WO2015184731A1 - 与Aβ斑块具有亲和力的含手性侧链取代的氟代2-芳基苯并杂环化合物、其制备方法及应用 - Google Patents

与Aβ斑块具有亲和力的含手性侧链取代的氟代2-芳基苯并杂环化合物、其制备方法及应用 Download PDF

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WO2015184731A1
WO2015184731A1 PCT/CN2014/090536 CN2014090536W WO2015184731A1 WO 2015184731 A1 WO2015184731 A1 WO 2015184731A1 CN 2014090536 W CN2014090536 W CN 2014090536W WO 2015184731 A1 WO2015184731 A1 WO 2015184731A1
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formula
compound
reaction
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solvent
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张志勇
崔孟超
林春平
刘伯里
国毓智
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北京智博高科生物技术有限公司
张志勇
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Priority to EP14894174.3A priority Critical patent/EP3153507A4/en
Priority to US14/890,922 priority patent/US9809585B2/en
Priority to JP2017516015A priority patent/JP6634075B2/ja
Publication of WO2015184731A1 publication Critical patent/WO2015184731A1/zh

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    • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
    • C07D417/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings
    • C07D417/04—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K49/00—Preparations for testing in vivo
    • A61K49/06—Nuclear magnetic resonance [NMR] contrast preparations; Magnetic resonance imaging [MRI] contrast preparations
    • A61K49/08—Nuclear magnetic resonance [NMR] contrast preparations; Magnetic resonance imaging [MRI] contrast preparations characterised by the carrier
    • A61K49/10—Organic compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
    • A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
    • A61K51/04—Organic compounds
    • A61K51/041—Heterocyclic compounds
    • A61K51/044—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine, rifamycins
    • A61K51/0453—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine, rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
    • A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
    • A61K51/04—Organic compounds
    • A61K51/041—Heterocyclic compounds
    • A61K51/044—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine, rifamycins
    • A61K51/0455—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine, rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D263/00—Heterocyclic compounds containing 1,3-oxazole or hydrogenated 1,3-oxazole rings
    • C07D263/52—Heterocyclic compounds containing 1,3-oxazole or hydrogenated 1,3-oxazole rings condensed with carbocyclic rings or ring systems
    • C07D263/54—Benzoxazoles; Hydrogenated benzoxazoles
    • C07D263/56—Benzoxazoles; Hydrogenated benzoxazoles with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached in position 2
    • C07D263/57—Aryl or substituted aryl radicals
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D277/00—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings
    • C07D277/60—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings condensed with carbocyclic rings or ring systems
    • C07D277/62—Benzothiazoles
    • C07D277/64—Benzothiazoles with only hydrocarbon or substituted hydrocarbon radicals attached in position 2
    • C07D277/66—Benzothiazoles with only hydrocarbon or substituted hydrocarbon radicals attached in position 2 with aromatic rings or ring systems directly attached in position 2
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D413/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms
    • C07D413/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings
    • C07D413/04—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2123/00—Preparations for testing in vivo

Definitions

  • the present invention relates to a chiral side chain-substituted fluoro-2-arylbenzoheterocyclic compound having affinity with A ⁇ plaque, a preparation method and application thereof.
  • AD Alzheimer's Disease
  • AD is a progressive developmental fatal neurodegenerative disease with clinical manifestations of decreased cognitive and memory functions, decreased daily living ability, and a variety of neuropsychiatric symptoms and behaviors. obstacle. AD has become a major disease that seriously threatens the physical and mental health of the elderly after cancer, heart disease and stroke.
  • SPs senile plaques
  • NFTs neurofibrillary tangles
  • AD positron emission tomography
  • PIB is the most widely used A ⁇ imaging agent, which can clearly distinguish between normal and AD patients.
  • a ⁇ imaging agents labeled with long half-life nuclide F-18 are currently in development, such as the analog of PIB [ 18 F]GE-067 (Koole M et al. Journal of Nuclear Medicine., 2009, 50: 818-22) and stilbene-derived [ 18 F]AV-45 (Wong D F et al. Journal of Nuclear Medicine., 2010, 51: 913-20) are currently approved by the FDA and entered the commercial development stage.
  • a chiral side chain-substituted fluoro-2-arylbenzoheterocyclic compound having affinity with A ⁇ plaque of the present invention has a structural formula of the formula (I). Show:
  • R 1 is a 5- or 6-member substituent, and R 1 is F is 19 F or 18 F;
  • R 2 is NHCH 3 or N(CH 3 ) 2 .
  • the invention also provides a preparation method of the compound represented by the formula (I).
  • the preparation method of the compound comprises the following steps:
  • the preparation method of the compound comprises the following steps:
  • R is N(CH 3 ) 2 ;
  • the present invention also provides the use of the compound for the preparation of an A ⁇ plaque imaging agent, wherein in the compound of the formula (I), F is 18 F.
  • the present invention provides for the first time a new class of 2-arylbenzoheterocyclic compounds having a chiral side chain substitution.
  • the in vitro competition binding experiments show that the molecules have higher affinity with A ⁇ 1-42 aggregates; Biodistribution experiments showed that some F-18-labeled imaging agents have the advantages of high initial brain uptake and rapid clearance. It is expected to become a new positron A ⁇ plaque imaging agent for clinical imaging.
  • Figure 1 is a schematic view showing the synthesis of a chiral side chain-substituted 2-arylbenzoheterocyclic compound of the present invention; wherein a represents K 2 CO 3 , 18-crown-6 and acetone, reflux; b represents TBAF and toluene. , reflux.
  • Figure 2 is a schematic diagram of the preparation of the optically pure chiral side chain substituted 2-arylbenzoheterocyclic compound F-18 labeled intermediate of the present invention; wherein a represents NaOH, EtOH and H 2 O, 90 ° C; b represents TBDMSCl, CH 2 Cl 2 and imidazole; c represents (Boc) 2 O and THF, reflux; d represents TBAF and THF, reflux; e represents TsCl and pyridine; f represents PPTS, CH 2 Cl 2 and 3,4- Dihydro-2H-pyran, reflux.
  • a represents NaOH, EtOH and H 2 O, 90 ° C
  • b represents TBDMSCl, CH 2 Cl 2 and imidazole
  • c represents (Boc) 2 O and THF, reflux
  • d represents TBAF and THF, reflux
  • e represents TsCl and pyridine
  • f represents PPTS, CH 2 Cl 2 and 3,4- Dihydro-2H-
  • Figure 3 is a schematic diagram of the preparation of the F-18-labeled compound of the present invention; wherein a represents 18 F - , K 2 CO 3 , Kryptofix-2.2.2 and acetonitrile, the reaction temperature is 100 ° C; b represents HCl (1 M), the reaction temperature 100 ° C.
  • FIG. 4 of the present invention the compound prepared in Example 84 in the embodiment [18 F] 29s AD and AD patients and gene transfected mouse brain sections autoradiographic image.
  • FIG. 5 of the present invention the compound of Preparation 83 in embodiment [18 F] 29r AD and AD patients and gene transfected mouse brain sections autoradiographic image.
  • A is a brain slice of an AD patient; B is a brain slice of an AD transgenic mouse; C is a normal human brain slice; and D is a normal mouse brain slice.
  • the intermediate 8s was obtained by the reaction of the compound 1, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 23.0%.
  • the intermediate 9r was obtained by the reaction of the compound 2, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 16.4%.
  • the intermediate 10r was obtained by the reaction of the compound 3, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 38.3%.
  • the intermediate 10s was obtained by the reaction of the compound 3, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as in Example 1, and the yield was 31.6%.
  • the intermediate 11r was obtained by the reaction of the compound 4, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 58.8%.
  • the intermediate 11s was obtained by the reaction of the compound 4, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 70.7%.
  • the intermediate 12r was obtained by the reaction of the compound 5, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 55.3%.
  • the intermediate 12s was obtained by the reaction of the compound 5, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as in Example 1, and the yield was 51.9%.
  • the intermediate 13r was obtained by the reaction of the compound 6, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 43.5%.
  • the target compound 14r was obtained by the reaction of the compound 8s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 7.4%.
  • the target compound 15s was obtained by the reaction of the intermediate 9r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 5.8%.
  • the target compound 15r was obtained by the reaction of the intermediate 9s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 14.8%.
  • the target compound 16s was obtained by the reaction of the intermediate 10r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 21.0%.
  • the target compound 16r was obtained by the reaction of the intermediate 10s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 14.1%.
  • the target compound 17s was obtained by the reaction of the intermediate 11r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 38.6%.
  • the target compound 17r was obtained by the reaction of the intermediate 11s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 35.7%.
  • the target compound 18s was obtained by the reaction of the intermediate 12r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 62.4%.
  • the target compound 18r was obtained by the reaction of the intermediate 12s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 51.4%.
  • the target compound 19s was obtained by the reaction of the intermediate 13r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 61.1%.
  • the target compound 19r was obtained by the reaction of the intermediate 13s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 63.3%.
  • the intermediate 24r was obtained by the reaction of the compound 20, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as in Example 1, and the yield was 31.7%.
  • the intermediate 24s was obtained by the reaction of the compound 20, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as in Example 1, and the yield was 16.2%.
  • the intermediate 25r was obtained by the reaction of the compound 21, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 52.9%.
  • the intermediate 25s was obtained by the reaction of the compound 21, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 72.9%.
  • the intermediate 26r was obtained by the reaction of the compound 22, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 34.6%.
  • the intermediate 26s was obtained by the reaction of the compound 22, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 33.3%.
  • the intermediate 27r was obtained by the reaction of the compound 23, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 71.7%.
  • the intermediate 27s was obtained by the reaction of the compound 23, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 1, and the yield was 61.8%.
  • the target compound 28s was obtained by the reaction of the intermediate 24r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 16.6%.
  • the target compound 28r was obtained by the reaction of Intermediate 24s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 31.3%.
  • HRMS (EI): m/z calcd for C 17 H 18 N 2 O 3 F317.1301; found 317.1295 (M+H) + .
  • the target compound 29s was obtained by the reaction of Intermediate 25r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 53.0%.
  • the target compound 29r was obtained by the reaction of Intermediate 25s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 28.0%.
  • the target compound 30s was obtained by the reaction of the intermediate 26r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 22.6%.
  • the target compound 30r was obtained by the reaction of Intermediate 26s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 28.2%.
  • the target compound 31s was obtained by the reaction of the intermediate 27r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 19.5%.
  • the target compound 31r was obtained by the reaction of Intermediate 27s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 13, and the yield was 29.2%.
  • the compound 33s (129.0 mg, 0.4 mmol) was dissolved in 30 mL of CH 2 Cl 2 solution, and imidazole (237.9 mg, 3.5 mmol) and TBDMSCl (536.2 mg, 3.6 mmol) were added thereto, and the above solution was subjected to an oil bath at 40 ° C. The reaction was stirred and refluxed. After the reaction was completed, the precipitate was removed by suction, and CH 2 Cl 2 was removed. The residue was purified by column chromatography to afford 154.2 mg of Intermediate 34.
  • the intermediate 34s was obtained by the reaction of the intermediate 33r, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 43, and the yield was 24.2%.
  • the intermediate 35s was obtained by the reaction of the intermediate 34s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 45, and the yield was 88.0%.
  • the intermediate 36r was obtained by the reaction of the intermediate 35s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 47, and the yield was 60.1%.
  • the intermediate 37s was obtained by the reaction of the intermediate 36r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in the example 49, and the yield was 62.3%.
  • the labeled precursor 38s was obtained by the reaction of Intermediate 37s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 51, and the yield was 79.4%.
  • the intermediate 39s was obtained by reacting the compound 1 and the compound 32s, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in the example 41, and the yield was 80.9%.
  • the intermediate 39r was obtained by the reaction of the compound 1 and the compound 32r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in the example 41, and the yield was 49.0%.
  • the intermediate 40r was obtained by the reaction of the intermediate 39s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in the example 43, and the yield was 63.8%.
  • the intermediate 40s was obtained by the reaction of the intermediate 39r, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 43, and the yield was 90.4%.
  • the labeled intermediate 41r was obtained by the reaction of the intermediate 40r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 45, and the yield was 86.0%.
  • the intermediate 41s was obtained by the reaction of the intermediate 40s, and the ratio of the raw materials, the solvent, the reaction conditions and the like were the same as those in Example 45, and the yield was 92.9%.
  • the intermediate 42s was obtained by the reaction of the intermediate 41r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 47, and the yield was 83.4%.
