[go: up one dir, main page]

WO2020215998A1 - 嘧啶并五元杂环类化合物及其作为突变型idh2抑制剂的用途 - Google Patents

嘧啶并五元杂环类化合物及其作为突变型idh2抑制剂的用途 Download PDF

Info

Publication number
WO2020215998A1
WO2020215998A1 PCT/CN2020/082016 CN2020082016W WO2020215998A1 WO 2020215998 A1 WO2020215998 A1 WO 2020215998A1 CN 2020082016 W CN2020082016 W CN 2020082016W WO 2020215998 A1 WO2020215998 A1 WO 2020215998A1
Authority
WO
WIPO (PCT)
Prior art keywords
compound
substituted
unsubstituted
amine
methyl
Prior art date
Application number
PCT/CN2020/082016
Other languages
English (en)
French (fr)
Inventor
王猛
朱卫星
Original Assignee
上海仕谱生物科技有限公司
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 上海仕谱生物科技有限公司 filed Critical 上海仕谱生物科技有限公司
Priority to CN202080029927.1A priority Critical patent/CN114127063B/zh
Priority to US17/605,205 priority patent/US20220204509A1/en
Priority to JP2021562110A priority patent/JP2022530371A/ja
Priority to EP20794845.6A priority patent/EP3967691A4/en
Publication of WO2020215998A1 publication Critical patent/WO2020215998A1/zh

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D473/00—Heterocyclic compounds containing purine ring systems
    • C07D473/26—Heterocyclic compounds containing purine ring systems with an oxygen, sulphur, or nitrogen atom directly attached in position 2 or 6, but not in both
    • C07D473/32—Nitrogen atom
    • C07D473/34—Nitrogen atom attached in position 6, e.g. adenine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D473/00—Heterocyclic compounds containing purine ring systems
    • C07D473/26—Heterocyclic compounds containing purine ring systems with an oxygen, sulphur, or nitrogen atom directly attached in position 2 or 6, but not in both
    • C07D473/32—Nitrogen atom
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems

Definitions

  • the invention relates to the field of medicinal chemistry, in particular to a pyrimido five-membered heterocyclic compound and its use as a mutant IDH2 inhibitor.
  • Isocitrate dehydrogenase is a key rate-limiting enzyme that catalyzes the production of alpha-ketoglutarate ( ⁇ -KG) from isocitrate in the tricarboxylic acid cycle.
  • IDH1 is mainly located in the cytoplasmic matrix and peroxisomes, while IDH2 and IDH3 are mainly located in mitochondria.
  • IDH1 and IDH2 are in the form of homodimers and use nicotinamide adenine dinucleotide phosphate (NADP+) as a coenzyme to perform enzymatic catalytic function.
  • NADP+ nicotinamide adenine dinucleotide phosphate
  • IDH3 is a heterotetramer composed of 2 ⁇ subunits, 1 ⁇ subunit and 1 ⁇ subunit, and uses nicotinamide adenine dinucleotide (NAD+) as a coenzyme to catalyze the production of ⁇ -KG from isocitrate At the same time, NADH is produced to regulate the redox reaction in the cell.
  • NAD+ nicotinamide adenine dinucleotide
  • IDH1/2 have mutations in many different types of tumors. Including brain tumors, leukemia, chondrosarcoma, cholangiocarcinoma, etc. Compared with IDH1, IDH2 has a mutation rate of 8.7%-19% in acute myelocytic leukemia (AML). The mutation sites are mainly concentrated in R140Q and R172K. Mutated IDH2 (IDH2m) can cause loss of its normal function and convert ⁇ -KG into the carcinogenic metabolite 2-hydroxyglutarate (2-HG), allowing 2-HG to accumulate in the mutant tumor cells.
  • IDH2m can cause loss of its normal function and convert ⁇ -KG into the carcinogenic metabolite 2-hydroxyglutarate (2-HG), allowing 2-HG to accumulate in the mutant tumor cells.
  • ⁇ -KG is similar in structure to 2-HG, and 2-HG will competitively bind ⁇ -KG-dependent dioxygenase activities (such as DNA demethylase and histone demethylase), resulting in some Nucleosomes and/or DNA in key regions of the genome are highly methylated. This epigenetic change is thought to interfere with normal cell differentiation, leading to excessive proliferation of immature cells, thereby causing cancer.
  • ⁇ -KG-dependent dioxygenase activities such as DNA demethylase and histone demethylase
  • mutant IDH2 inhibitors can specifically bind to the mutant enzyme protein, effectively inhibit the activity of the mutant protease, and reduce the carcinogenic metabolite 2-HG in the body, thereby inducing the demethylation of histones and/or DNA To achieve the effect of promoting tumor cell differentiation and inhibiting tumor development.
  • Companies that take IDH2 mutants as targets and participate in new drug research and development are mainly represented by the United States Agios Pharmaceuticals.
  • the drug Enasidenib which targets the IDH2-R140Q mutation, was approved by the FDA on August 1, 2017 and was successfully marketed.
  • the diversification of IDH2 inhibitors is insufficient, so there is an urgent need to develop new, highly effective and low-toxicity drugs. IDH2m inhibitor.
  • the purpose of the present invention is to provide a class of mutant IDH2 inhibitors with high selectivity, high efficiency and low toxicity.
  • the first aspect of the present invention provides a pyrimido five-membered heterocyclic compound represented by formula I, or its stereoisomer or tautomer, or a pharmaceutically acceptable salt, hydrate or solvate,
  • R 1 is selected from none, hydrogen, halogen, -CN, substituted or unsubstituted C 1 -C 8 alkyl, substituted or unsubstituted C 2 -C 8 alkenyl, substituted or unsubstituted C 2 -C 8 alkyne Group, substituted or unsubstituted C 3 -C 10 cycloalkyl;
  • R 2 is selected from hydrogen, halogen, -CN, substituted or unsubstituted C 1 -C 8 alkyl, substituted or unsubstituted C 2 -C 8 alkenyl, substituted or unsubstituted C 2 -C 8 alkynyl, Substituted or unsubstituted C 3 -C 10 cycloalkyl group, substituted or unsubstituted C 1 -C 8 alkoxy group, substituted or unsubstituted C 1 -C 8 carboxyl group, substituted or unsubstituted C 2 -C 20 Ester group, substituted or unsubstituted C 6 -C 10 aryl group or substituted or unsubstituted 5-10 membered heteroaryl group having 1-3 heteroatoms selected from N, S and O;
  • X is selected from N, O, S or CR 5 ; wherein R 5 is hydrogen, halogen, -CN, substituted or unsubstituted C 1 -C 8 alkyl, substituted or unsubstituted C 2 -C 8 alkenyl, substituted Or unsubstituted C 2 -C 8 alkynyl, or substituted or unsubstituted C 3 -C 10 cycloalkyl;
  • n 1 is 0, 1, 2, 3, or 4; each L is independently selected from none, O, S, -CO-, -NH- or -CH 2 -;
  • n 2 is 0, 1 or 2; each Z is independently selected from none, O, S, -CO-, -NH- or -CH 2 -;
  • R 3 is selected from hydrogen, halogen, -CN, substituted or unsubstituted C 1 -C 8 alkyl, substituted or unsubstituted C 2 -C 8 alkenyl, substituted or unsubstituted C 2 -C 8 alkynyl, A substituted or unsubstituted C 3 -C 10 cycloalkyl group, a substituted or unsubstituted C 6 -C 10 aryl group, a substituted or unsubstituted 5- C with 1-3 heteroatoms selected from N, S and O 10-membered heteroaryl group, substituted or unsubstituted 4-8 membered heterocyclic group with 1-3 heteroatoms selected from N, S and O;
  • R 4 is selected from hydrogen, halogen, CN, substituted or unsubstituted C 1 -C 8 alkyl, substituted or unsubstituted C 2 -C 8 alkenyl, substituted or unsubstituted C 2 -C 8 alkynyl, substituted or Unsubstituted C 3 -C 10 cycloalkyl, substituted or unsubstituted C 6 -C 10 aryl, substituted or unsubstituted 5-10 member with 1-3 heteroatoms selected from N, S and O Heteroaryl
  • substituted refers to being substituted by one or more (for example, 2, 3, 4, etc.) substituents selected from the following group: halogen, C 1 -C 6 alkyl, halogen C 1 -C 6 alkyl, C 1 -C 6 alkoxy, halogenated C 1 -C 6 alkoxy, C 3 -C 8 cycloalkyl, halogenated C 3 -C 8 cycloalkane Group, oxo, -CN, hydroxyl, amino, carboxyl, benzyl, C 6 -C 10 aryl, halogenated C 6 -C 10 aryl, with 1-3 heterocyclic groups selected from N, S and O Atomic 5-10 membered heteroaryl group, halogenated 5-10 membered heteroaryl group having 1-3 heteroatoms selected from N, S and O.
  • substituents selected from the following group: halogen, C 1 -C 6 alkyl, halogen C 1 -C 6 alkyl, C 1
  • substitution refers to one or more (for example, 2, 3, 4, etc.) substituents selected from the group consisting of halogen, CN, hydroxyl, substitution or Unsubstituted C 1 -C 6 alkyl, C 1 -C 6 haloalkyl or substituted or unsubstituted C 1 -C 6 alkoxy.
  • substitution refers to substitution by one or more (for example, 2, 3, 4, etc.) substituents selected from the group consisting of halogen, CN, hydroxyl, substitution or Unsubstituted C 1 -C 6 alkyl or substituted or unsubstituted C 1 -C 6 haloalkyl.
  • the ester group includes -(substituted or unsubstituted C 1 -C 6 alkylene)-C(O)-O-(substituted or unsubstituted C 1 -C 6 alkyl ).
  • X is O or S, and R 1 is none.
  • the compound has the structure shown in formula Ia:
  • R 1 , R 2 , R 3 , R 4 , L, and m 1 are as defined above.
  • m 1 is 2, and -(L)m 1 -is -NH-CH 2 or -CH 2 -NH-.
  • m 2 is 2, and -(Z)m 2 -is -NH-CH 2 or -CH 2 -NH-.
  • L is NH
  • m 1 is 1, and Z is none, and m 2 is 0.
  • R 3 is a C 3 -C 6 cycloalkyl group.
  • R 2 is methyl or trifluoromethyl.
  • R 4 is a fluorine-substituted phenyl group.
  • X is CR 5 .
  • R 5 is H, C 1 -C 4 alkyl, or C 3 -C 4 cycloalkyl.
  • the compound is the compound #1, #2, #3, #4, #5, #6, #7, #8, #9, #10, #11, #12, #13, #14, #15, #16, #17, #18, #19, #20, #21, #22, #23, #24, #25, #26, #27, #28 , #29, #30, #31, #32, #33, #34, #35, #36, #37, #38, #39, #40, #41, #42, #43, #44, # 45, #46, #47, #48, #49, #50, or #51, or a pharmaceutically acceptable salt thereof.
  • the compound is compound #28, #48, #49 or #51 in Table 1, or a pharmaceutically acceptable salt thereof.
  • the compound is selected from:
  • the second aspect of the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising: (1) the compound according to the first aspect of the present invention, or its stereoisomers or tautomers, or pharmaceutically acceptable salts , Hydrate or solvate; (2) pharmaceutically acceptable carrier.
  • the third aspect of the present invention provides a compound as described in the first aspect of the present invention, or a stereoisomer or tautomer, or a pharmaceutically acceptable salt, hydrate or solvate thereof, or
  • the use of the pharmaceutical composition according to the second aspect of the invention is for preparing a medicine for preventing and/or treating a disease mediated by mutant IDH2.
  • the mutant IDH2 mediated disease is cancer; preferably, the cancer is selected from bladder cancer, breast cancer, kidney cancer, liver cancer, lung cancer (including small cell lung cancer), esophageal cancer, Gallbladder cancer, ovarian cancer, pancreatic cancer, gastric cancer, cervical cancer, thyroid cancer, prostate cancer and skin cancer (including squamous cell carcinoma); hematopoietic tumors of the lymphatic system, such as leukemia, acute lymphoblastic leukemia, acute lymphoblastic leukemia , B-cell lymphoma, T-cell lymphoma, Hodgkin’s lymphoma, non-Hodgkin’s lymphoma, hair cell lymphoma, and Burkitt’s lymphoma; tumors derived from mesenchymal cells, such as fibrosarcoma, rhabdomen Sarcoma; hematopoietic tumors of the myeloid lineage, including, for example, acute and chronic myelogenous leukemia
  • R 1 , R 2 , R 3 , R 4 , L, Z, m 1 , and m 2 are defined as described in the first aspect of the present invention.
  • R 1 , R 2 , R 3 , R 4 , L and m 1 are defined as described in the first aspect of the present invention.
  • the sixth aspect of the present invention provides a mutant IDH2 inhibitor, which comprises the compound of the first aspect of the present invention, or a stereoisomer or tautomer, or a pharmaceutically acceptable salt thereof, Hydrate or solvate or the pharmaceutical composition according to the second aspect of the invention.
  • a method for inhibiting the proliferation of tumor cells containing mutant IDH2 in vitro which comprises the steps of: combining the compound according to the first aspect of the present invention, or its stereoisomers or tautomers , Or a pharmaceutically acceptable salt, hydrate or solvate or the pharmaceutical composition according to the second aspect of the present invention is contacted with the mutant IDH2, thereby inhibiting the activity of the mutant IDH2.
  • the eighth aspect of the present invention provides a method for preventing and/or treating a disease mediated by mutant IDH2, comprising the steps of: administering the compound according to the first aspect of the present invention, or a stereoisomer thereof, to a desired subject Or a tautomer, or a pharmaceutically acceptable salt, hydrate or solvate, or the pharmaceutical composition according to the second aspect of the present invention.
  • the subject includes mammals, such as humans.
  • the compound of the present invention has excellent selectivity for mutant IDH2, extremely low toxicity to normal cells, and has good druggability and pharmacokinetic activity. On this basis, the present invention has been completed.
  • alkyl includes linear or branched alkyl groups.
  • C 1 -C 8 alkyl means a straight or branched alkyl group having 1-8 carbon atoms (preferably, 1-6, more preferably, 1-3), such as methyl, ethyl , Propyl, isopropyl, butyl, isobutyl, tert-butyl, etc.
  • alkenyl includes straight or branched chain alkenyl.
  • C 2 -C 8 alkenyl refers to a linear or branched alkenyl having 2-8 carbon atoms (preferably, 2-4), such as vinyl, allyl, 1-propenyl, isopropenyl Group, 1-butenyl, 2-butenyl, or similar groups.
  • alkynyl includes linear or branched alkynyl groups.
  • C 2 -C 8 alkynyl refers to a linear or branched alkynyl group having 2-8 carbon atoms (preferably, 2-4), such as ethynyl, propynyl, butynyl, or the like Group.
  • C 3 -C 10 cycloalkyl refers to a cycloalkyl having 3-10 carbon atoms (preferably, 3-6). It may be a monocyclic ring, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, or similar groups. It can also be in the form of a bicyclic ring, such as a bridged ring or a spiro ring.
  • C 1 -C 8 alkoxy refers to a linear or branched alkane having 1-8 carbon atoms (preferably, 1-6, more preferably, 1-3).
  • Oxy group for example, methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, tert-butoxy and the like.
  • the term "3-10 membered heterocyclic group having 1-3 heteroatoms selected from the group consisting of N, S, and O" refers to a group having 3-10 ring atoms and 1-3 ring atoms It is a saturated or partially saturated cyclic group of heteroatoms selected from the following group of N, S and O. It can be monocyclic, bicyclic or polycyclic, such as bridged or spirocyclic. Representative examples include but are not limited to: oxetane, azetidine, tetrahydro-2H-pyranyl, piperidinyl, tetrahydrofuranyl, morpholinyl, pyrrolidinyl, and the like.
  • C 6 -C 10 aryl group refers to an aryl group having 6-10 carbon atoms, for example, a phenyl group or a naphthyl group and the like.
  • the term "5-10 membered heteroaryl group having 1-3 heteroatoms selected from the group consisting of N, S, and O" refers to those having 5-10 ring atoms and wherein 1-3 ring atoms are A cyclic aromatic group selected from the group consisting of N, S and O heteroatoms. It may be a monocyclic ring or a condensed ring form.
  • pyridyl pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, pyrrolyl, pyrazolyl, imidazolyl, (1,2,3)-triazolyl and (1,2, 4)-Triazolyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, etc.
  • substituents selected from the following group: one or more selected from the group (for example 2, 3, 4, 5, etc.) Substituents are substituted: halogen, C 1 -C 6 alkyl, halogenated C 1 -C 6 alkyl, C 1 -C 6 alkoxy, halogenated C 1 -C 6 alkoxy, C 3 -C 8 cycloalkyl, halogenated C 3 -C 8 cycloalkyl, oxo, -CN, hydroxyl, amino, carboxy, benzyl, C 6 -C 10 aryl groups, halogenated C 6 -C 10 aryl groups, 5-10 membered heteroaryl groups having 1-3 heteroatoms selected from N, S and O, halogenated having 1-3 heteroatoms selected from N , S and O heteroatoms of 5-10 membered heteroaryl groups.
  • halogen or halogen atom refers to F, Cl, Br, and I. More preferably, the halogen or halogen atom is selected from F, Cl and Br. "Halogenated” refers to substitution with an atom selected from F, Cl, Br, and I.
  • the structural formula described in the present invention is intended to include all isomeric forms (such as enantiomers, diastereomers and geometric isomers (or conformational isomers)): for example, containing asymmetry The R and S configuration of the center, the (Z) and (E) isomers of the double bond, etc. Therefore, a single stereochemical isomer of the compound of the present invention or a mixture of its enantiomers, diastereomers or geometric isomers (or conformational isomers) belongs to the scope of the present invention.
  • tautomers means that structural isomers with different energies can exceed the low energy barrier to convert into each other.
  • proton tautomers ie, proton transfer
  • Valence tautomers include interconversion through some bond-forming electron recombination.
  • solvate refers to a complex in which the compound of the present invention coordinates with solvent molecules to form a specific ratio.
  • hydrate refers to a complex formed by coordination of the compound of the present invention with water.
  • the compound of the present invention refers to the compound represented by formula I, and also includes the isomer, racemate, crystal form or amorphous, pharmaceutically acceptable salt, hydrate or solvate of the compound of formula I. Things.
  • pharmaceutically acceptable salt refers to a salt formed by a compound of the present invention and an acid or base suitable for use as a medicine.
  • Pharmaceutically acceptable salts include inorganic salts and organic salts.
  • a preferred class of salts are the salts of the compounds of this invention with acids.
  • Acids suitable for salt formation include but are not limited to: hydrochloric acid, hydrobromic acid, hydrofluoric acid, sulfuric acid, nitric acid, phosphoric acid and other inorganic acids, formic acid, acetic acid, propionic acid, oxalic acid, malonic acid, succinic acid, fumaric acid, Organic acids such as maleic acid, lactic acid, malic acid, tartaric acid, citric acid, picric acid, methanesulfonic acid, toluenesulfonic acid, and benzenesulfonic acid; and acidic amino acids such as aspartic acid and glutamic acid.
  • a preferred class of salts are the salts of the compounds of this invention with bases.
  • Suitable bases for salt formation include, but are not limited to, inorganic bases such as sodium hydroxide, potassium hydroxide, sodium carbonate, sodium bicarbonate, and sodium phosphate, and organic bases such as ammonia, triethylamine, and diethylamine.
  • the compound of the present invention may be amorphous, crystalline, or a mixture thereof.
  • Certain compounds of the invention can exist in unsolvated as well as solvated forms, including hydrated forms.
  • the solvated form is usually equivalent to the unsolvated form and should be included in the scope of the present invention.
  • Certain compounds of the invention may exist in polymorphic or amorphous forms. Generally, as far as the application considered in the present invention is concerned, all physical forms are equivalent and should be included in the scope of the present invention.
  • the compounds of the present invention may also contain unnatural proportions of atomic isotopes at one or more of the isotopic atoms constituting such compounds.
  • the unnatural proportion of a certain isotope can be defined as the amount from the naturally found amount of the atom in question to 100% of that atom.
  • the compound can incorporate radioactive isotopes, such as tritium ( 3 H), iodine-125 ( 125 I), or carbon-14 ( 14 C), or non-radioactive isotopes, such as deuterium ( 2 H) or carbon-13 ( 13 C ).
  • radioactive isotopes such as tritium ( 3 H), iodine-125 ( 125 I), or carbon-14 ( 14 C)
  • non-radioactive isotopes such as deuterium ( 2 H) or carbon-13 ( 13 C ).
  • such isotopic variants may provide additional uses.
  • isotopic variants of the compounds of the invention may have additional uses, including, but not limited to, as diagnostic and/or imaging reagents, or as cytotoxic/radiotoxic therapeutic agents.
  • isotopic variants of the compounds of the invention may have altered pharmacokinetic and pharmacodynamic characteristics, thereby helping to increase safety, tolerability or efficacy during treatment. Regardless of whether it is radioactive or not, all isotopic variants of the compounds of the present invention should be included within the scope of the present invention.
  • the R 1 , R 2 , R 3 , R 4 , L, Z, m 1 , and m 2 are each independently a group corresponding to each compound in Table 1.
  • a method for preparing a compound of formula I comprising the steps:
  • R 1 , R 2 , R 3 , R 4 , L, Z, m 1 , and m 2 are as defined above.
  • a method for preparing the compound represented by formula Ia comprising the steps:
  • R 1 , R 2 , R 3 , R 4 , L, and m 1 are as defined above.
  • the compound of the present invention has excellent mutant IDH2 inhibitory activity and high selectivity
  • the compound of the present invention and the pharmaceutical composition containing the compound of the present invention as the main active ingredient can be used to prevent and/or treat (stabilize, reduce or cure) mutations Type IDH2-mediated related diseases.
  • Representative diseases include but are not limited to: bladder cancer, breast cancer, kidney cancer, liver cancer, lung cancer (including small cell lung cancer), esophageal cancer, gallbladder cancer, ovarian cancer, pancreatic cancer, gastric cancer, cervical cancer, thyroid cancer, prostate cancer Carcinoma and skin cancer (including squamous cell carcinoma); hematopoietic tumors of the lymphatic system, for example, including leukemia, acute lymphoblastic leukemia, acute lymphoblastic leukemia, B-cell lymphoma, T-cell lymphoma, Hodgkin’s lymphoma, Non-Hodgkin’s lymphoma, pilocytic lymphoma and Burkitt’s lymphoma; mesenchymal cell-derived tumors, such as fibrosarcoma and rhabdomyosarcoma; myeloid hematopoietic tumors, such as acute and chronic myelogenous leukemia, myeloproliferation Abnormal syndromes and prom
  • the pharmaceutical composition of the present invention contains a safe and effective amount of the compound of the present invention and a pharmaceutically acceptable excipient or carrier.
  • the "safe and effective amount” refers to: the amount of the compound is sufficient to significantly improve the condition without causing serious side effects.
  • the pharmaceutical composition contains 1-2000 mg of the compound of the present invention/agent, more preferably, 10-200 mg of the compound of the present invention/agent.
  • the "one dose” is a capsule or tablet.
  • “Pharmaceutically acceptable carrier” refers to: one or more compatible solid or liquid fillers or gel substances, which are suitable for human use, and must have sufficient purity and sufficiently low toxicity. "Compatibility” here means that each component of the composition can be blended with the compound of the present invention and between them without significantly reducing the efficacy of the compound.
  • pharmaceutically acceptable carriers include cellulose and its derivatives (such as sodium carboxymethyl cellulose, sodium ethyl cellulose, cellulose acetate, etc.), gelatin, talc, and solid lubricants (such as stearic acid).
  • Magnesium stearate calcium sulfate, vegetable oils (such as soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (such as propylene glycol, glycerin, mannitol, sorbitol, etc.), emulsifiers (such as Tween) ), wetting agents (such as sodium lauryl sulfate), coloring agents, flavoring agents, stabilizers, antioxidants, preservatives, pyrogen-free water, etc.
  • vegetable oils such as soybean oil, sesame oil, peanut oil, olive oil, etc.
  • polyols such as propylene glycol, glycerin, mannitol, sorbitol, etc.
  • emulsifiers such as Tween
  • wetting agents such as sodium lauryl sulfate
  • coloring agents such as sodium lauryl sulfate
  • flavoring agents such as pepperminophen, sorbitol, etc.
  • antioxidants
  • the method of administration of the compound or pharmaceutical composition of the present invention is not particularly limited, and representative administration methods include (but are not limited to): oral, parenteral (intravenous, intramuscular, or subcutaneous).
  • Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules.
  • the active compound is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or mixed with the following ingredients: (a) fillers or compatibilizers, for example, Starch, lactose, sucrose, glucose, mannitol and silicic acid; (b) binders such as hydroxymethyl cellulose, alginate, gelatin, polyvinylpyrrolidone, sucrose and gum arabic; (c) humectants, For example, glycerin; (d) disintegrants, such as agar, calcium carbonate, potato starch or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (e) slow solvents, such as paraffin; (f) Absorption accelerators, such as quaternary amine compounds; (g) wetting agents, such as cetyl alcohol and glycty
  • Solid dosage forms such as tablets, sugar pills, capsules, pills and granules can be prepared with coatings and shell materials, such as enteric coatings and other materials known in the art. They may contain opacifying agents, and the active compound or the release of the compound in such a composition may be released in a certain part of the digestive tract in a delayed manner. Examples of embedding components that can be used are polymeric substances and waxes. If necessary, the active compound can also be formed into microcapsules with one or more of the above-mentioned excipients.
  • Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups or tinctures.
  • the liquid dosage form may contain inert diluents conventionally used in the art, such as water or other solvents, solubilizers and emulsifiers, for example, ethanol, isopropanol, ethyl carbonate, ethyl acetate, propylene glycol, 1 , 3-Butanediol, dimethylformamide and oils, especially cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil or mixtures of these substances.
  • composition may also contain adjuvants such as wetting agents, emulsifying and suspending agents, sweetening agents, flavoring agents and perfumes.
  • adjuvants such as wetting agents, emulsifying and suspending agents, sweetening agents, flavoring agents and perfumes.
  • the suspension may contain suspending agents, for example, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methoxide and agar, or mixtures of these substances, and the like.
  • suspending agents for example, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methoxide and agar, or mixtures of these substances, and the like.
  • composition for parenteral injection may contain physiologically acceptable sterile aqueous or non-aqueous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions.
  • Suitable aqueous and non-aqueous carriers, diluents, solvents or excipients include water, ethanol, polyols and suitable mixtures thereof.
  • the compound of the present invention can be administered alone or in combination with other pharmaceutically acceptable compounds (for example, anticancer agents).
  • pharmaceutically acceptable compounds for example, anticancer agents.
  • the pharmaceutical composition When administered in combination, the pharmaceutical composition also includes one or more (2, 3, 4, or more) other pharmaceutically acceptable compounds (for example, anticancer agents).
  • One or more (2, 3, 4, or more) of the other pharmaceutically acceptable compounds (for example, anticancer agents) can be used simultaneously, separately or sequentially with the compound of the present invention Prevent and/or treat diseases mediated by mutant IDH2.
  • a safe and effective amount of the compound of the present invention is applied to a mammal (such as a human) in need of treatment, wherein the dosage is the pharmaceutically effective dosage considered to be administered.
  • the daily The dosage is usually 1 to 2000 mg, preferably 20 to 500 mg.
  • the specific dosage should also consider factors such as the route of administration, the patient's health status, etc., which are within the skill range of a skilled physician.
  • the compound of the present invention has a novel structure and excellent inhibitory effect on mutant IDH2, and the compound of the present invention has almost no activity on wild-type IDH2 (IDH2/WT) and has good selectivity.
  • the compound of the present invention has very low toxicity to normal cells, so it can be applied to a subject in a larger dose range.
  • the compound of the present invention has good medicamentability and can be easily prepared into a pharmaceutically acceptable salt, thereby helping to further form a preparation.
  • the compound of the present invention and the pharmaceutical composition containing the compound of the present invention as the main active ingredient can be used to prevent and/or treat diseases mediated by mutant IDH2.
  • the reference compound in the examples is Enasidenib (AG-221), CAS: 1446502-11-9, and the structural formula is as follows:
  • Step 2 9-isopropyl-N-(2-methoxyphenyl)-2-phenyl-9H-purin-6-amine (3 EPT60049)
  • Step 2 9-isopropyl-2-phenyl-N-(2-(trifluoromethyl)pyridin-4-yl)-9H-purin-6-amine (3 EPT60050)
  • Step 2 9-isopropyl-2-(6-(trifluoromethyl)pyridin-2-yl)-N-(2-(trifluoromethyl)pyridin-4-yl)-9H-purine-6 -Amine (3 EPT60061)
  • Step 1 9-isopropyl-2-(pyridin-3-yl)-N-(2-(trifluoromethyl)pyridin-4-yl)-9H-purin-6-amine (2 EPT60062)
  • Step 1 9-isopropyl-2-(pyridin-4-yl)-N-(2-(trifluoromethyl)pyridin-4-yl)-9H-purin-6-amine (2 EPT60072)
  • Step 1 9-isopropyl-2-(pyridin-2-yl)-N-(2-(trifluoromethyl)pyridin-4-yl)-9H-purin-6-amine (2 EPT60073)
  • reaction mixture and silica gel are concentrated by evaporation to obtain a powder residue.
  • Step 2 9-isopropyl-N,6-diphenyl-9H-purin-2-amine (EPT60086)
  • Step 3 N-Cyclopropyl-6-methyl-2-(naphthalene-2-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (7)
  • the product is a yellow solid N-cyclopropyl-6-methyl-2-(naphthalene-2-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (7,100 mg, 82.18% yield).
  • Step 4 N-cyclopropyl-6-methyl-2-(naphthalene-2-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine (EPT60187)
  • Step 1 N-isopropyl-6-methyl-2-(pyridin-2-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine (2 EPT60101)
  • Step 1 N-isopropyl-6-methyl-2-(pyridin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine (EPT60102)
  • Step 1 N-isopropyl-6-methyl-2-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine (EPT60103)
  • Step 3 6-Methyl-2-phenyl-7-tosyl-N-(2-(trifluoromethyl)pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidine- 4-amine (4)
  • Step 4 6-Methyl-2-phenyl-N-(2-(trifluoromethyl)pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine (5 EPT60121 )
  • Step 2 N-(2-fluoropyridin-4-yl)-6-methyl-2-phenyl-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (3 )
  • N-(2-fluoropyridine-4-acyl)-6-methyl-2-phenyl-7-tosyl-7H-pyrroline[2,3-d]pyrimidin-4-amine 22mg, 0.046 mmol was dissolved in CH 3 OH (3 mL), and CH 3 ONa (5.4 mol/L, 0.5 mL) was added. The reaction mixture was stirred at 60°C for 2h. After the reaction was completed, the reaction was quenched with saturated NH 4 Cl aqueous solution (1 mL), and the reaction was concentrated by evaporation to obtain oily residue.
  • Step 1 6-Methyl-N 2 , N 4 -diphenyl-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine (EPT60123)
  • Step 1 N-cyclopropyl-2-(2-fluorophenyl)-6-methyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (EPT60134)
  • Step 2 N-(Cyclopropylmethyl)-6-methyl-2-phenyl-7H-pyrrole[2,3-d]pyrimidin-4-amine (EPT60135) was added to compound 2 (68.5mg, 0.29mmol ) K 3 PO 4 (216.5mg, 1.02mmol), Pd(dppf)Cl 2 (47.4mg, 0.058mmol) and phenylboronic acid (176.9) were added to the dioxane (5mL) and H 2 O (1mL) solutions respectively. mg, 1.45mmol). The reaction mixture was stirred at 100° C. for 16 h (overnight). Under Ar conditions, LCMS (EPN18040-012-1) showed that the reaction was complete and 30% of SM remained.
  • Step 1 N-cyclopropyl-2-(3-fluorophenyl)-6-methyl-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (7)
  • Step 2 N-cyclopropyl-2-(3-fluorophenyl)-6-methyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (EPT60152)
  • Step 1 1-((2-Chloro-6-methyl-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)-2-methylpropan-2- Alcohol (2)
  • Step 2 2-Methyl-1-((6-methyl-2-phenyl-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)propan-2 -Alcohol (3)
  • Step 3 2-methyl-1-((6-methyl-2-phenyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)propan-2-ol (EPT60153)
  • Step 1 N-cyclopropyl-2-(4-fluorophenyl)-6-methyl-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(2)
  • reaction mixture and silica gel are concentrated by evaporation to obtain a powder residue.
  • Step 1 N-cyclopropyl-6-methyl-2-(o-tolyl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (3)
  • Step 4 2-(2-Phenyl-4-(phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-6-yl) ethyl acetate (6,EPT60168)
  • Step 1 N-isopropyl 2,6-diphenyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (5 EPT60181)
  • Step 1 4-(4-(Cyclopropylamino)-6-methyl-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-yl)phenol(2)
  • Step 1 N-isopropyl-2-phenyl-6-(trifluoromethyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine (2 EPT60184)
  • Step 1 N-cyclopentyl-2-(2-fluorophenyl)-6-methyl-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (3)
  • the inhibitory activity of the compound on IDH2/R140Q was determined by measuring the reduction of the cofactor NADPH.
  • the compound was incubated with IDH2/R140Q and NADPH, and then the reaction was initiated by adding ⁇ -KG. After a certain period of reaction under linear conditions, lipoamide dehydrogenase and the corresponding substrate resazurin were added for detection.
  • Lipoamide dehydrogenase terminates the IDH2/R140Q reaction by subtracting the available cofactor NADPH. It oxidizes NADPH to NADP and reduces resazurin to highly fluorescent resorufin. By detecting the production of resorufin To quantify the amount of cofactor NADPH remaining after a specific reaction time.
  • the specific operation method is: in a 96-well plate, the total reaction volume is 50 ⁇ L, 1.2nM IDH2/R140Q, a mixture of serially diluted inhibitors and 5 ⁇ M NADPH in a mixture containing Tris-HCl 50mM (pH 7.5), 150mM NaCl, 10mM MgCl 2. Pre-incubate at 25°C for 16 hours in a reaction buffer of 0.05% BSA, 10% Glycerol and 2.5mM ⁇ -mercaptoethanol. Add ⁇ -KG to 1mM, and react at 25°C for 40 minutes.
  • the termination mixture (lipoamide dehydrogenase 36 ⁇ g/mL, resazurin 30 ⁇ M) prepared by the above reaction buffer was added to convert resazurin to resorufin to measure the remaining NADPH. After incubating at 25°C for 10 minutes, the fluorescence value was measured by Tecan INFINITE F FLEX under Ex544/Em590. Each compound was tested for enzyme activity at 10 concentrations, and multiple background holes without enzyme and full enzyme activity holes without compound were set up in the reaction. The concentration of dimethyl sulfoxide in the system is less than or equal to 2%.
  • IDBS Software XLFit5 software
  • the multi-well plate of this example was purchased from Thermo Fisher Scientific, NADPH, ⁇ -KG, lipoamide dehydrogenase and resazurin were purchased from Shenggong Bioengineering Co., Ltd., and IDH2/R140Q was purchased from Abcam (ab198153).
  • the selected compounds of the present invention were analyzed, and the results are shown in Table 2.
  • “A” in Table 2 refers to the inhibitory activity against IDH2/R140Q with an IC 50 ⁇ 100nM
  • “B” refers to the inhibitory activity against IDH2/R140Q with an IC 50 ⁇ 1 ⁇ M
  • “C” refers to the inhibitory activity against IDH2/R140Q 1 ⁇ M ⁇ IC 50 ⁇ 10 ⁇ M inhibitory activity
  • D refers to IDH2/R140Q IC 50 >10 ⁇ M inhibitory activity.
  • the compound was incubated with IDH2/WT and NADP, and then the reaction was started by adding isocitrate. After a period of reaction under linear conditions, lipoamide dehydrogenase and resazurin were added to detect the amount of fluorescent substances.
  • NADP is reduced to NADPH.
  • the latter reduces resazurin to high-fluorescence resorufin under the action of lipoamide dehydrogenase.
  • the formation of resorufin is detected to quantify the cofactors generated after a specific reaction time.
  • the amount of NADPH can be used to calculate the compound's inhibitory effect on IDH2/WT.
  • the specific operation method is: in a 96-well plate, the total reaction volume is 50 ⁇ L, 0.6nM IDH2/WT, a mixture of serially diluted inhibitors and 50 ⁇ M NADP containing Tris-HCl 20mM (pH 7.5), 150mM NaCl, 10mM MgCl 2.
  • a reaction buffer of 10% Glycerol, 0.03% BSA and 2.5 mM ⁇ -mercaptoethanol pre-incubate at 25°C for 16 hours. Add isocitrate to 50 ⁇ M and react at 25°C for 30 minutes.
  • IDBS Software XLFit5 software
  • the multi-well plate was purchased from Thermo Fisher Scientific, NADP, ⁇ -KG, lipoamide dehydrogenase, and resazurin were purchased from Shenggong Bioengineering Co., Ltd., and IDH2/WT was purified by E. coli overexpression.
  • Table 3 The test results of some compounds are shown in Table 3.
  • 2-Hydroxyglutarate dehydrogenase (2HGDH) can reduce NAD+ to NADH in the presence of 2-HG.
  • the latter can be quantitatively determined by diaphorase and its substrate Resazurin (resazurin).
  • Glioma cell U87MG overexpressing IDH2/R140Q mutation cultured in 1% sodium pyruvate high-sugar MEM, 10% FBS, placed in a CO 2 incubator (37°C, 5% CO 2 , 95% air) .
  • the cells were trypsinized and seeded in a 96-well plate at a density of 1 ⁇ 10 4 with a medium of 200 ⁇ L and cultured overnight in a 37°C incubator.
  • the test compound was added the next day, and the final concentration of DMSO was 0.1%.
  • Add 50uL CellTiter-Glo purchased from Promega
  • reaction buffer 50mM Tris pH7.5, 100mM NaCl, 20mM MgCl 2 , 0.05% BSA
  • the final concentration of NAD+ is 40 ⁇ M
  • the final concentration of 2HGDH is 20nM
  • the test sample is added to 5 ⁇ L of culture medium;
  • the reaction solution was mixed and centrifuged, and reacted for 1 hour at 25°C in the dark;
  • color development buffer 50mM Tris pH7.5, 100mM NaCl, 20mM MgCl 2 , 0.05% BSA
  • the final concentration of diaphorase is 36 ⁇ g/mL
  • the final concentration of resazurin sodium It is 3 ⁇ M
  • 2-HG standard curve preparation Dilute the 2-HG stock solution to 20 ⁇ M with reaction buffer, and then perform a 2-fold gradient dilution for a total of 6 points. Then, the above-mentioned 2-HG was measured according to the extracellular 2-HG measurement system, and a standard curve was calculated and drawn.
  • the fluorescence value obtained in the extracellular 2-HG assay system was calculated using the 2-HG standard curve to calculate the content of 2-HG in the culture medium, and DMSO was used as a negative control to calculate the compound's inhibition of IDH2/R140Q mutation producing 2-HG activity.
  • A in Table 4 refers to the inhibitory activity against IDH2/R140Q at the cellular level with an IC 50 ⁇ 100nM;
  • B refers to the cellular level against IDH2/R140Q with an inhibitory activity of 100nM ⁇ IC 50 ⁇ 1 ⁇ M;
  • C It refers to the inhibitory activity of IDH2/R140Q at the cellular level with 1 ⁇ M ⁇ IC 50 ⁇ 10 ⁇ M;
  • D refers to the inhibitory activity at the cellular level with IC 50 of IDH2/R140Q >10 ⁇ M.
  • the results show that the tested compound can inhibit IDH2/R140Q mutant cells from producing 2-HG at a lower concentration, showing the compound's inhibitory effect on the activity of mutant IDH2 at the cellular level.

