CN104231023A - Tricyclic fused-heterocyclic nucleoside phosphoramidate compound and preparation method and applications of tricyclic fused-heterocyclic nucleoside phosphoramidate compound - Google Patents

Tricyclic fused-heterocyclic nucleoside phosphoramidate compound and preparation method and applications of tricyclic fused-heterocyclic nucleoside phosphoramidate compound Download PDF

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CN104231023A
CN104231023A CN201410245982.0A CN201410245982A CN104231023A CN 104231023 A CN104231023 A CN 104231023A CN 201410245982 A CN201410245982 A CN 201410245982A CN 104231023 A CN104231023 A CN 104231023A
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alkylamino
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hydroxyl
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CN104231023B (en
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王勇
赵立文
毕胜
高毅平
王德忠
陈宏雁
刘阳
张仓
张文萍
南阳
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Nanjing Huicheng Pharmaceutical Co., Ltd.
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Nanjing Sanhome Pharmaceutical Co Ltd
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Abstract

The invention provides a tricyclic fused-heterocyclic nucleoside phosphoramidate compound, a preparation method thereof and a composition comprising the tricyclic fused-heterocyclic nucleoside phosphoramidate compound, and applications of the compound or the composition as a medicament for treating virus infected diseases, especially an application of a medicament for treating viral hepatitis.

Description

Three ring fused heterocycle type nucleoside phosphoramidate compounds, its preparation method and application
Technical field
The invention belongs to medicinal chemistry arts, be specifically related to a class three ring fused heterocycle type nucleoside phosphoramidate compound, its preparation method, composition containing this nucleoside phosphoramidate compound, and described compound or composition are as the purposes of disease of viral infection medicine, particularly as the purposes of viral hepatitis treatment medicine.
Background technology
Hepatitis C virus (HCV) is the main pathogens causing most of non-A non-B hepatitis.Infection with hepatitis C virus causes chronic hepatopathy, as the major health concern of liver cirrhosis and liver cancer.According to the data of the World Health Organization, there is the infected population more than 200,000,000 in the whole world, once infect, only have an appointment 20% people can remove virus, others will carry HCV.
HCV is a kind of virus belonging to flaviviridae, and being has enveloped positive strand RNA viruses in flaviviridae family.Length about 9500 Nucleotide of strand HCV rna gene group, containing single open reading frame (ORF), about 3000 amino acid whose large polymeric proteins of encoding.
HCV and two other genus in flaviviridae and pestivirus similar with the gene structure of Flavivirus.At present, treat the standard method of HCV infection and have Interferon, rabbit and Interferon, rabbit and ribavirin combination therapy.But, only the curer of 50% responds to the method, and Interferon, rabbit has obvious side effect, and such as flu-like symptoms, body weight lower and fatigue and weak, Interferon, rabbit and ribavirin combination therapy then produce sizable side effect, comprise haemolysis, anemia and tired.
For a long time, owing to lacking the cell model of suitable external virus replication, the anti-HCV medicament of development of new is made slow progress.Until 1999, Lohmann establishes in liver cancer cell can the sub-genome duplication model (replicon) of high-level self-replicating, this model is selectivity bicistronic mRNA subgene group HCV rna replicon subsystem, and it is acknowledged as the milestone that the model research of HCV RNA replication in vitro obtains breakthrough development.Replicon can high-caliber self-replicating in human hepatoma cell strain Huh7 cell strain.Research finds that replicon copies in cell and has no significant effect host cell growth and metabolism, and high level copies relevant with the receptivity of virus genomic adaptive mutation and host cell.These results of study are that the research of selectivity HCV complete sequence subgenomic replicons is laid a good foundation.
Summary of the invention
One object of the present invention is three ring fused heterocycle type nucleoside phosphoramidate compounds, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, the solvate shown in the general formula (I) that are provided for treating and/or preventing HCV infection
P* represents chiral phosphorus atom,
Wherein, (1) R 1be selected from H and alkyl, described alkyl is optionally replaced by one or more alkyl, alkoxyl group, alkylamino, halogen, hydroxyl, amino, nitro, cyano group, alkyl acyl, aminoacyl, alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl;
(2) R 2be selected from H, alkyl, aminoacyl, aryl and heteroaryl, described alkyl is optionally replaced by one or more alkyl, alkoxyl group, halogen, hydroxyl, amino, single alkylamino, two alkylamino, aryl or heteroaryl;
(3) R 3for not existing, or be selected from alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, alkylamino, alkyl sulphonyl, alkyl sulfonyl amino, sulfamoyl alkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, amido, ester group ,-CN, cycloalkyl, Heterocyclylalkyl, aryl, heteroaryl, aralkyl;
(4) D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-C (R c1r c2the R that)-, is wherein said a1, R a2be hydrogen, alkyl, halogen, haloalkyl, thiazolinyl or haloalkenyl group separately, or R a1, R a2form cycloalkyl together with the C that they connect, described m is 0 or 1, described R bfor not existing, hydrogen, alkyl, alkyl sulphonyl or alkyl-carbonyl, described R c1, R c2be hydrogen, halogen, alkyl, haloalkyl, thiazolinyl or haloalkenyl group separately, or work as R c1, R c2when being alkyl, C can with R c1, R c2form cycloalkyl;
(5) D 1be selected from oxygen, sulphur ,-SO-,-SO 2-,-CO-,-N (R d)-,-(CH) n-, wherein said R dfor not existing, hydrogen, alkyl or haloalkyl, described n is 0,1 or 2, described-(CH) n-optional alkoxy, alkylamino, halogen, hydroxyl, amino, nitro, cyano group, alkyl acyl, aminoacyl, alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl replaces;
(6) G, G 1be-N-or-CH-separately;
(7) for singly-bound or double bond, wherein two be singly-bound, or one for singly-bound, and another is double bond; With
(8) ring T is selected from:
A, aromatic ring or hetero-aromatic ring, described aromatic ring or hetero-aromatic ring can by R 4replace, wherein R 4for alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, alkyl acyl, aminoacyl, alkylamino acyl group, amido ,-CN, aryl or heteroaryl; With
B, naphthenic hydrocarbon or heterocycloalkane, described naphthenic hydrocarbon or heterocycloalkane can by R 4replace, wherein R 4for alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, alkyl acyl, aminoacyl, alkylamino acyl group, amido ,-CN, aryl or heteroaryl.Another object of the present invention is to provide the method for the nucleoside phosphoramidate compound of preparation general formula of the present invention (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization.
Another object of the present invention be providing package contain the nucleoside phosphoramidate compound of general formula of the present invention (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate or solvate and drug effect acceptable carrier composition and comprise the composition of the nucleoside phosphoramidate compound of general formula of the present invention (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate or solvate and another kind of antiviral drug.
An also object of the present invention is the nucleoside phosphoramidate compound providing general formula of the present invention (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate treats and/or prevents the method for infection with hepatitis C virus and the nucleoside phosphoramidate compound of general formula of the present invention (I) and steric isomer thereof, salt, hydrate, solvate or crystallization are for the preparation of the application treated and/or prevented in the medicine of infection with hepatitis C virus.