  • the labeled intermediate 42r was obtained by the reaction of the intermediate 41s.
  • the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 47, and the yield was 59.2%.
  • the intermediate 43r was obtained by the reaction of the intermediate 42s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 49, and the yield was 14.0%.
  • the intermediate 43s was obtained by the reaction of the intermediate 42r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 49, and the yield was 14.4%.
  • the labeled precursor 44r was obtained by the reaction of the intermediate 43r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 51, and the yield was 66.8%.
  • the labeled precursor 44s was obtained by the reaction of the intermediate 43s, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 51, and the yield was 72.1%.
  • the intermediate 45r was obtained by the reaction of the compound 2, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in the example 41, and the yield was 54.1%.
  • the intermediate 45s was obtained by the reaction of the compound 2, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in the example 41, and the yield was 89.2%.
  • the labeled precursor 47s was obtained by the reaction of the intermediate 46s, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 51, and the yield was 35.5%.
  • the labeled precursor 47r was obtained by the reaction of the intermediate 46r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as in Example 57, and the yield was 94.0%.
  • the intermediate 48r was obtained by the reaction of the compound 21, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in the example 41, and the yield was 56.0%.
  • the intermediate 48s was obtained by the reaction of the compound 21, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as in the example 41, and the yield was 52.2%.
  • the intermediate 49s was obtained by the reaction of the compound 48r, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 49, and the yield was 7.8%.
  • the intermediate 49r was obtained by the reaction of the compound 48s, and the ratio of the starting materials, the solvent, the reaction conditions and the like were the same as those in Example 49, and the yield was 25.8%.
  • the labeled precursor 50s was obtained by the reaction of the compound 49s, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 51, and the yield was 83.8%.
  • the labeled precursor 50r was obtained by the reaction of the compound 49r, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as those in Example 51, and the yield was 91.2%.
  • the label [ 18 F] 14b having a purity greater than 98% was isolated by HPLC.
  • the labeling rate is approximately 5-40%.
  • the retention time is 9.09 min.
  • [ 18 F]14s was obtained by the reaction of the labeled precursor 44s, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as in Example 77, and the labeling ratio was about 5 to 40%.
  • Example 47r labeling precursor from the reaction of [18 F] 15r the same starting material ratio of the reaction, the solvent, etc.
  • the reaction conditions of Example 77 labeled rate of about 5-40%.
  • the retention time is 15.23 min.
  • [ 18 F]15s was obtained by the reaction of the labeled precursor 47s, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as in Example 77, and the labeling ratio was about 5 to 40%.
  • Example 38r 77 same as Example 38r prepared by the reaction of the labeling precursor to obtain [18 F] 28r, the reaction ratio of the starting material, solvent, reaction conditions, etc., the mark rate is about 5-40%.
  • [ 18 F]28s was obtained by the reaction of the labeled precursor 38s, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as in Example 77, and the labeling ratio was about 5 to 40%.
  • Example 50r 77 same as Example 50r prepared by the reaction of the labeling precursor to obtain [18 F] 29r, the reaction ratio of the starting material, solvent, reaction conditions, etc., the mark rate is about 5-40%.
  • [ 18 F]29s was obtained by the reaction of the labeled precursor 50s, and the ratio of the raw materials, the solvent, the reaction conditions and the like of the reaction were the same as in Example 77, and the labeling ratio was about 5-40%.
  • the compounds of the present invention were evaluated for affinity by competitive binding experiments and autoradiography experiments, and their high affinity to A? plaques was sufficiently verified.
  • the initial brain uptake and brain clearance evaluation of the labeled compounds were performed by biodistribution experiments in normal mice.
  • K i determination a certain concentration of A ⁇ 1-42 aggregate protein binds to a certain concentration of radioligand [ 125 I]IMPY, and different concentrations of test compounds are added to the reaction system (respectively The target compound in the inventive example competes with [ 125 I]IMPY, and after separation, the complex is separated to calculate the inhibition constant (K i ) by measuring the radioactivity.
  • Radioligand [ 125 I] IMPY was prepared according to the prior art. [ 125I ]IMPY is formulated into an aqueous solution of 100000 cpm/100 ⁇ L;
  • the receptor A ⁇ 1-42 protein was prepared according to a conventional method. Dilute it into an aqueous solution of about 30 nM;
  • the glass fiber filter membrane is immersed in a PBS solution containing 0.1% (by volume fraction) of polyethyleneimine for 0.5 h;
  • the multi-head cell harvester collects the reaction solution and rinses three times with PBS for 3 mL each time;
  • mice The pharmacokinetic properties in mice, especially initial brain uptake and brain clearance, were studied by in vivo distribution experiments.
  • the experimental results are shown in Table 2.
  • the 18 F-labeled compounds of the present invention can smoothly pass through the blood-brain barrier, and the brain volume is higher in 2 minutes, especially the initial brain uptake of [ 18 F] 29s is high and The brain was cleared quickly in normal mice, and the brain uptake ratio was 27.8 at 2 minutes and 60 minutes.

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Abstract

本发明提供一种与Aβ斑块具有亲和力的含手性侧链取代的氟代2-芳基苯并杂环化合物，所述化合物的结构通式如式(I)所示：其中，X为N；Y为S或O；Z为N或CH； R1为5位或6位取代基，R1为式(a)或式(b)， F为19F或18F； R2为NHCH3或N(CH3)2。本发明所述化合物与Aβ斑块具有很高的亲和力，可以制备成相应放射性核素标记的探针，用于阿尔茨海默症的早期诊断。

Description

与Aβ斑块具有亲和力的含手性侧链取代的氟代2-芳基苯并杂环化合物、其制备方法及应用 技术领域
本发明涉及一种与Aβ斑块具有亲和力的含手性侧链取代的氟代2-芳基苯并杂环化合物、其制备方法及应用。
背景技术
阿尔茨海默症(Alzheimer's Disease，AD)是一种进行性发展的致死性神经退行性疾病，临床表现为认知和记忆功能下降，日常生活能力减退，并伴有各种神经精神症状和行为障碍。AD已成为继肿瘤、心脏病和中风之后，严重威胁老年人身心健康的主要疾病。统计资料显示，我国AD的患病率在65岁以上人群中平均为6.6％，发病率随年龄的增长而增加，75-80岁为11％，80岁以上高达22％。目前，我国正在快速步入老龄化社会，预计到2050年，我国60岁及以上老人占比将超过30％。因此，AD的防治工作任务艰巨，研究AD的早期诊断方法具有极其重要的意义。