Landscapes

  • Organic Chemistry (AREA)
  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • General Chemical & Material Sciences (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Animal Behavior & Ethology (AREA)
  • Medicinal Chemistry (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Hematology (AREA)
  • Oncology (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Nitrogen Condensed Heterocyclic Rings (AREA)

Abstract

本发明涉及一种嘧啶并五元杂环类化合物及其作为突变型IDH2抑制剂的用途，具体地，本发明公开了一种可作为突变型IDH2抑制剂的嘧啶并五元杂环类化合物、或其立体异构体或互变异构体、或药学上可接受的盐、水合物或溶剂化物。本发明还涉及包含上述化合物的药物组合物及其用于制备预防和/或治疗突变型IDH2介导的疾病的药物的用途。

Description

嘧啶并五元杂环类化合物及其作为突变型IDH2抑制剂的用途 技术领域
本发明涉及医药化学领域，具体涉及一种嘧啶并五元杂环类化合物及其作为突变型IDH2抑制剂的用途。
背景技术
异柠檬酸脱氢酶(isocitrate dehydrogenase,IDH)是三羧酸循环中催化异柠檬酸生成alpha-酮戊二酸(α-KG)的关键限速酶。高等哺乳动物存在3种亚型，分别是IDH1、IDH2、IDH3。其中IDH1主要位于细胞质基质和过氧化物酶体，而IDH2和IDH3则主要定位在线粒体。IDH1，IDH2以同源二聚体的形式并以烟酰胺腺嘌呤二核苷酸磷酸(NADP+)作为辅酶行使酶催化功能。IDH3由2个α亚基、1个β亚基和1个γ亚基组成的异源四聚体，并以烟酰胺腺嘌呤二核苷酸(NAD+)为辅酶催化异柠檬酸生成α-KG，同时伴随产生NADH调控细胞内的氧化还原反应。
研究发现IDH1/2在许多不同类型的肿瘤中都存在突变。包括脑瘤、白血病、软骨肉瘤、胆管癌等。相比IDH1，IDH2在急性髓系白血病(acute myelocytic leukemia,AML)中的突变率率为8.7％-19％。其突变位点主要集中在R140Q和R172K。突变的IDH2(IDH2m)会导致其正常功能缺失，并将α-KG转化为致癌代谢物2-羟基戊二酸(2-HG)，使2-HG在突变的肿瘤细胞中累积。研究表明α-KG与2-HG结构相似，2-HG会竞争性结合α-KG依赖的双加氧酶的活性(如DNA去甲基化酶和组蛋白去甲基化酶)，导致一些基因组关键区域的核小体和/或DNA高度甲基化，这种表观遗传改变被认为干扰了正常的细胞分化，导致未成熟细胞过度增殖，从而引发癌症。
在含IDH2突变的肿瘤细胞中，突变型IDH2抑制剂可特异结合于突变酶蛋白，有效抑制突变蛋白酶活，使体内致癌代谢物2-HG减少，从而诱导组蛋白和/或DNA的去甲基化，达到促进肿瘤细胞分化，抑制肿瘤发展的效果。以IDH2突变体为靶点参与新药研发的公司主要以美国Agios制药公司为代表。其研发的靶向IDH2-R140Q突变的药物Enasidenib已于2017年8月1日获过FDA批准并成功上市，但IDH2抑制剂的研发化学型的多样化不足，因此急需开发新型的、高效低毒IDH2m抑制剂。
发明内容
本发明的目的是提供一类高选择性、高效低毒的突变型IDH2抑制剂。
本发明第一方面提供了一种式I所示的嘧啶并五元杂环类化合物、或其立体异构体或互变异构体、或药学上可接受的盐、水合物或溶剂化物，
Figure PCTCN2020082016-appb-000001
其中，
R 1选自无、氢、卤素、-CN、取代或未取代的C 1-C 8烷基、取代或未取代的C 2-C 8烯基、取代或未取代的C 2-C 8炔基、取代或未取代的C 3-C 10环烷基；
R 2选自氢、卤素、-CN、取代或未取代的C 1-C 8烷基、取代或未取代的C 2-C 8烯基、取代或未取代的C 2-C 8炔基、取代或未取代的C 3-C 10环烷基、取代或未取代的C 1-C 8烷氧基、取代或未取代的C 1-C 8羧基、取代或未取代的C 2-C 20酯基、取代或未取代的C 6-C 10芳基或取代或未取代的具有1-3个选自N、S和O的杂原子的5-10元杂芳基；
X选自N、O、S或CR 5；其中R 5为氢、卤素、-CN、取代或未取代的C 1-C 8烷基、取代或未取代的C 2-C 8烯基、取代或未取代的C 2-C 8炔基、或取代或未取代的C 3-C 10环烷基；
m 1为0、1、2、3、或4；各个L独立地选自无、O、S、-CO-、-NH-或-CH 2-；
m 2为0、1或2；各个Z独立地选自无、O、S、-CO-、-NH-或-CH 2-；
R 3选自氢、卤素、-CN、取代或未取代的C 1-C 8烷基、取代或未取代的C 2-C 8烯基、取代或未取代的C 2-C 8炔基、取代或未取代的C 3-C 10环烷基、取代或未取代的C 6-C 10芳基、取代或未取代的具有1-3个选自N、S和O的杂原子的5-10元杂芳基、取代或未取代的具有1-3个选自N、S和O的杂原子的4-8元杂环基；
R 4选自氢、卤素、CN、取代或未取代C 1-C 8烷基、取代或未取代的C 2-C 8烯基、取代或未取代的C 2-C 8炔基、取代或未取代的C 3-C 10环烷基、取代或未取代的C 6-C 10芳基、取代或未取代的具有1-3个选自N、S和O的杂原子的5-10元杂芳基；
除非特别说明，所述的“取代”是指被选自下组的一个或多个(例如2个、3个、4个等)取代基所取代：卤素、C 1-C 6烷基、卤代的C 1-C 6烷基、C 1-C 6烷氧基、卤代的C 1-C 6烷氧基、C 3-C 8环烷基、卤代的C 3-C 8环烷基、氧代、-CN、羟基、氨基、羧基、苄基、C 6-C 10芳基、卤代的C 6-C 10芳基、具有1-3个选自N、S和O的杂原子的5-10元杂芳基、卤代的具有1-3个选自N、S和O的杂原子的5-10元杂芳基。
在另一优选例中，对于R 3，所述取代是指被选自下组的一个或多个(例如2个、3个、4个等)取代基取代：卤素、CN、羟基、取代或未取代的C 1-C 6烷基、C 1-C 6卤代烷基或取代或未取代的C 1-C 6烷氧基。
在另一优选例中，对于R 4，所述取代是指被选自下组的一个或多个(例如2个、3个、4个等)取代基取代：卤素、CN、羟基、取代或未取代C 1-C 6烷基或取代或未取代C 1-C 6卤代烷基。
在另一优选例中，所述的酯基包括-(取代或未取代的C 1-C 6亚烷基)-C(O)-O-(取代或未取代的C 1-C 6烷基)。
在另一优选例中，X为O或S，且R 1为无。
在另一优选例中，所述化合物具有式Ia所示的结构：
Figure PCTCN2020082016-appb-000002
式中，R 1、R 2、R 3、R 4、L、和m 1的定义如上所述。
在另一优选例中，m 1为2，且-(L)m 1-为-NH-CH 2或-CH 2-NH-。
在另一优选例中，m 2为2，且-(Z)m 2-为-NH-CH 2或-CH 2-NH-。
在另一优选例中，L为NH，m 1为1，并且Z为无，m 2为0。
在另一优选例中，R 3为C 3-C 6环烷基。
在另一优选例中，R 2为甲基或三氟甲基。
在另一优选例中，R 4为氟取代的苯基。
在另一优选例中，X为CR 5。
在另一优选例中，R 5为H、C 1-C 4烷基、或C 3-C 4环烷基。
在另一优选例中，所述的化合物为表1中的化合物#1、#2、#3、#4、#5、#6、#7、#8、#9、#10、#11、#12、#13、#14、#15、#16、#17、#18、#19、#20、#21、#22、#23、#24、#25、#26、#27、#28、#29、#30、#31、#32、#33、#34、#35、#36、#37、#38、#39、#40、#41、#42、#43、#44、#45、#46、#47、#48、#49、#50、或#51，或其药学上可接受的盐。
在另一优选例中，所述化合物为表1中的化合物#28、#48、#49或#51，或其药学上可接受的盐。
在另一优选例中，所述化合物选自：
Figure PCTCN2020082016-appb-000003
Figure PCTCN2020082016-appb-000004
Figure PCTCN2020082016-appb-000005
本发明第二方面，提供了一种药物组合物，包含：(1)如本发明第一方面所述的化合物、或其立体异构体或互变异构体、或药学上可接受的盐、水合物或溶剂化物；(2)药学上可接受的载体。
本发明第三方面，提供了一种如本发明第一方面所述的化合物、或其立体异构体或互变异构体、或药学上可接受的盐、水合物或溶剂化物或如本发明第二方面所述的药物组合物的用途，用于制备预防和/或治疗突变型IDH2介导的疾病的药物。
在另一优选例中，所述突变型IDH2介导的疾病为癌症；较佳地，所述癌症选自膀胱癌、乳腺癌、肾癌、肝癌、肺癌(包括小细胞肺癌)、食道癌、胆囊癌、卵巢癌、胰腺癌、胃癌、宫颈癌、甲状腺癌、前列腺癌和皮肤癌(包括鳞状细胞癌)；淋巴系的造血肿瘤，例如包括白血病、急性淋巴细胞白血病、急性淋巴母细胞白血病、B细胞淋巴瘤、T-细胞淋巴瘤、霍奇金淋巴瘤、非霍奇金淋巴瘤、毛细胞淋巴瘤和伯基特淋巴瘤；间充质细胞来源的肿瘤，例如包括纤维肉瘤、横纹肌肉瘤；髓系的造血肿瘤，例如包括急慢性骨髓性白血病、骨髓增生异常综合征和前髓细胞白血病；中枢和周围神经系统肿瘤，例如包括星形细胞瘤、成神经细胞瘤、神经胶质瘤和神经鞘瘤；和其它肿瘤，例如包括黑素瘤、精原细胞瘤、畸胎癌、骨肉瘤、色性干皮病、角化棘皮瘤、甲状腺滤泡癌和卡波济氏肉瘤。
本发明第四方面，提供了一种式I化合物的制备方法，所述方法包括步骤：
Figure PCTCN2020082016-appb-000006
(a)将式(1)化合物与H-(L)m 1-R 3反应，从而制得式(2)化合物，其中H-(L)m 1-R 3为R 3取代的胺类化合物或者硼酸类化合物或者硼酸酯类化合物；和
Figure PCTCN2020082016-appb-000007
(b)将式(2)化合物与H-(Z)m 2-R 4反应，从而制得式(I)化合物，其中H-(Z)m 2-R 4为R 4取代的胺类化合物或者硼酸类化合物或者硼酸酯类化合物或者有机锡化合物；
式中，R 1、R 2、R 3、R 4、L、Z、m 1、m 2的定义如本发明第一方面所述。
本发明第五方面，提供了一种式Ia所示的化合物的制备方法，所述的方法包括步骤：
Figure PCTCN2020082016-appb-000008
(a1)将式(1a)的化合物与N-溴代丁二酰亚胺或者S-(三氟甲基)二苯并噻吩嗡四氟硼酸盐(Umemoto's reagents)反应，制得式(2a)化合物；和
(b1)将式(2a)化合物与硼酸类化合物R 2-B(OH) 2反应，从而制得式(Ia)所示的化合；
式中，R 1、R 2、R 3、R 4、L、m 1的定义如本发明第一方面所述。
本发明的第六方面，提供了一种突变型IDH2抑制剂，所述抑制剂包含本发明第一方面化合物、或其立体异构体或互变异构体、或药学上可接受的盐、水合物或溶剂化物或本发明第二方面所述的药物组合物。
本发明的第七方面，提供了一种体外抑制含突变型IDH2的肿瘤细胞增殖的方法，包括步骤：将本发明第一方面所述的化合物、或其立体异构体或互变异构体、或药学上可接受的盐、水合物或溶剂化物或本发明第二方面所述的药物组合物与突变型IDH2接触，从而抑制突变型IDH2的活性。
本发明的第八方面，提供了一种预防和/或治疗突变型IDH2介导的疾病的方法，包括步骤：向所需对象施用本发明第一方面所述的化合物、或其立体异构体或互变异构体、或药学上可接受的盐、水合物或溶剂化物或本发明第二方面所述的药物组合物。
在另一优选例中，所述对象包括哺乳动物，如人。
应理解，在本发明范围内中，本发明的上述各技术特征和在下文(如实施例)中具体描述的各技术特征之间都可以互相组合，从而构成新的或优选的技术方案。限于篇幅，在此不再一一累述。
具体实施方式
本发明人经过广泛而深入的研究，首次开发了一类对突变型IDH2具有优异抑制活性的新型化合物。本发明的化合物对于突变型IDH2具有优异的选择性，对正常细胞的毒性极低，并且具有具有良好的成药性和药代活性。在此基础上，完成了本发明。
定义
如本文所用，术语“烷基”包括直链或支链的烷基。例如C 1-C 8烷基表示具有1-8个碳原子(优选地，1-6个，更优选地，1-3个)的直链或支链的烷基，例如甲基、乙基、丙基、异丙基、丁基、异丁基、叔丁基等。
如本文所用，术语“烯基”包括直链或支链的烯基。例如C 2-C 8烯基指具有2-8个碳原子(优选地，2-4个)的直链或支链的烯基，例如乙烯基、烯丙基、1-丙烯基、异丙烯基、1-丁烯基、2-丁烯基、或类似基团。
如本文所用，术语“炔基”包括直链或支链的炔基。例如C 2-C 8炔基是指具有2-8个碳原子(优选地，2-4个)的直链或支链的炔基，例如乙炔基、丙炔基、丁炔基、或类似基团。
如本文所用，术语“C 3-C 10环烷基”指具有3-10个碳原子(优选地，3-6个)的环烷基。其可以是单环，例如环丙基、环丁基、环戊基、环己基、或类似基团。也可以是双环形式，例如桥环或螺环形式。
如本文所用，术语“C 1-C 8烷氧基”是指具有1-8个碳原子(优选地，1-6个，更优选地，1-3个)的直链或支链的烷氧基；例如，甲氧基、乙氧基、丙氧基、异丙氧基、丁氧基、异丁氧基、叔丁氧基等。
如本文所用，术语“具有1-3个选自下组N、S和O的杂原子的3-10元杂环基”是指具有3-10个环原子的且其中1-3个环原子为选自下组N、S和O的杂原子的饱和或部分饱和的环状基团。其可以是单环，也可以是双环或多环形式，例如桥环或螺环形式。代表性的实例包括但并不限于：氧杂环丁烷、氮杂环丁烷、四氢-2H-吡喃基、哌啶基、四氢呋喃基、吗啉基和吡咯烷基等。
如本文所用，术语“C 6-C 10芳基”是指具有6-10个碳原子的芳基，例如，苯基或萘基等类似基团。
如本文所用，术语“具有1-3个选自下组N、S和O的杂原子的5-10元杂芳基”指具有5-10个环原子的且其中1-3个环原子为选自下组N、S和O的杂原子的环状芳香基团。其可以是单环，也可以是稠环形式。具体的实例可以为吡啶基、哒嗪基、嘧啶基、吡嗪基、三嗪基、吡咯基、吡唑基、咪唑基、(1,2,3)-三唑基以及(1,2,4)-三唑基、四唑基、呋喃基、噻吩基、异噁唑基、噻唑基、噁唑基等。
本发明所述的基团除非特别说明是“取代的或未取代的”，否则本发明的基团均可被选自下组的取代基所取代：选自下组的一个或多个(例如2个、3个、4个、5个等)取代基所取代：卤素、C 1-C 6烷基、卤代的C 1-C 6烷基、C 1-C 6烷氧基、卤代的C 1-C 6烷氧基、C 3-C 8环烷基、卤代的C 3-C 8环烷基、氧代、-CN、羟基、氨基、羧基、苄基、C 6-C 10芳基、卤代的C 6-C 10芳基、具有1-3个选自N、S和O的杂原子的5-10元杂芳基、卤代的具有1-3个选自N、S和O的杂原子的5-10元杂芳基。
如本文所用，“卤素”或“卤原子”指F、Cl、Br、和I。更佳地，卤素或卤原子选自F、Cl和Br。“卤代的”是指被选自F、Cl、Br、和I的原子所取代。
除非特别说明，本发明所描述的结构式意在包括所有的同分异构形式(如对映异构，非对映异构和几何异构体(或构象异构体))：例如含有不对称中心的R、S构型，双键的(Z)、(E)异构体等。因此，本发明化合物的单个立体化学异构体或其对映异构体、非对映异构体或几何异构体(或构象异构体)的混合物都属于本发明的范围。
如本文所用，术语“互变异构体”表示具有不同能量的结构同分异构体可以超过低能垒，从而互相转化。比如，质子互变异构体(即质子移变)包括通过质子迁移进行互变，如1H-吲唑与2H-吲唑。化合价互变异构体包括通过一些成键电子重组而进行互变。
如本文所用，术语“溶剂合物”是指本发明化合物与溶剂分子配位形成特定比例的配合物。
如本文所用，术语“水合物”是指本发明化合物与水进行配位形成的配合物。
活性成分
如本文所用，“本发明化合物”指式I所示的化合物，并且还包括式I化合物的异构体、消旋体、晶型或无定形、药学上可接受的盐、水合物或溶剂合物。
如本文所用，“药学上可接受的盐”指本发明化合物与酸或碱所形成的适合用作药物的盐。药学上可接受的盐包括无机盐和有机盐。一类优选的盐是本发明化合物与酸形成的盐。适合形成盐的酸包括但并不限于：盐酸、氢溴酸、氢氟酸、硫酸、硝酸、磷酸等无机酸，甲酸、乙酸、丙酸、草酸、丙二酸、琥珀酸、富马酸、马来酸、乳酸、苹果酸、酒石酸、柠檬酸、苦味酸、甲磺酸、苯甲磺酸，苯磺酸等有机酸；以及天冬氨酸、谷氨酸等酸性氨基酸。一类优选的盐是本发明化合物与碱形成的盐。适合形成盐的碱包括但并不限于：氢氧化钠、氢氧化钾、碳酸钠、碳酸氢钠、磷酸钠等无机碱，氨水、三乙胺、二乙胺等有机碱。
本发明化合物可以是无定形的、晶型或其混合物。
本发明的某些化合物可以非溶剂化形式以及溶剂化形式存在，包括水化形式。溶剂化形式通常与非溶剂化形式等价，应包括在本发明范围内。本发明的某些化合物可以多晶型或无定形形式存在。通常，就本发明所考虑的应用而言，所有物理形式是等价的，应包括在本发明范围内。
本发明化合物还可在构成此类化合物的一个或多个同位素原子处含有非天然比例的原子同位素。某同位素的非天然比例可以定义为从所讨论原子的天然发现的量到100％该原子的量。例如，化合物可以掺入放射性同位素，例如氚( 3H)、碘-125( 125I)或碳-14( 14C)，或非放射性同位素，例如氘( 2H)或碳-13( 13C)。除了本申请所述的那些用途，此类同位素变体可提供额外的用途。例如，本发明化合物的同位素变体可以有额外的用途，包括但不限于作为诊断的和/或成像试剂，或作为细胞毒性/放射毒性治疗剂。另外，本发明化合物的同位素变体可具有改变的药代动力学和药效学特征，从而有助于增加治疗期间的安全性、耐受性或疗效。无论是否有放射性，本发明化合物的所有同位素变体均应包括在本发 明范围内。
在另一优选例中，所述的R 1、R 2、R 3、R 4、L、Z、m 1、m 2各自独立地为表1中各个化合物所对应的基团。
优选的本发明化合物如表1所示：
表1
Figure PCTCN2020082016-appb-000009
Figure PCTCN2020082016-appb-000010
Figure PCTCN2020082016-appb-000011
Figure PCTCN2020082016-appb-000012
制备方法
一种式I化合物的制备方法，所述方法包括步骤：
Figure PCTCN2020082016-appb-000013
(a)将式(1)化合物与H-(L)m 1-R 3反应，从而制得式(2)化合物，其中H-(L)m 1-R 3为R 3取代的胺类化合物或者硼酸类化合物或者硼酸酯类化合物；和
Figure PCTCN2020082016-appb-000014
(b)将式(2)化合物与H-(Z)m 2-R 4反应，从而制得式(I)化合物，其中H-(Z)m 2-R 4为R 4取代的胺类化合物或者硼酸类化合物或者硼酸酯类化合物或者有机锡化合物；
式中，R 1、R 2、R 3、R 4、L、Z、m 1、m 2的定义如上所述。
一种式Ia所示的化合物的制备方法，所述的方法包括步骤：
Figure PCTCN2020082016-appb-000015
Figure PCTCN2020082016-appb-000016
(a1)将式(1a)的化合物与N-溴代丁二酰亚胺或者S-(三氟甲基)二苯并噻吩嗡四氟硼酸盐(Umemoto's reagents)反应，制得式(2a)化合物；和
(b1)将式(2a)化合物与硼酸类化合物R 2-B(OH) 2反应，从而制得式(Ia)所示的化合；
式中，R 1、R 2、R 3、R 4、L、m 1的定义如上所述。
药物组合物和施用方法
由于本发明化合物具有优异的突变型IDH2抑制活性和高选择性，因此本发明化合物，以及含有本发明化合物为主要活性成分的药物组合物可用于预防和/或治疗(稳定、减轻或治愈)突变型IDH2介导的相关疾病。代表性的疾病包括但并不限于：膀胱癌、乳腺癌、肾癌、肝癌、肺癌(包括小细胞肺癌)、食道癌、胆囊癌、卵巢癌、胰腺癌、胃癌、宫颈癌、甲状腺癌、前列腺癌和皮肤癌(包括鳞状细胞癌)；淋巴系的造血肿瘤，例如包括白血病、急性淋巴细胞白血病、急性淋巴母细胞白血病、B细胞淋巴瘤、T-细胞淋巴瘤、霍奇金淋巴瘤、非霍奇金淋巴瘤、毛细胞淋巴瘤和伯基特淋巴瘤；间充质细胞来源的肿瘤，例如包括纤维肉瘤、横纹肌肉瘤；髓系的造血肿瘤，例如包括急慢性骨髓性白血病、骨髓增生异常综合征和前髓细胞白血病；中枢和周围神经系统肿瘤，例如包括星形细胞瘤、成神经细胞瘤、神经胶质瘤和神经鞘瘤；和其它肿瘤，例如包括黑素瘤、精原细胞瘤、畸胎癌、骨肉瘤、色性干皮病、角化棘皮瘤、甲状腺滤泡癌和卡波济氏肉瘤。
本发明的药物组合物包含安全有效量范围内的本发明化合物及药学上可以接受的赋形剂或载体。其中“安全有效量”指的是：化合物的量足以明显改善病情，而不至于产生严重的副作用。通常，药物组合物含有1-2000mg本发明化合物/剂，更佳地，含有10-200mg本发明化合物/剂。较佳地，所述的“一剂”为一个胶囊或药片。
“药学上可接受的载体”指的是：一种或多种相容性固体或液体填料或凝胶物质，它们适合于人使用，而且必须有足够的纯度和足够低的毒性。“相容性”在此指的是组合物中各组份能和本发明化合物以及它们之间相互掺和，而不明显降低化合物的药效。药学上可以接受的载体部分例子有纤维素及其衍生物(如羧甲基纤维素钠、乙基纤维素钠、纤维素乙酸酯等)、明胶、滑石、固体润滑剂(如硬脂酸、硬脂酸镁)、硫酸钙、植物油(如豆油、芝麻油、花生油、橄榄油等)、多元醇(如丙二醇、甘油、甘露醇、山梨醇等)、乳化剂(如吐温
Figure PCTCN2020082016-appb-000017
)、润湿剂(如十二烷基硫酸钠)、着色剂、调味剂、稳定剂、抗氧化剂、防腐剂、无热原水等。
本发明化合物或药物组合物的施用方式没有特别限制，代表性的施用方式包括(但并不限于)：口服、肠胃外(静脉内、肌肉内或皮下)。
用于口服给药的固体剂型包括胶囊剂、片剂、丸剂、散剂和颗粒剂。在这些固体剂型中，活性化合物与至少一种常规惰性赋形剂(或载体)混合，如柠檬酸钠或磷酸二钙，或与下述成分混合：(a)填料或增容剂，例如，淀粉、乳糖、蔗糖、葡萄糖、甘露醇和硅酸； (b)粘合剂，例如，羟甲基纤维素、藻酸盐、明胶、聚乙烯基吡咯烷酮、蔗糖和阿拉伯胶；(c)保湿剂，例如，甘油；(d)崩解剂，例如，琼脂、碳酸钙、马铃薯淀粉或木薯淀粉、藻酸、某些复合硅酸盐、和碳酸钠；(e)缓溶剂，例如石蜡；(f)吸收加速剂，例如，季胺化合物；(g)润湿剂，例如鲸蜡醇和单硬脂酸甘油酯；(h)吸附剂，例如，高岭土；和(i)润滑剂，例如，滑石、硬脂酸钙、硬脂酸镁、固体聚乙二醇、十二烷基硫酸钠，或其混合物。胶囊剂、片剂和丸剂中，剂型也可包含缓冲剂。
固体剂型如片剂、糖丸、胶囊剂、丸剂和颗粒剂可采用包衣和壳材制备，如肠衣和其它本领域公知的材料。它们可包含不透明剂，并且，这种组合物中活性化合物或化合物的释放可以延迟的方式在消化道内的某一部分中释放。可采用的包埋组分的实例是聚合物质和蜡类物质。必要时，活性化合物也可与上述赋形剂中的一种或多种形成微胶囊形式。
用于口服给药的液体剂型包括药学上可接受的乳液、溶液、悬浮液、糖浆或酊剂。除了活性化合物外，液体剂型可包含本领域中常规采用的惰性稀释剂，如水或其它溶剂，增溶剂和乳化剂，例知，乙醇、异丙醇、碳酸乙酯、乙酸乙酯、丙二醇、1,3-丁二醇、二甲基甲酰胺以及油，特别是棉籽油、花生油、玉米胚油、橄榄油、蓖麻油和芝麻油或这些物质的混合物等。
除了这些惰性稀释剂外，组合物也可包含助剂，如润湿剂、乳化剂和悬浮剂、甜味剂、矫味剂和香料。
除了活性化合物外，悬浮液可包含悬浮剂，例如，乙氧基化异十八烷醇、聚氧乙烯山梨醇和脱水山梨醇酯、微晶纤维素、甲醇铝和琼脂或这些物质的混合物等。
用于肠胃外注射的组合物可包含生理上可接受的无菌含水或无水溶液、分散液、悬浮液或乳液，和用于重新溶解成无菌的可注射溶液或分散液的无菌粉末。适宜的含水和非水载体、稀释剂、溶剂或赋形剂包括水、乙醇、多元醇及其适宜的混合物。
本发明化合物可以单独给药，或者与其他药学上可接受的化合物(例如抗癌剂)联合给药。
联合给药时，所述药物组合物还包括与一种或多种(2种，3种，4种，或更多种)其他药学上可接受的化合物(例如抗癌剂)。该其他药学上可接受的化合物(例如抗癌剂)中的一种或多种(2种，3种，4种，或更多种)可与本发明的化合物同时、分开或顺序地用于预防和/或治疗突变型IDH2介导的疾病。
使用药物组合物时，是将安全有效量的本发明化合物适用于需要治疗的哺乳动物(如人)，其中施用时剂量为药学上认为的有效给药剂量，对于60kg体重的人而言，日给药剂量通常为1～2000mg，优选20～500mg。当然，具体剂量还应考虑给药途径、病人健康状况等因素，这些都是熟练医师技能范围之内的。
本发明的主要优点包括：
(1)本发明的化合物结构新颖且具有优异的突变型IDH2抑制作用，且本发明的化合物 对野生型的IDH2(IDH2/WT)几乎没有活性，具有很好的选择性。
(2)本发明的化合物对正常细胞的毒性非常低，因而可以在较大的剂量范围内应用于治疗对象。
(3)本发明化合物具有良好的成药性，极易制成药学上可接受的盐，因而有助于进一步形成制剂。
(4)本发明化合物以及含有本发明化合物为主要活性成分的药物组合物可用于预防和/或治疗突变型IDH2介导的疾病。
下面结合具体实施例，进一步阐述本发明。应理解，这些实施例仅用于说明本发明而不用于限制本发明的范围。下列实施例中未注明具体条件的实验方法，通常按照常规条件，例如Sambrook等人，分子克隆：实验室手册(New York:Cold Spring Harbor Laboratory Press,1989)中所述的条件，或按照制造厂商所建议的条件。除非另外说明，否则百分比和份数是重量百分比和重量份数。本发明实施例中所用原料或仪器，若非特别说明，均市售可得。
实施例中的对照化合物为Enasidenib(AG-221)，CAS：1446502-11-9，结构式如下：
Figure PCTCN2020082016-appb-000018
实施例1
Figure PCTCN2020082016-appb-000019
Step 1:2-氯-9-异丙基-N-苯基-9H-嘌呤-6-胺(2)
在一个25毫升的圆底烧瓶中加入2,6-二氯-9-异丙基-9H-嘌呤(138mg,0.60mmol)和8毫升正丁醇。加入Et 3N(96mg,0.96mmol)和苯胺(67mg,0.72mmol)。反应混合物在100℃下搅拌16h(过夜)，并在真空中浓缩。采用自动闪柱柱色谱(硅胶，DCM:MeOH＝20:1)对残留进行纯化，得到2-氯-9-异丙基-N-苯基-9H-嘌呤-6-胺(144mg,83％收率)为白色固体。
ESI m/z:415.2(M+H) +.
Step 2:9异丙基-N，2-二苯基-9H-嘌呤-6-胺(3)
以2-氯-9-异丙基-N-苯基-9H-嘌呤-6-胺(100mg,0.35mmol)、苯基硼酸(50mg,0.4mmol)、Pd(PPh3) 4(40mg,0.03mmol)、K 2CO 3(140mg,1mmol)、甲苯(5mL)、水(1mL)充入10ml密封管。反应混合物在100℃惰性气体中搅拌3h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝1:1)对产物进行纯化，得到产物92mg，收率80％，为无色固体。
ESI m/z:330.2(M+H)+.1H NMR(DMSO-d6,400MHz)δ9.91(s,1H),8.41-8.44(m,3H),8.06(d,J＝7.6Hz 2H),7.38-7.54(m,3H),7.36(m,2H),7.06(t,J＝7.2Hz 1H),4.88-4.95(m,J＝6.8Hz,1H),1.63(d,J＝6.4Hz,6H)ppm.
实施例2
Figure PCTCN2020082016-appb-000020
Step 1:2-氯-9-异丙基-N-(2-甲氧基苯基)-9H-嘌呤-6-胺(2)
用2,6-二氯-9-异丙基-9H-嘌呤(115mg,0.50mmol)和5mL正丁醇充入10mL密封管。加入Et 3N(80mg,0.80mmol)和2-甲氧基苯胺(74mg,0.60mmol)。反应混合物在100℃下搅拌4h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝3:2)对产物进行纯化，得到产物为白色固体(95mg，收率60％)。
分子式:C 15H 16ClN 5O，分子量:317.78，ESI m/z:318.1(M+H) +.
Step 2:9-异丙基-N-(2-甲氧基苯基)-2-苯基-9H-嘌呤-6-胺(3 EPT60049)
以2-氯-9-异丙基N-(2-甲氧基苯基)-9H-嘌呤-6-胺(95mg,0.3mmol)、苯基硼酸(50mg,0.4mmol)、Pd(PPh3) 4(34mg,0.03mmol)、K 2CO 3(139mg,1mmol)、二氧六环(5mL)、水(1mL)充入10ml密封管。反应混合物在惰性气体中以100℃加热3h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱层析法(硅胶，EA:PE＝1:1)对产物进行纯化，得到产物(58mg,54％收率)为白色溶胶。
分子式:C 21H 21N 5O，分子量:359.43，(ESI)m/z＝360.2(M+H) +. 1H NMR(400MHz,DMSO-d 6)8.63-8.66(m,1H),8.41(m,4H),7.48-7.55(m,3H),7.10-7.17(m,3H),4.91(m,J＝6.8Hz,1H)3.94(s,3H)1.63(d,J＝6.8Hz,6H)ppm.
实施例3
Figure PCTCN2020082016-appb-000021