A further object of the present invention is to provide and suppresses RNA RNA-dependent varial polymerases, particularly suppresses the method for HCVNS5B polysaccharase.
The present invention also aims to be provided in treatment RNA RNA-dependent virus replication, particularly treat HCV copy in application.
Another object of the present invention is also to be provided for treatment RNA RNA-dependent virus infection, particularly treats the application in HCV infection.
For foregoing invention object, the invention provides following technical scheme:
First aspect, the invention provides a kind of three ring fused heterocycle type nucleoside phosphoramidate compounds, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, described compound is as shown in general formula (I):
P* represents chiral phosphorus atom,
Wherein,
(1) R 1be selected from H and alkyl, described alkyl is optionally replaced by one or more alkyl, alkoxyl group, alkylamino, halogen, hydroxyl, amino, nitro, cyano group, alkyl acyl, aminoacyl, alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl;
(2) R 2be selected from H, alkyl, aminoacyl, aryl and heteroaryl, described alkyl is optionally replaced by one or more alkyl, alkoxyl group, halogen, hydroxyl, amino, single alkylamino, two alkylamino, aryl or heteroaryl;
(3) R 3for not existing, or be selected from alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, alkylamino, alkyl sulphonyl, alkyl sulfonyl amino, sulfamoyl alkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, amido, ester group ,-CN, cycloalkyl, Heterocyclylalkyl, aryl, heteroaryl, aralkyl;
(4) D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-C (R c1r c2the R that)-, is wherein said a1, R a2be hydrogen, alkyl, halogen, haloalkyl, thiazolinyl or haloalkenyl group separately, or R a1, R a2form cycloalkyl together with the C that they connect, described m is 0 or 1, described R bfor not existing, hydrogen, alkyl, alkyl sulphonyl or alkyl-carbonyl, described R c1, R c2be hydrogen, halogen, alkyl, haloalkyl, thiazolinyl or haloalkenyl group separately, or work as R c1, R c2when being alkyl, C can with R c1, R c2form cycloalkyl;
(5) D 1be selected from oxygen, sulphur ,-SO-,-SO 2-,-CO-,-N (R d)-,-(CH) n-, wherein said R dfor not existing, hydrogen, alkyl or haloalkyl, described n is 0,1 or 2, described-(CH) n-optional alkoxy, alkylamino, halogen, hydroxyl, amino, nitro, cyano group, alkyl acyl, aminoacyl, alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl replaces;
(6) G, G 1be-N-or-CH-separately;
(7) for singly-bound or double bond, wherein two be singly-bound, or one for singly-bound, and another is double bond; With
(8) ring T is selected from:
A, aromatic ring or hetero-aromatic ring, described aromatic ring or hetero-aromatic ring can by R 4replace, wherein R 4for alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, alkyl acyl, aminoacyl, alkylamino acyl group, amido ,-CN, aryl or heteroaryl; With
B, naphthenic hydrocarbon or heterocycloalkane, described naphthenic hydrocarbon or heterocycloalkane can by R 4replace, wherein R 4for alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, alkyl acyl, aminoacyl, alkylamino acyl group, amido ,-CN, aryl or heteroaryl.
In some embodiments of first aspect, the invention provides the nucleoside phosphoramidate compound of above-mentioned general formula (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein
D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-C (R c1r c2the R that)-, is wherein said a1, R a2be hydrogen, C separately 1-6alkyl, halogen, halo C 1-6alkyl, C 2-6thiazolinyl or halo C 2-6thiazolinyl, or R a1, R a2c is formed together with the C that they connect 3-8cycloalkyl, described m is 0 or 1, described R bfor not existing, hydrogen, C 1-6alkyl, C 1-6alkyl sulphonyl or C 1-6alkyl-carbonyl, described R c1, R c2be hydrogen, halogen, C separately 1-6alkyl, halo C 1-6alkyl, C 2-6thiazolinyl or halo C 2-6thiazolinyl, or work as R c1, R c2when being alkyl, C can with R c1, R c2form C 3-8cycloalkyl;
D 1be selected from oxygen, sulphur ,-SO-,-SO 2-,-CO-,-N (R d)-,-(CH) n-, wherein said R dfor not existing, hydrogen, C 1-6alkyl, halo C 1-6alkyl, described n is 0,1 or 2, described-(CH) n-optionally by C 1-6alkoxyl group, C 1-6alkylamino, halogen, hydroxyl, amino, nitro, cyano group, C 1-6alkyl acyl, aminoacyl, C 1-6alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl replace; With
Ring T is selected from:
A, C 5-10aromatic ring or hetero-aromatic ring, described aromatic ring or hetero-aromatic ring can by R 4replace, wherein R 4for C 1-6alkyl, C 2-6thiazolinyl, C 2-6alkynyl, C 1-6alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino, C 1-6alkyl acyl, aminoacyl, C 1-6alkylamino acyl group, amido ,-CN, aryl or heteroaryl; With
B, C 3-10naphthenic hydrocarbon or heterocycloalkane, described naphthenic hydrocarbon or heterocycloalkane can by R 4replace, wherein R 4for C 1-6alkyl, C 2-6thiazolinyl, C 2-6alkynyl, C 1-6alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino, C 1-6alkyl acyl, aminoacyl, C 1-6alkylamino acyl group, amido ,-CN, aryl or heteroaryl.
In other embodiments of first aspect, the invention provides the nucleoside phosphoramidate compound of above-mentioned general formula (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein
R 1be selected from H and C 1-6alkyl, described alkyl is optionally by one or more C 1-6alkyl, C 1-6alkoxyl group, C 1-6alkylamino, halogen, hydroxyl, amino, nitro, cyano group, C 1-6alkyl acyl, aminoacyl, C 1-6alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl replace;
R 2be selected from H, C 1-6alkyl, aminoacyl, aryl and heteroaryl, described alkyl is optionally by one or more C 1-6alkyl, C 1-6alkoxyl group, halogen, hydroxyl, amino, single C 1-6alkylamino, two C 1-6alkylamino, aryl or heteroaryl replace; With
R 3for not existing, or be selected from C 1-6alkyl, C 2-6thiazolinyl, C 2-6alkynyl, C 1-6alkoxyl group, halogen, halo C 1-6alkyl, C 1-6alkylamino, C 1-6alkyl sulphonyl, C 1-6alkyl sulfonyl is amino, sulfamyl C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino, amido, ester group ,-CN, C 3-8cycloalkyl, C 3-8heterocyclylalkyl, aryl, heteroaryl, fragrant C 1-6alkyl.