AD脑内沉积在神经细胞外的老年斑(SPs)和神经细胞内的神经纤维缠结(NFTs)是AD的两大主要病理特征，但其确切的致病机理目前尚不清楚(Hardy J et al.Science.,2002,297:353-356)。研究表明，脑内的Aβ斑块沉积在AD发病前10-20年就已经开始(Braak,H et al.Acta Neuropathol.,1991,82:239-259)。目前，临床上还很难准确诊断AD，主要通过评价病人的认知功能损伤来诊断。AD的确诊只能通过病人死后尸检脑内的SPs和NFTs来判断。因此，尽管目前AD病因还不明确，但利用脑内的Aβ斑块为靶点，开发与之具有高亲和性和选择性的分子探针，通过正电子发射断层(PET)扫描显像，可以灵敏、无创伤的从分子水平上早期诊断AD(Cai L S et al.Curr Med Chem.,2007,14:19-52)。
在过去10年里，已经有不少正电子Aβ分子探针进入了临床试验阶段，如C-11标记的PIB是目前使用最为广泛的Aβ显像剂，它可以明显的区分正常人和AD病人(Klunk W E et al.Annals of Neurology.,2004,55:306-319)。然而，C-11核素短半衰期(20.4min)的性质限制了其在临床上的应用。因此，采用长半衰期核素F-18标记的Aβ显像剂是目前发展的趋势，如PIB的类似物[18F]GE-067(Koole M et al.Journal of Nuclear Medicine.,2009,50:818-22)以及二苯乙烯衍生[18F]AV-45(Wong D F et al.Journal of Nuclear Medicine.,2010,51:913-20)目前均已被FDA批准，进入了商业开发阶段。
发明内容
本发明的目的是提供一种与AD病人脑部Aβ斑块具有高亲和力的含手性侧链取代的2-芳基苯并杂环化合物、其制备方法及应用。
为了实现本发明目的，本发明的一种与Aβ斑块具有亲和力的含手性侧链取代的氟代2-芳基苯并杂环化合物，所述化合物的结构通式如式(I)所示：
Figure PCTCN2014090536-appb-000001
其中，
X为N；Y为S或O；Z为N或CH；
R1为5位或6位取代基，R1为
Figure PCTCN2014090536-appb-000002
F为19F或18F；
R2为NHCH3或N(CH3)2。
本发明还提供式(I)所示化合物的制备方法，当R1中F为19F时，所述化合物的制备方法包括以下步骤：
(1)将1mmol
Figure PCTCN2014090536-appb-000003
与1.5mmol
Figure PCTCN2014090536-appb-000004
以及3mmol K2CO3和催化剂量的18-crown-6溶于30mL无水丙酮中，在80℃油浴条件下搅拌回流反应；反应完毕后，除去溶剂丙酮，经柱层析分离得到如式(II)或式(III)所示的化合物：
Figure PCTCN2014090536-appb-000005
(2)将0.2mmol式(II)或式(III)所示化合物溶于15mL甲苯中，向其中加入含1M TBAF的THF溶液1mL，在80℃油浴条件下搅拌回流反应；反应完毕后，除去溶剂，用CH2Cl2萃取后，最后经柱层析分离得到如式(IV)或式(V)所示的化合物：
Figure PCTCN2014090536-appb-000006
其中，X为N；Y为S或O；Z为N或CH；R为NHCH3或N(CH3)2。
前述的化合物，当R1中F为18F，且R2为N(CH3)2时，所述化合物的制备方法包括以下步骤：
(1)将1mmol
Figure PCTCN2014090536-appb-000007
溶于30mL无水乙醇中，向其中加入200g/L NaOH溶液0.3mL，在80℃油浴条件下搅拌回流反应1h后，向其中加入1.5mmol
Figure PCTCN2014090536-appb-000008
反应完毕后，除去溶剂，经柱层析分离得到如式(VI)或式(VII)所示的化合物：
Figure PCTCN2014090536-appb-000009
(2)当R为NHCH3时，将1mmol式(VI)或式(VII)所示化合物、8mmol咪唑以及8mmol叔丁基二甲基氯硅烷溶于30mL CH2Cl2溶液中，在40℃油浴条件下搅拌回流反应5h，反应完毕后除去CH2Cl2，经柱层析分离得到如式(VIII)或式(IX)所示的化合物：
Figure PCTCN2014090536-appb-000010
(3)将1mmol式(VIII)或式(IX)所示化合物溶于30mL THF溶液中，向其中加入过量的(Boc)2O,在80℃油浴条件下搅拌回流反应过夜，反应完毕后除去THF，经柱层析分离得到如式(X)或式(XI)所示的化合物：
Figure PCTCN2014090536-appb-000011
(4)将1mmol式(X)或式(XI)所示化合物溶于30mL THF溶液中，向其中加入7mmol TBAF，在30℃油浴条件下搅拌回流反应过夜，反应完毕后除去THF，经柱层析分离得到如式(XII)或式(XIII)所示的化合物：
Figure PCTCN2014090536-appb-000012
(5)将1mmol式(VI)或式(VII)所示化合物(R＝NMe2)以及1mmol式(XII)或式(XIII)所示化合物分别溶于5mL吡啶溶液中，向其中加入1.5mmol TsCl，在0℃冰浴条件下搅拌反应，反应完毕后除去吡啶，经柱层析分离得到如式(XIV)或式(XV)所示的化合物：
Figure PCTCN2014090536-appb-000013
其中，R为N(CH3)2；
(6)将1mmol式(XIV)或式(XV)所示化合物、4mmol3,4-dihydro-2H-pyran以及0.2mmol PPTS分别溶于30mL CH2Cl2溶液中，在40℃油浴条件下搅拌回流反应过夜，反应完毕后除去CH2Cl2，经柱层析分离得到如式(XVI)或式(XVII)所示的化合物：
Figure PCTCN2014090536-appb-000014
(7)将10mg式(XVI)或式(XVII)所示化合物加入2mL乙腈溶解。将其加入到已除水的具有一定活度的含K222/K2CO3的18F-的反应管中，于100℃标记12min，冷却后加入1M HCl0.15mL，涡旋后继续反应5min中，冷却后加入少量水用NaHCO3中和至偏碱性，最后过C18反相柱用水冲洗除去盐和剩余的18F-，然后用乙腈淋洗，N2吹干后，用HPLC分离得到如式(XVI)或式(XVII)所示的化合物：
Figure PCTCN2014090536-appb-000015
本发明还提供所述化合物在制备Aβ斑块显像剂中的应用，其中式(I)所示的化合物中，F为18F。
本发明首次提供一类新结构含手性侧链取代的2-芳基苯并杂环化合物，体外竞争结合实验表明，该类分子与Aβ1-42聚集体的亲和力较高；正常小鼠体内生物分布实验表明，部分F-18标记的显像剂具有初始脑摄取高，清除快的优点。有望成为一种新的用于临床显像的正电子Aβ斑块显像剂。
附图说明
图1为本发明含手性侧链取代的2-芳基苯并杂环类化合物的合成示意图；其中，a表示K2CO3、18-crown-6和丙酮，回流；b表示TBAF和甲苯，回流。
图2为本发明含光学纯手性侧链取代的2-芳基苯并杂环类化合物F-18标记中间体的制备路线图；其中，a表示NaOH、EtOH和H2O，90℃；b表示TBDMSCl、CH2Cl2和咪唑；c表示(Boc)2O和THF，回流；d表示TBAF和THF，回流；e表示TsCl和吡啶；f表示PPTS、CH2Cl2和3,4-dihydro-2H-pyran，回流。
图3为本发明F-18标记化合物的制备路线图；其中，a表示18F-、K2CO3、 Kryptofix-2.2.2和乙腈，反应温度100℃；b表示HCl(1M)，反应温度100℃。
图4为本发明实施例84中制备的化合物[18F]29s与AD病人及AD转基因小鼠脑切片的放射自显影图片。
图5为本发明实施例83中制备的化合物[18F]29r与AD病人及AD转基因小鼠脑切片的放射自显影图片。
图4和图5中：A是AD病人脑切片；B是AD转基因小鼠脑切片；C是正常人脑切片；D是正常小鼠脑切片。
具体实施方式
以下实施例用于说明本发明，但不用来限制本发明的范围。若未特别指明，实施例中所用的技术手段为本领域技术人员所熟知的常规手段，所用原料均为市售商品。
含手性侧链取代的2-芳基苯并杂环类化合物的合成。合成反应式如图1所示，本发明以下实施例中的化合物编号与图1所示反应式中编号统一。
含光学纯手性侧链取代的2-芳基苯并杂环类化合物F-18标记中间体的制备路线如图2所示。
实施例1合成中间体8r
将化合物1(257.2mg,1.0mmol)、K2CO3(435.0mg,3.2mmol)以及催化剂量的18-crown-6溶于60mL丙酮溶液中，向其中加入化合物7r(139.0mg,1.5mmol)，将上述混合物在80℃油浴条件下搅拌回流反应12h，反应完毕后，除去丙酮，经柱层析分离得到35.0mg中间体8r，结构如下，产率11.2％。1H NMR(400MHz,CDCl3)δ7.87(d,J＝8.7Hz,2H),7.86(d,J＝9.0Hz,1H),7.35(d,J＝2.5Hz,1H),7.06(dd,J＝8.9,2.6Hz,1H),6.64(d,J＝8.7Hz,2H),4.29(dd,J＝11.0,3.1Hz,1H),4.03(dd,J＝11.0,5.7Hz,1H),3.42–3.37(m,1H),2.95–2.93(m,1H),2.91(s,3H),2.79(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/z calcd for C17H16N2O2S312.09,found313.2(M+H)+。
Figure PCTCN2014090536-appb-000016
实施例2合成中间体8s
由化合物1反应制得中间体8s，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率23.0％。1H NMR(400MHz,CDCl3)δ7.87(d,J＝8.7Hz,2H),7.86(d,J＝8.9Hz,1H),7.34(d,J＝2.5Hz,1H),7.06(dd,J＝8.9,2.5Hz,1H),6.64(d,J＝8.7Hz,2H),4.29(dd,J＝11.0,3.1Hz,1H),4.02(dd,J＝11.0,5.7Hz,1H),3.41–3.36(m,1H),2.94–2.91(m,1H),2.90(s,3H),2.78(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/zcalcd for C17H16N2O2S312.09,found313.1(M+H)+。
Figure PCTCN2014090536-appb-000017
实施例3合成中间体9r
由化合物2反应制得中间体9r，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率16.4％。1H NMR(400MHz,CDCl3)δ7.91(d,J＝8.8Hz,2H),7.87(d,J＝8.9Hz,1H),7.34(d,J＝2.5Hz,1H),7.06(dd,J＝8.9,2.5Hz,1H),6.74(d,J＝8.9Hz,2H),4.29(dd,J＝11.0,3.1Hz,1H),4.02(dd,J＝11.0,5.7Hz,1H),3.42–3.37(m,1H),3.05(s,6H),2.93(t,J＝4.5Hz,1H),2.79(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/z calcd for C18H18N2O2S326.11,found327.2(M+H)+。
Figure PCTCN2014090536-appb-000018
实施例4合成中间体9s
由化合物2反应制得中间体9s，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率34.0％。1H NMR(400MHz,CDCl3)δ7.91(d,J＝8.9Hz,2H),7.87(d,J＝8.9Hz,1H),7.34(d,J＝2.5Hz,1H),7.06(dd,J＝8.9,2.5Hz,1H),6.74(d,J＝8.9Hz,2H),4.29(dd,J＝11.0,3.1Hz,1H),4.02(dd,J＝11.0,5.7Hz,1H),3.42–3.37(m,1H),3.05(s,3H),2.93(t,J＝4.6Hz,1H),2.78(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/z calcd for C18H18N2O2S326.11,found327.1(M+H)+。
Figure PCTCN2014090536-appb-000019
实施例5合成中间体10r
由化合物3反应制得中间体10r，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率38.3％。1H NMR(400MHz,CDCl3)δ8.71(d,J＝2.1Hz,1H),8.12(dd,J＝8.8,2.3Hz,1H),7.88(d,J＝8.9Hz,1H),7.35(d,J＝2.5Hz,1H),7.09(dd,J＝8.9,2.5Hz,1H),6.48(d,J＝8.8Hz,1H),5.06(d,J＝4.2Hz,1H),4.31(dd,J＝11.0,3.0Hz,1H),4.02(dd,J＝11.0,5.8Hz,1H),3.43–3.39(m,1H),3.01(d,J＝5.1Hz,3H),2.94(t,J＝4.5Hz,1H),2.80(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/zcalcd for C16H15N3O2S313.09,found314.2(M+H)+。
Figure PCTCN2014090536-appb-000020
实施例6合成中间体10s
由化合物3反应制得中间体10s，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率31.6％。1H NMR(400MHz,CDCl3)δ8.71(d,J＝2.1Hz,1H),8.12(dd,J＝8.8,2.2Hz,1H),7.88(d,J＝8.9Hz,1H),7.36(d,J＝2.4Hz,1H),7.09(dd,J＝8.9,2.5Hz,1H),6.48(d,J＝8.8Hz,1H),5.05(d,J＝4.0Hz,1H),4.32(dd,J＝11.0,3.0Hz,1H),4.02(dd,J＝11.0,5.8Hz,1H),3.43–3.38(m,1H),3.01(d,J＝5.1Hz,3H),2.95(t,J＝4.5Hz,1H),2.80(dd,J＝4.8,2.6Hz,1H),MS(ESI):m/zcalcd for C16H15N3O2S313.09,found314.1(M+H)+。
Figure PCTCN2014090536-appb-000021
