated saline ( 10 mL), dried over anhydrous sodium sulfate, filtered, and spin-dried. Ethyl acetate: Petroleum ether = 1:5 (5V) for pulping, filtering, and spin-drying. Compound 9-4 was obtained.
MS–ESI m/z:[M+H] +＝377.3。 MS-ESI m/z: [M+H] + = 377.3.
步骤6：化合物9的合成Step 6: Synthesis of compound 9
将化合物9-4(95.59mg,253.98μmol)溶于DMF(5mL)中，加入DIEA(65.65mg,507.95μmol,88.48μL)后，在25℃下搅拌5分钟。然后向反应液中加入HATU(289.71mg,761.93μmol)和6-3的盐酸盐(83mg,253.98μmol)的DMF(3mL)溶液，加入反应完毕后，向反应液中加入水，用乙酸乙酯(50mL x 3)萃取。合并有机相，用饱和食盐水(30mL×3)洗涤，无水硫酸镁干燥，过滤，滤液减压浓缩除去溶剂。所得残余物经高效液相制备分离[柱型:Phenomenex C18 80*40mm*3μm；流动相:[水(含0.1％(氨水+碳酸氢铵))-乙腈]；乙腈％:43％-73％,8min]得到化合物9。Compound 9-4 (95.59 mg, 253.98 μmol) was dissolved in DMF (5 mL), DIEA (65.65 mg, 507.95 μmol, 88.48 μL) was added, and stirred at 25° C. for 5 minutes. Then add HATU (289.71mg, 761.93μmol) and DMF (3mL) solution of hydrochloride (83mg, 253.98μmol) to the reaction solution, after adding the reaction, add water to the reaction solution, wash with ethyl acetate Ester (50mL x 3) extraction. The organic phases were combined, washed with saturated brine (30 mL×3), dried over anhydrous magnesium sulfate, filtered, and the filtrate was concentrated under reduced pressure to remove the solvent. The obtained residue was prepared and separated by high performance liquid phase [column type: Phenomenex C18 80*40mm*3 μm; mobile phase: [water (containing 0.1% (ammonium water + ammonium bicarbonate))-acetonitrile]; acetonitrile%: 43%-73% ,8min] Compound 9 was obtained.
MS–ESI m/z:[M+H] +＝649.1。 MS-ESI m/z: [M+H] + = 649.1.
1H NMR(400MHz,DMSO-d 6)δppm 8.83(br d,J＝3.51Hz,1H)8.78(br d,J＝7.78Hz,1H)8.65(d,J＝8.28Hz,1H)7.79-7.70(m,1H)7.66(s,1H)5.59(br t,J＝8.41Hz,1H)4.64(br s,1H)4.22(s,1H)3.25-3.15(m,1H)3.14-3.07(m,1H)2.67(br s,1H)2.44-2.21(m,2H)2.13-1.94(m,2H)1.89-1.62(m,4H)1.59-1.44(m,1H)1.23(br s,2H)1.04–0.83(m,9H)0.78(br s,1H)0.67(br d,J＝3.26Hz,1H)0.52-0.31(m,2H)。 1 H NMR (400MHz, DMSO-d 6 ) δppm 8.83 (br d, J = 3.51Hz, 1H) 8.78 (br d, J = 7.78Hz, 1H) 8.65 (d, J = 8.28Hz, 1H) 7.79-7.70 (m,1H)7.66(s,1H)5.59(br t,J＝8.41Hz,1H)4.64(br s,1H)4.22(s,1H)3.25-3.15(m,1H)3.14-3.07(m, 1H)2.67(br s,1H)2.44-2.21(m,2H)2.13-1.94(m,2H)1.89-1.62(m,4H)1.59-1.44(m,1H)1.23(br s,2H)1.04– 0.83 (m, 9H) 0.78 (br s, 1H) 0.67 (br d, J=3.26Hz, 1H) 0.52-0.31 (m, 2H).