In other embodiments of first aspect, the invention provides the nucleoside phosphoramidate compound of above-mentioned general formula (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein
(1) R 1be selected from H and C 1-6alkyl, described alkyl is optionally by one or more C 1-6alkoxyl group, C 1-6alkylamino, halogen, hydroxyl, amino, nitro, cyano group, C 1-6alkyl acyl, aminoacyl, C 1-6alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl replace;
(2) R 2be selected from H, C 1-6alkyl, aminoacyl, aryl and heteroaryl, described alkyl is optionally by one or more C 1-6alkoxyl group, halogen, hydroxyl, amino, single C 1-6alkylamino, two C 1-6alkylamino, aryl or heteroaryl replace;
(3) R 3for not existing, or be selected from C 1-6alkyl, C 1-6alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino, amido, ester group ,-CN, C 3-8cycloalkyl, assorted C 3-8cycloalkyl, aryl, heteroaryl, fragrant C 1-6alkyl;
(4) D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-C (R c1r c2the R that)-, is wherein said a1, R a2be hydrogen, C separately 1-6alkyl, halogen, halo C 1-6alkyl, C 2-6thiazolinyl or halo C 2-6thiazolinyl, or R a1, R a2c is formed together with the C that they connect 3-8cycloalkyl, described m is 0 or 1, described R bfor not existing, hydrogen, alkyl, alkyl sulphonyl or alkyl-carbonyl, described R c1, R c2be hydrogen, halogen, alkyl, thiazolinyl or haloalkenyl group separately, or work as R c1, R c2when being alkyl, C can with R c1, R c2form cycloalkyl;
(5) D 1be selected from oxygen, sulphur ,-SO-,-SO 2-,-CO-,-N (R d)-,-(CH) n-, described R dfor not existing, hydrogen, C 1-6alkyl or halo C 1-6alkyl, described n is 0,1 or 2, described-(CH) n-optionally by C 1-6alkoxyl group, C 1-6alkylamino, halogen, hydroxyl, amino, nitro, cyano group, C 1-6alkyl acyl, aminoacyl, C 1-6alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl replace;
(6) G, G 1be-N-or-CH-separately;
(7) for singly-bound or double bond, wherein two be singly-bound, or one for singly-bound, and another is double bond, and
A, as G, G 1be-CH-, and G and G 1be connected during for double bond, D and G is connected for singly-bound;
B, when G be-CH-,-N-, G 1for-N-time, G and G 1be connected for singly-bound, D and G is connected for singly-bound; Just when G is-CH-, G 1for-N-, and when D is-N-, G and G 1be connected for singly-bound, D and G is connected for double bond;
C, when G be N, G 1for-CH-time, G and G 1be connected for singly-bound, D and G is connected for singly-bound;
(8) ring T is selected from:
A, five yuan and hexa-atomic aromatic ring or hetero-aromatic ring, described five yuan and hexa-atomic aromatic ring or hetero-aromatic ring can by R 4replace, wherein R 4for alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, alkyl acyl, aminoacyl, alkylamino acyl group, amido ,-CN, aryl or heteroaryl; With
B, C 3-8naphthenic hydrocarbon or heterocycloalkane, described naphthenic hydrocarbon or heterocycloalkane can by R 4replace, wherein R 4for C 1-6alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, alkyl acyl, aminoacyl, alkylamino acyl group, amido ,-CN, aryl or heteroaryl.
In some preferred embodiments, the invention provides three ring fused heterocycle type nucleoside phosphoramidate compounds of general formula (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein
R 1be selected from H and C 1-6alkyl;
R 2be selected from C 1-6alkyl, benzyl;
R 3for not existing, or be selected from C 1-6alkyl, C 1-6alkoxyl group, fluorine, chlorine, bromine, halo C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino, amido, ester group ,-CN, C 3-6cycloalkyl, C 3-6heterocyclylalkyl, aryl, heteroaryl, aralkyl;
D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-C (R c1r c2the R that)-, is wherein said a1, R a2be hydrogen or C separately 1-6alkyl; Described m is 0 or 1; Described R bfor not existing, hydrogen, C 1-6alkyl, C 1-6alkyl sulphonyl or C 1-6alkyl-carbonyl; Described R c1, R c2be hydrogen, halogen, C separately 1-6alkyl, C 2-6thiazolinyl or halo C 2-6thiazolinyl, or work as R c1, R c2all be selected from C 1-3during alkyl, C can with R c1, R c2form C 3-7cycloalkyl;
D 1be selected from oxygen, sulphur ,-SO-,-SO 2-,-CO-,-N (R d)-,-(CH) n-, described R dfor not existing, hydrogen, C 1-6alkyl; Described n is 0,1 or 2;
G, G 1be-N-or-CH-separately; And
Ring T is selected from:
(a), at least containing heteroatomic five yuan of hetero-aromatic rings or six-membered Hetero-aromatic or phenyl ring, described five yuan or six-membered Hetero-aromatic or phenyl ring can by R 4replace, wherein R 4for C 1-6alkyl, C 1-6alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino, amido or-CN; With
(b), C 3-8naphthenic hydrocarbon or heterocycloalkane, described naphthenic hydrocarbon or heterocycloalkane can by R 4replace, wherein R 4for C 1-6alkyl, C 1-6alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino, amido or-CN.
In other preferred embodiments, the invention provides three ring fused heterocycle type nucleoside phosphoramidate compounds of general formula (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein
R 1be selected from H and C 1-4alkyl;
R 2be selected from methyl, ethyl, sec.-propyl, n-propyl, cyclopropyl, cyclopentyl, benzyl;
R 3for not existing, or be selected from C 1-4alkyl, C 1-4alkoxyl group, fluorine, chlorine, bromine and phenyl;
D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-C (R c1r c2the R that)-, is wherein said a1, R a2be hydrogen, methyl or ethyl separately; Described m is 0 or 1; Described R bfor not existing, hydrogen, methyl, ethyl, propyl group, sec.-propyl, cyclopropyl, methyl sulphonyl or methyl carbonyl; Described R c1, R c2be hydrogen, fluorine, methyl, ethyl, C separately 2-4thiazolinyl or halo C 2-4thiazolinyl, or work as R c1, R c2when being methyl, C and R c1, R c2form cyclopropyl;
D 1be selected from oxygen, sulphur ,-SO-,-SO 2-,-CO-,-N (R d)-,-(CH) n-, wherein said R dfor not existing, hydrogen or C 1-4alkyl; Described n is 0 or 1;
G, G 1be N or-CH-separately; And
Ring T is selected from:
(a), at least containing the heteroatomic five yuan of hetero-aromatic rings of N, O or S or six-membered Hetero-aromatic or phenyl ring, described five yuan or six-membered Hetero-aromatic or phenyl ring can by R 4replace, wherein R 4for C 1-6alkyl, C 1-6alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino, formamido group, kharophen or-CN; With
(b), pentamethylene, hexanaphthene, suberane or at least containing the heteroatomic heterocycle pentane of N, O or S, azacyclohexane, a trioxepane.