实施例7合成中间体11r
由化合物4反应制得中间体11r，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率58.8％。1H NMR(400MHz,CDCl3)δ8.76(d,J＝2.3Hz,1H),8.13(dd,J＝8.9,2.2Hz,1H),7.87(d,J＝8.9Hz,1H),7.35(d,J＝2.4Hz,1H),7.08(dd,J＝8.9,2.4Hz,1H),6.59(d,J＝9.0Hz,1H),4.31(dd,J＝11.0,3.0Hz,1H),4.01(dd,J＝11.0,5.7Hz,1H),3.43–3.38(m,1H),3.18(s,6H),2.94(t,J＝4.5Hz,1H),2.80(dd,J＝4.8,2.6Hz,1H),MS(ESI):m/z calcd for C17H17N3O2S327.10,found328.2(M+H)+。
Figure PCTCN2014090536-appb-000022
实施例8合成中间体11s
由化合物4反应制得中间体11s，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率70.7％。1H NMR(400MHz,CDCl3)δ8.76(d,J＝2.3Hz,1H),8.13(dd,J＝9.0,2.4Hz,1H),7.87(d,J＝8.9Hz,1H),7.35(d,J＝2.5Hz,1H),7.08(dd,J＝8.9,2.5Hz,1H),6.59(d,J＝9.0Hz,1H),4.31(dd,J＝11.0,3.0Hz,1H),4.01(dd,J＝11.0,5.8Hz,1H),3.43–3.38(m,1H),3.18(s,6H),2.95(t,J＝4.5Hz,1H),2.80(dd,J＝4.8,2.6Hz,1H),MS(ESI):m/z calcd for C17H17N3O2S327.10,found328.1(M+H)+。
Figure PCTCN2014090536-appb-000023
实施例9合成中间体12r
由化合物5反应制得中间体12r，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率55.3％。1H NMR(400MHz,CDCl3)δ8.02(d,J＝8.8Hz,2H),7.56(d,J＝8.7Hz,1H),7.10(d,J＝2.3Hz,1H),6.93(dd,J＝8.7,2.4Hz,1H),6.66(d,J＝8.8Hz,2H),4.29(dd,J＝11.0,3.1Hz,1H),4.00(dd,J＝11.0,5.7Hz,1H),3.42–3.38(m,1H),2.99–2.93(m,1H),2.92(s,3H),2.80(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/zcalcd for C17H16N2O3296.12,found297.0(M+H)+。
Figure PCTCN2014090536-appb-000024
实施例10合成中间体12s
由化合物5反应制得中间体12s，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率51.9％。1H NMR(400MHz,CDCl3)δ8.03(d,J＝8.8Hz,2H),7.57(d,J＝8.7Hz,1H),7.11(d,J＝2.4Hz,1H),6.93(dd,J＝8.7,2.4Hz,1H),6.67(d,J＝ 8.8Hz,2H),4.29(dd,J＝11.0,3.1Hz,1H),4.01(dd,J＝11.0,5.7Hz,1H),3.43–3.38(m,1H),2.97–2.93(m,1H),2.92(s,3H),2.80(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/z calcd for C17H16N2O3296.12,found296.9(M+H)+。
Figure PCTCN2014090536-appb-000025
实施例11合成中间体13r
由化合物6反应制得中间体13r，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率43.5％。1H NMR(400MHz,CDCl3)δ8.06(d,J＝9.0Hz,2H),7.57(d,J＝8.7Hz,1H),7.11(d,J＝2.3Hz,1H),6.93(dd,J＝8.7,2.4Hz,1H),6.77(d,J＝9.0Hz,2H),4.29(dd,J＝11.0,3.1Hz,1H),4.01(dd,J＝11.0,5.7Hz,1H),3.43–3.38(m,1H),3.07(s,6H),2.98–2.91(m,1H),2.80(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/zcalcd for C18H18N2O3310.13,found310.8(M+H)+。
Figure PCTCN2014090536-appb-000026
实施例12合成中间体13s
由化合物6反应制得中间体13s，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率28.1％。1H NMR(400MHz,CDCl3)δ8.06(d,J＝8.7Hz,2H),7.57(d,J＝8.6Hz,1H),7.11(d,J＝1.7Hz,1H),6.93(dd,J＝8.6,1.8Hz,1H),6.77(d,J＝8.7Hz,2H),4.29(dd,J＝10.9,2.7Hz,1H),4.01(dd,J＝10.9,5.7Hz,1H),3.43–3.38(m,1H),3.07(s,6H),2.95(t,J＝4.4Hz,1H),2.88–2.73(m,1H),MS(ESI):m/z calcd for C18H18N2O3310.13,found310.8(M+H)+。
Figure PCTCN2014090536-appb-000027
实施例13合成目标化合物14s
将中间体8r(54.1mg,0.17mmol)溶于15mL甲苯溶液中，向其中加入TBAF的THF溶液(1mL，1.0mmol)，将上述混合物在80℃油浴条件下搅拌回流反应12h，反应完毕后，除去甲苯，残余物经柱层析分离得到8.4mg目标化合物14s，结构如下，产率14.9％。1H NMR(400MHz,DMSO-d6)δ7.77(d,J＝9.0Hz,1H),7.74(d,J＝8.7Hz,2H),7.62(d,J＝2.4Hz,1H),7.05(dd,J＝8.9,2.4Hz,1H),6.62(d,J＝8.7Hz,2H),5.47(d,J＝4.8Hz,1H),4.59–4.50(m,1H),4.47–4.39(m,1H),4.09–3.98(m,3H),2.73(d,J＝4.8Hz,3H),HRMS(EI):m/z calcd for C17H18N2O2FS333.1073；found333.1072(M+H)+。
Figure PCTCN2014090536-appb-000028
实施例14合成目标化合物14r
由化合物8s反应制得目标化合物14r，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率7.4％。1H NMR(400MHz,DMSO-d6)δ7.77(d,J＝9.0Hz,1H),7.74(d,J＝8.8Hz,2H),7.62(d,J＝2.4Hz,1H),7.05(dd,J＝8.9,2.5Hz,1H),6.62(d,J＝8.7Hz,2H),5.47(d,J＝5.1Hz,1H),4.60–4.50(m,1H),4.47–4.39(m,1H),4.09–4.01(m,3H),2.73(d,J＝4.9Hz,3H),HRMS(EI):m/z calcd for C17H18N2O2FS333.1073,found333.1079(M+H)+。
Figure PCTCN2014090536-appb-000029
实施例15合成目标化合物15s
由中间体9r反应制得目标化合物15s，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率5.8％。1H NMR(400MHz,CDCl3)δ7.91(d,J＝8.8Hz,2H),7.88(d,J＝9.0Hz,1H),7.33(d,J＝2.4Hz,1H),7.04(dd,J＝8.9,2.5Hz,1H),6.74(d,J＝8.8Hz,2H),4.72–4.64(m,1H),4.60–4.52(m,1H),4.35–4.24(m,1H),4.17–4.10(m,2H),3.05(s,6H),HRMS(EI):m/z calcd for C18H20N2O2FS347.1230,found347.1234(M+H)+。
Figure PCTCN2014090536-appb-000030
实施例16合成目标化合物15r
由中间体9s反应制得目标化合物15r，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率14.8％。1H NMR(400MHz,CDCl3)δ7.91(d,J＝8.8Hz,2H),7.88(d,J＝8.9Hz,1H),7.33(d,J＝2.4Hz,1H),7.04(dd,J＝8.9,2.5Hz,1H),6.74(d,J＝8.9Hz,2H),4.72–4.64(m,1H),4.61–4.52(m,1H),4.35–4.24(m,1H),4.15–4.13(m,2H),3.05(s,6H),HRMS(EI):m/z calcd for C18H20N2O2FS347.1230,found347.1223(M+H)+。
Figure PCTCN2014090536-appb-000031
实施例17合成目标化合物16s
由中间体10r反应制得目标化合物16s，其反应的原料比例、溶剂、反应条件等均 与实施例13相同，产率21.0％。1H NMR(400MHz,CDCl3)δ8.71(d,J＝1.8Hz,1H),8.12(dd,J＝8.8,2.2Hz,1H),7.88(d,J＝8.9Hz,1H),7.35(d,J＝2.4Hz,1H),7.07(dd,J＝8.9,2.5Hz,1H),6.48(d,J＝8.8Hz,1H),5.04(s,1H),4.73–4.65(m,1H),4.61–4.53(m,1H),4.36–4.25(m,1H),4.17–4.15(m,2H),3.01(d,J＝5.1Hz,3H),HRMS(EI):m/z calcd for C16H17N3O2FS334.1026,found334.1033(M+H)+。
Figure PCTCN2014090536-appb-000032
实施例18合成目标化合物16r
由中间体10s反应制得目标化合物16r，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率14.1％。1H NMR(400MHz,CDCl3)δ8.68(d,J＝2.0Hz,1H),8.15(dd,J＝8.8,2.3Hz,1H),7.88(d,J＝8.9Hz,1H),7.36(d,J＝2.5Hz,1H),7.08(dd,J＝8.9,2.5Hz,1H),6.51(d,J＝8.8Hz,1H),5.27(s,1H),4.73–4.68(m,1H),4.61–4.54(m,1H),4.34–4.27(m,1H),4.17–4.15(m,2H),3.01(d,J＝5.1Hz,3H),HRMS(EI):m/zcalcd for C16H17N3O2FS334.1026,found334.1022(M+H)+。
Figure PCTCN2014090536-appb-000033
实施例19合成目标化合物17s
由中间体11r反应制得目标化合物17s，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率38.6％。1H NMR(400MHz,CDCl3)δ8.76(d,J＝2.0Hz,1H),8.14(d,J＝8.8Hz,1H),7.88(d,J＝8.9Hz,1H),7.35(d,J＝2.2Hz,1H),7.06(dd,J＝8.9,2.2Hz,1H),6.59(d,J＝9.0Hz,1H),4.73–4.65(m,1H),4.64–4.53(m,1H),4.33–4.28(m,1H),4.19–4.11(m,2H),3.19(s,6H),HRMS(EI):m/z calcd for C17H19N3O2FS348.1182,found348.1188(M+H)+。
Figure PCTCN2014090536-appb-000034
实施例20合成目标化合物17r
由中间体11s反应制得目标化合物17r，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率35.7％。1H NMR(400MHz,CDCl3)δ8.76(s,1H),8.13(d,J＝8.9Hz,1H),7.87(d,J＝8.9Hz,1H),7.35(d,J＝1.9Hz,1H),7.06(dd,J＝8.8,1.9Hz,1H),6.59(d,J＝9.0Hz,1H),4.73–4.65(m,1H),4.61–4.53(m,1H),4.34–4.26(m,1H),4.16–4.11(m,2H),3.19(s,6H),HRMS(EI):m/z calcd for C17H19N3O2FS348.1182,found348.1189(M+H)+。
Figure PCTCN2014090536-appb-000035
实施例21合成目标化合物18s
由中间体12r反应制得目标化合物18s，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率62.4％。1H NMR(400MHz,CDCl3)δ8.06(d,J＝8.7Hz,2H),7.59(d,J＝8.7Hz,1H),7.08(d,J＝2.3Hz,1H),6.93(dd,J＝8.7,2.4Hz,1H),6.77(d,J＝8.4Hz,2H),4.75–4.64(m,1H),4.63–4.52(m,1H),4.39–4.25(m,1H),4.19–4.01(m,2H),2.94(s,3H)。
Figure PCTCN2014090536-appb-000036
实施例22合成目标化合物18r
由中间体12s反应制得目标化合物18r，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率51.4％。1H NMR(400MHz,CDCl3)δ8.03(d,J＝8.8Hz,2H),7.57(d,J＝8.7Hz,1H),7.09(d,J＝2.3Hz,1H),6.91(dd,J＝8.7,2.4Hz,1H),6.68(d,J＝8.6Hz,2H),4.75–4.64(m,1H),4.63–4.52(m,1H),4.38–4.24(m,1H),4.19–4.06(m,2H),2.92(s,3H)。
Figure PCTCN2014090536-appb-000037
实施例23合成目标化合物19s