生物测试：Biological test:
实验例1：评价受试化合物的体外抗新型冠状病毒Mpro蛋白酶活性Experimental example 1: Evaluation of the in vitro anti-new coronavirus Mpro protease activity of the test compound
1.实验材料：1. Experimental materials:
1.1试剂与耗材：1.1 Reagents and consumables:
表1 试剂、耗材及其品牌Table 1 Reagents, consumables and their brands
Figure PCTCN2022140910-appb-000059
Figure PCTCN2022140910-appb-000059
1.2仪器：1.2 Instruments:
表2 仪器及其品牌Table 2 Instruments and their brands
 the 仪器instrument 品牌brand
11 SpectraMax M2e酶标仪SpectraMax M2e microplate reader Molecular DevicesMolecular Devices
22 Echo 655液体工作站Echo 655 Liquid Workstation LabcyteLabcyte
33 台式高速离心机Desktop high speed centrifuge EppendorfEppendorf
2.实验方法：2. Experimental method:
化合物溶于DMSO中，根据待测浓度要求使用Echo655按照3倍梯度稀释，10个浓度点，每个浓度双复孔，加入384孔板中。用测试缓冲液(100mM NaCl，20mM Tris-HCL，1mM EDTA)将Mpro蛋白和底物进行稀释，并将Mpro蛋白加入到384孔测试板中，和化合物室温孵育30min，然后加入底物，Mpro蛋白的测试浓度为25nM，底物的测试浓度为25μM。在30℃恒温培养箱中孵育60分钟。然后用酶标仪检测Ex/Em＝340nm/490nm荧光信号值。同时检测含底物和化合物但不含Mpro蛋白的背景孔为对照。The compound was dissolved in DMSO, and was diluted with Echo655 according to the 3-fold gradient according to the concentration to be tested, and 10 concentration points were added to the 384-well plate in duplicate wells for each concentration. Dilute Mpro protein and substrate with test buffer (100mM NaCl, 20mM Tris-HCL, 1mM EDTA), add Mpro protein to a 384-well test plate, and incubate with compound for 30min at room temperature, then add substrate, Mpro protein The test concentration was 25 nM for , and the test concentration for the substrate was 25 μM. Incubate for 60 minutes in a 30°C incubator. Then a microplate reader was used to detect the fluorescence signal value of Ex/Em=340nm/490nm. A background well containing both substrate and compound but no Mpro protein was detected as a control.
3.数据分析：3. Data analysis:
1)使用以下公式计算抑制率：1) Calculate the inhibition rate using the following formula:
抑制率％＝[(化合物-BG 化合物)-(ZPE-BG ZPE)]/[(HPE-BG HPE)-(ZPE-BG ZPE)]*100％ Inhibition rate%=[(Compound-BG Compound )-(ZPE-BG ZPE )]/[(HPE-BG HPE )-(ZPE-BG ZPE )]*100%
#HPE:100％抑制作用对照，包含25nM Mpro蛋白+25μM底物+1μM GC376 # HPE: 100% inhibition control, containing 25nM Mpro protein + 25μM substrate + 1μM GC376
ZPE:无抑制作用对照，包含25nM Mpro蛋白+25μM底物，不含化合物ZPE: no inhibition control, containing 25nM Mpro protein + 25μM substrate, no compound
化合物:测试化合物孔，包含25nM Mpro蛋白+25μM底物+化合物Compound: test compound well, containing 25nM Mpro protein + 25μM substrate + compound
BG:背景对照孔，包含25μM底物+化合物，不含Mpro蛋白BG: Background control wells containing 25 μM substrate+compound without Mpro protein
2)使用GraphPad Prism软件对化合物的抑制率数据(抑制率％)进行log(agonist)vs.res–nse--Variable slope非线性拟合分析，得到化合物的IC 50值。实验结果如表3所示： 2) Use GraphPad Prism software to perform log(agonist) vs. res–nse--Variable slope nonlinear fitting analysis on the inhibition rate data (inhibition rate%) of the compound to obtain the IC 50 value of the compound. The experimental results are shown in Table 3:
表3：受试化合物的体外抗新型冠状病毒Mpro蛋白酶活性Table 3: In vitro anti-new coronavirus Mpro protease activity of test compounds
化合物编号Compound number IC 50(nM) IC 50 (nM)
11 7474
22 3838
33 15.115.1
44 1010
55 1414
77 1111
88 4747
99 1010
结论：本发明化合物具有较好的体外抗新型冠状病毒Mpro蛋白酶活性。Conclusion: The compound of the present invention has better anti-new coronavirus Mpro protease activity in vitro.