In some further preferred embodiments, the invention provides three ring fused heterocycle type nucleoside phosphoramidate compounds of general formula (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein R 1be selected from H and C 1-6alkyl; R 2be selected from C 1-6alkyl, benzyl; R 3for not existing, or be selected from C 1-6alkyl, C 1-6alkoxyl group, fluorine, chlorine, bromine, halo C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino ,-CN; D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-(CR c1r c2the R that)-, is wherein said a1, R a2be hydrogen or C separately 1-6alkyl, described m is 0 or 1, described R bfor not existing, hydrogen, C 1-6alkyl, C 1-6alkyl sulphonyl or C 1-6alkyl-carbonyl, described R c1, R c2be hydrogen, halogen, C separately 1-6alkyl, C 2-6thiazolinyl or halo C 2-6thiazolinyl, or work as R c1, R c2all be selected from C 1-3during alkyl, C and R c1, R c2form C 3-7cycloalkyl; D 1be selected from oxygen, sulphur ,-SO-,-SO 2-,-CO-,-NR d-,-(CH) n-, wherein said R dfor not existing, hydrogen or C 1-6alkyl, described n is 0,1 or 2; G, G 1be-N-or-CH-separately; And ring T is selected from: (a), at least containing heteroatomic five yuan of hetero-aromatic rings or six-membered Hetero-aromatic, phenyl ring, wherein said five yuan or six-membered Hetero-aromatic or phenyl ring can by R 4replace, wherein R 4for C 1-6alkyl, C 1-6alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino, amido or-CN; (b) C 3-8naphthenic hydrocarbon or heterocycloalkane, described naphthenic hydrocarbon or heterocycloalkane can by R 4replace, wherein R 4for R 4for C 1-6alkyl, C 1-6alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino, amido or-CN.
In some further preferred embodiments, the invention provides three ring fused heterocycle type nucleoside phosphoramidate compounds of general formula (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein R 1be selected from H and C 1-4alkyl; R 2be selected from methyl, ethyl, sec.-propyl, n-propyl, cyclopropyl, cyclopentyl, benzyl; R 3for not existing, or be selected from C 1-4alkyl, C 1-4alkoxyl group, fluorine, chlorine, bromine; D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-C (R c1r c2the R that)-, is wherein said a1, R a2be hydrogen, methyl or ethyl separately, described m is 0 or 1, described R bfor not existing, hydrogen, methyl, ethyl, propyl group, sec.-propyl, cyclopropyl, methyl sulphonyl or methyl carbonyl, described R c1, R c2be hydrogen, fluorine, methyl, ethyl, C separately 2-4thiazolinyl or halo C 2-4thiazolinyl, or work as R c1, R c2when being methyl, C and R c1, R c2form cyclopropyl; D 1be selected from oxygen, sulphur ,-SO-,-SO 2-,-CO-,-N (R d)-,-(CH) n-, wherein said R dfor not existing, hydrogen or C 1-4alkyl, described n is 0 or 1; G, G 1be-N-or-CH-separately; And ring T is selected from: (a), at least containing the heteroatomic five yuan of hetero-aromatic rings of N, O or S or six-membered Hetero-aromatic, phenyl ring, described five yuan or six-membered Hetero-aromatic or phenyl ring can by R 4replace, wherein R 4for C 1-6alkyl, C 1-6alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino, formamido group, kharophen or-CN; (b), pentamethylene, hexanaphthene, suberane or at least containing the heteroatomic heterocycle pentane of N, O or S, hexanaphthene, a suberane.
According to a first aspect of the invention, in some preferred embodiments, D is worked as 1for-(CH) n-, wherein n is 0, i.e. D 1when not existing, D, G, G 1the phenyl ring be connected with them forms benzo five-membered or six-ring, as shown in general formula (II)
Wherein
(1) R 1be selected from H and C 1-6alkyl, described alkyl is optionally replaced by one or more alkyl, alkoxyl group, alkylamino, halogen, hydroxyl, amino, nitro, cyano group, alkyl acyl, aminoacyl, alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl;
(2) R 2be selected from H, C 1-6alkyl, aminoacyl, aryl and heteroaryl, described alkyl is optionally replaced by one or more alkyl, alkoxyl group, halogen, hydroxyl, amino, single alkylamino, two alkylamino, aryl or heteroaryl, such as halo C 1-6alkyl, benzyl;
(3) R 3for not existing, or be selected from alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, alkylamino, alkyl sulphonyl, alkyl sulfonyl amino, sulfamoyl alkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, amido, ester group ,-CN, cycloalkyl, Heterocyclylalkyl, aryl, heteroaryl, aralkyl;
(4) D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-C (R c1r c2the R that)-, is wherein said a1, R a2be hydrogen, alkyl, halogen, haloalkyl, thiazolinyl or haloalkenyl group separately, or R a1, R a2form cycloalkyl together with the C that they connect, described m is 0 or 1, described R bfor not existing, hydrogen, alkyl, alkyl sulphonyl or alkyl-carbonyl, described R c1, R c2be hydrogen, halogen, alkyl, haloalkyl, thiazolinyl or haloalkenyl group separately, or work as R c1, R c2when being alkyl, C can with R c1, R c2form cycloalkyl;
(5) G, G 1be-N-or-CH-separately;
(6) for singly-bound or double bond, wherein two be singly-bound, or one for singly-bound, and another is double bond;
(7) ring T is selected from:
A, aromatic ring or assorted virtue, can be such as five yuan and hexa-atomic aromatic ring or hetero-aromatic ring, described aromatic ring or hetero-aromatic ring can by R 4replace, wherein R 4for alkyl, alkoxyl group, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, amido or-CN; With
B, naphthenic hydrocarbon or heterocycloalkane, described naphthenic hydrocarbon or heterocycloalkane can by R 4replace, wherein R 4for alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, alkyl acyl, aminoacyl, alkylamino acyl group, amido ,-CN, aryl or heteroaryl.
In some embodiments, the invention provides three ring fused heterocycle type nucleoside phosphoramidate compounds of general formula (II), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein
R 1be selected from H and C 1-6alkyl;
R 2be selected from methyl, ethyl, n-propyl, sec.-propyl, normal-butyl, isobutyl-, the tertiary butyl and sec-butyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, benzyl;
R 3for not existing, or be selected from C 1-4alkyl, C 1-4alkoxyl group, fluorine, chlorine, bromine, phenyl;
D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-C (R c1r c2the R that)-, is wherein said a1, R a2be hydrogen, methyl or ethyl separately, described m is 0 or 1, described R bfor not existing, hydrogen, methyl, ethyl, propyl group, sec.-propyl, cyclopropyl, methyl sulphonyl or methyl carbonyl, described R c1, R c2be hydrogen, fluorine, chlorine, bromine, methyl, ethyl, C separately 2-6thiazolinyl or halo C 2-6thiazolinyl, or work as R c1, R c2when being methyl, C can with R c1, R c2form cyclopropyl;
G, G 1be-N-or-CH-separately;
for singly-bound or double bond, wherein two be singly-bound, or one for singly-bound, and another is double bond,
A, as G, G 1be-CH-, and G and G 1be connected during for double bond, D and G is connected for singly-bound, and
B, when G be-CH-,-N-, G 1for-N-time, G and G 1be connected for singly-bound, D and G is connected for singly-bound; Just when G is-CH-, G 1for-N-, and when D is-N-, G and G 1be connected for singly-bound, D and G is connected for double bond; With
Ring T is selected from:
A, at least containing the heteroatomic five yuan of hetero-aromatic rings of N, O or S or six-membered Hetero-aromatic, phenyl ring, described five yuan or six-membered Hetero-aromatic or phenyl ring can by R 4replace, wherein R 4for C 1-4alkyl, C 1-4alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-4alkoxyl group, hydroxyl, nitro, amino, single C 1-4alkylamino, two C 1-4alkylamino, amido or-CN; With
B, pentamethylene, hexanaphthene, suberane or at least containing the heteroatomic heterocycle pentane of N, O or S, azacyclohexane, a trioxepane.