由中间体13r反应制得目标化合物19s，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率61.1％。1H NMR(400MHz,CDCl3)δ8.07(d,J＝8.9Hz,2H),7.58(d,J＝8.7Hz,1H),7.10(d,J＝2.3Hz,1H),6.92(dd,J＝8.7,2.4Hz,1H),6.78(d,J＝8.8Hz,2H),4.73–4.65(m,1H),4.62–4.53(m,1H),4.38–4.22(m,1H),4.17–4.07(m,1H),3.08(s,6H)。
Figure PCTCN2014090536-appb-000038
实施例24合成目标化合物19r
由中间体13s反应制得目标化合物19r，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率63.3％。1H NMR(400MHz,CDCl3)δ8.06(d,J＝8.5Hz,2H),7.58(d,J＝8.7Hz,1H),7.10(d,J＝2.3Hz,1H),6.92(dd,J＝8.7,2.4Hz,1H),6.78(d,J ＝7.9Hz,2H),4.73–4.65(m,1H),4.62–4.53(m,1H),4.40–4.22(m,1H),4.19–4.06(m,1H),4.17–4.09(m,1H),3.07(s,6H)。
Figure PCTCN2014090536-appb-000039
实施例25合成中间体24r
由化合物20反应制得中间体24r，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率31.7％。1H NMR(400MHz,CDCl3)δ8.06(d,J＝8.8Hz,2H),7.40(d,J＝8.8Hz,1H),7.21(d,J＝2.5Hz,1H),6.91(dd,J＝8.8,2.5Hz,1H),6.67(d,J＝8.8Hz,2H),4.27(dd,J＝10.9,3.2Hz,1H),4.01(dd,J＝10.9,5.7Hz,1H),3.42–3.37(m,1H),2.93(s,3H),2.94–2.91(m,1H),2.79(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/zcalcd for C17H16N2O3296.12,found297.2(M+H)+。
Figure PCTCN2014090536-appb-000040
实施例26合成中间体24s
由化合物20反应制得中间体24s，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率16.2％。1H NMR(400MHz,CDCl3)δ8.04(d,J＝8.8Hz,2H),7.39(d,J＝8.8Hz,1H),7.20(d,J＝2.4Hz,1H),6.90(dd,J＝8.8,2.5Hz,1H),6.66(d,J＝8.8Hz,2H),4.26(dd,J＝10.9,3.2Hz,1H),4.00(dd,J＝10.9,5.7Hz,1H),3.42–3.37(m,1H),2.92(s,3H),2.94–2.90(m,1H),2.79(dd,J＝4.9,2.7Hz,1H),MS(ESI):m/zcalcd for C17H16N2O3296.12,found297.2(M+H)+。
Figure PCTCN2014090536-appb-000041
实施例27合成中间体25r
由化合物21反应制得中间体25r，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率52.9％。1H NMR(400MHz,CDCl3)δ8.08(d,J＝9.0Hz,2H),7.39(d,J＝8.8Hz,1H),7.21(d,J＝2.4Hz,1H),6.90(dd,J＝8.8,2.5Hz,1H),6.76(d,J＝9.0Hz,2H),4.26(dd,J＝10.9,3.2Hz,1H),4.01(dd,J＝10.9,5.6Hz,1H),3.42–3.37(m,1H),3.07(s,6H),2.92(t,J＝4.5Hz,1H),2.79(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/z calcd for C18H18N2O3310.13,found311.2(M+H)+。
Figure PCTCN2014090536-appb-000042
实施例28合成中间体25s
由化合物21反应制得中间体25s，其反应的原料比例、溶剂、反应条 件等均与实施例1相同，产率72.9％。1H NMR(400MHz,CDCl3)δ8.08(d,J＝9.0Hz,2H),7.39(d,J＝8.8Hz,1H),7.21(d,J＝2.4Hz,1H),6.90(dd,J＝8.8,2.5Hz,1H),6.76(d,J＝9.0Hz,2H),4.26(dd,J＝10.9,3.2Hz,1H),4.01(dd,J＝10.9,5.6Hz,1H),3.42–3.37(m,1H),3.07(s,6H),2.92(t,J＝4.5Hz,1H),2.79(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/z calcd for C18H18N2O3310.13,found311.2(M+H)+。
Figure PCTCN2014090536-appb-000043
实施例29合成中间体26r
由化合物22反应制得中间体26r，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率34.6％。1H NMR(400MHz,CDCl3)δ8.93(d,J＝2.0Hz,1H),8.22(dd,J＝8.8,2.2Hz,1H),7.42(d,J＝8.8Hz,1H),7.21(d,J＝2.4Hz,1H),6.93(dd,J＝8.8,2.5Hz,1H),6.50(d,J＝8.8Hz,1H),5.18(s,1H),4.28(dd,J＝10.9,3.1Hz,1H),4.00(dd,J＝10.9,5.7Hz,1H),3.43–3.38(m,1H),3.02(d,J＝5.1Hz,3H),2.94(t,J＝4.5Hz,1H),2.80(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/z calcd for C16H15N3O3297.11,found298.2(M+H)+。
Figure PCTCN2014090536-appb-000044
实施例30合成中间体26s
由化合物22反应制得中间体26s，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率33.3％。1H NMR(400MHz,CDCl3)δ8.93(d,J＝1.6Hz,1H),8.22(dd,J＝8.8,2.1Hz,1H),7.42(d,J＝8.8Hz,1H),7.21(d,J＝2.4Hz,1H),6.93(dd,J＝8.8,2.5Hz,1H),6.50(d,J＝8.9Hz,1H),5.19(s,1H),4.28(dd,J＝10.9,3.1Hz,1H),4.00(dd,J＝10.9,5.7Hz,1H),3.42–3.39(m,1H),3.02(d,J＝5.1Hz,3H),2.94(t,J＝4.5Hz,1H),2.80(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/z calcd for C16H15N3O3297.11,found298.2(M+H)+。
Figure PCTCN2014090536-appb-000045
实施例31合成中间体27r
由化合物23反应制得中间体27r，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率71.7％。1H NMR(400MHz,CDCl3)δ8.98(d,J＝2.2Hz,1H),8.21(d,J＝9.0Hz,1H),7.42(d,J＝8.8Hz,1H),7.21(d,J＝2.4Hz,1H),6.92(dd,J＝8.8,2.5Hz,1H),6.61(d,J＝9.0Hz,1H),4.28(dd,J＝10.9,3.1Hz,1H),4.00(dd,J＝10.9,5.7Hz,1H),3.43–3.38(m,1H),3.21(s,6H),2.93(t,J＝4.5Hz,1H),2.80(dd,J＝4.9,2.6Hz,1H),MS(ESI):m/z calcd for C17H17N3O3311.13,found312.2(M+H)+。
Figure PCTCN2014090536-appb-000046
实施例32合成中间体27s
由化合物23反应制得中间体27s，其反应的原料比例、溶剂、反应条件等均与实施例1相同，产率61.8％。1H NMR(400MHz,CDCl3)δ8.87(d,J＝2.1Hz,1H),8.09(dd,J＝8.9,1.9Hz,1H),7.31(d,J＝8.8Hz,1H),7.09(d,J＝2.4Hz,1H),6.81(dd,J＝8.8,2.5Hz,1H),6.49(d,J＝9.0Hz,1H),4.17(dd,J＝10.9,3.1Hz,1H),3.88(dd,J＝10.9,5.7Hz,1H),3.32–3.27(m,1H),3.09(s,6H),2.82(t,J＝4.5Hz,1H),2.69(dd,J＝4.8,2.6Hz,1H),MS(ESI):m/z calcd for C17H17N3O3311.13,found312.2(M+H)+。
Figure PCTCN2014090536-appb-000047
实施例33合成目标化合物28s
由由中间体24r反应制得制得目标化合物28s，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率16.6％。1H NMR(400MHz,CDCl3)δ8.05(d,J＝8.6Hz,2H),7.40(d,J＝8.8Hz,1H),7.22(d,J＝2.1Hz,1H),6.88(dd,J＝8.8,2.1Hz,1H),6.67(d,J＝8.6Hz,2H),4.73–4.64(m,1H),4.61–4.52(m,1H),4.34–4.24(m,1H),4.16–4.09(m,2H),2.93(s,3H),HRMS(EI):m/z calcd for C17H18N2O3F317.1301；found317.1293(M+H)+。
Figure PCTCN2014090536-appb-000048
实施例34合成目标化合物28r
由中间体24s反应制得目标化合物28r，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率31.3％。1H NMR(400MHz,CDCl3)δ8.05(d,J＝8.7Hz,2H),7.40(d,J＝8.8Hz,1H),7.22(d,J＝2.5Hz,1H),6.88(dd,J＝8.8,2.5Hz,1H),6.67(d,J＝8.8Hz,2H),4.72–4.64(m,1H),4.60–4.52(m,1H),4.34–4.24(m,1H),4.16–4.07(m,2H),2.92(s,3H).HRMS(EI):m/z calcd for C17H18N2O3F317.1301；found317.1295(M+H)+。
Figure PCTCN2014090536-appb-000049
实施例35合成目标化合物29s
由中间体25r反应制得目标化合物29s，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率53.0％。1H NMR(400MHz,CDCl3)δ8.08(d,J＝8.5Hz,2H),7.40(d,J ＝8.6Hz,1H),7.22(s,1H),6.87(d,J＝8.0Hz,1H),6.77(d,J＝8.3Hz,2H),4.68–4.66(m,1H),4.57–4.54(m,1H),4.32–4.27(m,1H),4.13(s,2H),3.07(s,6H),HRMS(EI):m/z calcd for C18H20N2O3F331.1458,found331.1448(M+H)+。
Figure PCTCN2014090536-appb-000050
实施例36合成目标化合物29r
由中间体25s反应制得目标化合物29r，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率28.0％。1H NMR(400MHz,CDCl3)δ8.09(d,J＝8.6Hz,2H),7.40(d,J＝8.8Hz,1H),7.22(s,1H),6.88(d,J＝8.7Hz,1H),6.78(d,J＝8.6Hz,2H),4.72–4.64(m,1H),4.60–4.52(m,1H),4.33–4.25(m,1H),4.13(s,2H),3.08(s,6H),HRMS(EI):m/z calcd for C18H20N2O3F331.1458,found331.1454(M+H)+。
Figure PCTCN2014090536-appb-000051
实施例37合成目标化合物30s
由中间体26r反应制得目标化合物30s，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率22.6％。1H NMR(400MHz,CDCl3)δ8.92(d,J＝1.6Hz,1H),8.23(dd,J＝8.8,1.7Hz,1H),7.43(d,J＝8.8Hz,1H),7.22(d,J＝2.3Hz,1H),6.91(dd,J＝8.8,2.3Hz,1H),6.51(d,J＝8.9Hz,1H),5.26(s,1H),4.74–4.65(m,1H),4.62–4.54(m,1H),4.34–4.27(m,1H),4.14–4.12(m,2H),3.02(d,J＝5.1Hz,3H),HRMS(EI):m/zcalcd for C16H17N3O3F318.1254,found318.1251(M+H)+。
Figure PCTCN2014090536-appb-000052
实施例38合成目标化合物30r
由中间体26s反应制得目标化合物30r，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率28.2％。1H NMR(400MHz,CDCl3)δ8.93(s,1H),8.21(d,J＝8.8Hz,1H),7.42(d,J＝8.8Hz,1H),7.21(d,J＝2.0Hz,1H),6.90(dd,J＝8.8,2.2Hz,1H),6.49(d,J＝8.8Hz,1H),5.09(s,1H),4.73–4.64(m,1H),4.61–4.52(m,1H),4.35–4.24(m,1H),4.13(s,2H),3.01(d,J＝5.1Hz,3H),HRMS(EI):m/z calcd for C16H17N3O3F318.1254,found318.1246(M+H)+。
Figure PCTCN2014090536-appb-000053
实施例39合成目标化合物31s