Step 1:2-氯-9-异丙基-N-(2-(三氟甲基)吡啶-4-基)-9H-嘌呤-6-胺(2)
用2,6-二氯-9-异丙基-9H-嘌呤(200mg,0.86mmol)和10mL DMSO充入10mL密封管。加入t-BuOK(150mg,1.33mmol)和2-(三氟甲基)吡啶-4-胺(180mg,1.11mmol)。反应混合物在100℃微波加热1h。反应后将混合物加水(50mL)。混合料用EA(20mL 3)萃取，有机层复合，在无水碳酸钠上干燥，过滤。然后用硅胶(100-200目)对有机层进行浓缩蒸发，得到粉状残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝7:3)对残留进行纯化，得到标题化合物(225mg，收率73％)为黄色固体。
分子式:C 14H 12ClF 3N 6,分子量:356.74,(ESI)m/z＝357.1(M+H) +.
Step 2:9-异丙基-2-苯基-N-(2-(三氟甲基)吡啶-4-基)-9H-嘌呤-6-胺(3 EPT60050)
用2-氯-9-异丙基N-(2-(三氟甲基)吡啶-4-基)-9H-嘌呤-6-胺(50mg,0.14mmol)、苯基硼酸(18mg,0.15mmol)、Pd(PPh3) 4(15mg,0.014mmol)、K 2CO 3(56mg,0.4mmol)、二氧六环(5mL)、水(1mL)分别充入10-mL密封管。反应混合物在惰性气体中加热到100摄氏度，加热1.5h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝4:1)对产物进行纯化，得到产物为白色固体(15mg，收率26.7％)。
分子式:C 20H 17F 3N 6,分子量:398.39,(ESI)m/z＝399.2(M+H) +. 1H NMR(400MHz,DMSO-d 6)10.84(s,1H),8.93(d,J＝2.0Hz,1H),8.61(d,J＝5.6Hz,1H),8.55(s,1H),8.39-8.41(m,2H),8.19-8.21(m,1H),7.48-7.54(m,3H),4.92(m,J＝6.8Hz,1H),1.61(d,J＝6.8Hz,6H)ppm.
实施例4
Figure PCTCN2020082016-appb-000022
Step 1(6-(三氟甲基)吡啶-2-基)硼酸(2)
用2-溴-6-(三氟甲基)吡啶(226mg,1.0mmol)、4、4、4′，4′，5、5、5′，5′，5′，5′，5′，5′，5′-八甲基-2,2′-bi(1,3,2-二噁硼烷)(280mg,1.1mmol)、Pd(dppf)Cl 2(140mg,0.2mmol)、AcOK(400mg,4.0mmol)分别充入10mL密封管。反应混合物在惰性气体中以90c加热4h。采用LCMS对反应进行监测，(ESI)m/z＝192.09，没有残留SM-1，下一步使用粗产物，未进一步纯化。
Step 2:9-异丙基-2-(6-(三氟甲基)吡啶-2-基)-N-(2-(三氟甲基)吡啶-4-基)-9H-嘌呤-6-胺(3 EPT60061)
用2-氯-9-异丙基N-(2-(三氟甲基)吡啶-4-基)-9H-嘌呤-6-胺(50mg,0.14mmol)，(6-(三氟甲基)吡啶-2-基)硼酸(2.5mL二氧六环原液)，Pd(dppf)Cl 2(10mg,0.014mmol)，K 2CO 3(56mg,0.4mmol)，加入水(0.5mL)，密封管20ml。反应混合物在惰性气体中加热到90度，加热3h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。渣油通过蒸发浓缩而成黑色油渣。采用自动闪柱柱色谱法(硅胶，MeOH:DCM＝1:24)对产物进行纯化，得到产物为白色固体(30mg，收率46％)。
分子式:C 20H 15F 6N 7,分子量:467.38,(ESI)m/z＝468.2(M+H) +. 1HNMR(400MHz,DMSO-d6)10.93(s,1H),9.01(d,J＝2.0Hz,1H),8.70(d,J＝8.0Hz,1H),8.66(s,1H),8.53-8.54(d,J＝5.6Hz,1H),8.42-8.44(m,1H),8.26(t,J＝8.0Hz 1H),8.00(d,J＝7.6Hz,1H)4.95(m,J＝6.8Hz,1H),1.60-1.62(d,J＝6.8Hz,6H)ppm.
实施例5
Figure PCTCN2020082016-appb-000023
Step 1:9-异丙基-2-(吡啶-3-基)-N-(2-(三氟甲基)吡啶-4-基)-9H-嘌呤-6-胺(2 EPT60062)
以2-氯-9-异丙基-N-(2-(三氟甲基)吡啶-4-酰基)-9H-嘌呤-6-胺(36mg,0.10mmol)、吡啶-3-基硼酸(37mg,0.30mmol)、Pd(dppf)Cl 2(7mg,0.01mmol)、K 3PO 4(84mg,0.4mmol)、二氧六环(2.5mL)、水(0.5mL)分别充入10ml密封管。反应混合物在惰性气体中以100℃加热4h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，MeOH:DCM＝1:10)对产物进行纯化，得到产物为白色固体(20mg，收率50％)。
分子式:C 19H 16F 3N 7,分子量:399.38,(ESI)m/z＝400.2(M+H) +. 1HNMR(400MHz,DMSO-d 6)10.89(s,1H),9.52(d,J＝1.6Hz,1H),8.82(d,J＝2.0Hz,1H),8.58-8.68(m,4H),8.21-8.23(m,1H),7.53-7.56(m,1H),4.92(m,J＝6.8Hz,1H),1.60-1.62(d,J＝7.2Hz,6H)ppm.
实施例6
Figure PCTCN2020082016-appb-000024
Step 1:2-氯-N-苯基-9H-嘌呤-6-胺(2)
一个25毫升的圆底烧瓶中装有2,6-二氯-9H-嘌呤(2,589mg,3.11mmol)和10毫升正丁醇。加入Et 3N(944mg,9.35mmol)和苯胺(348mg,3.74mmol)。反应混合物在100℃下搅拌16h(过夜)，LCMS分析确认反应完成。然后用MeOH对沉淀物进行过滤和洗涤，得到的产物产率为66％(505mg)，为灰白色固体。ESI m/z:246.1(M+H) +. 1H NMR(DMSO-d 6,400MHz)δ7.08(d,J＝7.2Hz,1H),7.36(t,J＝7.6Hz,2H),7.84(d,J＝7.6Hz,2H),8.30(s,1H),10.18(s,1H),13.29(s,1H)ppm.
Step 2:N，2-二苯基-9H-嘌呤-6-胺(3,EPT60063)
在一个10毫升的微波管中加入化合物2(65mg,0.26mmol)和二氧六环(2.5mL)和水(0.5mL)的混合物。加入K 2CO 3(108mg,0.78mmol)、苯基硼酸(39mg,0.32mmol)和Pd(dppf)Cl 2(19mg,0.026mmol)。反应混合物在100℃氩气微波下搅拌4小时。通过LCMS分析证实了反应的完成。将混合物冷却至室温(20℃)，形成沉淀，过滤收集。粗品经硅胶柱层析(DCM/MeOH＝10/1)纯化得到所需的白色固体(38mg，产率54％)。ESI m/z:288.16(M+H) +. 1H NMR(DMSO-d 6,400MHz)δ7.07(d,J＝7.2Hz,1H),7.40(t,J＝7.6Hz,2H),7.53-7.43(m,3H),8.05(d,J＝8.0Hz,2H),8.30(s,1H),8.39-8.37(m,2H),9.84(s,1H),13.20(s,1H)ppm.
实施例7
Figure PCTCN2020082016-appb-000025
Step 1:2-氯-6-甲基-4-苯基-7H-吡咯并[2,3-d]嘧啶(2)
将2,4-二氯-6-甲基-7H-吡啶(1)(150mg,0.743mmol)、苯基硼酸(145mg,1.19mmol)、碳酸钾(513mg,3.71mmol)、二(三苯基膦)氯化钯(II)(84mg,0.12mmol)在二氧六环/水(10/1,13mL)中，在80℃氮气气氛下搅拌过夜。冷却至室温后，用乙酸乙酯(30ml)和水(30ml)稀释。分离有机层，用乙酸乙酯(30ml)萃取水层。采用C 18柱(乙腈/水浓度为55％～60％)对合成的有机层进行浓缩纯化，得到标题化合物2(110mg，产率61％)为灰白色固体。
LC-MS[流动相:在2.5min内，从95％水和5％CH 3CN到5％水和95％CH 3CN],Rt＝1.64min；MS计算值:243.1；MS实测值:244.0[M+H] +.
1H NMR(400MHz,DMSO-d 6)δ12.32(s,1H),8.13-8.11(m,2H),7.61-7.56(m,3H),6.67(s,1H),2.45(s,3H).
Step 2:6-甲基-N，4-二苯基-7H-吡咯并[2,3-d]嘧啶-2-胺(3)
将2-氯-6-甲基-4-苯基-7H-吡咯[2,3-d]嘧啶(2)(90mg,0.37mmol)和苯胺(103mg,1.11mmol)溶于乙二醇(3mL)中，加一滴浓盐酸水溶液。混合物在150℃微波和氮气气氛下，在密封管中搅拌10小时。冷却至室温后，将混合物倒入水中(30ml)，用乙酸乙酯萃取(15ml×2)，用盐水(30ml)冲洗并浓缩。采用C 18柱(乙腈/水从60％～70％)对残留进行纯化，得到标题化合物(90mg，收率82％)为灰白色固体。
LC-MS纯度:95.84％(214nm),98.12％(254nm)；MS计算值:300.1；MS实测值:301.0[M+H] +.
1H NMR(400MHz,DMSO-d 6)δ11.55(s,1H),9.24(s,1H),8.13(d,J＝7.2Hz,2H),7.90(d,J＝7.6Hz,2H),7.59-7.50(m,3H),7.27(t,J＝7.6Hz,2H),6.87(t,J＝7.6Hz,1H),6.40(s,1H),2.37(s,3H).
实施例8
Figure PCTCN2020082016-appb-000026
Step1：2-氯-6-甲基-N-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺
在乙二醇(4ml)中2,4-二氯-6-甲基-7H-吡咯[2,3-d]嘧啶(1)(100mg,0.495mmol)和苯胺(125mg,1.34mmol)溶液中加入4滴浓缩盐酸水溶液。混合物在100℃氮气气氛下搅拌过夜。冷却至室温后，将混合物倒入水中(30ml)，用乙酸乙酯萃取(15ml×2)，用盐水(30ml)冲洗并浓缩。用C 18柱(乙腈/水从50％到60％)对残渣进行纯化，得到标题化合物2(80mg，收率63％)为灰白色固体。
LC-MS[流动相:在2.5min内，从70％水和30％CH 3CN到5％水和95％CH 3CN],Rt＝1.30min；MS计算值:258.1；MS实测值:259.0[M+H] +.
1H NMR(400MHz,DMSO-d 6)δ11.75(s,1H),9.48(s,1H),7.76-7.74(dd,J＝8.4,1.2Hz,2H),7.36(dd,J＝8.4,3.2Hz,2H),7.06(t,J＝7.6Hz,1H),6.40(s,1H),2.34(s,3H).
Step 2：6-甲基-N，2-二苯基-7H-吡咯并[2,3-d]嘧啶-4-胺
将2-氯-6-甲基-N-苯基-7H-吡咯[2,3-d]嘧啶-4-胺(2)(130mg,0.502mmol)、苯基硼酸(123mg,1.01mmol)、碳酸钾(347mg,2.51mmol)和四价(三苯基膦)钯(58mg,0.05mmol)在1,2-二甲氧基乙烷/水(5/1,3.6mL)中，在100℃微波氮气中搅拌3.5小时。冷却至室温后，将混合物倒入水中(25ml)，用乙酸乙酯萃取(25ml×2)，用盐水(30ml)冲洗并浓缩。用C 18柱(乙腈/水从50％到60％)对残留进行纯化，得到标题化合物(70mg，收率47％)为灰白色固体。
LC-MS(ESI):R T＝2.253min,MS计算值C 19H 16N 4 300.1,m/z实测值301.0[M+H] +. 1H NMR(400MHz,DMSO-d 6)δ11.69(s,1H),9.22(s,1H),8.37(d,J＝7.2Hz,2H),7.98(d,J＝7.6Hz,2H),7.50-7.37(m,5H),7.03(t,J＝7.2Hz,1H),6.49(s,1H),2.39(s,3H).
实施例9
Figure PCTCN2020082016-appb-000027
Step 1:9-异丙基-2-(吡啶-4-基)-N-(2-(三氟甲基)吡啶-4-基)-9H-嘌呤-6-胺(2 EPT60072)
向10mL密封管中加入2-氯-9-异丙基-N-(2-(三氟甲基)吡啶-4-基)-9H-嘌呤-6-胺(50mg，0.14mmol)，吡啶-4加入-硼酸(50mg，0.40mmol)，Pd(dppf)Cl 2(10mg，0.014mmol)，K 3PO 4(84mg，0.4mmol)，然后加入二氧六环(2.5mL)和水(0.5mL)。将混合物在惰性气氛下在100℃加热4小时。反应完成后，将反应混合物用硅胶(100-200mesh)浓缩，蒸发，得到粉末残余物。通过自动快速柱色谱法(硅胶，MeOH：DCM＝1:10)纯化残余物，得到标题化合物(40mg，71％产率)，为白色固体。
分子式:C 19H 16F 3N 7,分子量:399.38,(ESI)m/z＝400.2(M+H) +. 1HNMR(400MHz,DMSO-d 6)10.97(s,1H),8.85(d,J＝1.6Hz,1H),8.77-8.79(dd,J 1＝1.6Hz,J 2＝14.8Hz,2H),8.66-8.68(m,2H),8.27-8.30(m,3H),4.9(m,J＝6.8Hz,1H),1.65(d,J＝6.8Hz,6H)ppm.
实施例10
Figure PCTCN2020082016-appb-000028
Step 1:9-异丙基-2-(吡啶-2-基)-N-(2-(三氟甲基)吡啶-4-基)-9H-嘌呤-6-胺(2 EPT60073)
用2-氯-9-异丙基N-(2-(三氟甲基)吡啶-4-酰基)-9H-嘌呤-6-胺(107mg,0.30mmol)，2-(三丁基锡基)吡啶(310mg,0.84mmol)，Pd(PPh3) 4(35mg,0.03mmol)，加入二氧六环(5.0mL)，密封管10-mL。反应混合物在100℃惰性气体中加热8h。反应完成后，用饱和KF水溶液(2mL)对反应混合物进行骤冷，用硅胶(100-200mesh)对有机相进行浓缩蒸发，得到粉体残渣。采用自动闪柱柱色谱法(硅胶，MeOH:DCM＝1:19)对产物进行纯化，得到标题化合物(55mg，收率46％)为黄色固体。
分子式:C 19H 16F 3N 7,分子量:399.38,(ESI)m/z＝400.2(M+H) +. 1H NMR(500MHz,DMSO-d 6)10.93(s,1H),9.22(d,J＝2.5Hz,1H),8.78-8.80(m,1H),8.66(s,1H),8.62(d,J＝5.5Hz,1H),8.44(d,J＝8.0Hz,1H),8.35-8.36(m,1H),7.98-8.02(m,1H),7.51-7.55(m,1H),4.98(m,1H),1.63-1.65(d,J＝7.0Hz,6H)ppm.
实施例11
Figure PCTCN2020082016-appb-000029
Step 1:2-氯-7-甲基-N-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
用2,4-二氯-7-甲基-7H-吡啶[2,3-d]嘧啶(101mg,0.50mmol)、t-BuOK(85mg,0.75mmol)、苯胺(70mg,0.75mmol)分别充入20mL密封管，加入THF(5mL)。反应混合物在rt条件下搅拌1.5h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，PE:EA＝7:3)对产物进行纯化，得到产物为白色固体(63mg，收率49％)。
分子式:C 13H 11ClN 4,分子量:258.71,(ESI)m/z＝259.1(M+H) +.
Step 2:7-甲基-N，2-二苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(3 EPT60083)
用2-氯-7-甲基-N-苯基-7H-吡咯并[2,3-d]封闭管充入2-氯-7-甲基-N-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(63mg,0.25mmol)、苯基硼酸(45mg,0.37mmol)、Pd(dppf)Cl 2(18mg,0.025mmol)、K 3PO 4(110mg,0.5mmol)、二氧六环(5mL)、水(1mL)。反应混合物在惰性气体中以100℃加热2h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱层析(硅胶，PE:EA＝7:3)对产物进行纯化，得到产物为白色固体(20mg，收率27％)。
分子式:C 19H 16N 4，分子量:300.37，(ESI)m/z＝301.2(M+H) +. 1H NMR(500MHz,DMSO-d 6)9.43(s,1H),8.43-8.45(m,2H),7.99(d,J＝7.5Hz,2H),7.39-7.51(m,5H),7.32(d,J＝3.5Hz,1H),7.06(t,J＝7.5Hz,1H),6.83(d,J＝3.5Hz,1H),3.84(s,3H)ppm.
实施例12
Figure PCTCN2020082016-appb-000030
Step 12-氯-9-异丙基-6-苯基-9H-嘌呤(3)
将2,6-二氯-9-异丙基-9H-嘌呤(1,100mg,0.43mmol)、苯基硼酸(2,63mg,0.52mmol)、Pd(dppf)Cl 2(3mg,0.0043mmol)和K 2CO 3(120mg,0.86mmol)分别装入25ml圆底烧瓶中。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃氮气气氛下搅拌2h。减压除去溶剂，用自动闪柱柱层析(硅胶，PE:EA＝5:1)纯化得到2-氯-9-异丙基-6-苯基-9H-嘌呤(3,100mg,84.73％收率)为白色固体。
LCMS:(ESI)m/z＝273.08(M+H) +；RT＝1.75min.
Step 2:9-异丙基-N，6-二苯基-9H-嘌呤-2-胺(EPT60086)
一个25ml圆底烧瓶中加入2-氯-9-异丙基-6-苯基-9H-嘌呤(330毫克0.108mmol),苯胺(415.6毫克0.162mmol),Pd(OAc)2(0.24毫克,1.08μmol)BINAP(1.2毫克,2.16μmol)和Cs 2CO 3(108毫克,0.33mmol)。混合物悬浮在二氧六环(5ml)中。反应在100℃氮气气氛下搅拌2h。减压除去溶剂，采用自动闪柱柱层析(硅胶，PE:EA＝5:1)纯化得到9-异丙基n,6-二苯基-9H-嘌呤-2-胺(EPT60086,20mg,55.2％收率)为黄色固体。
LCMS:(ESI)m/z＝330.21(M+H) +；RT＝1.84min.
1H NMR(500MHz,DMSO)δ9.59(s,1H),8.81(dd,J＝8.1,1.5Hz,2H),8.43(s,1H),7.91(d,J＝7.7Hz,2H),7.62–7.54(m,3H),7.35–7.30(m,2H),6.95(t,J＝7.3Hz,1H),4.81(s,1H),1.61(d,J＝6.8Hz,6H).
实施例13
Figure PCTCN2020082016-appb-000031
Step 1:2-氯-N-异丙基-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(3)
在一个25毫升的圆底烧瓶中加入2,4-二氯-6-甲基-7H-吡咯[2,3-d]嘧啶(1,202mg,1mmol)、propaN-2-amine(2,89mg,1.5mmol)和DIPEA(258mg,2mmol)。反应混合物是溶解在二氧六环在70℃(5毫升)和搅拌2h。与EA(10毫升x 3)提取后,用盐水洗净(10毫升x 3),干燥无水硫酸钠和集中在真空,2-氯-N-异丙基-6-甲基-7H-吡咯并[2,3-d]嘧啶-N-4-胺(220毫克,收率98.9％)获得了作为一个棕色固体。
LCMS:(ESI)m/z＝225.18(M+H) +；RT＝1.44min.
Step 2:N-异丙基-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60087)
将2-氯-N-异丙基-6-甲基-7H-吡啶醇[2,3-d]4-氨基吡啶(3,50mg,0.22mmol)、苯基硼酸(4,32mg,0.27mmol)、Pd(dppf)Cl 2(2mg,0.0022mmol)和K 3PO 4(93mg,0.44mmol)分别装入25ml圆底烧瓶中。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃氮气气氛下进行了2小时的搅拌。减压除去溶剂，用自动闪柱柱层析(硅胶，PE:EA＝1:1)纯化得到N-异丙基-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60087,10mg,16.87％收率)为黄色固体。
LCMS:(ESI)m/z＝267.2(M+H) +；RT＝1.30min.
1H NMR(500MHz,DMSO)δ11.35(s,1H),8.36–8.33(m,2H),7.45–7.35(m,3H),6.95(d,J＝7.6Hz,1H),6.25(dd,J＝1.9,1.0Hz,1H),4.51(d,J＝6.9Hz,1H),2.32(d,J＝0.7Hz,3H),1.28(d,J＝6.5Hz,6H).
实施例14
Figure PCTCN2020082016-appb-000032
Step 1:2-氯-8-甲基-N-苯基-9H-嘌呤-6-胺(2)
一个20毫升的密封管被充入2,6-二氯-8-甲基-9H-嘌呤(101毫克，0.50mmol)和5毫升正丁醇。加入DIPEA(129mg,1.0mmol)和苯胺(70mg,0.75mmol)。反应混合物在100℃下搅拌4h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，MeOH:DCM＝1:10)对产物进行纯化，得到产物为白色固体(100mg,77.5％收率)。
分子式:C 12H 10ClN 5,分子量:259.70,(ESI)m/z＝260.1(M+H) +.
Step 2:8-甲基-N，2-二苯基-9H-嘌呤-6-胺(3 EPT60088)
将2-氯-8-甲基-N-苯基-9H-嘌呤-6-胺(50mg,0.2mmol)、苯基硼酸(125mg,1.0mmol)、Pd(dppf)Cl 2(30mg,0.04mmol)、K 3PO 4(212mg,1.0mmol)分别充入10-mL密封管，然后加入2,2,5ml的二氧六环和1mL的水。反应混合物在100℃惰性气体中加热22h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱层析(硅胶，DCM:MeOH＝19:1)对产物进行纯化，得到标题化合物(40mg，收率66％)为黄色固体。
分子式:C 18H 15N 5,分子量:301.35,(ESI)m/z＝3020.2(M+H) +. 1H NMR(400MHz,DMSO-d 6)12.91(s,1H),9.67(s,1H),8.33(d,J＝7.2Hz,2H),8.02(d,J＝8.0Hz,2H),7.32-7.48(m,5H),7.01(m,1H),2.50(s,3H)ppm.
实施例15
Figure PCTCN2020082016-appb-000033
Step 1:2,4-二氯-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶(3)
到25ml圆底烧瓶中加入2-氯-N-异丙基-6-甲基-7H-吡咯并[2,3-d]嘧啶-N-4-胺(500毫克,2.47mmol),对甲苯磺酰氯(705毫克,3.71mmol)DIPEA(957毫克,7.42mmol)和深度贴图(30毫克,0.25mmol),反应混合物溶解在DCM(5毫升),搅拌2h RT。残渣集中在真空和净化自动闪光柱层析法(硅胶,PE:EA＝1:1)将2,4-二氯-6-甲基-7-甲苯磺酰-7H-吡啶[2,3-d]嘧啶(3,800mg，产率91.24％)制成黄色固体。LCMS:(ESI)m/z＝356.06(M+H) +；RT＝1.88min.
Step 2:2-氯-N-环丙基-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(5)
在一个25毫升的圆底烧瓶中加入2,4-二氯-6-甲基-7-甲苯磺酰-7H-吡啶[2,3-d]3,100mg,0.28mmol，环丙胺(4,31mg,0.56mmol)和DIPEA(108mg,0.84mmol)。反应混合物溶解在二氧六环(5ml)中，70℃搅拌4h，残渣浓缩于真空中，用自动闪柱柱色谱(硅胶，PE:EA＝85:15)纯化得到2-氯-N-环丙基-6-甲基-7-甲苯磺酰-7H-吡咯并[2,3-d]嘧啶-4-胺(5,100mg,94.98％收率)为白色固体。LCMS:(ESI)m/z＝377.89(M+H) +；RT＝1.76min.
Step 3:N-环丙基-6-甲基-2-(萘-2-基)-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(7)
在25ml圆底烧瓶中加入2-氯-N-环丙基-6-甲基-7-甲苯磺酰-7-h-吡咯并[2,3-d]嘧啶-4-胺(5,100mg,0.26mmol)，萘-1-基硼酸(6,134mg,0.78mmol)，Pd(dppf)Cl 2(19mg,0.026mmol)和K 3PO 4(165mg,0.78mmol)。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃的氮气气氛下进行了一夜。该产品为黄色固体N-环丙基-6-甲基-2-(萘-2-基)-7-甲苯磺酰-7H-吡咯并[2,3-d]嘧啶-4-胺(7,100mg,82.18％收率)。
LCMS:(ESI)m/z＝469.29(M+H) +；RT＝2.18min
Step 4:N-环丙基-6-甲基-2-(萘-2-基)-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60187)
到25ml圆底烧瓶中加入N-环丙基-6-甲基-2-(萘-N-2-基) 7-甲基磺酰基-7H-吡咯并[2,3-d]嘧啶-N-4-胺(100毫克,0.21mmol)溶解在MeONa(2毫升,5.4年的甲醇)和甲醇(10毫升)和搅拌60℃ 4h。残渣集中在真空和净化自动闪光柱层析法(硅胶、PE:EA＝5:1)得到N-环丙基-6-甲基-2-(萘-2-基)-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60187 50毫克,产率为75.82％)为白色固体。
LCMS:(ESI)m/z＝315.21(M+H) +；RT＝1.51min.
1H NMR(500MHz,DMSO)δ11.49(s,1H),8.89(s,1H),8.55(d,J＝8.4Hz,1H),8.04–7.89(m,3H),7.57–7.48(m,2H),7.43(d,J＝3.1Hz,1H),6.32(s,1H),3.08(s,1H),2.35(s,3H),0.88–0.83(m,2H),0.66–0.60(m,2H).
实施例16
Figure PCTCN2020082016-appb-000034
Step 1:N-异丙基-6-甲基-2-(吡啶-2-基)-7H-吡咯并[2,3-d]嘧啶-4-胺(2 EPT60101)
用2-氯-N-异丙基-6-甲基-7H-吡咯[2,3-d]嘧啶-4-胺(100mg,0.44mmol)，2-(三丁基锡基)吡啶(300mg,0.81mmol)，Pd(PPh3) 4(50mg,0.04mmol)，再加入二氧六环(5.0mL)，充入10-mL密封管。反应混合物在100℃惰性气体中加热16h。反应完成后，用饱和KF水溶液(2mL)对反应混合物进行骤冷，用硅胶(100-200mesh)对有机相进行浓缩蒸发，得到粉体残渣。采用自动闪柱柱色谱法(硅胶，MeOH:DCM＝1:19)对残留进行纯化，得到标题化合物(12mg，收率9％)为黄色固体。
分子式:C 15H 17N 5,分子量:267.34,(ESI)m/z＝268.2(M+H) +. 1H NMR(400MHz,DMSO-d 6)11.37(s,1H) 8.60(m,1H),8.28(d,J＝8.0Hz,1H),7.80-7.84(m,1H),7.31-7.34(m,1H),6.95(d,J＝8.0Hz,1H),6.26(s,1H),4.46-4.52(m,1H),2.30(s,3H),1.29(m,6H)ppm.
实施例17
Figure PCTCN2020082016-appb-000035
Step 1:N-异丙基-6-甲基-2-(吡啶-3-基)-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60102)