In some preferred embodiments, the invention provides three ring fused heterocycle type nucleoside phosphoramidate compounds of general formula (II), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein
R 1for H, C 1-4alkyl;
R 2be selected from methyl, ethyl, n-propyl, sec.-propyl, normal-butyl, isobutyl-, the tertiary butyl and sec-butyl, cyclopropyl, cyclobutyl, cyclopentyl, benzyl;
R 3for not existing, or be selected from fluorine, chlorine, bromine, phenyl;
D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-C (R c1r c2the R that)-, is wherein said a1, R a2be hydrogen, methyl or ethyl separately, described m is 0 or 1, described R bfor not existing, hydrogen, methyl, ethyl, propyl group, sec.-propyl, cyclopropyl, described R c1, R c2be hydrogen, fluorine, methyl, ethyl, C separately 2-6thiazolinyl or halo C 2-6thiazolinyl, or work as R c1, R c2be separately methyl or ethyl time, C can with R c1, R c2form cycloalkyl;
G, G 1be N or-CH-separately;
for singly-bound or double bond, wherein two be singly-bound, or one for singly-bound, and another is double bond,
A, as G, G 1be-CH-, and G and G 1be connected during for double bond, D and G is connected for singly-bound, and
B, when G be-CH-, N, G 1during for N, G and G 1be connected for singly-bound, D and G is connected for singly-bound; Just when G is-CH-, G 1for N, and when D is-N-, G and G 1be connected for singly-bound, D and G is connected for double bond; With
Ring T is selected from:
A, at least containing the heteroatomic five yuan of hetero-aromatic rings of N, O or S or six-membered Hetero-aromatic, phenyl ring, described five yuan or six-membered Hetero-aromatic or phenyl ring can by R 4replace, wherein R 4for C 1-4alkyl, C 1-4alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-4alkoxyl group, hydroxyl, nitro, amino, single C 1-4alkylamino, two C 1-4alkylamino, formamido group, kharophen or-CN; With
B, pentamethylene, hexanaphthene, suberane or at least containing the heteroatomic heterocycle pentane of N, O or S, azacyclohexane, a trioxepane, described cycloalkyl or Heterocyclylalkyl can by R 4replace, wherein R 4for C 1-4alkyl, C 1-4alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-4alkoxyl group, hydroxyl, nitro, amino, single C 1-4alkylamino, two C 1-4alkylamino, formamido group, kharophen or-CN.
In other preferred embodiments, the invention provides three ring fused heterocycle type nucleoside phosphoramidate compounds of general formula (II), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein
R 1for H, methyl, ethyl;
R 2be selected from methyl, ethyl, sec.-propyl, n-propyl, cyclopentyl, benzyl;
R 3for not existing, or be selected from fluorine, chlorine, bromine;
D is selected from-O-,-S-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-C (R c1r c2the R that)-, is wherein said bfor not existing, hydrogen, methyl, ethyl, propyl group, sec.-propyl, described R c1, R c2be hydrogen, fluorine, methyl, ethyl, C separately 2-4thiazolinyl, or work as R c1, R c2be separately methyl or ethyl time, C and R c1, R c2form cycloalkyl;
G, G 1be-N-or-CH-separately;
for singly-bound or double bond, wherein two be singly-bound, or one for singly-bound, and another is double bond,
A, as G, G 1be-CH-, and G and G 1be connected during for double bond, D and G is connected for singly-bound, and
B, when G be-CH-,-N-, G 1during for N, G and G 1be connected for singly-bound, D and G is connected for singly-bound; Just when G is-CH-, G 1for-N-, and when D is-N-, G and G 1be connected for singly-bound, D and G is connected for double bond; With
Ring T is phenyl ring or hexanaphthene, and wherein said phenyl ring or hexanaphthene can by R 4replace, wherein R 4for C 1-4alkyl, C 1-4alkoxyl group, halogen, halo C 1-6alkyl, halo C 1-4alkoxyl group, hydroxyl, nitro, amino, single C 1-4alkylamino, two C 1-4alkylamino, formamido group, kharophen or-CN.
The invention provides following particular compound and non-corresponding isomer, prodrug, pharmacy acceptable salt, hydrate, solvate:
On the other hand, the invention provides the preparation method of general formula of the present invention (1) compound, the method comprises the following steps:
A) compound of formula (1) in the basic conditions with phosphorus oxychloride reaction after, add the compound reaction of formula (2), then add the compound that Pentafluorophenol is obtained by reacting formula (3); With
B), under cold condition, the compound of formula (3) and the compound of formula (4) react, and obtain the target compound of formula (5).
Wherein, R 1, R 2, R 3, D, D 1, G, G 1, ring T has above definition.
The third aspect, the invention provides pharmaceutical composition, and it comprises compound, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization shown in general formula of the present invention (I).
In some embodiments, the invention provides pharmaceutical composition, it comprises compound shown in general formula (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, also comprise one or more HCV-Ab IgG therapeutical agents being selected from following composition: HCV NS3 proteinase inhibitor, HCV NS5B RNA RNA-dependent AG14361, nucleoside analog, interferon alpha, the Interferon, rabbit of Pegylation, ribavirin, Levovirin, the pyridine of Wei rummy, TLR7 agonist, TLR9 agonist, cyclophilin inhibitor, α glucosidase inhibitor, NS5A inhibitor and NS3 helicase inhibitors.
Compound general formula of the present invention (I) Suo Shi, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization and pharmaceutically acceptable carrier, thinner or mixed with excipients can be prepared into pharmaceutical preparation, to be suitable for per os or parenteral admin.Medication includes, but are not limited to intracutaneous, intramuscular, intraperitoneal, intravenously, subcutaneous, nose interior and peroral route.Described preparation can be used by any approach, such as, by infusion or inject, is used by the approach absorbed through epithelium or mucocutaneous (such as oral mucosa or rectum etc.).Administration can be whole body or local.The example of oral administration preparation comprises solid or liquid dosage form, specifically, comprises tablet, pill, granula, pulvis, capsule, syrup, emulsion, suspensoid etc.Described preparation is prepared by methods known in the art, and comprises conventional carrier, thinner or the vehicle used of field of pharmaceutical preparations.