由中间体27r反应制得目标化合物31s，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率19.5％。1H NMR(400MHz,CDCl3)δ8.98(d,J＝1.8Hz,1H),8.20(dd,J＝9.0,2.0Hz,1H),7.42(d,J＝8.8Hz,1H),7.21(d,J＝2.3Hz,1H),6.89(dd,J＝8.8,2.3Hz,1H),6.60(d,J＝9.0Hz,1H),4.74–4.65(m,1H),4.62–4.53(m,1H),4.35–4.26(m,1H),4.16–4.09(m,2H),3.20(s,6H),HRMS(EI):m/z calcd for C17H19N3O3F332.1410,found332.1415(M+H)+。
Figure PCTCN2014090536-appb-000054
实施例40合成目标化合物31r
由中间体27s反应制得目标化合物31r，其反应的原料比例、溶剂、反应条件等均与实施例13相同，产率29.2％。1H NMR(400MHz,CDCl3)δ9.00(d,J＝2.0Hz,1H),8.23(d,J＝7.9Hz,1H),7.44(d,J＝8.8Hz,1H),7.23(d,J＝2.4Hz,1H),6.92(dd,J＝8.8,2.5Hz,1H),6.64(d,J＝8.9Hz,1H),4.75–4.66(m,1H),4.63–4.55(m,1H),4.37–4.27(m,1H),4.18–4.11(m,2H),3.23(s,6H),HRMS(EI):m/z calcd for C17H19N3O3F332.1410,found332.1403(M+H)+。
Figure PCTCN2014090536-appb-000055
实施例41合成标记中间体33s
将化合物20(240.0mg，1mmol)溶于20mL乙醇溶液中，向其中加入NaOH水溶液(69.0mg，2mL)，上述溶液在80℃油浴条件下搅拌回流反应1h后，向其中加入化合物32s(135.5mg，1.2mmol)，继续搅拌回流反应3h，反应完毕后经柱层析分离得到146.0mg中间体33s，产率46.5％。1H NMR(400MHz,d6-DMSO)δ7.90(d,J＝7.9Hz,2H),7.55(d,J＝8.2Hz,1H),7.20(s,1H),6.89(d,J＝8.7Hz,1H),6.68(d,J＝7.9Hz,2H),6.55(s,1H),4.96(d,J＝3.4Hz,1H),4.68(s,1H),4.03(s,1H),3.97–3.85(m,1H),3.82(s,1H),3.47(s,2H),2.76(d,J＝2.7Hz,3H),MS(ESI):m/z calcd for C17H18N2O4314.13,found315.2(M+H)+。
Figure PCTCN2014090536-appb-000056
实施例42合成标记中间体33r
由化合物20反应制得中间体33r，其反应的原料比例、溶剂、反应条件等均与实施例41相同，产率39.6％。1H NMR(400MHz,d6-DMSO)δ7.90(d,J＝8.8Hz,2H),7.54(d,J＝8.8Hz,1H),7.20(d,J＝2.5Hz,1H),6.88(dd,J＝8.8,2.5Hz,1H),6.67(d,J＝8.8Hz,2H),6.53(d,J＝5.0Hz,1H),4.04(dd,J＝9.9,4.2Hz,1H),3.91(dd,J＝9.9,6.1Hz,1H),3.82(dd,J＝10.7,5.0Hz,1H),3.47(d,J＝5.7Hz,2H),2.76(d,J＝4.1Hz,3H),MS(ESI):m/z calcd for C17H18N2O4314.13,found315.3(M+H)+。
Figure PCTCN2014090536-appb-000057
实施例43合成标记中间体34r
将化合物33s(129.0mg，0.4mmol)溶于30mL CH2Cl2溶液中，向其中加入咪唑(237.9mg，3.5mmol)以及TBDMSCl(536.2mg，3.6mmol)，上述溶液在40℃油浴条件下搅拌回流反应，反应完毕后，抽滤除去沉淀，除去CH2Cl2，残余物经柱层析分离得到154.2mg中间体34r，产率71.1％。1H NMR(400MHz,CDCl3)δ7.96(d,J＝8.7Hz,2H),7.28(d,J＝8.8Hz,1H),7.10(d,J＝2.4Hz,1H),6.77(dd,J＝8.8,2.5Hz,1H),6.57(d,J＝8.8Hz,2H),4.08–3.94(m,2H),3.80(dd,J＝8.4,5.6Hz,1H),3.57(d,J＝5.7Hz,2H),2.82(s,3H),0.80(s,18H),MS(ESI):m/z calcd for C29H46N2O4Si2542.30,found543.7(M+H)+。
Figure PCTCN2014090536-appb-000058
实施例44合成标记中间体34s
由中间体33r反应制得中间体34s，其反应的原料比例、溶剂、反应条件等均与实施例43相同，产率24.2％。1H NMR(400MHz,CDCl3)δ7.97(d,J＝8.8Hz,2H),7.30(d,J＝8.8Hz,1H),7.12(d,J＝2.4Hz,1H),6.79(dd,J＝8.8,2.5Hz,1H),6.59(d,J＝8.8Hz,2H),4.06–3.99(m,2H),3.85–3.81(m,1H),3.60(d,J＝5.7Hz,2H),2.84(s,3H),0.83(s,18H),MS(ESI):m/z calcd for C29H46N2O4Si2542.30,found543.1(M+H)+。
Figure PCTCN2014090536-appb-000059
实施例45合成标记中间体35r
将化合物34r(245.7mg，0.45mmol)溶于30mL THF溶液中，向其中加入过量的(Boc)2O，上述溶液在85℃油浴条件下搅拌回流反应，反应完毕后，除去THF，残余物经柱层析分离得到279.2mg中间体35r，产率95.9％。1H NMR(400MHz,CDCl3)δ8.11(d,J＝8.5Hz,2H),7.36(t,J＝8.6Hz,3H),7.17(d,J＝2.3Hz,1H),6.88(dd,J＝ 8.9,2.2Hz,1H),4.10–3.96(m,2H),3.84(dd,J＝8.8,6.2Hz,1H),3.60(d,J＝5.8Hz,2H),3.26(s,3H),1.41(s,9H),0.83(s,18H),MS(ESI):m/z calcd for C34H54N2O6Si2642.35,found643.6(M+H)+。
Figure PCTCN2014090536-appb-000060
实施例46合成标记中间体35s
由中间体34s反应制得中间体35s，其反应的原料比例、溶剂、反应条件等均与实施例45相同，产率88.0％。1H NMR(400MHz,CDCl3)δ8.10(d,J＝8.7Hz,2H),7.35(t,J＝8.7Hz,3H),7.17(d,J＝2.4Hz,1H),6.87(dd,J＝8.9,2.5Hz,1H),4.13–3.98(m,2H),3.84(dd,J＝8.9,6.2Hz,1H),3.70–3.53(m,2H),3.25(s,3H),1.41(s,9H),0.83(s,18H),MS(ESI):m/z calcd for C34H54N2O6Si2642.35,found643.4(M+H)+。
Figure PCTCN2014090536-appb-000061
实施例47合成标记中间体36s
将化合物35r(273.2mg，0.37mmol)溶于20mL THF溶液中，向其中加入TBAF的THF溶液2.5mL，上述溶液在30℃油浴条件下搅拌回流反应，反应完毕后，除去THF，残余物经柱层析分离得到78.6mg中间体36s，产率37.4％。1H NMR(400MHz,CDCl3)δ8.13(d,J＝8.5Hz,2H),7.38(dd,J＝15.0,8.7Hz,3H),7.23(d,J＝2.2Hz,1H),6.88(dd,J＝8.8,2.4Hz,1H),4.20–4.12(m,1H),4.07(s,2H),3.86(dd,J＝11.4,3.5Hz,1H),3.78(dd,J＝11.4,5.7Hz,1H),3.31(d,J＝2.7Hz,3H),1.49(s,9H),MS(ESI):m/z calcd for C22H26N2O6414.18,found415.3(M+H)+。
Figure PCTCN2014090536-appb-000062
实施例48合成标记中间体36r
由中间体35s反应制得中间体36r，其反应的原料比例、溶剂、反应条件等均与实施例47相同，产率60.1％。1H NMR(400MHz,CDCl3)δ8.17(d,J＝8.8Hz,2H),7.43(t,J＝8.6Hz,3H),7.25(s,1H),6.94(dd,J＝8.8,2.5Hz,1H),4.21-4.13(m,1H),4.13-4.05(m,2H),3.87(dd,J＝11.4,3.8Hz,1H),3.79(dd,J＝11.4,5.4Hz,1H),3.33(s,3H),1.49(s,9H)。
Figure PCTCN2014090536-appb-000063
实施例49合成标记中间体37r
将化合物36s(79.6mg，0.19mmol)溶于5mL吡啶溶液中，向其中加入TsCl(55.2mg，0.29mmol)，在0℃冰浴条件下搅拌反应，反应完毕后，除去吡啶，残余物经柱层析分离得到30.0mg中间体37r，产率27.8％。1H NMR(400MHz,CDCl3)δ8.17(t,J＝5.6Hz,2H),7.79(d,J＝8.2Hz,2H),7.43(d,J＝8.7Hz,3H),7.30(d,J＝8.1Hz,2H),7.15(d,J＝2.4Hz,1H),6.86(dd,J＝8.9,2.5Hz,1H),4.34–4.13(m,3H),4.03(d,J＝4.3Hz,2H),3.33(s,3H),2.40(s,3H),1.49(s,9H),MS(ESI):m/zcalcd for C29H32N2O8S568.19,found569.6(M+H)+。
Figure PCTCN2014090536-appb-000064
实施例50合成标记中间体37s
由中间体36r反应制得中间体37s，其反应的原料比例、溶剂、反应条件等均与实施例49相同，产率62.3％。1H NMR(400MHz,CDCl3)δ8.18(d,J＝8.7Hz,2H),7.81(d,J＝8.2Hz,2H),7.44(dd,J＝8.7,3.8Hz,3H),7.32(d,J＝8.0Hz,2H),7.16(d,J＝2.3Hz,1H),6.87(dd,J＝8.9,2.4Hz,1H),4.34-4.20(m,3H),4.04(d,J＝4.1Hz,2H),3.34(s,3H),2.41(s,3H),1.49(s,9H)。
Figure PCTCN2014090536-appb-000065
实施例51合成标记前体38r
将化合物37r(30.0mg，0.05mmol)溶于10mL CH2Cl2溶液中，向其中加入3,4-dihydro-2H-pyran(78.2mg，0.93mmol)以及PPTS(59.2mg，0.24mmol),上述溶液在40℃油浴条件下搅拌回流反应，反应完毕后，除去CH2Cl2，残余物经柱层析分离得到19.9mg标记前体38r，产率61.0％。1H NMR(400MHz,CDCl3)δ8.18(d,J＝8.5Hz,2H),7.78(dd,J＝8.2,5.5Hz,2H),7.42(dd,J＝8.7,3.3Hz,3H),7.33–7.25(m,2H),7.13(dd,J＝8.5,2.4Hz,1H),6.83(ddd,J＝8.8,3.8,2.6Hz,1H),4.78(dt,J＝6.8,3.8Hz,1H),4.43–4.18(m,3H),4.18–3.96(m,2H),3.95–3.69(m,1H),3.58–3.41(m,1H),3.33(s,3H),2.38(s,3H),1.88–1.62(m,3H),1.49(s,9H),MS(ESI):m/z calcd for C34H40N2O9S652.25,found653.6(M+H)+。
Figure PCTCN2014090536-appb-000066
实施例52合成标记前体38s
由中间体37s反应制得标记前体38s，其反应的原料比例、溶剂、反应条件等均与 实施例51相同，产率79.4％。1H NMR(400MHz,CDCl3)δ8.18(d,J＝8.5Hz,2H),7.78(dd,J＝8.2,5.4Hz,2H),7.42(dd,J＝8.6,2.9Hz,3H),7.28(s,2H),7.13(dd,J＝8.4,2.4Hz,1H),6.89-6.80(m,1H),4.86-4.69(m,1H),4.42-4.17(m,3H),4.12-3.96(m,2H),4.00-3.76(m,1H),3.51-3.47(m,1H),3.33(s,3H),2.39(d,J＝1.4Hz,3H),1.87-1.63(m,3H),1.64-1.52(m,3H),1.49(s,9H)。
Figure PCTCN2014090536-appb-000067
实施例53合成标记中间体39s
由化合物1和化合物32s反应制得中间体39s，其反应的原料比例、溶剂、反应条件等均与实施例41相同，产率80.9％。1H NMR(400MHz,d6-DMSO)δ7.77(t,J＝8.6Hz,3H),7.60(d,J＝2.5Hz,1H),7.05(dd,J＝8.9,2.5Hz,1H),6.64(d,J＝8.8Hz,2H),6.39(d,J＝4.9Hz,1H),4.98(d,J＝5.1Hz,1H),4.68(t,J＝5.6Hz,1H),4.07(dd,J＝9.8,4.2Hz,1H),3.94(dd,J＝9.9,6.1Hz,1H),3.88–3.76(m,1H),3.47(t,J＝5.7Hz,2H),2.75(d,J＝4.9Hz,3H),MS(ESI):m/z calcd for C17H1 8N2O3S330.10,found330.8(M+H)+。
Figure PCTCN2014090536-appb-000068
实施例54合成标记中间体39r
由化合物1和化合物32r反应制得中间体39r，其反应的原料比例、溶剂、反应条件等均与实施例41相同，产率49.0％。1H NMR(400MHz,d6-DMSO)δ7.78(t,J＝8.7Hz,3H),7.60(d,J＝2.5Hz,1H),7.05(dd,J＝8.8,2.5Hz,1H),6.63(d,J＝8.8Hz,2H),6.38(d,J＝4.9Hz,1H),4.97(s,1H),4.68(s,1H),4.07(dd,J＝9.9,4.2Hz,1H),3.94(dd,J＝9.9,6.1Hz,1H),3.84–3.81(m,1H),3.47(d,J＝5.5Hz,2H),2.75(d,J＝4.9Hz,3H),MS(ESI):m/z calcd for C17H18N2O3S330.10,found330.8(M+H)+。
Figure PCTCN2014090536-appb-000069
实施例55合成标记中间体40r