将2-氯-N-异丙基-6-甲基-7H-吡啶-4-胺(1,50mg,0.22mmol)、吡啶-3-基硼酸(2,81mg,0.66mmol)、Pd(dppf)Cl 2(16mg,0.022mmol)和K 3PO 4(93mg,0.44mmol)分别装入25ml圆底烧瓶中。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃氮气气氛下搅拌过夜。减压除去溶剂，采用自动闪柱柱层析(硅胶，DCM:MeOH＝95:5)纯化得到N- 异丙基-6-甲基-2-(吡啶-3-酰基)-7H-吡啶-4-胺(EPT60102,54mg,91.93％收率)为黄色固体。
LCMS:(ESI)m/z＝268.28(M+H) +；RT＝1.21min.
1H NMR(500MHz,DMSO)δ11.46(s,1H),9.47(d,J＝1.4Hz,1H),8.60–8.54(m,2H),7.49–7.44(m,1H),7.08(d,J＝7.6Hz,1H),6.28(d,J＝0.8Hz,1H),4.51(dq,J＝13.1,6.6Hz,1H),2.33(s,3H),1.28(d,J＝6.5Hz,6H).
实施例18
Figure PCTCN2020082016-appb-000036
Step 1:N-异丙基-6-甲基-2-(吡啶-4-基)-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60103)
将2-氯-N-异丙基-6-甲基-7H-吡啶-4-胺(1,50mg,0.22mmol)、吡啶-4-乙基硼酸(2,81mg,0.66mmol)、Pd(dppf)Cl 2(16mg,0.022mmol)和K 3PO 4(93mg,0.44mmol)分别装入25ml圆底烧瓶中。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃氮气气氛下搅拌过夜。减压除去溶剂，采用自动闪柱柱色谱(硅胶，DCM:MeOH＝95:5)进行纯化，得到N-异丙基-6-甲基-2-(吡啶-4-酰基)-7H-吡啶-4-胺(EPT60103,50mg,85.12％收率)为黄色固体。
LCMS:(ESI)m/z＝268.28(M+H) +；RT＝1.21min.
1H NMR(500MHz,DMSO)δ11.54(s,1H),8.64(dd,J＝4.5,1.5Hz,2H),8.20(dd,J＝4.5,1.6Hz,2H),7.13(d,J＝7.6Hz,1H),6.31(d,J＝0.8Hz,1H),4.52(dd,J＝13.5,6.7Hz,1H),2.34(s,3H),1.28(d,J＝6.5Hz,6H).
实施例19
Figure PCTCN2020082016-appb-000037
Step 1:2-氯-N-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
将化合物1(100mg,0.54mmol)在正丁醇(5mL)溶液中加入Et 3N(86.9mg,0.86mmol)和苯胺(60.5mg,0.65mmol)。反应混合物在100℃下搅拌16小时(过夜)。LCMS(EPN18040-002-1)显示反应完成，SM残留10％。溶剂在真空中被除去。未纯化的粗化合物直接用于下一步。(ESI)m/z＝245.31(M+H) +
Step 2:N，2-二苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60098)
将化合物2(60mg,0.25mmol)在二氧六环(5mL)和H 2O(1mL)溶液中分别加入Pd(dppf)Cl 2(36.6mg,0.05mmol)、K 3PO 4(182.6mg,0.86mmol)和苯基硼酸(149.5mg,1.23mmol)。反应混合物在100℃下搅拌16h(过夜)，Ar下LCMS(EPN18040-005-1)显示反应完成，SM残留10％。溶剂在真空中浓缩。采用自动闪柱柱色谱法(硅胶，PE/EA＝8:1)对残留进行纯化，得到粗品。用饱和K 2CO 3溶液(100mL)稀释残渣，用CH 2Cl 2(50mL×3)萃取混合物，再用饱和NaCl(100mL)洗涤。所得到的有机层用无水Na 2SO 4干燥，在真空中除去溶剂，得到所需的产物(40mg,57.1％收率)为白色固体。(ESI)m/z＝287.12(M+H) +.1H NMR(500MHz,DMSO-d6)11.82(s,1H),9.40(s,1H),8.38-8.40(m,2H),7.99-8.01(m,2H),7.48-7.51(m,2H),7.40-7.45(m,3H),7.27-7.28(m,1H),7.04-7.07(m,1H),6.82-6.83(m,1H)ppm.
实施例20
Figure PCTCN2020082016-appb-000038
Step 1:2,4-二氯-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶(2)
取50ml圆底烧瓶，加入2,4-二氯-6-甲基-7H-吡啶[2,3-d]400mg,2.0mmol,4-甲基苯磺酰氯(570mg,3.0mmol)，DMAP(24mg,0.2mmoL)，DIPEA(1mL)，再加入DCM(20mL)。反应混合物在室温下搅拌1h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝1:10)对产物进行纯化，得到标题化合物(565mg，收率80％)为白色固体。
分子式:C 14H 11Cl 2N 3O 2S,分子量:356.22,(ESI)m/z＝356.1(M+H) +.
Step 2:2-氯-6-甲基-7-甲苯磺酰基-N-(2-(三氟甲基)吡啶-4-基)-7H-吡咯并[2,3-d]嘧啶-4-胺(3)
取2,4-二氯-6-甲基-7-甲苯磺酰-7H-吡咯啉[2,3-d]嘧啶(140mg,0.4mmol)，Cs 2CO 3(156mg,0.48mmol)，2-(三氟甲基)吡啶-4-胺(100mg,0.60mmol)，加入DMSO(4ml)，密封管20mL。反应混合物rt搅拌18h,70℃搅拌2h。反应由LCMS监测。反应完成后，加入水(20mL)，用EA(20mL 2)萃取反应混合物。采用硅胶(100-200目)蒸发浓缩有机相， 得到粉体残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝3:2)对残留进行纯化，得到标题化合物(64mg，收率33％)为白色固体。分子式:C 20H 15ClF 3N 5O 2S,分子量:481.88,(ESI)m/z＝482.2(M+H) +.
Step 3:6-甲基-2-苯基-7-甲苯磺酰基-N-(2-(三氟甲基)吡啶-4-基)-7H-吡咯并[2,3-d]嘧啶-4-胺(4)
用2-氯-6-甲基-7-甲苯磺酰-N-(2-(三氟甲基)吡啶-4-酰基)-7H-吡咯啉[2,3-d]苯基硼酸(12mg,0.1mmol)，Pd(dppf)Cl 2(5mg,0.008mmol)，K 3PO 4(30mg,0.15mmol)，然后加入二氧六环(2mL)和水(0.4mL)，充入10-mL密封管。反应混合物在惰性气体中加热到100c，加热1.5h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱层析法(硅胶，EA:PE＝1:1)对产物进行纯化，得到标题化合物(20mg，收率77％)为白色固体。
分子式:C 26H 20F 3N 5O 2S,分子量:523.53,(ESI)m/z＝524.3(M+H) +.
Step 4:6-甲基-2-苯基-N-(2-(三氟甲基)吡啶-4-基)-7H-吡咯并[2,3-d]嘧啶-4-胺(5 EPT60121)
将6-甲基-2-苯基-7-甲苯磺酰-N-(2-(三氟甲基)吡啶-4-yl)-7H-吡咯[2,3-d]嘧啶-4-胺(20mg,0.04mmol)溶于CH 3OH(3mL)中，加入CH 3ONa(5.4mol/L,0.5mL)。反应混合物在55℃下搅拌2h。反应完成后，用饱和NH 4Cl水溶液(1mL)对反应进行骤冷，蒸发浓缩反应得到油渣，反相柱层析(C 18,H 2O/MeCN＝2/3)纯化得到标题化合物EPN18033-070-A(4mg,28％收率)为白色固体。分子式:C 19H 14F 3N 5,分子量:369.35,(ESI)m/z＝370.2(M+H) +. 1H NMR(400MHz,DMSO-d 6)11.92(s,1H),10.05(s,1H),8.84(d,J＝1.6Hz,1H),8.58(d,J＝5.6Hz,1H),8.34(d,J＝6.8Hz,2H),8.06(m,1H),7.41-7.49(m,3H),6.52(s,1H),2.29(d,J＝3.6Hz,3H)ppm.
实施例21
Figure PCTCN2020082016-appb-000039
Step 1:2-氯-N-(2-氟吡啶-4-基)-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
取2,4-二氯-6-甲基-7-甲苯磺酰-7H-吡啶[2,3-d]嘧啶(66mg,0.2mmol)、Cs 2CO 3(65mg,0.2mmol)、2-氟吡啶-4-胺(45mg,0.40mmol)分别充入20ml密封管，加入DMSO(2mL)。反应混合物rt搅拌18h,70℃搅拌2h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝3:2)对产物进行纯化，得到标题化合物(26mg，产率32％)为白色固体。
分子式:C 19H 15ClFN 5O 2S,分子量:431.87(ESI)m/z＝432.2(M+H) +.
Step 2:N-(2-氟吡啶-4-基)-6-甲基-2-苯基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(3)
用2-氯-N-(2-氟吡啶-4-酰基)-6-甲基-7-甲苯磺酰-7H-吡咯并[2,3-d]吡啶-4-胺(26mg,0.06mmol)、苯基硼酸(12mg,0.1mmol)、Pd(dppf)Cl 2(5mg,0.008mmol)、K 3PO 4(30mg,0.15mmol)、二氧六环(3mL)、水(0.5mL)充入10-mL密封管。反应混合物在惰性气体中加热到100℃，加热5h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝1:4)对产物进行纯化，得到标题化合物(22mg，收率78％)为白色固体。
分子式:C 25H 20FN 5O 2S,分子量:473.53,(ESI)m/z＝474.3(M+H) +.
Step 3:N-(2-氟吡啶-4-基)-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(4 EPT60122)
将N-(2-氟吡啶-4-酰基)-6-甲基-2-苯基-7-甲苯磺酰-7H-吡咯啉[2,3-d]嘧啶-4-胺(22mg,0.046mmol)溶于CH 3OH(3mL)中，加入CH 3ONa(5.4mol/L,0.5mL)。反应混合物在60℃下搅拌2h。反应完成后，用饱和NH 4Cl水溶液(1mL)对反应进行骤冷，蒸发浓缩反应得到油渣，反相柱层析(C 18,H 2O/MeCN＝2/3)纯化产物为白色固体(5mg，产率32％)。分子式:C 18H 14FN 5,分子量:319.34(ESI)m/z＝320.2(M+H) +. 1H NMR(400MHz,DMSO-d 6)11.90(s,1H),9.92(s,1H),8.31(d,J＝7.2Hz,2H),8.07(d,J＝5.6Hz,1H),7.93(s,1H),7.74(d,J＝4.8Hz,1H),7.42-7.51(m,3H),6.52(s,1H),2.38(s,3H)ppm.
实施例22
Figure PCTCN2020082016-appb-000040
Step 1:6-甲基-N 2，N 4-二苯基-7H-吡咯并[2,3-d]嘧啶-2,4-二胺(EPT60123)
一个25ml圆底烧瓶中加入2,4-二氯-6-甲基-7H-吡咯并[2,3-d]嘧啶(0.25 1 50毫克mmol),苯胺(69毫克,0.74mmol),Pd(OAc)2(2.8毫克,12.5μmol)BINAP(15毫克,25μmol)和Cs 2CO 3(245毫克,0.75mmol)。混合物悬浮在二氧六环(2ml)中。反应在100℃氮气气氛下搅拌4h。减压除去溶剂，首先用自动闪柱柱层析(硅胶，DCM:MeOH＝95:5)纯化 得到粗品，然后用反相柱层析(NH 4HCO 3,a.q.，0.5％)得到白色固体6-甲基-n2,n4-二苯基-7H-吡啶-2,4-二胺(EPT60123,4mg,5.08％收率)得到n4-二苯基-2,4-二胺[2,3-d]。
LCMS:(ESI)m/z＝316.25(M+H) +；RT＝1.42min.
1H NMR(500MHz,DMSO)δ11.07(s,1H),8.94(s,1H),8.75(s,1H),7.91(d,J＝7.7Hz,2H),7.81(d,J＝7.7Hz,2H),7.31(t,J＝7.9Hz,2H),7.21(t,J＝7.9Hz,2H),6.99(t,J＝7.3Hz,1H),6.84(t,J＝7.3Hz,1H),6.31(s,1H),2.29(s,3H).
实施例23
Figure PCTCN2020082016-appb-000041
Step 1:N-环丙基-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(3)
在25毫升的圆底烧瓶中加入2,4-二氯-6-甲基-7H-吡啶[2,3-d],100mg,0.5mmol，环丙胺(2,42mg,0.75mmol)和DIPEA(129mg,1mmol)。反应混合物溶解在二甲氧基(2ml)中，70℃搅拌2h,EA(10ml x 3)萃取后，用盐水(10ml x 3)冲洗，无水硫酸钠干燥，浓缩得到2-氯-N-环丙基-6-甲基-7H-吡啶-4-胺(3,110mg，产率99.1％)黄色固体。
LCMS:(ESI)m/z＝223.22(M+H) +；RT＝1.32min.
Step 2:N-环丙基-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60124)
将2-氯-N-环丙基-6-甲基-7H-吡啶醇[2,3-d]4-氨基吡啶(3,50mg,0.22mmol)、苯基硼酸(4,80mg,0.66mmol)、Pd(dppf)Cl 2(16mg,0.022mmol)和K 3PO 4(93mg,0.44mmol)分别装入25ml圆底烧瓶中。
混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃的氮气气氛下进行了一夜。减压除去溶剂，首先用自动闪柱柱层析(硅胶，DCM:MeOH＝95:5)纯化得到粗品，然后用反相柱层析(NH 4HCO 3,a.q.0.5％):MeCN＝70:30得到白色固体N-环丙基-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60124,20mg,34.3％收率)。LCMS:(ESI)m/z＝265.24(M+H) +；RT＝1.20min.
1H NMR(500MHz,DMSO)δ11.41(s,1H),8.37(d,J＝7.1Hz,2H),7.46–7.30(m,4H),6.28(s,1H),3.02(dt,J＝10.0,3.3Hz,1H),2.32(d,J＝0.7Hz,3H),0.84–0.78(m,2H),0.65–0.55(m,2H).
实施例24
Figure PCTCN2020082016-appb-000042
Step 1:2-氯-9-甲基-N-苯基-9H-嘌呤-6-胺(3)
在一个25毫升的圆底烧瓶中加入2,6-二氯-9-甲基-9H-嘌呤(1,100mg,0.49mmol)，苯胺(2,69mg,0.74mmol)和DIPEA(191mg,1.48mmol)。反应混合物溶解在二氧六环(5ml)中，70℃搅拌4h，残渣浓缩于真空中，用自动闪柱柱色谱(硅胶，DCM:MeOH＝95:5)纯化得到2-氯-9-甲基-N-苯基-9H-嘌呤-6-胺(3,120mg,94.55％收率)为off白色固体。
LCMS:(ESI)m/z＝260.19(M+H) +；RT＝1.52min.
Step 2:9-甲基-N，2-二苯基-9H-嘌呤-6-胺(EPT60125)
将2-氯-9-甲基-N-苯基-9H-嘌呤-6-胺(3,50mg,0.19mmol)、苯基硼酸(4,69mg,0.57mmol)、Pd(dppf)Cl 2(14mg,0.019mmol)和K 3PO 4(80mg,0.38mmol)分别装入25ml圆底烧瓶中。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃氮气气氛下搅拌过夜。减压除去溶剂，用自动闪柱柱层析(硅胶，PE:EA＝1:1)纯化得到9-甲基-n,2-二苯基-9H-嘌呤-6-胺(EPT60125,40mg,69.94％收率)为黄色固体。
LCMS:(ESI)m/z＝302.25(M+H) +；RT＝1.76min.
1H NMR(500MHz,DMSO)δ9.90(s,1H),8.43(d,J＝7.5Hz,2H),8.30(s,1H),8.05(d,J＝7.9Hz,2H),7.55–7.46(m,3H),7.40(t,J＝7.7Hz,2H),7.07(t,J＝7.1Hz,1H),3.87(s,3H).
实施例25
Figure PCTCN2020082016-appb-000043
Step 1:2-(2-氟苯基)-N-异丙基-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60126)
将化合物1(50mg,0.22mmol)加入到2,5mL二氧六环(5ml)和H 2O(1ml)溶液中，加入K 3PO 4(163.4mg,0.77mmol)，Pd(dppf)Cl 2(32.7mg,0.04mmol)和2-氟苯硼酸(154.0mg,1.1mmol)。反应混合物在100℃条件下搅拌16h(过夜)。LCMS(EPN18040-009-1)显示反应已经完成，SM残留20％。溶剂在真空中浓缩。采用自动闪柱柱层析(硅胶，PE/EA＝8:1)对残留进行纯化，再用闪柱(C18柱层析，H 2O(NH 4HCO 3,0.8g/L)/CH 3CN＝70/30)进行纯化，得到标题化合物(8.5mg,13.4％收率)为白色固体。(ESI)m/z＝285.24(M+H) +. 1H NMR(500MHz,DMSO-d6)11.42(s,1H),9.94-9.98(m,1H),7.38-7.43(m,1H),7.20-7.26(m,2H),6.98-6.99(d,J＝7.5Hz,1H),6.26(s,1H),4.38-4.42(m,1H),3.32(s,3H),1.23-1.25(d,J＝6.5Hz,6H)ppm.
实施例26
Figure PCTCN2020082016-appb-000044
Step 1:6-甲基-2,4-二苯基-7H-吡咯并[2,3-d]嘧啶(EPT60132)
在一个25毫升的圆底烧瓶中加入2,4-二氯-6-甲基-7H-吡啶[2,50mg,0.25mmol]，苯基硼酸(2,90mg,0.75mmol)，Pd(dppf)Cl 2(18mg,0.025mmol)和K 3PO 4(159mg,0.75mmol)。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃的氮气气氛下进行了一夜。采用自动闪柱柱色谱法(硅胶，PE:EA＝1:1)将残渣浓缩在真空中纯化，得到6-甲基-2,4-二苯基-7H-吡咯啉[2,3-d]嘧啶(EPT60132,40mg,56.14％得率)为白色固体。LCMS:(ESI)m/z＝286.20(M+H) +；RT＝1.83min.
1H NMR(500MHz,DMSO)δ12.17(s,1H),8.56–8.50(m,2H),8.31–8.26(m,2H),7.65–7.44(m,6H),6.65(s,1H),2.48(s,3H).
实施例27
Figure PCTCN2020082016-appb-000045
Step 1:N-(叔丁基)-2-氯-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺
将2,4-二氯-6-甲基-7-甲苯磺酰-7H-吡啶[2,3-d]嘧啶(100mg,0.28mmol)，2-甲基丙基-2-胺(31mg,0.42mmol)和DIPEA(108mg,0.84mmol)分别装入25ml圆底烧瓶中。反应混合物溶解于二氧六环(5ml)中，70℃搅拌4h，残渣浓缩于真空中，用自动闪柱柱色谱(硅胶，PE:EA＝85:15)纯化得到N-(叔丁基)-2-氯-6-甲基-7-甲苯磺酰-7H-吡咯并[2,3-d]嘧啶-4-胺(80mg,72.89％收率)为白色固体。LCMS:(ESI)m/z＝393.27(M+H) +；RT＝1.93min.
Step 2:N-(叔丁基)-6-甲基-2-苯基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺
在一个25毫升的圆底烧瓶中加入N-(叔丁基)-2-氯-6-甲基-7-甲苯磺酰-7H-吡咯并[2,3-d]嘧啶-4-胺(80mg,0.2mmol)，苯基硼酸(6,73mg,0.6mmol)，Pd(dppf)Cl 2(17mg,0.02mmol)和K 3PO 4(127mg,0.6mmol)。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。 反应在100℃氮气气氛下搅拌过夜。减压除去溶剂，采用自动闪柱柱层析(硅胶，PE:EA＝5:1)纯化得到N-(叔丁基)-6-甲基-2-苯基-7-甲苯磺酰-7H-吡咯[2,3-d]嘧啶-4-胺(7,60mg,69.12％)。
LCMS:(ESI)m/z＝435.32(M+H) +；RT＝2.02min.
Step 3:N-(叔丁基)-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60133)
将装有N-(叔丁基)-6-甲基-2-苯基-7-甲苯磺酰-7H-吡咯[2,3-d]嘧啶-4-胺(60mg,0.14mmol)溶于MeONa(0.5ml,5.4M in MeOH)和MeOH(5ml)中，在50c下搅拌过夜。减压除去溶剂，用自动闪柱柱层析(硅胶，PE:EA＝5:1)纯化得到N-(叔丁基)-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60133,20mg,51.02％收率)为白色固体。
LCMS:(ESI)m/z＝281.20(M+H) +；RT＝1.49min.
1H NMR(500MHz,DMSO)δ11.33(s,1H),8.36–8.33(m,2H),7.45(dd,J＝10.3,4.6Hz,2H),7.40–7.36(m,1H),6.53(s,1H),6.35(dd,J＝1.9,1.0Hz,1H),2.31(d,J＝0.6Hz,3H),1.58(s,9H).
实施例28
Figure PCTCN2020082016-appb-000046
Step 1:N-环丙基-2-(2-氟苯基)-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60134)
将2-氯-N-环丙基-6-甲基-7H-吡啶醇[2,3-d]4-氨基吡啶(1,50mg,0.22mmol)，(2-氟苯基)硼酸(2,92mg,0.66mmol)，Pd(dppf)Cl 2(16mg,0.022mmol)和K 3PO 4(93mg,0.44mmol)分别装入25ml圆底烧瓶中。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃的氮气气氛下进行了一夜。减压除去溶剂，首先用自动闪柱柱层析(硅胶，PE:EA＝1:1)纯化得到粗品，然后用反相柱层析(NH 4HCO 3,a.q.0.5％)得到白色固体N-环丙基-2-(2-氟苯基)-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60134,6mg,9.67％收率):MeCN＝55:45。
LCMS:(ESI)m/z＝283.16(M+H) +；RT＝1.13min.
1H NMR(500MHz,DMSO)δ11.44(d,J＝35.0Hz,1H),8.02–7.89(m,1H),7.44–7.34(m,2H),7.28–7.19(m,2H),6.27(d,J＝34.6Hz,1H),2.98–2.94(m,1H),2.33(s,3H),0.80–0.73(m,2H),0.62–0.55(m,2H).
实施例29
Figure PCTCN2020082016-appb-000047
Step 1:2-氯-N-(环丙基甲基)-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
向EtOH(3ml)中化合物1(100mg,0.50mmol)溶液中加入Et 3N(202.2mg,2.0mmol)、氨基甲基环丙烷(142.1mg,2.0mmol)。反应混合物在80℃下搅拌2小时。LCMS(EPN18040-010-1)显示反应完成，无SM残留。溶剂在真空中浓缩。未纯化的粗化合物直接用于下一步。(ESI)m/z＝237.21(m+H) +。
步骤2:N-(环丙基甲基)-6-甲基-2-苯基-7H-吡咯[2,3-d]嘧啶-4-胺(EPT60135)将化合物2(68.5mg,0.29mmol)在二氧六环(5mL)和H 2O(1mL)溶液中分别加入K 3PO 4(216.5mg,1.02mmol)、Pd(dppf)Cl 2(47.4mg,0.058mmol)和苯硼酸(176.9mg,1.45mmol)。反应混合物在100℃下搅拌16h(过夜)，Ar条件下LCMS(EPN18040-012-1)显示反应完成，SM残留30％。溶剂在真空中浓缩。采用自动闪柱柱层析(硅胶，PE/EA＝8:1)对残留进行纯化，再用闪柱(C-18柱层析，H 2O(NH 4HCO 3,0.8g/L)/CH 3CN＝70/30)进行纯化，得到标题化合物(7.5mg,9.3％收率)为淡黄色固体。(ESI)m/z＝279.26(M+H) +. 1H NMR(500MHz,DMSO-d6)11.37(s,1H),8.34-8.36(m,2H),7.42-7.45(m,2H),7.31-7.39(m,2H),6.24(s,1H),3.44-3.50(m,2H),2.33(s,3H),1.18-1.21(m,1H),0.46-0.48(m,2H),0.33-0.36(m,2H)ppm.
实施例30
Figure PCTCN2020082016-appb-000048
Step 1:N-苄基-2-氯-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
在EtOH(5mL)中加入化合物1(100mg,0.50mmol)的溶液中加入Et 3N(101.16mg,1.0mmol)和苄胺(107.1mg,1.0mmol)。反应混合物在80℃下搅拌2小时。LCMS(EPN18040-014-1)显示反应完成，无SM残留。溶剂在真空中浓缩。未纯化的粗化合物直接用于下一步。(ESI)m/z＝273.28(M+H) +.
Step 2:N-苄基-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60136)
将化合物2(55mg,0.2mmol)在二噁烃(5mL)和H 2O(1mL)溶液中分别加入K 3PO 4(150.2mg,0.71mmol)、Pd(dppf)Cl 2(32.7mg,0.04mmol)和苯硼酸(122.1mg,1.0mmol)。反应混合物在100℃下搅拌16h(过夜)，Ar下LCMS(EPN18040-017-1)显示反应完成，SM残留20％。溶剂在真空中浓缩。采用自动闪柱柱层析(硅胶，PE/EA＝8:1)纯化残渣，再用闪柱(C-18柱层析，H 2O(NH 4HCO 3,0.8g/L)/CH 3CN＝70/30)纯化得到标题化合物(4.4mg，7％产率)。(ESI)m/z＝315.15(M+H) +. 1H NMR(500MHz,DMSO-d6)11.43(s,1H),8.31-8.33(m,2H),7.82-7.84(m,1H),7.39-7.43(m,4H),7.35-7.37(m,1H),7.29-7.32(m,2H),7.20-7.23(m,1H),6.25(s,1H),4.81-4.82(d,J＝6.0Hz,2H),2.33(s,3H)ppm.
实施例31
Figure PCTCN2020082016-appb-000049
Step 1:2-氯-N-(3-氟苯基)-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
将化合物1(100mg,0.50mmol)在二氧六环(3ml)溶液中加入tBuOK(101.0mg,1.0mmol)，pd(OAc) 2(11.3mg,0.05mmol)，BINAP(62.3mg,0.1mmol)和3-氟苯胺(134.2mg,1.1mmol)。反应混合物在100℃下搅拌16h(过夜)，Ar下LCMS(EPN18040-015-1)显示反应完成，SM残留20％。溶剂在真空中浓缩。采用自动闪柱柱色谱法(硅胶，PE/EA＝8:1)对残留进行纯化，得到标题化合物(60mg,44.2％收率)为淡黄色固体。(ESI)m/z＝277.21(M+H) +.
Step 2N-(3-氟苯基)-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60137)