Fourth aspect, the invention provides compound shown in general formula of the present invention (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, the method of the curee of solvate or crystallization or medicine composite for curing flaviviridae infections of the present invention, comprise the compound using general formula (I) to described curee, steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization or comprise the compound of general formula (I), its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, effectively to reduce the amount of the virus load of virus described in described curee.In some embodiments, the invention provides the method being used for the treatment of and/or preventing RNA viruses such as flaviviridae infections, comprising to there being the individuality of these needs to give compound of the present invention, its steric isomer, salt, hydrate, solvate or crystallization or its pharmaceutical composition.In other embodiments, the invention provides the method suppressing RNA viruses such as flaviviridae infections, comprise make described virus and treatment significant quantity compound of the present invention, its steric isomer, salt, hydrate, solvate or crystallization or its pharmaceutical composition thereof.
" flaviviridae " refers to any virus of flaviviridae, comprises those viruses infecting people and non-human animal, such as flavivirus, pestivirus and hepatitis C virus.Compound of the present invention and composition can be used in particular for treatment or prophylactic treatment HCV.
On the other hand, the invention provides general formula of the present invention (I) compound, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization and be used for prevention or treatment viral infection, especially the application of flaviviridae infections disease, and the application in the medicine preparing preventing/treating viral infectious diseases, particularly preparing preventing/treating HCV virus infection, as the application in the medicine of HCV viral hepatitis disease.
Term definition
Except as otherwise noted, all technology used herein and scientific terminology have the identical implication usually understood with those skilled in the art.
As used herein, term " steric isomer " refers to by the molecule Atom spatially different isomer produced of arrangement mode, comprises cis-trans-isomer, enantiomer and conformer.All steric isomers all belong to scope of the present invention.
As used herein, term " prodrug " refers to that compound of the present invention is when being applied to any pharmaceutically acceptable form (such as ester, amide compound) that Mammals provides this compound of active compound.
As used herein, term " salt " refers to the pharmacy acceptable salt that compound of the present invention is formed with acid, described acid such as can be selected from, but is not limited to: phosphoric acid, sulfuric acid, hydrochloric acid, Hydrogen bromide, citric acid, toxilic acid, propanedioic acid, amygdalic acid, succsinic acid, fumaric acid, acetic acid, lactic acid, nitric acid, sulfonic acid, tosic acid, oxysuccinic acid, methanesulfonic or its analogue.
As used herein, term " solvate " refers to the form of the compounds of this invention by forming solid-state or liquid title complex with solvent molecule coordination.Hydrate is the special shape of solvate, wherein with water generation coordination.Within the scope of the present invention, solvate is preferably hydrate.
As used herein, term " crystallization " refers to the various solid form of compound formation of the present invention, comprises crystal formation, amorphous.
As used herein, term " alkyl " refers to the saturated hydrocarbyl of straight chain, side chain or ring-type, " C 1-6alkyl " refer to and the saturated hydrocarbyl of below 6 carbon atoms comprise straight or branched C 1-6alkyl and C 1-6cycloalkyl.Such as, suitable C 1-6alkyl group includes but not limited to methyl, ethyl, n-propyl, sec.-propyl, cyclopropyl, normal-butyl, isobutyl-, the tertiary butyl, cyclobutyl, n-pentyl, isopentyl, cyclopentyl, cyclohexyl, n-hexyl.This term comprises substituted or unsubstituted alkyl, and described alkyl can be optionally selected from following group replace by one or more: alkyl, alkoxyl group, aryloxy, alkylamino, arylamino, halogen, hydroxyl, amino, nitro, cyano group, alkyl acyl, aminoacyl, alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl.
As used herein, term " thiazolinyl " refers to the straight or branched unsaturated alkyl containing one or more carbon-carbon double bond (C=C), thiazolinyl preferably containing 2 to 6 carbon atoms, the thiazolinyl more preferably containing 2 to 4 carbon atoms, the thiazolinyl most preferably containing 2 carbon atoms.Term " C 2-6thiazolinyl " to refer to containing 1 or 2 carbon-carbon double bonds and containing the unsaturated alkyl of 2 to 6 carbon atoms.Term " C 2-4thiazolinyl " to refer to containing 1 or 2 carbon-carbon double bonds and containing the unsaturated alkyl of 2 to 4 carbon atoms, suitable alkenyl group includes but not limited to vinyl, propenyl.Similarly, term " alkynyl " refers to the straight or branched unsaturated alkyl containing one or more carbon carbon triple bond (C ≡ C), the alkynyl preferably containing 2 to 6 carbon atoms, the alkynyl more preferably containing 2 to 4 carbon atoms.
As used herein, term " alkoxyl group " refers to-O-alkyl, preferred C 1-6alkoxyl group, suitable alkoxy base includes but not limited to methoxyl group, oxyethyl group etc.
As used herein, term " halogen " refers to fluorine, chlorine, bromine, iodine.
As used herein, term " haloalkyl " refers at least by the alkyl that a halogen atom replaces, preferred halo C 1-6alkyl.
As used herein, term " alkyl sulphonyl " expression " alkyl-SO 2-", preferred C 1-6alkyl-SO 2-, suitable alkylsulfonyl radicals includes but not limited to methyl sulphonyl, ethylsulfonyl.
As used herein, term " alkyl-carbonyl " expression " C 1-6alkyl-C (=O)-", suitable alkylcarbonyl-residues includes but not limited to methyl carbonyl, ethylcarbonyl group.
As used herein, term " aromatic ring " includes but not limited to phenyl ring, naphthalene nucleus, cyclohexyl biphenyl, anthracene nucleus, tetrahydric naphthalene ring, fluorenes ring, indane ring, sub-cyclohexyl biphenyl and acenaphthene ring, preferred phenyl ring.This term comprises replacement and unsubstituted group.Described aromatic ring can be optionally selected from following group replace by one or more: alkyl, thiazolinyl, alkynyl, alkoxyl group, aryloxy, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, arylamino, alkyl acyl, aminoacyl, alkylamino acyl group, amido ,-CN, aryl or heteroaryl.
As used herein, term " hetero-aromatic ring " refers to the heteroatomic 3-10 unit heteroaromatic ring system being selected from N, O and S containing 1-4, preferred C 3-7unit's heteroaromatic ring system, as thiophene, pyridine, imidazoles, furans, pyrroles, thiazole, 1,2,3-triazole, 1,2,4-triazole, 1,2,3-thiadiazoles ， oxazole, 1,2,4-oxadiazole, 1,3,4-oxadiazole etc., suitable hetero-aromatic ring includes but not limited to thiophene, pyridine, imidazoles.This term comprises replacement and unsubstituted group.Described hetero-aromatic ring can be optionally selected from following group replace by one or more: alkyl, thiazolinyl, alkynyl, alkoxyl group, aryloxy, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, arylamino, alkyl acyl, aminoacyl, alkylamino acyl group, amido ,-CN, aryl or heteroaryl.