由中间体39s反应制得中间体40r，其反应的原料比例、溶剂、反应条件等均与实施例43相同，产率63.8％。1H NMR(400MHz,CDCl3)δ7.84–7.75(m,3H),7.22(d,J＝2.5Hz,1H),6.95(dd,J＝8.9,2.5Hz,1H),6.55(d,J＝8.7Hz,2H),4.11–4.01(m,1H),4.03–3.94(m,1H),3.83(dd,J＝9.5,6.5Hz,1H),3.63–3.51(m,2H),2.81(s ,3H),0.81(s,9H),0.80(s,9H),MS(ESI):m/z calcd for C29H46N2O3SSi2558.28,found559.1(M+H)+。
Figure PCTCN2014090536-appb-000070
实施例56合成标记中间体40s
由中间体39r反应制得中间体40s，其反应的原料比例、溶剂、反应条件等均与实施例43相同，产率90.4％。1H NMR(400MHz,CDCl3)δ7.81(d,J＝8.3Hz,3H),7.24(d,J＝2.4Hz,1H),6.97(dd,J＝8.9,2.5Hz,1H),6.57(d,J＝8.6Hz,2H),4.08(dd,J＝9.5,3.6Hz,1H),4.03–3.97(m,1H),3.86(dd,J＝9.5,6.5Hz,1H),3.65–3.51(m,2H),2.83(s,3H),0.83(s,9H),0.82(s,9H),MS(ESI):m/z calcd for C29H46N2O3SSi2558.28,found559.0(M+H)+。
Figure PCTCN2014090536-appb-000071
实施例57合成标记中间体41r
由中间体40r反应制得标记中间体41r，其反应的原料比例、溶剂、反应条件等均与实施例45相同，产率86.0％。1H NMR(400MHz,CDCl3)δ7.92(d,J＝8.6Hz,2H),7.86(d,J＝8.9Hz,1H),7.29(d,J＝8.6Hz,3H),7.02(dd,J＝8.9,2.5Hz,1H),4.10(dd,J＝9.5,3.5Hz,1H),4.06–3.97(m,1H),3.87(dd,J＝9.5,6.6Hz,1H),3.65–3.52(m,2H),3.24(s,3H),1.40(s,9H),0.83(s,9H),0.82(s,9H),MS(ESI):m/z calcd for C34H54N2O5SSi2658.33,found659.37(M+H)+。
Figure PCTCN2014090536-appb-000072
实施例58合成标记中间体41s
由中间体40s反应制得中间体41s，其反应的原料比例、溶剂、反应条件等均与实施例45相同，产率92.9％。1H NMR(400MHz,CDCl3)δ7.91(d,J＝8.6Hz,2H),7.85(d,J＝8.9Hz,1H),7.33–7.25(m,3H),7.01(dd,J＝9.0,2.5Hz,1H),4.10(dd,J＝9.5,3.5Hz,1H),4.07–3.96(m,1H),3.87(dd,J＝9.5,6.5Hz,1H),3.66–3.49(m,2H),3.23(s,3H),1.40(s,9H),0.83(s,9H),0.82(s,9H),MS(ESI):m/z calcd for C34H54N2O5SSi2658.33,found659.1(M+H)+。
Figure PCTCN2014090536-appb-000073
实施例59合成标记中间体42s
由中间体41r反应制得中间体42s，其反应的原料比例、溶剂、反应条件等均与实施例47相同，产率83.4％。1H NMR(400MHz,CDCl3)δ8.01(d,J＝8.6Hz,2H),7.96(d,J＝8.9Hz,1H),7.43–7.34(m,3H),7.10(dd,J＝8.9,2.5Hz,1H),4.22–4.09(m,3H),3.88(dd,J＝11.4,3.7Hz,1H),3.79(dd,J＝11.4,5.1Hz,1H),3.32(s,3H),1.49(s,9H),MS(ESI):m/z calcd for C22H26N2O5S430.16,found431.20(M+H)+。
Figure PCTCN2014090536-appb-000074
实施例60合成标记中间体42r
由中间体41s反应制得标记中间体42r，其反应的原料比例、溶剂、反应条件等均与实施例47相同，产率59.2％。1H NMR(400MHz,CDCl3)δ8.00(d,J＝8.7Hz,2H),7.95(d,J＝8.9Hz,1H),7.37(dd,J＝5.5,2.9Hz,3H),7.10(dd,J＝8.9,2.5Hz,1H),4.24–4.09(m,3H),3.88(dd,J＝11.4,3.6Hz,1H),3.79(dd,J＝11.3,5.1Hz,1H),3.32(s,3H),1.48(s,9H),MS(ESI):m/z calcd for C22H26N2O5S430.16,found431.21(M+H)+。
Figure PCTCN2014090536-appb-000075
实施例61合成标记中间体43r
由中间体42s反应制得中间体43r，其反应的原料比例、溶剂、反应条件等均与实施例49相同，产率14.0％。1H NMR(400MHz,CDCl3)δ7.99(d,J＝8.6Hz,2H),7.90(d,J＝8.9Hz,1H),7.79(d,J＝8.2Hz,2H),7.37(d,J＝8.5Hz,2H),7.34–7.27(m,3H),6.98(dd,J＝8.9,2.4Hz,1H),4.35–4.15(m,3H),4.05(s,2H),3.32(s,3H),2.39(s,3H),1.49(s,9H),MS(ESI):m/z calcd for C29H32N2O7S2584.17,found584.9(M+H)+。
Figure PCTCN2014090536-appb-000076
实施例62合成标记中间体43s
由中间体42r反应制得中间体43s，其反应的原料比例、溶剂、反应条件等均与实施例49相同，产率14.4％。1H NMR(400MHz,CDCl3)δ7.99(d,J＝8.7Hz,2H),7.91(d,J＝8.9Hz,1H),7.79(d,J＝8.3Hz,2H),7.37(d,J＝8.6Hz,2H),7.33–7.26(m,3H),6.98(dd,J＝8.9,2.5Hz,1H),4.35–4.16(m,3H),4.12–4.00(m,2H),3.32(s,3H),2.39(s,3H),1.49( s,9H),MS(ESI):m/z calcd for C29H32N2O7S2584.17,found584.8(M+H)+。
Figure PCTCN2014090536-appb-000077
实施例63合成标记前体44r
由中间体43r反应制得标记前体44r，其反应的原料比例、溶剂、反应条件等均与实施例51相同，产率66.8％。1H NMR(400MHz,CDCl3)δ8.00(d,J＝8.6Hz,2H),7.90(d,J＝9.0Hz,1H),7.77(dd,J＝8.2,6.2Hz,2H),7.38(d,J＝8.6Hz,2H),7.30–7.26(m,2H),7.25(s,1H),7.03–6.93(m,1H),4.77(dt,J＝6.9,3.7Hz,1H),4.44–4.15(m,3H),4.15–3.96(m,2H),3.96–3.63(m,1H),3.60–3.44(m,1H),3.32(s,3H),2.37(s,3H),1.89–1.62(m,3H),1.49(s,9H),MS(ESI):m/z calcd for C34H40N2O8S2668.22,found669.29(M+H)+。
Figure PCTCN2014090536-appb-000078
实施例64合成标记前体44s
由中间体43s反应制得标记前体44s，其反应的原料比例、溶剂、反应条件等均与实施例51相同，产率72.1％。1H NMR(400MHz,CDCl3)δ8.00(d,J＝8.5Hz,2H),7.91(d,J＝8.9Hz,1H),7.77(dd,J＝8.1,6.3Hz,2H),7.37(d,J＝8.5Hz,2H),7.32–7.26(m,2H),7.26(d,J＝3.8Hz,2H),6.96(dt,J＝8.9,3.0Hz,1H),4.88–4.69(m,1H),4.40–4.14(m,3H),4.14–3.92(m,2H),3.90–3.68(m,1H),3.59–3.43(m,1H),3.32(s,3H),2.38(s,3H),1.87–1.63(m,3H),1.48(s,9H),MS(ESI):m/z calcd for C34H40N2O8S2668.22,found669.1(M+H)+。
Figure PCTCN2014090536-appb-000079
实施例65合成标记中间体45r
由化合物2反应制得中间体45r，其反应的原料比例、溶剂、反应条件等均与实施例41相同，产率54.1％。1H NMR(400MHz,d6-DMSO)δ7.82(t,J＝9.3Hz,3H),7.61(s,1H),7.07(d,J＝8.9Hz,1H),6.81(d,J＝8.8Hz,2H),4.98(d,J＝5.2Hz,1H),4.68(s,1H),4.06(s,1H),3.95(s,1H),3.82(s,1H),3.47(s,2H),3.01(s,6H),MS(ES I):m/z calcd for C18H20N2O3S344.12,found345.2(M+H)+。
Figure PCTCN2014090536-appb-000080
实施例66合成标记中间体45s
由化合物2反应制得中间体45s，其反应的原料比例、溶剂、反应条件等均与实施例41相同，产率89.2％。1H NMR(400MHz,d6-DMSO)δ7.82(s,3H),7.62(s,1H),7.06(d,J＝9.0Hz,1H),6.81(d,J＝6.1Hz,2H),5.00(s,1H),4.70(s,1H),4.05(s,1H),3.94(s,1H),3.82(s,1H),3.47(s,2H),3.01(s,6H),MS(ESI):m/z calcd for C18H20N2O3S344.12,found345.2(M+H)+。
Figure PCTCN2014090536-appb-000081
实施例67合成标记中间体46s
由中间体45r反应制得中间体46s，其反应的原料比例、溶剂、反应条件等均与实施例49相同，产率8.8％,1H NMR(400MHz,CDCl3)δ7.92(d,J＝8.5Hz,2H),7.79(d,J＝8.3Hz,3H),7.29(d,J＝8.1Hz,2H),7.23(d,J＝2.4Hz,1H),6.93(dd,J＝8.9,2.4Hz,1H),6.75(d,J＝8.8Hz,2H),4.34–4.16(m,3H),4.04(d,J＝4.2Hz,2H),3.06(s,6H),2.39(s,3H),MS(ESI):m/z calcd for C25H26N2O5S2498.13,found499.4(M+H)+。
Figure PCTCN2014090536-appb-000082
实施例68合成标记中间体46r
由中间体45s反应制得中间体46r，其反应的原料比例、溶剂、反应条件等均与实施例49相同，产率7.0％,1H NMR(400MHz,CDCl3)δ7.92(d,J＝8.2Hz,2H),7.85(d,J＝8.8Hz,1H),7.79(d,J＝7.9Hz,2H),7.34–7.28(m,2H),7.24(s,1H),6.95(dd,J＝14.2,9.1Hz,1H),6.75(d,J＝8.5Hz,2H),4.24(s,3H),4.05(s,2H),3.06(s,6H),2.39(s,3H),MS(ESI):m/z calcd for C25H26N2O5S2498.13,found499.5(M+H)+。
Figure PCTCN2014090536-appb-000083
实施例69合成标记前体47s
由中间体46s反应制得标记前体47s，其反应的原料比例、溶剂、反应条件等均与实施例51相同，产率35.5％。1H NMR(400MHz,CDCl3)δ7.91(d,J＝8.5Hz,2H),7.83(d,J＝8.9Hz,1H),7.77(dd,J＝8.1,6.1Hz,2H),7.29(d,J＝9.0Hz,1H),7.25–7.17(m,2H),6.90(ddd,J＝8.9,4.3,2.6Hz,1H),6.75(d,J＝8.9Hz,2H),4.86– 4.70(m,1H),4.43–4.11(m,4H),4.13–3.96(m,2H),3.95–3.66(m,2H),3.65–3.40(m,2H),3.05(s,6H),2.37(d,J＝1.2Hz,3H),MS(ESI):m/z calcd for C30H34N2O6S2582.19,found582.9(M+H)+。
Figure PCTCN2014090536-appb-000084
实施例70合成标记前体47r
由中间体46r反应制得标记前体47r，其反应的原料比例、溶剂、反应条件等均与实施例57相同，产率94.0％。1H NMR(400MHz,CDCl3)δ7.92(d,J＝8.5Hz,2H),7.83(d,J＝8.8Hz,1H),7.80–7.70(m,2H),7.35(s,1H),7.25–7.14(m,2H),6.90(d,J＝7.0Hz,1H),6.75(d,J＝8.7Hz,2H),4.42–3.96(m,5H),3.97–3.68(m,3H),3.64–3.35(m,2H),3.05(s,6H),2.37(s,3H),1.89–1.38(m,4H),MS(ESI):m/z calcd for C30H34N2O6S2582.19,found583.5(M+H)+。
Figure PCTCN2014090536-appb-000085
实施例71合成标记中间体48r
由化合物21反应制得中间体48r，其反应的原料比例、溶剂、反应条件等均与实施例41相同，产率56.0％。1H NMR(400MHz,d6-DMSO)δ7.97(d,J＝9.0Hz,2H),7.57(d,J＝8.8Hz,1H),7.22(d,J＝2.4Hz,1H),6.90(dd,J＝8.8,2.5Hz,1H),6.85(d,J＝9.1Hz,2H),4.04(dt,J＝7.3,3.7Hz,1H),3.91(dd,J＝9.9,6.1Hz,1H),3.86–3.74(m,1H),3.47(d,J＝5.7Hz,2H),3.03(s,6H),MS(ESI):m/z calcd for C18H20N2O4328.14,found329.17(M+H)+。
Figure PCTCN2014090536-appb-000086
实施例72合成标记中间体48s
由化合物21反应制得中间体48s，其反应的原料比例、溶剂、反应条件等均与实施例41相同，产率52.2％。1H NMR(400MHz,d6-DMSO)δ7.96(d,J＝5.7Hz,2H),7.57(d,J＝8.4Hz,1H),7.22(s,1H),6.94–6.75(m,3H),4.96(s,1H),4.68(s,1H),4.03(s,1H),3.91(s,1H),3.81(s,1H),3.47(s,2H),3.03(s,6H),MS(ESI):m/z calcd for C18H20N2O4328.14,found329.3(M+H)+。