将化合物2(66.0mg,0.22mmol)在二噁烃(5mL)和H 2O(1mL)溶液中分别加入Pd(dppf)Cl 2(32.7mg,0.04mmol)、K 3PO 4(163.4mg,0.77mmol)和苯基硼酸(134.2mg,1.1mmol)。反应混合物在100℃下搅拌16h(过夜)，Ar下LCMS(EPN18040-023-1)显示反应完成，SM残留5％。反应混合物集中在真空中。采用自动闪柱柱层析(硅胶，PE/EA＝8:1)对残渣进行纯化，得到粗品，用闪柱(C-18柱层析，H 2O(NH 4HCO 3,0.8g/L)/CH 3CN＝60/40)对固体进行进一步纯化，得到标题化合物(2.6mg,4.1％收率)为白色固体。
(ESI)m/z＝319.21(M+H) +. 1H NMR(500MHz,DMSO-d6)11.75(s,1H),9.44(s,1H),8.35-8.37(m,2H),8.09-8.13(m,1H),7.70(m,1H),7.49-7.52(m,2H),7.38-7.45(m,2H),6.81-6.85(m,1H),6.52(s,1H),2.4(s,3H)ppm.
实施例32
Figure PCTCN2020082016-appb-000050
Step 1:2-氯-N-异丙基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
取2,4-二氯-7H-吡咯并[2,3-d]嘧啶(1.0g,5.31mmol)，丙二胺(3.0mL)，DIPEA(1.0mL)分别充入20mL密封管，再加入二氧六环(10ml)。反应混合物在70℃下搅拌16h。反应完成后，混合物在常压真空蒸馏下浓缩。将残渣溶解于(EtOH) 5mL，加入水(20mL)，RT搅拌10min。对混合料进行过滤，收集固体，真空干燥，得到目标化合物(940mg，收率83％)为白色固体。
分子式:C 9H 11ClN 4,分子量:210.67,(ESI)m/z＝211.2(M+H) +.
Step 2:N-异丙基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(3 EPT60138)
以2-氯-N-异丙基-7H-吡咯[2,3-d]吡啶-4-胺(440mg,4.0mmol)、苯基硼酸(1.22g,10.0mmol)、Pd(dppf)Cl 2(140mg,0.2mmol)、K 3PO 4(2.12g,10.0mmol)充入10-mL密封管，然后加入二氧六环(15mL)和水(3.0mL)。反应混合物在惰性气体中以100℃加热22h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝1:3)对产物进行纯化，得到标题化合物224mg，收率44％)为白色固体，为白色固体。
分子式:C 15H 16N 4,分子量:252.32,(ESI)m/z＝253.2(M+H) +. 1HNMR(500MHz,DMSO-d 6)11.50(s,1H),8.37(m,2H),7.37-7.46(m,3H),7.19(d,J＝7.5Hz,1H),7.08(m,1H),6.60(m,1H)4.56(m,1H),1.30(d,J＝6.5Hz,6H)ppm.
实施例33
Figure PCTCN2020082016-appb-000051
Step 1:6-溴-N-异丙基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(2 EPT60139)
将N-异丙基-2-苯基-7H-吡啶醇[2,3-d]嘧啶-4-胺(200mg,0.8mmol)溶于DCM(30mL)中，然后将NBS(144mg溶于DCM(10mL)溶液中)于0-10oC缓慢滴入30min。反应混合物在rt下搅拌20min。反应由LCMS监测。反应完成后，将反应混合物与硅胶(100-200目)通 过蒸发浓缩，得到粉末残渣。采用自动闪柱柱层析法(硅胶，EA:PE＝1:8)对产物进行纯化，得到标题化合物(152mg，收率57.5％)为白色固体。
分子式:C 15H 15BrN 4,分子量:331.22,(ESI)m/z＝331.1(M+H) +. 1HNMR(500MHz,DMSO-d 6)12.03(s,1H),8.35-8.37(m,2H),7.42-7.49(m,3H),7.36(d,J＝2.5Hz,1H),5.99(d,J＝7.5Hz,1H),4.55(m,1H),1.34(d,J＝6.5Hz,6H)ppm.
实施例34
Figure PCTCN2020082016-appb-000052
Step 1:N-环丙基-2-(3-氟苯基)-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(7)
在一个25毫升的圆底烧瓶中加入2-氯-N-环丙基-6-甲基-7-甲苯磺酰-7H-吡咯并[2,3-d]嘧啶-4-胺(100mg,0.26mmol)，(3-氟苯基)硼酸(109mg,0.78mmol)，Pd(dppf)Cl 2(19mg,0.026mmol)和K 3PO 4(93mg,0.78mmol)。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃的氮气气氛下进行了一夜。减压除去溶剂，采用自动闪柱柱层析(硅胶，PE:EA＝1:1)纯化得到白色固体N-环丙基-2-(3-氟苯基)-6-甲基-7-甲苯磺酰-7H-吡咯并[2,3-d]嘧啶-4-胺(7,100mg,88.21％收率)。LCMS:(ESI)m/z＝437.32(M+H) +；RT＝1.13min.
Step 2:N-环丙基-2-(3-氟苯基)-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60152)
向25mL圆底烧瓶中加入N-环丙基-2-(3-氟苯基)-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(100mg)将0.23mmol(0.23mmol)溶于MeONa(1ml，5.4M的MeOH溶液)和MeOH(5ml)中，并在50℃下搅拌2小时。将残余物真空浓缩，并通过自动快速柱色谱法(硅胶，PE：EA＝4：1)纯化，得到灰白色固体N-环丙基-2-(3-氟苯基)-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60152,50mg，产率77.09％)。
LCMS:(ESI)m/z＝283.16(M+H) +；RT＝1.31min.
1H NMR(500MHz,DMSO)δ11.48(s,1H),8.21(d,J＝7.8Hz,1H),8.07(d,J＝10.0Hz,1H),7.48(td,J＝8.0,6.2Hz,1H),7.42(d,J＝3.1Hz,1H),7.21(d,J＝2.7Hz,1H),6.30(s,1H),3.02(dd,J＝6.6,3.4Hz,1H),2.33(s,3H),0.84–0.79(m,2H),0.63–0.57(m,2H).
实施例35
Figure PCTCN2020082016-appb-000053
Step 1:1-((2-氯-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-基)氨基)-2-甲基丙-2-醇(2)
在EtOH(5mL)中加入化合物1(100mg,0.28mmol)的溶液中加入Et 3N(56.6mg,0.56mmol)和1-氨基-2-甲基propaN-2-ol(49.8mg,0.56mmol)。反应混合物在80℃下搅拌2小时。LCMS(EPN18040-019-1)显示反应完成，无SM残留。溶剂在真空中浓缩。未纯化的粗化合物直接用于下一步(ESI)m/z＝409.28(M+H) +.
Step 2:2-甲基-1-((6-甲基-2-苯基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-基)氨基)丙-2-醇(3)
将化合物2(90mg,0.22mmol)在二氧六环(5mL)和H 2O(1mL)溶液中分别加入K 3PO 4(163.4mg,0.77mmol)、Pd(dppf)Cl 2(35.9mg,0.04mmol)和苯基硼酸(134.2mg,1.1mmol)。反应混合物在100℃下搅拌16h(过夜)，Ar下LCMS(EPN18040-021-1)显示反应完成，SM残留35％。溶剂在真空中浓缩。采用自动闪柱柱层析(硅胶，PE/EA＝8:1)对残留进行纯化，再用闪柱(C-18柱层析，H 2O(NH 4HCO 3,0.8g/L)/CH 3CN＝70/30)进行纯化，得到标题化合物(43.5mg,43.8％收率)为淡黄色固体。(ESI)m/z＝451.36(M+H) +.
Step 3:2-甲基-1-((6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-基)氨基)丙-2-醇(EPT60153)
在MeOH(5mL)中加入化合物3(43.5mg,0.10mmol)溶液中加入MeONa(0.5mL,2.7mmol，)。反应混合物在60℃下搅拌4小时。LCMS(EPN18040-025-1)显示反应完成，无SM残留。溶剂在真空中浓缩。用饱和NH 4Cl溶液(30ml)稀释残渣，用CH 2Cl 2(30ml×3)萃取混合物，再用饱和NaCl(30ml)洗涤。用无水Na 2SO 4干燥所得到的有机层，在真空中除去溶剂。用Flash(C-18柱层析，H 2O(NH 4HCO 3,0.8g/L)/CH 3CN＝40/60)对残留进行纯化，得到标题化合物(5.8mg,20.3％收率)为白色固体。
(ESI)m/z＝297.23(M+H) +. 1H NMR(500MHz,DMSO-d6)11.43(s,1H),8.32-8.34(m,2H),7.42-7.45(m,2H),7.36-7.39(m,1H),7.14(s,1H),6.31(s,1H),5.01(s,1H),3.58-3.59(d,J＝5.5Hz,2H),2.33(s,3H),1.17(s,6H)ppm.
实施例36
Figure PCTCN2020082016-appb-000054
Step 1:2-氯-N-环戊基-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
取2,4-二氯-6-甲基-7-甲苯磺酰-7H-吡啶[2,3-d]嘧啶(100mg,0.28mmol)、环戊胺(0.5mL)、DIPEA(0.5mL)分别充入20ml密封管，加入2,4-二氯-6-甲基-7-甲苯磺酰-7H-pyrrolo[2,3-d]嘧啶(100mg,0.28mmol)、环戊胺(0.5mL)、二氧环(5ml)。反应混合物在70℃下搅拌16h。反应由LCMS监测。分子式:C 19H 21ClN 4O 2S，分子量:404.91,(ESI) m/z＝405.3(M+H) +.No SM-1was remained,the crude product was used in the next step without further purification.
Step 2:N-环戊基-6-甲基-2-苯基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(3)
以2-氯-N-环戊基-6-甲基-7-甲苯磺酰-7H-吡咯并[2,3-d]2,3-d-嘧啶-4-胺(5mL二氧六环中粗品)、苯基硼酸(110mg,0.9mmol)、Pd(dppf)Cl 2(20mg,0.03mmol)、K 3PO 4(210mg,1.0mmol)充入10ml密封管，再加水(1.0mL)。反应混合物在100℃惰性气体中加热16h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝1:7)对产物进行纯化，得到产物为白色固体(100mg，收率80％)。
分子式:C 25H 26N 4O 2S,分子量:446.57,(ESI)m/z＝447.3(M+H) +.
Step 3:N-环戊基-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(4 EPT60155)
将N-环戊基-6-甲基-2-苯基-7-甲苯磺酰-7H-吡咯[2,3-d]嘧啶-4-胺(100mg,0.22mmol)溶于CH 3OH(5mL)中，加入CH 3ONa(5.4M,0.5mL)。反应混合物在55℃下搅拌20h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝1:3)对产物进行纯化，得到产物为白色固体(55mg，收率84％)。
分子式:C 18H 20N 4,分子量:292.39,(ESI)m/z＝293.2(M+H) +. 1HNMR(500MHz,DMSO-d 6) 11.37(s,1H),8.34-8.36(m,2H),7.35-7.44(m,3H),7.08(d,J＝7.0Hz,1H),6.27(s,1H),4.57(m,1H),2.31(s,3H),2.03-2.08(m,2H),1.73-1.74(m,2H),1.57-1.63(m,4H)ppm.
实施例37
Figure PCTCN2020082016-appb-000055
Step 1:2-氯-N-环己基-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
取2,4-二氯-6-甲基-7-甲苯磺酰-7H-吡啶[2,3-d]嘧啶(100mg,0.28mmol)、环己胺(0.5mL)、DIPEA(0.5mL)分别充入20ml密封管，加入5mL的二氧六环。反应混合物在70℃下搅拌16h。反应由LCMS监测。分子式:C 20H 23ClN 4O 2S,分子量:418.94(ESI)m/z＝419.3(M+H) +.没有残留SM-1，下一步使用粗品，没有进一步纯化。
Step 2:N-环己基-6-甲基-2-苯基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(3)
以2-氯-N-环己基-6-甲基-7-甲苯磺酰-7H-吡咯并[2,3-d]2,3-d-嘧啶-4-胺(5mL二氧六环中粗品)、苯基硼酸(110mg,0.9mmol)、Pd(dppf)Cl 2(20mg,0.03mmol)、K 3PO 4(210mg,1.0mmol)充入10ml密封管，再加水(1.0mL)。反应混合物在100℃惰性气体中加热14h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自 动闪柱柱层析法(硅胶，EA:PE＝1:9)对产物进行纯化，得到标题化合物(105mg，收率81％)为白色固体。
分子式:C 26H 28N 4O 2S,分子量:460.60(ESI)m/z＝461.3(M+H) +.
Step 3:N-环己基-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(4 EPT60156)
将N-环己基-6-甲基-2-苯基-7-甲苯磺酰-7H-吡咯[2,3-d]吡啶-4-胺(105mg,0.22mmol)溶于CH 3OH(5mL)中，加入CH 3ONa(5.4M,0.5mL)。反应混合物在55℃下搅拌20h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱层析(硅胶，EA:PE＝3:7)对产物进行纯化，得到产物为白色固体(60mg,85％收率)。
分子式:C 19H 22N 4,分子量:306.41,(ESI)m/z＝307.2(M+H) +. 1HNMR(500MHz,DMSO-d 6)11.34(s,1H),8.33-8.35(m,2H),7.41-7.44(m,2H),7.35-7.38(m,1H),6.95(d,J＝7.5Hz,1H),6.21(s,1H),4.15(m,1H),2.31(s,3H),2.03(m,2H),1.17-1.80(m,8H)ppm.
实施例38
Figure PCTCN2020082016-appb-000056
Step 1:N-环丙基-2-(4-氟苯基)-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
以2-氯-N-环丙基-6-甲基-7-甲苯磺酰-7H-吡咯烷[2,3-d]嘧啶-4-胺(75mg,0.2mmol)、2-(4-氟苯基)-4、4、5-四甲基-1,3,2-二噁硼烷(100mg,0.45mmol)、Pd(dppf)Cl 2(14mg,0.02mmol)、K 3PO 4(120mg,0.6mmol)、水(1.0mL)、二氧六环(5mL)充入10-mL密封管。反应混合物在90℃惰性气体中加热16h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱层析法(硅胶，EA:PE＝1:4)对产物进行纯化，得到标题化合物(71mg，收率81％)为白色固体。
分子式:C 23H 21FN 4O 2S,分子量:436.51(ESI)m/z＝437.3(M+H) +.
Step 3:N-环丙基-2-(4-氟苯基)-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60157)
将N-环丙基-2-(4-氟苯基)-6-甲基-7-甲苯磺酰-7H-吡咯[2,3-d]嘧啶-4-胺(71mg,0.16mmol)溶于THF(5mL)中，加入TBAF(1M,0.5mL)。反应混合物在55℃下搅拌1h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱层析法(硅胶，EA:PE＝1:4)对产物进行纯化，得到标题化合物(32mg，收率69.5％)为白色固体。分子式:C 16H 15FN 4,分子量:282.32(ESI)m/z＝283.3(M+H) +. 1HNMR(400MHz,DMSO-d 6)11.36(s,1H),8.34-8.37(m,2H),7.32(d,J＝3.2Hz,1H),7.18-7.22(m,2H),6.23(s,1H),2.97(m,1H),2.28(s,3H),0.74-0.83(m,2H),0.53-0.576(m,2H)ppm.
实施例39
Figure PCTCN2020082016-appb-000057
Step 1:2-氯-N-异丁基-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
在EtOH(3ml)中加入化合物1(100mg,0.28mmol)的溶液中加入Et 3N(56.6mg,0.56mmol)和2-甲基丙基-1-胺(40.9mg,0.56mmol)。反应混合物在80℃下搅拌4h,LCMS(EPN18040-040-1)表明反应已经完成，没有SM残留。溶剂在真空中浓缩。未纯化的粗化合物直接用于下一步。(ESI)m/z＝393.22(M+H) +.
Step 2:N-异丁基-6-甲基-2-苯基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(3)
将化合物2(90mg,0.23mmol)在二噁烃(5mL)和H 2O(1mL)溶液中分别加入K 3PO 4(163.5mg,0.81mmol)、Pd(dppf)Cl 2(35.9mg,0.046mmol)和苯基硼酸(134.2mg,1.15mmol)。反应混合物在100℃下搅拌16h(过夜)。LCMS(EPN18040-041-1)显示反应完成，SM残留10％。溶剂在真空中浓缩。采用自动闪柱柱层析(硅胶，PE/EA＝8:1)对残留进行纯化，再用闪柱(C-18柱层析，H 2O(NH 4HCO 3,0.8g/L)/CH 3CN＝70/30)进行纯化，得到标题化合物(15mg,15.1％收率)为淡黄色固体。
(ESI)m/z＝435.32(M+H) +.
Step 3:N-异丁基-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60163)
在MeOH(3ml)中加入化合物1(15mg,0.03mmol)溶液中加入MeONa(0.5mL,2.7mmol)。反应混合物在60℃下搅拌6h,LCMS(EPN18040-043-1)显示反应已经完成，没有SM残留。溶剂在真空中浓缩。反应混合物用饱和NH 4Cl溶液(30ml)稀释，用CH 2Cl 2(30ml×3)萃取，再用饱和NaCl(30ml)洗涤。用无水Na 2SO 4干燥所得到的有机层，在真空中除去溶剂。采用闪柱(C-18柱层析，H 2O(NH 4HCO 3,0.8g/L)/CH 3CN＝70/30)纯化得到标题化合物(8.0mg,87％收率)为白色固体。(ESI)m/z＝281.21(M+H) +. 1H NMR(500MHz,DMSO-d6)11.36(s,1H),8.34-8.35(d,J＝8.5Hz,2H),7.41-7.44(m,2H),7.37(m,1H),7.24-7.26(m,1H),6.25(s,1H),3.50(m,2H),2.30(s,3H),2.02-2.04(m,1H),0.78-0.95(m,6H)ppm.
实施例40
Figure PCTCN2020082016-appb-000058
Step 1:2-氯-6-甲基-4-苯基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶(2)
将化合物1(150mg,0.43mmol)在二氧六环(5mL)和H 2O(1mL)溶液中分别加入K 3PO 4(320.5mg,1.51mmol)、Pd(dppf)Cl 2(73.4mg,0.09mmol)和苯硼酸(52.5mg,1.0mmol)。反应混合物在80℃条件下搅拌16h(过夜)，Ar条件下LCMS(EPN18040-028-1)显示反应已经完成，没有SM残留。溶剂在真空中浓缩。采用自动闪柱柱色谱法(硅胶，PE/EA＝8:1)对残留进行纯化，得到标题化合物(90mg,54％收率)为淡黄色固体。(ESI)m/z＝398.20(M+H) +.
Step 2:N-异丙基-6-甲基-4-苯基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-2-胺(3)
在化合物2(85mg,0.21mmol)的二氧六环(3mL)溶液中加入tBuOK(47.1mg,0.42mmol)、pd(OAc) 2(4.5mg,0.02mmol)、BINAP(24.9mg,0.2mmol)和propaN-2-amine(25mg,0.42mmol)。反应混合物在Ar下60℃搅拌16h(过夜)，LCMS(EPN18040-034-1)显示反应已经完成，没有SM残留。溶剂在真空中浓缩。采用自动闪柱柱层析(硅胶，PE/EA＝8:1)对残留进行纯化，再用闪柱(C-18柱层析，H 2O(NH 4HCO 3,0.8g/L)/CH 3CN＝40/60)进行纯化，得到标题化合物(30mg,34％收率)为淡黄色固体。(ESI)m/z＝421.88(M+H) +.
Step 3:N-异丙基-6-甲基-4-苯基-7H-吡咯并[2,3-d]嘧啶-2-胺(EPT60164)
在MeOH(3ml)中加入化合物3(30mg,0.07mmol)溶液中加入MeONa(0.5mL,2.7mmol)。反应混合物在60℃下搅拌4h,LCMS(EPN18040-036-1)表明反应已经完成，没有SM残留。溶剂在真空中浓缩。反应混合物用饱和NH 4Cl溶液(30ml)稀释，用CH 2Cl 2(30ml×3)萃取，然后用饱和NaCl(30ml)洗涤。用无水Na 2SO 4干燥所得到的有机层，在真空中除去溶剂。经C-18柱层析，H 2O(NH 4HCO 3,0.8g/L)/CH 3CN＝70/30纯化，得到标题化合物(3.1mg,16.4％收率)为白色固体。(ESI)m/z＝267.24(M+H) +.1H NMR(400MHz,DMSO-d6)11.12(s,1H),8.00-8.02(d,J＝8.4Hz,2H),7.43-7.50(m,3H),6.22-6.26(m,2H),4.02(m,1H),2.26(s,3H),1.45-1.13(m,6H)ppm.
实施例41
Figure PCTCN2020082016-appb-000059
Step 1:2-氯-N-(3-氯苯基)-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
在DMSO(3ml)中加入化合物1(100mg,0.50mmol)，加入Cs 2CO 3(182.5mg,0.56mmol)和3-氯苯胺(71.1mg,0.56mmol)溶液。反应混合物在40℃下搅拌16小时(过夜)。LCMS(EPN18040-027-1)显示反应完成，SM残留5％。反应混合物用H 2O(30mL)稀释，用CH 2Cl 2(50mL×3)萃取，然后用饱和NaCl(30mL)洗涤。所得的有机层用无水Na 2SO 4干燥，在真空中除去溶剂，得到所需的产物(90mg,71.6％收率)为淡黄色固体。(ESI)m/z＝447.20(M+H) +.
Step 2:N-(3-氯苯基)-6-甲基-2-苯基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(3)
将化合物2(90mg,0.20mmol)在二噁烃(5mL)和H 2O(1mL)溶液中分别加入K 3PO 4(148.6mg,0.70mmol)、Pd(dppf)Cl 2(29.3mg,0.04mmol)和苯硼酸(122.0mg,1.0mmol)。反应混合物在100℃条件下搅拌16h(过夜)。LCMS(EPN18040-031-1)显示反应完成，SM残留20％。溶剂在真空中浓缩。采用自动闪柱柱层析(硅胶，PE/EA＝8:1)对残留进行纯化，再用闪柱(C-18柱层析，H 2O(NH 4HCO 3,0.8g/L)/CH 3CN＝70/30)进行纯化，得到标题化合物(20mg,20.3％收率)为白色固体。
(ESI)m/z＝489.25(M+H) +.
Step 3:N-(3-氯苯基)-6-甲基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60165)
在MeOH(3ml)中加入化合物3(20mg,0.04mmol)溶液中加入MeONa(0.5mL,2.7mmol)。反应混合物在60℃下搅拌5小时。LCMS(EPN18040-037-1)显示反应完成，无SM残留。溶剂在真空中浓缩。反应混合物用H 2O(30ml)稀释，用CH 2Cl 2(30ml×3)萃取，然后用饱和NaCl(30ml)洗涤。用无水Na 2SO 4干燥所得到的有机层，在真空中除去溶剂。反应混合物用H 2O(30ml)稀释，用CH 2Cl 2(30ml×3)萃取，然后用饱和NaCl(30ml)洗涤。用无水Na 2SO 4干燥所得到的有机层，在真空中除去溶剂。采用预薄层色谱法(PE/EA＝2/1)对残留进行纯化，得到所需产物(1.9mg,8.5％yiled)为白色固体。
(ESI)m/z＝335.22(M+H) +1H NMR(400MHz,DMSO-d6)11.73(s,1H),9.39(s,1H),8.32-8.34(m,3H),7.80-7.82(m,1H),7.35-7.47(m,4H),7.01-7.04(m,1H),6.47(m,1H),2.36(m,3H)ppm.
实施例42
Figure PCTCN2020082016-appb-000060
Step 1:N-环丙基-6-甲基-2-(间甲苯基)-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
用N-环丙基-6-甲基-2-(间甲苯基)-7-甲苯磺酰-7H-吡咯并[2,3-d]，间甲苯硼酸(40mg,0.3mmol)，Pd(dppf)Cl 2(14mg,0.02mmol)，K 3PO 4(63mg,0.3mmol)，加入水 (0.5mL)和二氧六环(3mL)，充入10-mL密封管。反应混合物在90℃惰性气体中加热16h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝1:3)对产物进行纯化，得到产物为白色固体(84mg,97％收率)。
分子式:C 24H 24N 4O 2S,分子量:432.54,(ESI)m/z＝433.3(M+H) +.
Step 2:N-环丙基-6-甲基-2-(间甲苯基)-7H-吡咯并[2,3-d]嘧啶-4-胺(3 EPT60166)
将N-环丙基-6-甲基-2-(间甲苯基)-7-甲苯磺酰-7H-吡咯啉[2,3-d]嘧啶-4-胺(84mg,0.20mmol)溶于THF(5mL)中，加入TBAF(2M,2.0mL)。反应混合物在55℃下搅拌24h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱层析法(硅胶，EA:PE＝1:3)对产物进行纯化，得到标题化合物(26mg，收率46％)为白色固体。
分子式:C 17H 18N 4,分子量:278.36,(ESI)m/z＝279.2(M+H) +. 1HNMR(500MHz,DMSO-d 6)11.39(s,1H),8.16-8.19(m,2H),7.29-7.32(m,2H)，7.18(d,J＝7.5Hz,1H),6.28(s,1H),3.01(s,1H),2.38(s,3H),2.32(s,3H),0.78-0.85(m,2H),0.58-0.61(m,2H)ppm.
实施例43
Figure PCTCN2020082016-appb-000061
Step 1:N-环丙基-6-甲基-2-(邻甲苯基)-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(3)
在一个25毫升的圆底烧瓶中加入2-氯-N-环丙基-6-甲基-7-甲苯磺酰-7H-吡咯啉[2,3-d]嘧啶-4-胺(50mg,0.13mmol)，邻甲苯硼酸(53mg,0.39mmol)，Pd(dppf)Cl 2(9mg,0.013mmol)和K 3PO 4(83mg,0.39mmol)。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃的氮气气氛下进行了一夜。减压除去溶剂，首先用自动闪柱柱层析(硅胶，PE:EA＝90:10)纯化得到N-环丙基-6-甲基-2-(o-tolyl)-7-甲苯磺酰-7H-吡咯并[2,3-d]嘧啶-4-胺(7,20mg,35.53％收率)为黄色固体。