As used herein, term " heterocycloalkane " refers at least containing the heteroatomic naphthenic hydrocarbon of N, O or S, preferred C 3-7heterocycloalkane, includes but not limited to heterocycle pentane, azacyclohexane, trioxepane.Naphthenic hydrocarbon herein or heterocycloalkane comprise replacement and unsubstituted group.Described naphthenic hydrocarbon or heterocycloalkane can be optionally selected from following group replace by one or more: alkyl, thiazolinyl, alkynyl, alkoxyl group, aryloxy, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, arylamino, alkyl acyl, aminoacyl, alkylamino acyl group, amido ,-CN, aryl or heteroaryl.
As used herein, term " amido " is " C 1-6alkyl-CONH 2", suitable amido groups includes but not limited to formamido group, kharophen.
Compound of the present invention contains multiple asymmetric center, therefore, can exist with the form of racemoid and racemic mixture, single enantiomer, non-corresponding isomer, non-corresponding isomer mixture and single non-corresponding isomer.
Embodiment
Below in conjunction with embodiment, the present invention is further elaborated, but the invention is not restricted to these embodiments.Unless otherwise indicated, the reagent that uses of the present invention and raw material are and are purchased gained.
Embodiment 1 (2S)-2-(((9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1, (2S)-2-(((penta fluoro benzene oxygen base) (9H-carbazole-4-base oxygen base) phosphoryl) is amino) isopropyl propionate
Phosphorus oxychloride (0.84g is added in reaction flask, 0.01mol), add methylene chloride (DCM, 30mL), be cooled to-60 DEG C, drip 4-hydroxycarbazole (1g, 0.01mol) with triethylamine (0.55g, dichloromethane solution 0.01mol), drip and finish, room temperature reaction spends the night.This mixed solution is cooled to-60 DEG C, add ALANINE isopropyl ester hydrochloride, drip the dichloromethane solution of triethylamine (1.38g, 0.025mol) after stirring 20min, dropwise, be warmed up to-5 DEG C, drip the dichloromethane solution of Pentafluorophenol (1g, 10mmol) and triethylamine (0.82g, 0.015mol), drip and finish, to stirring at room temperature.React completely, add water extraction, dry, concentrated, and silica gel chromatographic column is separated, and obtains title compound.
Step 2, (2S)-2-(((9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
(2S)-2-(((penta fluoro benzene oxygen base) (9H-carbazole-4-base oxygen base) phosphoryl) is amino) isopropyl propionate (26mg, 0.1mmol) is added, sealing, argon shield in reaction flask.Injection tetrahydrofuran (THF) (THF, 1.5mL), ice bath cooling hemostasis tertiary butyl magnesium chloride (0.3mL), room temperature reaction 3h.Under ice bath, the solution of the THF of the fluoro-2'-methyluridine (70mg, 0.15mmol) of injection (2'R)-2'-deoxidation-2'-, reaction 15h, reaction terminates.Under ice bath, with the 2N HCl cancellation of 6mL, extraction into ethyl acetate, saturated sodium bicarbonate solution washs, and dry, concentrating under reduced pressure, concentrated solution is separated through silica gel chromatographic column, and purifying obtains title compound.
1H?NMR(300MHz,DMSO-d 6)δppm:11.46(s,1H),11.43(s,1H),8.19(d,1H),7.49(d,1H),7.39(t,1H),7.32(t,1H),7.30(t,1H),7.16(d,1H),7.13(t,1H),6.18(t,1H),6.01(t,1H),5.83(d,1H),5.24(s,1H),4.86(q,1H),4.44-4.48(m,1H),4.32-4.33(m,1H),4.05(s,1H),4.02(q,1H),3.84-3.90(m,2H),1.24(s,3H),1.20(d,3H),1.13(d,6H)。
ESI-MS?m/z:[M+H] +＝619。
Embodiment 2 (2S)-2-(((9-methyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,4-hydroxyl-9-methyl-9H-carbazole
In reaction flask, 4-hydroxycarbazole (2g) is dissolved in DMF (l5mL), at 0 DEG C, slowly adds NaH (880mg), then to stirring at room temperature 30min.Add MeI (1.5g) again, continue at room temperature to react 1h, TLC display reaction terminates, ethyl acetate (100mL) is diluted, organic phase washed with water, the 10%LiCl aqueous solution and saturated common salt are washed, dry, and removing organic phase obtains crude product, crude product obtains title compound through silicagel column (PET:EA=10:1) purifying, faint yellow solid product.
1H?NMR(400MHz,DMSO-d 6)δppm:10.10(s,1H),8.16-8.18(d,1H),7.50-7.53(d,1H),7.38-7.41(t,1H),7.14-7.26(m,2H),6.99-7.01(d,1H),6.60-6.62(d,1H),3.81(s,3H)。
LC-MS?m/z:[M+H] +＝198。
Step 2, (2S)-2-(((9-methyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 4-hydroxyl-9-methyl-9H-carbazole, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, title compound can be obtained.
1H?NMR(400MHz,DMSO-d 6)δppm:11.51(s,1H),8.22-8.24(d,1H),7.61-7.63(m,1H),7.40-7.51(m,4H),7.17-7.24(m,2H),6.24-6.33(m,1H),5.90-6.05(m,2H),5.15(s,1H),4.83-4.89(m,1H),4.31-4.48(m,2H),3.93-4.14(m,1H),3.84-3.93(m,5H),1.06-1.22(m,12H)。
LC-MS?m/z:[M+H] +＝633。
Embodiment 3 (2S)-2-(((9-sec.-propyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,4-hydroxyl-9-sec.-propyl-9H-carbazole
In reaction flask, add 4-hydroxycarbazole (1g, 5.46mmol), THF (20mL), be cooled to-10 DEG C after dissolving, slowly add NaH (0.55g, 14mmol), finish, stir 30 minutes, add 2-N-PROPYLE BROMIDE 0.74g (6mmol), finish, to room temperature reaction 3h, add water cancellation, extraction into ethyl acetate, drying, concentrated, concentrated solution is through preparative chromatography purifying, PET:EA=10%, obtains title compound, beige liquid.
Step 2, (2S)-2-(((9-sec.-propyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 4-hydroxyl-9-sec.-propyl-9H-carbazole, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.
1H?NMR(500MHz,DMSO-d 6)δppm:11.46(s,1H),8.20(d,1H),7.73-7.13(m,7H),6.27-5.67(m,3H),5.14-4.31(m,4H),4.02-3.80(m,4H),1.63(d,6H),1.26-1.22(m,6H),1.16-1.11(m,6H)。
ESI-MS?m/z:[M+H] +＝661.2。
Embodiment 4 (2S)-2-(((9-ethyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,4-hydroxyl-9-ethyl-9H-carbazole
In reaction flask, add 4-hydroxycarbazole (1g, 5.46mmol), be dissolved in THF (20mL), be cooled to-10 DEG C, add NaH (0.55g, 14mmol), finish, stir after 30 minutes, add monobromethane (0.66g, 6mmol), finish, room temperature reaction 3h, reaction terminates, and add water cancellation, extraction into ethyl acetate, drying, concentrated, through preparative chromatography purifying, PET:EA=10%, obtains title compound, beige liquid.