实验例2：应用细胞病变模型评价化合物体外抗冠状病毒的活性Experimental example 2: Application of cytopathic model to evaluate the anti-coronavirus activity of compounds in vitro
1.实验材料1. Experimental materials
1.1.试剂与耗材1.1. Reagents and consumables
表4 试剂、耗材及其名称Table 4 Reagents, consumables and their names
 the 试剂与耗材名称Names of reagents and consumables 品牌brand
11 MEM培养基MEM medium SigmaSigma
22 L-谷氨酰胺(L-Glutamine)L-Glutamine GibcoGibco
33 非必需氨基酸non-essential amino acids GibcoGibco
44 双抗(Penicillin-Streptomycin Solution)Double Antibody (Penicillin-Streptomycin Solution) HyCloneHyClone
55 胎牛血清(FBS)Fetal bovine serum (FBS) ExCellExCell
66 磷酸盐缓冲液(DPBS)Phosphate buffered saline (DPBS) CorningCorning
77 0.25％胰酶0.25% trypsin GibcoGibco
88 CellTiter Glo细胞活性检测试剂盒CellTiter Glo Cell Viability Assay Kit PromegaPromega
99 瑞德西韦(Remdesivir)Remdesivir MCEMCE
1010 96孔板96-well plates GrenierGrenier
1.2.仪器1.2. Instruments
表5 仪器及其品牌Table 5 Instruments and their brands
 the 仪器instrument 品牌brand
11 酶标仪Microplate reader BioTekBioTek
22 细胞计数仪cell counter BeckmanBeckman
33 CO 2培养箱 CO2 incubator ThermoThermo
1.3.细胞和病毒1.3. Cells and viruses
MRC5细胞和冠状病毒HCoV OC43购自ATCC。MRC5 cells and coronavirus HCoV OC43 were purchased from ATCC.
MRC5细胞使用添加了10％胎牛血清(Excell)，1％双抗(Hyclone)，1％L-谷氨酰胺(Gibco)和1％非必需氨基酸(Gibco)的MEM(Sigma)培养液培养。添加了5％胎牛血清(Excell)，1％双抗(Hyclone)，1％L-谷氨酰胺(Gibco)和1％非必需氨基酸(Gibco)的MEM(Sigma)培养液为实验培养液。MRC5 cells were cultured in MEM (Sigma) medium supplemented with 10% fetal bovine serum (Excell), 1% double antibody (Hyclone), 1% L-glutamine (Gibco) and 1% non-essential amino acids (Gibco). The MEM (Sigma) medium supplemented with 5% fetal bovine serum (Excell), 1% double antibody (Hyclone), 1% L-glutamine (Gibco) and 1% non-essential amino acid (Gibco) was used as the experimental medium.