LCMS:(ESI)m/z＝433.12(M+H) +；RT＝1.91min.
Step 2:N-环丙基-6-甲基-2-(邻甲苯基)-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60167)
到25ml圆底烧瓶中加入N-cyclopropyl-6-methyl-2——(o-tolyl)7-甲苯磺酰-7H-pyrrolo[2,3-d]pyrimidiN-4-amine(0.05 7,20毫克,mmol)溶解在TBAF(1毫升,1.0年的四氢呋喃)和四氢呋喃(2毫升)和搅拌在50℃ 4h。残渣被自动闪光集中在真空和纯化柱层析法(硅胶,DCM:甲醇＝97:3)给黄色固体N-cyclopropyl-6-methyl-2-(o-tolyl)7H-pyrrolo[2,3-d]pyrimidiN-4-amine(EPT60167 6毫克,43.16％的收率)。
LCMS:(ESI)m/z＝279.24(M+H) +；RT＝1.23min.
1H NMR(500MHz,DMSO)δ11.35(s,1H),7.73(d,J＝6.4Hz,1H),7.31(d,J＝2.5Hz,1H),7.24(d,J＝5.8Hz,3H),6.27(s,1H),2.91(s,1H),2.55(s,3H),2.33(s,3H),0.75(d,J＝5.2Hz,2H),0.57(s,2H).
实施例44
Figure PCTCN2020082016-appb-000062
Step 1:2-(2,4-二羟基-7H-吡咯并[2,3-d]嘧啶-6-基)-乙酸乙酯(3)
在6-氨基嘧啶-2,4(1H,3H)-二酮(1)(15.0g,118mmol)水溶液(300mL)悬浮液中加入乙酸钠(10.7g,130mmol)。将混合物加热到95℃。滴加4-氯-3-草酸乙酯(2)(21.4g,130mmol)，静置30min,95℃搅拌过夜，冷却过滤。蛋糕用水(30ml×3)和丙酮(30ml×2)洗涤，真空干燥，得到标题化合物(5.0g，收率18％)为黄色固体。
1H NMR(400MHz,DMSO-d 6):δ11.49(s,1H),11.15(s,1H),10.46(s,1H),6.05(s,1H),4.08(q,J＝7.2Hz,2H),3.60(s,2H),1.20(t,J＝7.2Hz,3H).
Step 2:2-(2,4-二氯-7H-吡咯并[2,3-d]嘧啶-6-基)乙酸乙酯(4)
在甲苯(12ml)中加入2-(2-苯基-4-(苯基氨基)-7H-吡咯[2,3-d]嘧啶-6-基)乙酸乙酯(3)(5.00g,2.11mmol)，POCl 3(12.9g,8.44mmol)。将反应升温至70℃，滴加N、N-二异丙基乙胺(7.08g,5.49mmol)10min,110℃搅拌过夜。将混合物冷却至室温，倒入水中(50毫升)。加入DIEA(10mL)，搅拌30min，加入乙酸乙酯(50mL)，过滤。滤液用乙酸乙酯(50mL×2)萃取，混合有机物层用盐水(20mL)冲洗，Na 2SO 4烘干，过滤。滤液浓缩后，用硅胶柱层析法(石油醚:乙酸乙酯，5:1～3:1)纯化残渣，得到粗品1.1g。将原油与(石油醚:乙酸乙酯＝10:1,5mL)三聚氰胺(880mg，收率15％)反应得到黄色固体。
1H NMR(400MHz,DMSO-d 6):δ12.74(s,1H),6.54(s,1H),4.14(q,J＝6.8Hz,2H),3.95(s,2H),1.21(t,J＝6.8Hz,3H).
Step 3:2-(2-氯-4-(苯基氨基)-7H-吡咯并[2,3-d]嘧啶-6-基)乙酸乙酯(5)
在乙二醇(15ml)中乙酸乙酯(4)(880mg,3.21mmol)和苯胺(806mg,8.67mmol)溶液中加入2滴浓缩盐酸水溶液。将混合物加热到95摄氏度，搅拌5小时。冷却至室温后，倒入80毫升水搅拌10分钟，过滤。滤饼用水(10mL×5)和石油醚(10mL×5)洗涤，真空干燥，得到标题化合物(950mg，收率90％)为粉红色固体。
1H NMR(400MHz,DMSO-d 6):δ11.86(s,1H),9.61(s,1H),7.76(d,J＝7.6Hz,2H),7.36(t,J＝8.0Hz,2H),7.07(t,J＝7.6Hz,1H),6.61(s,1H),4.13(q,J＝7.2Hz,2H),3.81(s,2H),1.22(t,J＝7.2Hz,3H).
Step 4:2-(2-苯基-4-(苯基氨基)-7H-吡咯并[2,3-d]嘧啶-6-基)乙酸乙酯(6,EPT60168)
将2-乙基2-(2-氯-4-(苯基氨基)-7H-吡啶醇[2,3-d]]乙酸乙酯(5)(500mg,1.51mmol)，苯基硼酸(553mg,4.53mmol)，DIEA(974mg,7.55mmol)和Pd(PPh3) 4(349mg,0.30mmol)在DMA/水(5mL/1mL)中密封管中，在140℃微波下搅拌3小时。冷却到室温后,混合物注入水(30毫升),用乙酸乙酯提取(20毫升2)。合并后的有机层集中和残渣被C 18柱纯化(乙腈:水15分钟期间从60％到80％)给原油产品(160毫克)。用预薄层色谱法(石油醚:乙酸乙酯＝3:1)纯化粗品60mg，得到标题化合物(30mg，收率14％)为黄色固体。 1H NMR(400MHz,DMSO-d 6):δ11.79(s,1H),9.35(s,1H),8.37(d,J＝9.2Hz,2H),7.98(d,J＝10.4Hz,2H),7.51-7.40(m,5H),7.04(t,J＝10.0Hz,1H),6.69(s,1H),4.14(q,J＝9.6Hz,2H),3.85(s,2H),1.23(t,J＝9.6Hz,3H).
LC-MS[流动相:从80％水(0.02％NH 4OAc)和20％CH 3CN变为5％水(0.02％NH 4OAc)和95％CH 3CN，在6.5min内],Rt＝3.981min；纯度:94.35％(214nm),96.03％(254nm)；MS计算值:372.2；MS实测值:373.0[M+H] +.
实施例45
Figure PCTCN2020082016-appb-000063
Step 2:N-环丙基-6-甲基-2-(对甲苯基)-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
将化合物1(100mg,0.27mmol)在二氧六环(5mL)和H 2O(1mL)溶液中分别加入K 3PO 4(201.7mg,0.95mmol)、Pd(dppf)Cl 2(36.6mg,0.05mmol)和4-甲基苯硼酸(183.6mg,1.35mmol)。反应混合物在100℃下搅拌16h(过夜)，Ar条件下LCMS(EPN18040-044-1)显示反应完成，SM残留30％。溶剂在真空中浓缩。采用自动闪柱柱色谱法(硅胶，PE/EA＝20/1)对残留进行纯化，得到标题化合物(60mg,43.8％收率)为淡黄色固体。(ESI)m/z＝433.36(M+H) +.
Step 2:N-环丙基-6-甲基-2-(对甲苯基)-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60180)
在MeOH(3ml)中加入化合物2(60mg,0.14mmol)溶液中加入MeONa(0.5mL,2.7mmol)。反应混合物在60℃下搅拌4h,LCMS(EPN18040-046-1)表明反应已经完成，没有SM残留。溶剂在真空中浓缩。反应混合物用饱和NH 4Cl溶液(30ml)稀释，用CH 2Cl 2(30ml×3)萃取，然后用饱和NaCl(30ml)洗涤。用无水Na 2SO 4干燥所得到的有机层，在 真空中除去溶剂。经C-18柱层析，H 2O(NH 4HCO 3,0.8g/L)/CH 3CN＝70/30纯化，得到标题化合物(3.1mg,16.4％产率)白色固体。(ESI)m/z＝279.24(M+H) +. 1H NMR(500MHz,DMSO-d6)11.26(s,1H),8.25-8.26(d,J＝8.0Hz,2H),7.31-7.32(m,1H),7.22-7.24(m,2H),6.23(s,1H),2.99(s,1H),2.35(s,3H),2.31(s,3H),0.79-0.81(s,2H),0.58-0.59(s,2H)ppm.
实施例46
Figure PCTCN2020082016-appb-000064
Step 1:N-异丙基2,6-二苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(5 EPT60181)
取6-溴-N-异丙基-2-苯基-7H-吡咯[2,3-d]嘧啶-4-胺(50mg,0.15mmol)、苯基硼酸(61mg,0.5mmol)、Pd(dppf)Cl 2(10mg,0.015mmol)、K 3PO 4(110mg,0.5mmol)、水(1.0mL)、二氧六环(5mL)充入10ml密封管。反应混合物在90℃惰性气体中加热16h。反应由LCMS监测。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝35:65)对产物进行纯化，得到产物为白色固体(30mg，收率60％)。
分子式:C 21H 20N 4,分子量:328.42,(ESI)m/z＝329.3(M+H) +. 1H NMR(400MHz,DMSO-d6)11.86(s,1H),8.36-8.38(m,2H),7.33-7.50(m,8H),7.24(d,J＝2.4Hz,1H),5.06(d,J＝7.2Hz,1H),4.41-4.46(m,1H),1.17(d,J＝6.4Hz,6H)ppm.
实施例47
Figure PCTCN2020082016-appb-000065
Step 1:4-(4-(环丙基氨基)-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-2-基)苯酚(2)
10mL封管中加入2-氯-N-环丙基-6-甲基-7-甲苯磺酰-7-h-吡咯并[2,3-d]嘧啶-4-氨基(110mg,0.3mmol)、(4-羟基苯基)硼酸(84mg,0.6mmol)、Pd(dppf)Cl 2(20mg,0.03mmol)、K 3PO 4(124mg,0.6mmol)、水(1.0mL)、二氧六环(5mL)。反应混合物在90℃惰性气体中加热16h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱层析法(硅胶，EA:PE＝3:7)对产物进行纯化，得到标题化合物(88.8mg，收率73.6％)为白色固体。
分子式:C 23H 22N 4O 3S,分子量:434.51,(ESI)m/z＝435.3(M+H) +.
Step 2:4-(4-(环丙基氨基)-6-甲基-7H-吡咯并[2,3-d]嘧啶-2-基)苯酚(3 EPT60182)
将4-(4-(环丙基氨基)-6-甲基-7-甲苯磺酰-7H-吡咯[2,3-d]嘧啶-2-基)苯酚(50mg,0.115mmol)溶于CH 3OH(2mL)中，加入CH 3ONa(1.0mL)。反应混合物在55℃下搅拌16h。反应完成后，用饱和NH 4Cl水溶液(1mL)对反应进行骤冷，再用硅胶(100-200mesh)对反应混合物进行蒸发浓缩，得到粉体残渣。采用自动闪柱柱层析法(硅胶，EA:PE＝2:3)对产物进行纯化，得到标题化合物(8mg，收率25％)为白色固体。
分子式:C 16H 16N 4O,分子量:280.33(ESI)m/z＝281.2(M+H) +. 1H NMR(400MHz,DMSO-d 6)11.23(s,1H),9.54(s,1H),8.15(d,J＝8.4Hz,2H),7.18(d,J＝2.8Hz,1H),6.75(d,J＝8.8Hz,2H),6.20(s,1H),2.94-2.98(m,1H),2.26(s,3H),0.73-0.77(m,2H),0.52-0.56(m,2H)ppm.
实施例48
Figure PCTCN2020082016-appb-000066
Step 1:2-氯-N-环丁基-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
用2,4-二氯-6-甲基-7-甲苯磺酰-7-h-吡咯并[2,3-d]嘧啶(107mg,0.3mmol)、环丁胺(185mg,2.6mmol)分别充入20ml密封管，然后加入二氧六环(5mL)。反应混合物在70℃下搅拌3h。反应由LCMS监测。
分子式:C 18H 19ClN 4O 2S,分子量:390.89,(ESI)m/z＝391.2(M+H) +,下一步直接使用。
Step 2:N-环丁基-2-(2-氟苯基)-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(3)
用2-氯-N-环丁基-6-甲基-7-甲苯磺酰-7-H-吡咯并[2,3-d]吡啶-4-胺(5mL二氧六环中粗品)、(2-氟苯基)硼酸(140mg,1.0mmol)、Pd(dppf)Cl 2(20mg,0.03mmol)、K 3PO 4(210mg,1.0mmol)充入10ml密封管，再加水(1.0mL)。反应混合物在90℃惰性气体中加热16h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，EA:PE＝15:85)对产物进行纯化，得到标题化合物112mg，收率83％)为白色固体。
分子式:C 24H 23FN 4O 2S,分子量:450.53,(ESI)m/z＝451.3(M+H) +.
Step 4:N-环丁基-2-(2-氟苯基)-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(4 EPT60183)
将N-环丁基-2-(2-氟苯基)-6-甲基-7-甲苯磺酰-7H-吡咯[2,3-d]嘧啶-4-胺(112mg,0.25mmol)溶于THF(5mL)中，加入TBAF(2M,2.0mL)。反应混合物在55℃下搅拌1h。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自 动闪柱柱色谱法(硅胶，EA:PE＝35:65)对产物进行纯化，得到产物为白色固体(35mg，收率47.3％)。
分子式:C 17H 17FN 4,分子量:296.35,(ESI)m/z＝297.2(M+H) +. 1H NMR(400MHz,DMSO-d 6)11.41(s,1H),7.91-7.95(m,1H),7.34-7.39(m,2H),7.16-7.23(m,2H),6.21(s,1H),4.61-4.67(m,1H),2.24-2.32(m,5H),1.96-2.06(m,2H),1.64-1.70(m,2H)ppm.
实施例49
Figure PCTCN2020082016-appb-000067
Step 1:N-异丙基-2-苯基-6-(三氟甲基)-7H-吡咯并[2,3-d]嘧啶-4-胺(2 EPT60184)
以N-异丙基-2-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(50mg,0.2mmol)、梅莫托试剂(Umemoto's reagents)(136mg,0.4mmol)、4-甲基吗啉(41mg,0.4mmol)充入10ml密封管，加入DMF(1mL)。反应混合物在室温下搅拌3h。反应由LCMS监测。反应完成后，将反应混合物与硅胶(100-200目)通过蒸发浓缩，得到粉末残渣。采用自动闪柱柱色谱法(硅胶，DCM)对产物进行纯化，得到产物为黄色固体(18mg，收率28％)。分子式分子式:C 16H 15F 3N 4,分子量:320.32,(ESI)m/z＝321.2(M+H) +. 1HNMR(500MHz,DMSO-d 6) 12.78(s,1H),8.37-8.40(m,2H),7.67(d,J＝7.5Hz,1H),7.43-7.50(m,3H),7.22(s,1H),4.54-4.58(m,1H),1.31(m,6H)ppm.
实施例50
Figure PCTCN2020082016-appb-000068
Step 1:2-氯-6-甲基-N-苯基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(2)
在一个25毫升的圆底烧瓶中加入2,4-二氯-6-甲基-7-甲苯磺酰-7H-吡啶[2,3-d]嘧啶(50mg,0.14mmol)，苯胺(26mg,0.28mmol)和Cs 2CO 3(138mg,0.42mmol)。将混合物悬浮于DMSO(3ml)中，50℃搅拌1h，残渣在真空中浓缩，得到2-氯-6-甲基-N-苯基-7-甲苯磺酰-7H-吡咯并[2,3-d]嘧啶-4-胺(50mg，收率86.67％)为黄色固体。粗产物用于下一步没有进一步提纯。
LCMS:(ESI)m/z＝413.24(M+H) +；RT＝1.88min.
Step 2:2-(2-氟苯基)-6-甲基-N-苯基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(3)
将2-氯-6-甲基-N-苯基-7-甲苯磺酰-7H-吡啶醇[2,3-d]4-氨基嘧啶(50mg,0.12mmol)，(2-氟苯基)硼酸(50mg,0.36mmol)，Pd(dppf)Cl 2(9mg,0.012mmol)和K 3PO 4(51mg,0.24mmol)分别装入25ml圆底烧瓶中。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃的氮气气氛下进行了一夜。减压除去溶剂，采用自动闪柱柱层析(硅胶，PE:EA＝1:1)纯化得到产物为黄色固体2-(2-氟苯基)-6-甲基-N-苯基-7-甲苯磺酰-7H-吡咯并[2,3-d]嘧啶-4-胺(7,40mg,70.62％收率)。LCMS:(ESI)m/z＝473.33(M+H) +；RT＝1.96min.
Step 3:2-(2-氟苯基)-6-甲基-N-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60185)
到25ml圆底烧瓶中加入2-(2-氟苯基) 6-甲基-N-苯基-7-甲基磺酰基-7H-吡咯并[2,3-d]嘧啶-N-4-胺(40毫克,0.08mmol)溶解在TBAF(1毫升,1.0年的四氢呋喃)和四氢呋喃(2毫升)和搅拌在8小时50℃。残留物被自动闪光集中在真空和纯化柱层析法(硅胶,DCM:甲醇＝97:3)给了白色固体2-(2-氟苯基)-6-甲基-N-苯基-7H-吡咯并[2,3-d]嘧啶-4-胺(10毫克,EPT60185 39.31％收率)。
LCMS:(ESI)m/z＝319.21(M+H) +；RT＝1.52min.
1H NMR(400MHz,dmso)δ11.76(s,1H),9.23(s,1H),8.05–7.95(m,3H),7.46(dd,J＝12.5,5.9Hz,1H),7.30(dt,J＝8.2,6.7Hz,4H),6.98(t,J＝7.3Hz,1H),6.51(s,1H),2.39(s,3H).
实施例51
Figure PCTCN2020082016-appb-000069
Step 1:N-环戊基-2-(2-氟苯基)-6-甲基-7-甲苯磺酰基-7H-吡咯并[2,3-d]嘧啶-4-胺(3)
在一个25毫升的圆底烧瓶中加入2-氯-N-环戊基-6-甲基-7-甲苯磺酰-7H-吡咯并[2,3-d]嘧啶-4-胺(100mg,0.25mmol)，(2-氟苯基)硼酸(105mg,0.75mmol)，Pd(dppf)Cl 2(18mg,0.025mmol)和K 3PO 4(106mg,0.5mmol)。混合物悬浮在二氧六环(5ml)和H 2O(1ml)中。反应在100℃的氮气气氛下进行了一夜。减压除去溶剂，用自动闪柱柱层析(硅胶，PE:EA＝60:40)纯化得到淡黄色固体N-环戊基-2-(2-氟苯基)-6-甲基-7-甲苯磺酰-7H-吡咯并[2,3-d]嘧啶-4-胺(7,110mg,94.83％)。LCMS:(ESI)m/z＝465.37(M+H) +；RT＝1.99min.
Step 2:N-环戊基-2-(2-氟苯基)-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60186)
到25ml圆底烧瓶中加入N-环戊基-2-(2-氟苯基)-6-甲基-7-甲基磺酰基-7H-吡咯并[2,3-d]嘧啶-N-4-胺(40毫克,0.09mmol)溶解在TBAF(1毫升,1.0年的四氢呋喃)和四氢呋喃(2毫升)和搅拌在8小时50℃。残留物被自动闪光集中在真空和纯化柱层析法(硅 胶,DCM:甲醇＝97:3)给了白色固体N-环戊基-2-(2-氟苯基)-6-甲基-7H-吡咯并[2,3-d]嘧啶-4-胺(EPT60186,25毫克,38.4％收率)。
LCMS:(ESI)m/z＝311.21(M+H) +；RT＝1.34min.
1H NMR(500MHz,DMSO)δ11.42(s,1H),7.97(td,J＝7.8,1.7Hz,1H),7.44–7.37(m,1H),7.28–7.19(m,2H),7.09(d,J＝6.9Hz,1H),6.28(d,J＝0.9Hz,1H),4.48(dd,J＝13.8,6.9Hz,1H),2.32(s,3H),2.01(dd,J＝9.3,5.4Hz,2H),1.72(s,2H),1.56(d,J＝4.6Hz,4H).
生物学实施例
实施例52化合物对IDH2/R140Q抑制活性的测定
本实施例通过测定辅助因子NADPH的消减来测定化合物对IDH2/R140Q的抑制活性。将化合物与IDH2/R140Q和NADPH进行孵育，然后通过添加α-KG启动反应，线性条件下反应一定时间后添加硫辛酰胺脱氢酶和相应的底物刃天青进行检测。硫辛酰胺脱氢酶通过消减可供使用的辅助因子NADPH而终止IDH2/R140Q反应，它将NADPH氧化成NADP，并且将刃天青还原成高荧光的试卤灵，通过检测试卤灵的生成来量化特定反应时间之后剩余的辅助因子NADPH的量。
具体操作方式为：在96孔板中，反应总体积是50μL，1.2nM IDH2/R140Q、梯度稀释的抑制剂和5μM NADPH的混合液在包含Tris-HCl 50mM(pH 7.5)、150mM NaCl、10mM MgCl 2、BSA 0.05％、10％Glycerol和2.5mMβ-巯基乙醇的反应缓冲液中，在25℃预孵育16小时。加入α-KG至1mM,在25℃反应40分钟。后加入25μL上述反应缓冲液配置的终止混合液(硫辛酰胺脱氢酶36μg/mL，刃天青30μM)，使刃天青转化为试卤灵来测量剩余的NADPH。25℃孵育10分钟后通过Tecan INFINITE F FLEX在Ex544/Em590下进行荧光值测定。每个化合物分别在10个浓度下测定酶的活性，反应中设置多个不加酶的背景孔和不含化合物的全酶活性孔。体系中二甲亚砜浓度小于等于百分之二。IC 50的值使用XLFit5软件(IDBS Software)通过公式：Y＝100/(1+10^((LogIC 50-X)*HillSlope))获得。本实施例的多孔板购自Thermo Fisher Scientific公司，NADPH、α-KG、硫辛酰胺脱氢酶和刃天青购自生工生物工程有限公司，IDH2/R140Q购自Abcam公司(ab198153)。
根据本实施例所述的生物学方法对本发明所选的化合物进行分析，其结果如表2所示。其中，表2的“A”指对IDH2/R140Q的IC 50≤100nM的抑制活性；“B”指对IDH2/R140Q的100nM<IC 50≤1μM的抑制活性；“C”指对IDH2/R140Q的1μM<IC 50≤10μM的抑制活性；“D”指对IDH2/R140Q的IC 50>10μM的抑制活性。
表2优选化合物的IDH2/R140Q抑制活性
Figure PCTCN2020082016-appb-000070
Figure PCTCN2020082016-appb-000071
Figure PCTCN2020082016-appb-000072
Figure PCTCN2020082016-appb-000073
结果显示，本专利所列化合物具有很好的IDH2/R140Q抑制活性，其中，活性水平为A和B的化合物的抑制作用极为明显。
实施例53化合物对野生型IDH2(IDH2/WT)选择性的测定
将化合物与IDH2/WT和NADP进行孵育，然后通过添加异柠檬酸启动反应，线性条件下反应一段时间后加入硫辛酰胺脱氢酶和刃天青检测荧光物质的量。本实验将NADP还原成NADPH，后者在硫辛酰胺脱氢酶的作用下将刃天青还原成高荧光的试卤灵，通过检测试卤灵的生成来量化特定反应时间之后生成的辅助因子NADPH的量，从而来计算化合物对IDH2/WT的抑制作用。
具体操作方式为：在96孔板中，反应总体积是50μL，0.6nM IDH2/WT、梯度稀释的抑制剂和50μM NADP的混合液在包含Tris-HCl 20mM(pH 7.5)、150mM NaCl、10mM MgCl 2、10％Glycerol、0.03％BSA和2.5mMβ-巯基乙醇的反应缓冲液中，在25℃预孵育16小时。加入isocitrate至50μM,25℃反应30分钟。后加入25μL上述反应缓冲液配置的混合液(硫辛酰胺脱氢酶36μg/mL，刃天青30μM)，使刃天青转化为试卤灵来测量生成的NADPH。25℃孵育10分钟后通过Tecan INFINITE F FLEX在Ex544/Em590下进行荧光值测定。每个化合物分别在10个浓度下测定酶的活性，反应中设置多个不加酶的背景孔和不含化合物的全酶活性孔。体系中二甲亚砜浓度小于等于百分之二。IC 50的值使用XLFit5软件(IDBS Software)通过公式：Y＝100/(1+10^((LogIC 50-X)*HillSlope))获得。方法中多孔板购自Thermo Fisher Scientific公司，NADP、α-KG、硫辛酰胺脱氢酶和刃天青购自生工生物工程有限公司，IDH2/WT通过大肠杆菌过表达后纯化得到。部分化合物的测试结果见表3。
表3部分化合物对IDH2/WT的抑制活性
Figure PCTCN2020082016-appb-000074
结果显示，本发明的化合物对野生型的IDH2(IDH2/WT)几乎没有活性，具有很好的选择性。
实施例54细胞水平对突变型IDH2活性的抑制检测
2-羟基戊二酸脱氢酶(2HGDH)在2-HG存在的情况下，能够将NAD+还原成NADH。后者可以通过心肌黄酶及其底物Resazurin(刃天青)进行定量测定。
过表达IDH2/R140Q突变的胶质瘤细胞U87MG，培养在1％丙酮酸钠的高糖MEM，10％FBS中，置于CO 2培养箱(37℃，5％CO 2，95％空气)培养。
细胞通过胰酶消化并以1×10 4的密度接种于96孔板中，培养基为200μL，37℃培养箱培养过夜。第二天加入待测化合物，DMSO终浓度均为0.1％，在培养24小时后，吸取100μL培养基，使用10KD
Figure PCTCN2020082016-appb-000075
超滤管(购自PALL公司) 14000g离心10分钟，过滤培养基中存在的可能干扰结果的蛋白等成分，使用后续方法检测2-HG的含量。剩余100μL培养基的96孔板中，加入50uLCellTiter-Glo(购自Promega公司)，测定细胞存活情况；
细胞外2-HG测定体系：
(1)50μL反应体系：反应缓冲液(50mM Tris pH7.5，100mM NaCl，20mM MgCl 2，0.05％BSA)，其中NAD+终浓度为40μM，2HGDH终浓度为20nM，待测样品加入5μL培养液；反应液混匀离心，避光25℃反应1小时；
(2)25μL显色体系：显色缓冲液(50mM Tris pH7.5，100mM NaCl，20mM MgCl 2，0.05％BSA)，其中心肌黄酶的终浓度为36μg/mL，刃天青钠的终浓度为3μM；将上述25μL显色液加入(1)中的50μL反应体系中，混匀并离心，立即在Ex544/Em590下进行荧光值测定。
2-HG标准曲线制备：将2-HG储液使用反应缓冲液稀释至20μM，之后进行2倍梯度稀释，共计6个点。之后将上述2-HG按照细胞外2-HG测定体系测定，计算并绘制标准曲线。
细胞外2-HG含量计算：
细胞外2-HG测定体系中获得的荧光值使用2-HG标准曲线计算培养基中2-HG的含量，以DMSO作为阴性对照，计算化合物对IDH2/R140Q突变产生2-HG活性的抑制。
根据本实施例所述的方法对本发明所选的化合物进行分析，其结果如表3所示。其中，表4的“A”指对在细胞水平IDH2/R140Q的IC 50≤100nM的抑制活性；“B”指在细胞水平对IDH2/R140Q的100nM<IC 50≤1μM的抑制活性；“C”指在细胞水平对IDH2/R140Q的1μM<IC 50≤10μM的抑制活性；“D”指在细胞水平对IDH2/R140Q的IC 50>10μM的抑制活性。
表4优选化合物在细胞水平对突变型IDH2/R140Q活性的抑制
Figure PCTCN2020082016-appb-000076
Figure PCTCN2020082016-appb-000077
Figure PCTCN2020082016-appb-000078
Figure PCTCN2020082016-appb-000079
结果显示，所测试的化合物能在较低的浓度下抑制IDH2/R140Q突变的细胞产生2-HG，显示了化合物在细胞水平对突变型IDH2的活性抑制作用。
在本发明提及的所有文献都在本申请中引用作为参考，就如同每一篇文献被单独引用作为参考那样。此外应理解，在阅读了本发明的上述讲授内容之后，本领域技术人员可以对本发明作各种改动或修改，这些等价形式同样落于本申请所附权利要求书所限定的范围。