Figure PCTCN2014090536-appb-000087
实施例73合成标记中间体49s
由化合物48r反应制得中间体49s，其反应的原料比例、溶剂、反应条件等均与实施例49相同，产率7.8％。1H NMR(400MHz,CDCl3)δ8.07(d,J＝9.0Hz,2H),7.78(d,J＝8.3Hz,2H),7.36(d,J＝8.8Hz,1H),7.29(d,J＝8.1Hz,2H),7.10(d,J＝2.5Hz,1H),6.80–6.70(m,3H),4.35–4.18(m,3H),4.01(d,J＝4.6Hz,2H),3.06(s,6H),2.39(s,3H),MS(ESI):m/z calcd for C25H26N2O6S482.15,found482.8(M+H)+。
Figure PCTCN2014090536-appb-000088
实施例74合成标记中间体49r
由化合物48s反应制得中间体49r，其反应的原料比例、溶剂、反应条件等均与实施例49相同，产率25.8％。1H NMR(400MHz,CDCl3)δ8.05(d,J＝9.0Hz,2H),7.76(d,J＝8.3Hz,2H),7.33(d,J＝8.8Hz,1H),7.26(s,2H),7.08(d,J＝2.4Hz,1H),6.77–6.65(m,3H),4.35–4.17(m,3H),4.09–3.92(m,2H),3.04(s,6H),2.36(s,3H),MS(ESI):m/z calcd for C25H26N2O6S482.15,found483.4(M+H)+。
Figure PCTCN2014090536-appb-000089
实施例75合成标记前体50s
由化合物49s反应制得标记前体50s，其反应的原料比例、溶剂、反应条件等均与实施例51相同，产率83.8％。1H NMR(400MHz,CDCl3)δ8.07(d,J＝8.7Hz,2H),7.80–7.75(m,2H),7.35(d,J＝8.8Hz,1H),7.27(s,1H),7.26–7.24(m,1H),7.07(dd,J＝8.4,2.4Hz,1H),6.76(d,J＝8.7Hz,2H),4.90–4.70(m,1H),4.40–4.10(m,3H),4.10–3.90(m,2H),3.90–3.70(m,1H),3.60–3.40(m,1H),3.07(s,6H),2.37(d,J＝2.6Hz,3H),1.60–1.40(m,6H),MS(ESI):m/z calcd for C30H34N2O7S566.21,found566.9(M+H)+。
Figure PCTCN2014090536-appb-000090
实施例76合成标记中间体50r
由化合物49r反应制得标记前体50r，其反应的原料比例、溶剂、反应条件等均与实施例51相同，产率91.2％。1H NMR(400MHz,CDCl3)δ8.09(d,J＝8.8Hz,2H),7.78(dd,J＝ 8.2,5.2Hz,2H),7.36(d,J＝8.8Hz,1H),7.30–7.26(m,1H),7.27–7.20(m,2H),7.09(dd,J＝7.6,2.4Hz,1H),6.78(d,J＝8.9Hz,2H),4.88–4.70(m,1H),4.44–4.14(m,4H),3.96–3.75(m,5H),3.61–3.44(m,4H),3.08(s,6H),2.38(d,J＝2.3Hz,3H),MS(ESI):m/z calcd for C30H34N2O7S566.21,found567.5(M+H)+。
Figure PCTCN2014090536-appb-000091
实施例77标记化合物[18F]14r
称取2mg标记前体44r，加入1.7mL乙腈溶解。将其加入到已除水的含一定活度的含K222/K2CO3的18F-的反应管中，在100℃的条件下标记12min，冷却后加入0.20mL的HCl(1M)，涡旋后继续反应5min，冷却后加入少量水用NaHCO3中和至偏碱性，最后过C18反相小柱子用水冲洗除去盐和剩余的18F-，之后用乙腈淋洗得到最终标记产品，N2吹干后，用HPLC分离得到纯度大于98％的标记物[18F]14b。标记率约为5-40％。HPLC分离条件为：Waters，C-18半制备柱(10×250mm，5μm)；流动相：乙腈:水＝3:2(体积比)。保留时间为9.09min。
实施例78标记化合物[18F]14s
由标记前体44s反应制得[18F]14s，其反应的原料比例、溶剂、反应条件等均与实施例77相同，标记率约为5-40％。HPLC分离条件为：Waters，C-18半制备柱(10×250mm，5μm)；流动相：乙腈:水＝＝3:2(体积比)。保留时间为9.09min。
实施例79标记化合物[18F]15r
由标记前体47r反应制得[18F]15r，其反应的原料比例、溶剂、反应条件等均与实施例77相同，标记率约为5-40％。HPLC分离条件为：Waters，C-18半制备柱(10×250mm，5μm)；流动相：乙腈:水＝3:2(体积比)。保留时间为15.23min。
实施例80标记化合物[18F]15s
由标记前体47s反应制得[18F]15s，其反应的原料比例、溶剂、反应条件等均与实施例77相同，标记率约为5-40％。HPLC分离条件为：Waters，C-18半制备柱(10×250mm，5μm)；流动相：乙腈:水＝3:2(体积比)。保留时间为15.23min。
实施例81标记化合物[18F]28r
由标记前体38r反应制得[18F]28r，其反应的原料比例、溶剂、反应条件等均与实施例77相同，标记率约为5-40％。HPLC分离条件为：Waters，C-18半制备柱(10×250mm，5μm)；流动相：乙腈:水＝3:2(体积比)。保留时间为7.95min。
实施例82标记化合物[18F]28s
由标记前体38s反应制得[18F]28s，其反应的原料比例、溶剂、反应条件等均与实施例77相同，标记率约为5-40％。HPLC分离条件为：Waters，C-18半制备柱(10×250mm，5μm)；流动相：乙腈:水＝＝3:2(体积比)。保留时间为7.95min。
实施例83标记化合物[18F]29r
由标记前体50r反应制得[18F]29r，其反应的原料比例、溶剂、反应条件等均与实施例77相同，标记率约为5-40％。HPLC分离条件为：Waters，C-18半制备柱(10×250mm， 5μm)；流动相：乙腈:水＝＝3:2(体积比)。保留时间为12.58min。
实施例84标记化合物[18F]29s
由标记前体50s反应制得[18F]29s，其反应的原料比例、溶剂、反应条件等均与实施例77相同，标记率约为5-40％。HPLC分离条件为：Waters，C-18半制备柱(10×250mm，5μm)；流动相：乙腈:水＝＝3:2(体积比)。保留时间为12.58min。
实施例85标记化合物的效果实验
以下通过竞争结合实验和放射自显影实验对本发明所述化合物进行亲和力评价，充分验证它们与Aβ斑块的高亲和力。通过正常小鼠体内生物分布实验对标记化合物进行初始脑摄取及脑清除评价。
1.竞争结合实验
竞争结合实验(Ki测定)：一定浓度的Aβ1-42聚集体蛋白与一定浓度的放射性配基[125I]IMPY起结合反应，反应***中同时加不同浓度的待测化合物(分别为本发明实施例中的目标化合物与[125I]IMPY起竞争反应，平衡后分离复合物通过测定放射性来计算抑制常数(Ki)。
1.1实验步骤：
(1)配制pH＝7.4的PBS(0.2M)缓冲液4L；
(2)放射配基[125I]IMPY按照已有方法制备。将[125I]IMPY配制成100000cpm/100μL的水溶液；
(3)将待测化合物配制成10-3至10-9mol/L的一系列乙醇溶液；
(4)受体Aβ1-42蛋白按照已有方法制备。将其稀释成约30nM的水溶液；
(5)玻璃纤维滤膜用含0.1％(体积分数)聚乙烯亚胺的PBS溶液浸泡0.5h；
(6)在12×75mm硼硅玻璃管中分别加入100μL不同浓度待测化合物溶液和100μL[125I]IMPY溶液、700μL PBS及100μL Aβ1-42溶液。用封口膜封好，涡旋；
(7)在37℃恒温水浴中振荡孵育2h；
(8)多头细胞收集器收集反应液，用PBS冲洗三遍，每次3mL；
(9)用γ计数仪测量计数；
(10)数据处理。
1.2实验结果：
由竞争结合实验得到的半抑制常数(IC50)以及进一步根据公式计算出的抑制常数见表1。
表1标记化合物的抑制常数
Figure PCTCN2014090536-appb-000092
2.正常小鼠体内分布实验
通过体内分布实验研究了在小鼠体内的药代动力学性质，特别是初始脑摄取和脑清除情况。
2.1实验步骤
将5-10μCi标记化合物(100μL生理盐水溶液，含5％乙醇)由尾静脉注射入正常小鼠(ICR，雄性，20-22g)体内(n＝5)，分别于注射后2分钟、10分钟、30分钟和60分钟将其断头处死，解剖取出相关脏器，测量湿量及放射性计数。数据表示为脏器中放射性百分剂量(％ID/器官)和每克脏器中放射性百分剂量(％ID/g)。
2.2实验结果
实验结果如表2所示，本发明所述的18F标记化合物都可以顺利的通过血脑屏障，2分钟进脑量都较高，特别是[18F]29s的初始脑摄取较高而且其在正常小鼠脑部清除很快，2分钟与60分钟脑摄取比为27.8。
表2.18F标记化合物在正常小鼠体内生物分布实验(％ID/g)
Figure PCTCN2014090536-appb-000093
虽然，上文中已经用一般性说明及具体实施方案对本发明作了详尽的描述，但在本发明基础上，可以对之作一些修改或改进，这对本领域技术人员而言是显而易见的。因此，在不偏离本发明精神的基础上所做的这些修改或改进，均属于本发明要求保护的范围。

Claims (4)

  1. 与Aβ斑块具有亲和力的含手性侧链取代的氟代2-芳基苯并杂环化合物，其特征在于，所述化合物的结构通式如式(I)所示：
    Figure PCTCN2014090536-appb-100001
    其中，
    X为N；Y为S或O；Z为N或CH；
    R1为5位或6位取代基，R1为
    Figure PCTCN2014090536-appb-100002
    F为19F或18F；
    R2为NHCH3或N(CH3)2。
  2. 权利要求1所述的化合物的制备方法，其特征在于，当R1中F为19F时，所述化合物的制备方法包括以下步骤：
    (1)将1mmol
    Figure PCTCN2014090536-appb-100003
    与1.5mmol
    Figure PCTCN2014090536-appb-100004
    以及3mmolK2CO3和催化剂量的18-crown-6溶于30mL无水丙酮中，在80℃油浴条件下搅拌回流反应；反应完毕后，除去溶剂丙酮，经柱层析分离得到如式(II)或式(III)所示的化合物：
    Figure PCTCN2014090536-appb-100005
    (2)将0.2mmol式(II)或式(III)所示化合物溶于15mL甲苯中，向其中加入含1MTBAF的THF溶液1mL，在80℃油浴条件下搅拌回流反应；反应完毕后，除去溶剂，用CH2Cl2萃取后，最后经柱层析分离，得到如式(IV)或式(V)所示的化合物：
    Figure PCTCN2014090536-appb-100006
    其中，X为N；Y为S或O；Z为N或CH；R为NHCH3或N(CH3)2。
  3. 权利要求1所述的化合物的制备方法，其特征在于，当R1中F为18F，且R2为N(CH3)2时，所述化合物的制备方法包括以下步骤：
    (1)将1mmol
    Figure PCTCN2014090536-appb-100007
    溶于30mL无水乙醇中，向其中加入200g/L NaOH溶液0.3mL，在80℃油浴条件下搅拌回流反应1h后，向其中加入1.5mmol
    Figure PCTCN2014090536-appb-100008
    反应完毕后，除去溶剂，经柱层析分离得到如式(VI)或式(VII)所示的化合物：
    Figure PCTCN2014090536-appb-100009
    (2)当R为NHCH3时，将1mmol式(VI)或式(VII)所示化合物、8mmol咪唑以及8mmol叔丁基二甲基氯硅烷溶于30mL CH2Cl2溶液中，在40℃油浴条件下搅拌回流反应5h，反应完毕后除去CH2Cl2，经柱层析分离得到如式(VIII)或式(IX)所示的化合物：
    Figure PCTCN2014090536-appb-100010
    (3)将1mmol式(VIII)或式(IX)所示化合物溶于30mL THF溶液中，向其中加入过量的(Boc)2O,在80℃油浴条件下搅拌回流反应过夜，反应完毕后除去THF，经柱层析分离得到如式(X)或式(XI)所示的化合物：
    Figure PCTCN2014090536-appb-100011
    (4)将1mmol式(X)或式(XI)所示化合物溶于30mL THF溶液中，向其中加入7mmol TBAF，在30℃油浴条件下搅拌回流反应过夜，反应完毕后除去THF，经柱层析分离得到如式(XII)或式(XIII)所示的化合物：
    Figure PCTCN2014090536-appb-100012
    (5)将1mmol式(VI)或式(VII)所示化合物以及1mmol式(XII)或式(XIII)所示化合物分别溶于5mL吡啶溶液中，向其中加入1.5mmol TsCl，在0℃冰浴条件下搅拌反应，反应完毕后除去吡啶，经柱层析分离得到如式(XIV)或式(XV)所示的化合物：
    Figure PCTCN2014090536-appb-100013
    其中，R为N(CH3)2；
    (6)将1mmol式(XIV)或式(XV)所示化合物、4mmol3,4-dihydro-2H-pyran以及0.2mmol PPTS分别溶于30mL CH2Cl2溶液中，在40℃油浴条件下搅拌回流反应过夜，反应完毕后除去CH2Cl2，经柱层析分离得到如式(XVI)或式(XVII)所示的化合物：
    Figure PCTCN2014090536-appb-100014
    (7)将10mg式(XVI)或式(XVII)所示化合物加入2mL乙腈溶解。将其加入到已除水的具有一定活度的含K222/K2CO3的18F-的反应管中，于100℃标记12min，冷却后加入1M HCl0.15mL，涡旋后继续反应5min中，冷却后加入少量水用NaHCO3中和至偏碱性，最后过C18反相柱用水冲洗除去盐和剩余的18F-，然后用乙腈淋洗，N2吹干后，用HPLC分离得到如式(XVI)或式(XVII)所示的化合物：
    Figure PCTCN2014090536-appb-100015
  4. 权利要求1所述化合物在制备Aβ斑块显像剂中的应用，其中式(I)所示的化合物中，F为18F。
PCT/CN2014/090536 2014-06-06 2014-11-07 与Aβ斑块具有亲和力的含手性侧链取代的氟代2-芳基苯并杂环化合物、其制备方法及应用 WO2015184731A1 (zh)

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