2.实验方法2. Experimental method
表6 本研究所用病毒试验方法Table 6 Virus test methods used in this study
Figure PCTCN2022140910-appb-000060
Figure PCTCN2022140910-appb-000060
细胞以一定密度(表2)接种到96微孔板中并于5％CO 2、37℃培养箱中培养过夜。第二天，加入倍比稀释后的化合物(8个浓度点、双复孔)，50μL每孔。随后稀释好的病毒以每孔100TCID 50加入细胞，50μL每孔。设置细胞对照(细胞，无化合物处理或病毒感染)，病毒对照(细胞感染病毒，无化合物处理)和培养液对照(只有培养液)。该实验培养液终体积为200μL，培养液中DMSO的终浓度分别为0.5％。细胞于5％CO 2、33℃培养箱中培养5天。使用细胞活力检测试剂盒CellTiter Glo(Promega)检测细胞活力。细胞毒性实验与抗病毒实验条件相同，但无病毒感染。 Cells were seeded into 96 microwell plates at a certain density (Table 2) and cultured overnight in a 5% CO 2 , 37° C. incubator. On the second day, the compound after doubling dilution (8 concentration points, duplicate wells) was added, 50 μL per well. Then the diluted virus was added to the cells at 100 TCID 50 per well, 50 μL per well. Set up cell control (cells, no compound treatment or virus infection), virus control (cells infected with virus, no compound treatment) and culture medium control (only culture medium). The final volume of the experimental culture solution was 200 μL, and the final concentration of DMSO in the culture solution was 0.5%. The cells were cultured in a 5% CO 2 , 33°C incubator for 5 days. Cell viability was detected using the cell viability assay kit CellTiter Glo (Promega). Cytotoxicity experiments were performed under the same conditions as antiviral experiments, but without virus infection.
3.数据分析3. Data Analysis
化合物的抗病毒活性和细胞毒性分别由不同浓度下的化合物对病毒引起的细胞病变效应的抑制率(％)和细胞活率(％)表示。计算公式如下：The antiviral activity and cytotoxicity of the compound are represented by the inhibitory rate (%) and cell viability (%) of the compound on the cytopathic effect caused by the virus at different concentrations, respectively. Calculated as follows:
抑制率(％)＝(测试孔读值-病毒对照平均值)/(细胞对照平均值-病毒对照平均值)×100Inhibition rate (%)=(test hole reading value-virus control average value)/(cell control average value-virus control average value)×100
细胞活率(％)＝(测试孔读值-培养液对照平均值)/(细胞对照平均值-培养液对照平均值)×100Cell viability (%) = (reading value of test well - average value of culture medium control) / (average value of cell control - average value of culture medium control) × 100
使用GraphPad Prism对化合物的抑制率和细胞活率进行非线性拟合分析，计算化合物的半数有效浓度(EC 50)和 半数细胞毒性浓度(CC 50)值。 GraphPad Prism was used to conduct nonlinear fitting analysis on the inhibition rate and cell viability of the compound, and calculate the half effective concentration (EC 50 ) and half cytotoxic concentration (CC 50 ) of the compound.
结论：本发明化合物具有较好的细胞水平的体外抗冠状病毒的活性，且无细胞毒性。Conclusion: The compound of the present invention has good in vitro anti-coronavirus activity at the cellular level, and has no cytotoxicity.
实验例3：应用冠状病毒OC43病毒株乳鼠感染模型评价受试化合物的体内抗病毒药效Experimental Example 3: Evaluation of the in vivo antiviral efficacy of the test compound using the coronavirus OC43 strain suckling mouse infection model
C57BL/6J乳鼠通过滴鼻感染致死剂量的冠状病毒，在感染前2小时后开始用溶媒(5％DMSO+40％PEG400+55％water)、本发明化合物处理乳鼠。实验期间，每日监控乳鼠的体重、健康状态及存活情况，以评估不同剂量下本发明化合物对乳鼠的保护作用。C57BL/6J suckling mice were infected with a lethal dose of coronavirus by intranasal instillation, and the suckling mice were treated with vehicle (5% DMSO+40% PEG400+55% water) and the compound of the present invention 2 hours before infection. During the experiment, the body weight, health status and survival of the suckling mice were monitored daily to evaluate the protective effect of the compound of the present invention on the suckling mice at different doses.
溶媒组乳鼠在病毒接种第6天后体重持续下降，终点存活率为0％。化合物9(25mpk)在感染前2小时首次给药，终点存活率为83.33％。实验结果见表7。The body weight of suckling mice in the vehicle group continued to decrease on the 6th day after virus inoculation, and the terminal survival rate was 0%. Compound 9 (25mpk) was administered for the first time 2 hours before infection, and the terminal survival rate was 83.33%. The experimental results are shown in Table 7.