Claims (14)

  1. 一种式I所示的嘧啶并五元杂环类化合物、或其立体异构体或互变异构体、或药学上可接受的盐、水合物或溶剂化物，
    Figure PCTCN2020082016-appb-100001
    其中，
    R 1选自无、氢、卤素、-CN、取代或未取代的C 1-C 8烷基、取代或未取代的C 2-C 8烯基、取代或未取代的C 2-C 8炔基、取代或未取代的C 3-C 10环烷基；
    R 2选自氢、卤素、-CN、取代或未取代的C 1-C 8烷基、取代或未取代的C 2-C 8烯基、取代或未取代的C 2-C 8炔基、取代或未取代的C 3-C 10环烷基、取代或未取代的C 1-C 8烷氧基、取代或未取代的C 1-C 8羧基、取代或未取代的C 2-C 20酯基、取代或未取代的C 6-C 10芳基或取代或未取代的具有1-3个选自N、S和O的杂原子的5-10元杂芳基；
    X选自N、O、S或CR 5；其中R 5为氢、卤素、-CN、取代或未取代的C 1-C 8烷基、取代或未取代的C 2-C 8烯基、取代或未取代的C 2-C 8炔基、或取代或未取代的C 3-C 10环烷基；
    m 1为0、1、2、3、或4；各个L独立地选自无、O、S、-CO-、-NH-或-CH 2-；
    m 2为0、1或2；各个Z
    独立地选自无、O、S、-CO-、-NH-或-CH 2-；
    R 3选自氢、卤素、-CN、取代或未取代的C 1-C 8烷基、取代或未取代的C 2-C 8烯基、取代或未取代的C 2-C 8炔基、取代或未取代的C 3-C 10环烷基、取代或未取代的C 6-C 10芳基、取代或未取代的具有1-3个选自N、S和O的杂原子的5-10元杂芳基、取代或未取代的具有1-3个选自N、S和O的杂原子的4-8元杂环基；
    R 4选自氢、卤素、CN、取代或未取代C 1-C 8烷基、取代或未取代的C 2-C 8烯基、取代或未取代的C 2-C 8炔基、取代或未取代的C 3-C 10环烷基、取代或未取代的C 6-C 10芳基、取代或未取代的具有1-3个选自N、S和O的杂原子的5-10元杂芳基；
    除非特别说明，所述的“取代”是指被选自下组的一个或多个(例如2个、3个、4个等)取代基所取代：卤素、C 1-C 6烷基、卤代的C 1-C 6烷基、C 1-C 6烷氧基、卤代的C 1-C 6烷氧基、C 3-C 8环烷基、卤代的C 3-C 8环烷基、氧代、-CN、羟基、氨基、羧基、苄基、C 6-C 10芳基、卤代的C 6-C 10芳基、具有1-3个选自N、S和O的杂原子的5-10元杂芳基、卤代的具有1-3个选自N、S和O的杂原子的5-10元杂芳基。
  2. 如权利要求1所述的化合物，其特征在于，所述化合物具有式Ia所示的结构：
    Figure PCTCN2020082016-appb-100002
    式中，R 1、R 2、R 3、R 4、L、和m 1的定义如权利要求1所述。
  3. 如权利要求1所述的化合物，其特征在于，L为NH，m 1为1，并且Z为无，m 2为0。
  4. 如权利要求1所述的化合物，其特征在于，R 2为甲基或三氟甲基。
  5. 如权利要求1所述的化合物，其特征在于，R 4为氟取代的苯基。6.如权利要求1所述的化合物，其特征在于，X为CR 5，其中R 5选自下组：H、C 1-C 4烷基、或C 3-C 4环烷基。
  6. 如权利要求1所述的化合物，其特征在于，所述的化合物为表1中的化合物#1、#2、#3、#4、#5、#6、#7、#8、#9、#10、#11、#12、#13、#14、#15、#16、#17、#18、#19、#20、#21、#22、#23、#24、#25、#26、#27、#28、#29、#30、#31、#32、#33、#34、#35、#36、#37、#38、#39、#40、#41、#42、#43、#44、#45、#46、#47、#48、#49、#50、或#51，或其药学上可接受的盐。
  7. 如权利要求1所述的化合物，其特征在于，所述化合物为表1中的化合物#28、#48、#49或#51，或其药学上可接受的盐。
  8. 一种药物组合物，包含：(1)如权利要求1所述的化合物、或其立体异构体或互变异构体、或药学上可接受的盐、水合物或溶剂化物；(2)药学上可接受的载体。
  9. 一种如权利要求1所述的化合物、或其立体异构体或互变异构体、或药学上可接受的盐、水合物或溶剂化物或如权利要求9所述的药物组合物的用途，其特征在于，用于制备预防和/或治疗突变型IDH2介导的疾病的药物。
  10. 如权利要求10所述的用途，其特征在于，所述突变型IDH2介导的疾病为癌症；较佳地，所述癌症选自膀胱癌、乳腺癌、肾癌、肝癌、肺癌(包括小细胞肺癌)、食道癌、胆囊癌、卵巢癌、胰腺癌、胃癌、宫颈癌、甲状腺癌、前列腺癌和皮肤癌(包括鳞状细胞癌)；淋巴系的造血肿瘤，例如包括白血病、急性淋巴细胞白血病、急性淋巴母细胞白血病、B细胞淋巴瘤、T-细胞淋巴瘤、霍奇金淋巴瘤、非霍奇金淋巴瘤、毛细胞淋巴瘤和伯基特淋巴瘤；间充质细胞来源的肿瘤，例如包括纤维肉瘤、横纹肌肉瘤；髓系的造血肿瘤，例如包括急慢性骨髓性白血病、骨髓增生异常综合征和前髓细胞白血病；中枢和周围神经系统肿瘤，例如包括星形细胞瘤、成神经细胞瘤、神经胶质瘤和神经鞘瘤；和其它肿瘤，例如包括黑素瘤、精原细胞瘤、畸胎癌、骨肉瘤、色性干皮病、角化棘皮瘤、甲状腺滤泡癌和卡波济氏肉瘤。
  11. 一种式I化合物的制备方法，所述方法包括步骤：
    Figure PCTCN2020082016-appb-100003
    (a)将式(1)化合物与H-(L)m 1-R 3反应，从而制得式(2)化合物，其中H-(L)m 1-R 3为R 3取代的胺类化合物或者硼酸类化合物或者硼酸酯类化合物；和
    Figure PCTCN2020082016-appb-100004
    (b)将式(2)化合物与H-(Z)m 2-R 4反应，从而制得式(I)化合物，其中H-(Z)m 2-R 4为R 4取代的胺类化合物或者硼酸类化合物或者硼酸酯类化合物或者有机锡化合物；
    式中，R 1、R 2、R 3、R 4、L、Z、m 1、m 2的定义如权利要求1中所述。
  12. 一种式Ia所示的化合物的制备方法，其特征在于，所述的方法包括步骤：
    Figure PCTCN2020082016-appb-100005
    (a1)将式(1a)的化合物与N-溴代丁二酰亚胺或者S-(三氟甲基)二苯并噻吩嗡四氟硼酸盐(Umemoto's reagents)反应，制得式(2a)化合物；和
    (b1)将式(2a)化合物与硼酸类化合物R 2-B(OH) 2反应，从而制得式(Ia)所示的化合；
    式中，R 1、R 2、R 3、R 4、L、m 1的定义如权利要求1所述。
  13. 一种体外抑制含突变型IDH2的肿瘤细胞增殖的方法，其特征在于，包括步骤：将如权利要求1-8所述的化合物、或其立体异构体或互变异构体、或药学上可接受的盐、水合物或溶剂化物或如权利要求9所述的药物组合物与突变型IDH2接触，从而抑制突变型IDH2的活性。
  14. 一种预防和/或治疗突变型IDH2介导的疾病的方法，其特征在于，包括步骤：向所需对象施用本如权利要求1-8所述的化合物、或其立体异构体或互变异构体、或药学上可接受的盐、水合物或溶剂化物或如权利要求9所述的药物组合物。
PCT/CN2020/082016 2019-04-22 2020-03-30 嘧啶并五元杂环类化合物及其作为突变型idh2抑制剂的用途 WO2020215998A1 (zh)