表7.小鼠生存分析Table 7. Mice Survival Analysis
Figure PCTCN2022140910-appb-000061
Figure PCTCN2022140910-appb-000061
实验结论：本发明化合物具有良好的体内抗冠状病毒药效。Experimental conclusion: the compound of the present invention has good anti-coronavirus efficacy in vivo.

Claims (14)

  1. 式(II)所示化合物或其药学上可接受的盐，A compound represented by formula (II) or a pharmaceutically acceptable salt thereof,
    Figure PCTCN2022140910-appb-100001
    Figure PCTCN2022140910-appb-100001
    其中，in,
    R 1各自独立地选自卤素、CN、OH、NH 2、C 1-3烷基和C 1-3烷氧基，所述C 1-3烷基和C 1-3烷氧基各自独立地任选被1、2或3个卤素取代； Each R 1 is independently selected from halogen, CN, OH, NH 2 , C 1-3 alkyl and C 1-3 alkoxy, and each of the C 1-3 alkyl and C 1-3 alkoxy is independently optionally substituted by 1, 2 or 3 halogens;
    或者，or,
    两个R 1及它们相连的原子形成C 3-6环烷基，所述C 3-6环烷基任选被1、2或3个R a取代； Two R 1 and the atoms connected to them form a C 3-6 cycloalkyl group, and the C 3-6 cycloalkyl group is optionally substituted by 1, 2 or 3 R a ;
    R a各自独立地选自卤素和C 1-3烷基； R a is each independently selected from halogen and C 1-3 alkyl;
    n选自0、1、2和3；n is selected from 0, 1, 2 and 3;
    R 2选自叔丁基、C 3-10环烷基、3-10元杂环烷基和苯基，所述C 3-10环烷基、3-10元杂环烷基和苯基各自独立地任选被1、2或3个R b取代； R 2 is selected from tert-butyl, C 3-10 cycloalkyl, 3-10 membered heterocycloalkyl and phenyl, each of said C 3-10 cycloalkyl, 3-10 membered heterocycloalkyl and phenyl independently optionally substituted by 1, 2 or 3 R ;
    R b各自独立地选自卤素和C 1-3烷基； R b are each independently selected from halogen and C 1-3 alkyl;
    R 3选自C 1-3烷基、C 1-3烷氧基、-CH 2R 4和-CH 2OR 4，所述C 1-3烷基和C 1-3烷氧基各自独立地任选被1、2或3个卤素取代； R 3 is selected from C 1-3 alkyl, C 1-3 alkoxy, -CH 2 R 4 and -CH 2 OR 4 , and the C 1-3 alkyl and C 1-3 alkoxy are each independently optionally substituted by 1, 2 or 3 halogens;
    R 4选自苯基和5-6元杂芳基，所述苯基和5-6元杂芳基任选独立地被1、2或3个R取代； R is selected from phenyl and 5-6 membered heteroaryl, which are optionally substituted independently by 1, 2 or 3 R;
    R选自卤素和C 1-3烷基； R is selected from halogen and C 1-3 alkyl;
    T 1选自O和S； T1 is selected from O and S;
    A 1、A 2、A 3和A 4各自独立地选自N和CH； A 1 , A 2 , A 3 and A 4 are each independently selected from N and CH;
    环A选自
    Figure PCTCN2022140910-appb-100002
    Ring A is selected from
    Figure PCTCN2022140910-appb-100002
  2. 根据权利要求1所述的化合物或其药学上可接受的盐，其中，R 1选自F和甲基。 The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein R is selected from F and methyl.
  3. 根据权利要求1所述的化合物或其药学上可接受的盐，其中，两个R 1及它们相连的原子形成C 3-6环烷基。 The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein two R 1 and their connected atoms form a C 3-6 cycloalkyl group.
  4. 根据权利要求3所述的化合物或其药学上可接受的盐，其中，两个R 1及它们相连的原子形成环丙基。 The compound according to claim 3 or a pharmaceutically acceptable salt thereof, wherein two R 1 and their connected atoms form a cyclopropyl group.