Priority Applications (4)

Application Number Priority Date Filing Date Title
CN202080029927.1A CN114127063B (zh) 2019-04-22 2020-03-30 嘧啶并五元杂环类化合物及其作为突变型idh2抑制剂的用途
US17/605,205 US20220204509A1 (en) 2019-04-22 2020-03-30 Pyrimido five-membered heterocyclic compound and use thereof as mutant idh2 inhibitor
JP2021562110A JP2022530371A (ja) 2019-04-22 2020-03-30 ピリミド５員複素環化合物及び変異型ｉｄｈ２阻害薬としてのその使用
EP20794845.6A EP3967691A4 (en) 2019-04-22 2020-03-30 FIVE-MEMBERED HETEROCYCLIC COMPOUND BASED ON PYRIMIDO AND ITS USE AS A MUTANT IDH2 INHIBITOR

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
CN201910324679.2 2019-04-22
CN201910324679.2A CN111825674A (zh) 2019-04-22 2019-04-22 嘧啶并五元杂环类化合物及其作为突变型idh2抑制剂的用途

Publications (1)

Publication Number Publication Date
WO2020215998A1 true WO2020215998A1 (zh) 2020-10-29

Family

ID=72912323

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/CN2020/082016 WO2020215998A1 (zh) 2019-04-22 2020-03-30 嘧啶并五元杂环类化合物及其作为突变型idh2抑制剂的用途

Country Status (5)

Country Link
US (1) US20220204509A1 (zh)
EP (1) EP3967691A4 (zh)
JP (1) JP2022530371A (zh)
CN (2) CN111825674A (zh)
WO (1) WO2020215998A1 (zh)

Families Citing this family (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107556366B (zh) * 2016-06-30 2025-02-07 上海海和药物研究开发股份有限公司 具有突变型异柠檬酸脱氢酶抑制活性的化合物、其制备方法及用途
WO2024035771A2 (en) * 2022-08-09 2024-02-15 H. Lee Moffitt Cancer Center And Research Institute, Inc. Ulk3 inhibitors and uses thereof
CN116478172B (zh) * 2023-06-20 2023-09-05 英矽智能科技(上海)有限公司 吡咯并[3,2-d]嘧啶类化合物及其应用
WO2025096505A1 (en) * 2023-10-31 2025-05-08 Bristol-Myers Squibb Company Ubiquitin specific processing protease 1 (usp1) compounds

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2017035118A1 (en) * 2015-08-25 2017-03-02 Bristol-Myers Squibb Company Tgf beta receptor antagonists
WO2017069960A1 (en) * 2015-10-19 2017-04-27 The Scripps Research Institute Inhibitors of sulfur metabolism with potent bactericidal activity against mdr and xdr m. tuberculosis
WO2019024908A1 (zh) * 2017-08-04 2019-02-07 厦门大学 取代五元并六元杂环类化合物、其制备方法、药物组合及其用途

Family Cites Families (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1996040142A1 (en) * 1995-06-07 1996-12-19 Pfizer Inc. Heterocyclic ring-fused pyrimidine derivatives
US6897307B2 (en) * 2002-03-28 2005-05-24 Novartis Ag Process for preparing 2,6-diaminopurine derivatives
WO2004021979A2 (en) * 2002-09-06 2004-03-18 Smithkline Beecham Corporation PYRROLO[2, 3-d]PYRIMIDINE-4-YL AND PURIN-6-YL UREA COMPOUNDS
JP5046942B2 (ja) * 2004-09-30 2012-10-10 テイボテク・フアーマシユーチカルズ Ｈｃｖ阻害性の二環式ピリミジン類
EP2900670A1 (en) * 2012-09-26 2015-08-05 Bayer Pharma Aktiengesellschaft Substituted indazol-pyrrolopyrimidines useful in the treatment of hyperfoliferative disorders
CA2890934A1 (en) * 2012-11-15 2014-05-22 Pharmacyclics, Inc. Pyrrolopyrimidine compounds as kinase inhibitors
US11021468B2 (en) * 2015-04-01 2021-06-01 Rigel Pharmaceuticals, Inc. TGF-ß inhibitors
KR20180034548A (ko) * 2015-07-28 2018-04-04 브리스톨-마이어스 스큅 컴퍼니 Tgf 베타 수용체 길항제
WO2017140758A1 (en) * 2016-02-19 2017-08-24 Debiopharm International S.A. Derivatives of 2-amino-4-(2-oxazolidinon-3-yl)-pyrimidine fused with a five-membered heteroaromatic ring in 5,6-position which are useful for the treatment of various cancers

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2017035118A1 (en) * 2015-08-25 2017-03-02 Bristol-Myers Squibb Company Tgf beta receptor antagonists
WO2017069960A1 (en) * 2015-10-19 2017-04-27 The Scripps Research Institute Inhibitors of sulfur metabolism with potent bactericidal activity against mdr and xdr m. tuberculosis
WO2019024908A1 (zh) * 2017-08-04 2019-02-07 厦门大学 取代五元并六元杂环类化合物、其制备方法、药物组合及其用途

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
CAS, no. 1446502-11-9
DATABASE CAS 8 July 2015 (2015-07-08), retrieved from STN Database accession no. 1796852-65-7 *
SAMBROOK ET AL.: "Molecular Cloning: Laboratory Manual", 1989, COLD SPRING HARBOR LABORATORY PRESS
See also references of EP3967691A4

Also Published As

Publication number Publication date
JP2022530371A (ja) 2022-06-29
CN111825674A (zh) 2020-10-27
US20220204509A1 (en) 2022-06-30
EP3967691A1 (en) 2022-03-16
CN114127063B (zh) 2023-12-01
EP3967691A4 (en) 2022-12-28
CN114127063A (zh) 2022-03-01

Similar Documents

Publication Publication Date Title
CN113423703B (zh) 含氮杂环类衍生物调节剂、其制备方法和应用
WO2021088945A1 (zh) 作为shp2抑制剂的化合物及其应用
JP2023511337A (ja) ピリミジン－４（３ｈ）－オンヘテロ環式化合物、その調製方法、およびその医薬的使用
CN114787161B (zh) 吡唑并[1,5-a]吡啶类化合物及其制备方法和应用
WO2020215998A1 (zh) 嘧啶并五元杂环类化合物及其作为突变型idh2抑制剂的用途
WO2015127872A1 (zh) 2,4-二取代苯-1,5-二胺衍生物及其应用以及由其制备的药物组合物和药用组合物
WO2016026445A1 (zh) 作为fgfr激酶抑制剂的吲唑类化合物及其制备和应用
CN107072985A (zh) 治疗性抑制化合物
WO2021139775A1 (zh) 吡啶酮化合物及应用
WO2018010514A1 (zh) 作为fgfr抑制剂的杂环化合物
WO2016011979A1 (zh) 2，4-二取代7H-吡咯并[2，3-d]嘧啶衍生物、其制法与医药上的用途
WO2019034128A1 (zh) 吡咯并三嗪类衍生物、其制备方法及其用途
CN108727340B (zh) 氟取代的吲唑类化合物及其用途
EP3694330B1 (en) Indazolyl-spiro[2.2]pentane-carbonitrile derivatives as lrrk2 inhibitors, pharmaceutical compositions, and uses thereof
CN105884695B (zh) 杂环衍生物类酪氨酸激酶抑制剂
CN112457326B (zh) 一类芳香杂环并内酰胺类化合物、制备方法和用途
CN114401955A (zh) 细胞周期蛋白依赖性激酶的抑制剂
WO2024032661A1 (zh) Kif18a抑制剂及其用途
CN108690016B (zh) 吡唑并吡啶类化合物及其用途
TWI869674B (zh) 雜芳基衍生化合物及其用途
WO2023036252A1 (zh) 吡咯并嘧啶类或吡咯并吡啶类衍生物及其医药用途
TWI793877B (zh) 作為甘油二酯激酶抑制劑的雜環化合物及其用途
CN107428682B (zh) 酰胺类衍生物、其制备方法及其在医药上的用途
WO2022037691A1 (zh) 一类芳香环并内酰胺类化合物、其制备方法和用途
CN116478130A (zh) N-羟基喹啉甲酰胺化合物及其用途

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 20794845

Country of ref document: EP

Kind code of ref document: A1

ENP Entry into the national phase

Ref document number: 2021562110

Country of ref document: JP

Kind code of ref document: A

NENP Non-entry into the national phase

Ref country code: DE

ENP Entry into the national phase

Ref document number: 2020794845

Country of ref document: EP

Effective date: 20211122