  5. 根据权利要求1所述的化合物或其药学上可接受的盐，其中，结构单元
    Figure PCTCN2022140910-appb-100003
    选自
    Figure PCTCN2022140910-appb-100004
    Figure PCTCN2022140910-appb-100005
    The compound or pharmaceutically acceptable salt thereof according to claim 1, wherein the structural unit
    Figure PCTCN2022140910-appb-100003
    selected from
    Figure PCTCN2022140910-appb-100004
    Figure PCTCN2022140910-appb-100005
  6. 根据权利要求1所述的化合物或其药学上可接受的盐，其中，R 2选自叔丁基和金刚烷基，所述金刚烷基任选被1、2或3个R b取代。 The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein R 2 is selected from tert-butyl and adamantyl, and the adamantyl is optionally substituted by 1, 2 or 3 R b .
  7. 根据权利要求6所述的化合物或其药学上可接受的盐，其中，R 2选自叔丁基、
    Figure PCTCN2022140910-appb-100006
    The compound according to claim 6 or a pharmaceutically acceptable salt thereof, wherein R is selected from tert-butyl,
    Figure PCTCN2022140910-appb-100006
  8. 根据权利要求1所述的化合物或其药学上可接受的盐，其中，结构单元
    Figure PCTCN2022140910-appb-100007
    选自
    Figure PCTCN2022140910-appb-100008
    Figure PCTCN2022140910-appb-100009
    The compound or pharmaceutically acceptable salt thereof according to claim 1, wherein the structural unit
    Figure PCTCN2022140910-appb-100007
    selected from
    Figure PCTCN2022140910-appb-100008
    Figure PCTCN2022140910-appb-100009
  9. 根据权利要求1所述的化合物或其药学上可接受的盐，其中，R 3选自-CF 3。 The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein R 3 is selected from -CF 3 .
  10. 根据权利要求1-9任意一项所述的化合物或其药学上可接受的盐，其化合物选自式(II-1)、(II-2)和(II-3)所示的结构，The compound or pharmaceutically acceptable salt thereof according to any one of claims 1-9, wherein the compound is selected from the structures shown in formulas (II-1), (II-2) and (II-3),
    Figure PCTCN2022140910-appb-100010
    Figure PCTCN2022140910-appb-100010
    Figure PCTCN2022140910-appb-100011
    Figure PCTCN2022140910-appb-100011
    其中，in,
    t选自0、1和2；t is selected from 0, 1 and 2;
    R 1、R b、A 1、A 2、A 3、A 4、T 1和n如权利要求1-9任意一项所定义。 R 1 , R b , A 1 , A 2 , A 3 , A 4 , T 1 and n are as defined in any one of claims 1-9.
  11. 下式的化合物或其药学上可接受的盐，A compound of the following formula or a pharmaceutically acceptable salt thereof,
    Figure PCTCN2022140910-appb-100012
    Figure PCTCN2022140910-appb-100012
    Figure PCTCN2022140910-appb-100013
    Figure PCTCN2022140910-appb-100013
    Figure PCTCN2022140910-appb-100014
    Figure PCTCN2022140910-appb-100014
  12. 一种药物组合物，其含有治疗有效量的权利要求1-11任意一项所述的化合物或其药学上可接受的盐。A pharmaceutical composition, which contains a therapeutically effective amount of the compound of any one of claims 1-11 or a pharmaceutically acceptable salt thereof.
  13. 权利要求1-11任意一项所述的化合物或其药学上可接受的盐或权利要求12所述的组合物在制备治疗冠状病毒感染的药物中的应用。Application of the compound described in any one of claims 1-11 or a pharmaceutically acceptable salt thereof or the composition described in claim 12 in the preparation of a medicament for treating coronavirus infection.
  14. 根据权利要求13所述的应用，所述冠状病毒感染选自COVID-19。The application according to claim 13, the coronavirus infection is selected from COVID-19.
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