Step 2, (2S)-2-(((9-ethyl--9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 4-hydroxyl-9-ethyl-9H-carbazole, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.
1H?NMR(500MHz,DMSO-d 6)δppm:11.47(s,1H),7.61-8.23(m,3H),7.15-7.50(m,5H),5.96-6.02(m,2H),5.64-5.75(m,2H,),5.16-5.32(m,2H),4.84-4.88(m,1H),4.44-4.48(m,2H,),4.30-4.37(m,1H),4.05-408(m,1H),3.61-3.65(m,1H),1.21-1.33(m,9H),1.12-1.17(m,6H)。
ESI-MS?m/z:[M+H] +＝647.2。
Embodiment 5 (2S)-2-(((9-oxo-9H-fluorenes-2-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
With 2-hydroxyl-9-Fluorenone, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and (2 ' R)-2 '-deoxidation-2 '-fluoro-2 '-methyluridine for raw material, with the method for embodiment 1, obtained title compound.
1H?NMR(300MHz,DMSO-d 6)δ:11.43(s,1H),7.96(d,1H),7.81(d,1H),7.77(d,1H),7.59(t,2H),7.41-7.46(m,1H),7.36(t,1H),6.01(d,1H),5.62-5.66(m,2H),5.57(d,1H),4.82-4.85(m,1H),4.39-4.42(m,1H),4.26-4.29(m,1H),4.02-4.04(m,1H),3.80-3.90(m,2H),3.62-3.65(m,1H),1.27(d,3H),1.23(d,6H),1.13(dd,3H)。
ESI-MS?m/z:[M+Na] +＝654.2。
Embodiment 6 (2S)-2-(((6; 7; 8; 9-tetrahydrochysene dibenzo [b, d] furans-2-base oxygen base) (((2R, 3R; 4R; 5R)-5-(2,4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,2-hydroxyl-6,7,8,9-tetrahydrochysene dibenzo [b, d] furans
Benzoquinones (5g, 46mmol) is dissolved in toluene, and drip 4-(cyclohexyl-1-alkene-1-base) morpholine (8.5g, 51mmol) at 0 DEG C, 0 DEG C of stirring reaction spends the night, and filter, filter residue toluene fully washs rear dry 4h.Add 100mL water, drip HCl (20mL, l5M), stirring at room temperature 18h, filter, filter residue water fully washs, and add water after dissolving with methylene dichloride extraction, and dry organic layer, column purification obtains title compound.
Step 2, (2S)-2-(((6; 7; 8; 9-tetrahydrochysene dibenzo [b, d] furans-2-base oxygen base) (((2R, 3R; 4R; 5R)-5-(2,4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 2-hydroxyl-6,7,8,9-tetrahydrochysene dibenzo [b, d] furans, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-be raw material, with the method for embodiment 1, obtained title compound.
1H?NMR(300MHz,DMSO-d 6)δppm:11.51(s,1H),7.52(s,1H),7.43(d,1H),7.29(s,1H),7.04(d,1H),6.02(d,1H),5.97(d,1H),5.84(s,1H),5.49(d,1H),4.37-4.39(m,1H),4.23-4.24(m,1H),4.01-4.05(m,2H),3.79-3.84(m,2H),2.70(m,2H),2.49(m,2H),1.86-1.87(m,2H),1.77-1.78(m,2H),1.25(d,3H),1.21(d,3H),1.13(t,6H)。
ESI-MS?m/z:[M+H] +＝624.3。
Embodiment 7 (2S)-2-(((dibenzo [b; d] thiophene-2-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,2-hydroxyl-dibenzo [b, d] thiophene
The preparation of 1.12-(dibenzo [b, d] thiophene-2-base)-4,4,5,5-tetramethyl--1,3,2-dioxo borines
In the reaction flask of drying, add the bromo-dibenzo [b of 2-, d] thiophene (2.64g, 10.0mmol), connection boric acid pinacol ester (6.09g, 24.0mmol), Potassium ethanoate (2.94g, 30.0mmol), 1,4-dioxane (50ml) and [two (diphenylphosphine) ferrocene of 1,1'-] palladium chloride (Pd (dppf) Cl 2, 364mg, 0.5mmol).Be heated to 90 DEG C under nitrogen protection, reaction 24h, TLC detect.Reaction terminates, and after reaction system is down to room temperature, adds water (100mL) wherein, extracts (60mL × 3) three times by ethyl acetate, merges organic phase, washs once with saturated aqueous common salt (60mL).Drying, filters, concentrated, obtains title compound, yellow oily through silica column purification.
1H?NMR(500MHz,DMSO-d 6)δppm：8.61(s,1H),8.25-8.23(m,1H),7.88-7.82(m,3H),7.45-7.43(m,2H),1.39-1.33(m,12H)。
The preparation of 1.22-hydroxyl-dibenzo [b, d] thiophene
In the reaction flask of drying, add the compound 2-(dibenzo [b, d] thiophene-2-base)-4 that above-mentioned steps 1.1 is obtained, 4,5,5-tetramethyl--1,3,2-dioxo borine (2.0g, 6.44mmol), ethanol (20ml), hydroxylamine hydrochloride (1.34g, 19.32mmol) with sodium hydrate solid (1.03g, 25.76mmol).Stirring at room temperature is after 10 minutes under nitrogen protection, is heated to 60 DEG C of reactions 24h, TLC and detects.After reaction system is down to room temperature, add water (100mL) wherein, extract (60mL) three times by ethyl acetate, merge organic phase, then use saturated aqueous common salt (60mL) to wash once.Drying, filters, concentrated, obtains title compound, yellow oily through silica column purification.
1H?NMR(500MHz,DMSO-d 6)δppm:8.06-8.05(m,1H),7.83-7.81(m,1H),7.69-7.37(m,1H),7.59-7.57(m,1H),7.45-7.41(m,2H),7.01-6.99(m,1H),4.79(brs,1H)。
Step 2, (2S)-2-(((dibenzo [b; d] thiophene-2-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 2-hydroxyl-dibenzo [b, the d] thiophene of preparation, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.
1HNMR(500MHz,DMSO)δppm:11.47(s,1H),8.29(d,1H),8.19(s,1H),8.02(m,2H),7.58-7.56(m,1H),7.54-7.50(m,2H),7.41-7.40(d,1H),6.13-6.03(m,2H),5.85(d,1H),5.55(d,1H),4.87-4.82(m,1H),4.44-4.38(m,1H),4.30-4.28(m,1H),4.05-4.01(m,1H),3.90-3.86(m,2H),1.28-1.23(m,6H),1.18-1.12(m,6H)。
ESI-MS?m/z:[M+H] +＝636.0。
Embodiment 8 (2S)-2-(((9-oxo-9H-Thiaxanthene-2-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,2-hydroxyl-9-oxo-9H-Thiaxanthene
Thiosalicylic acid (9g), phenol (24.7g), drops in reaction flask, then slowly adds the vitriol oil (85mL), react and be heated to 2h at 75 DEG C, and TLC monitors, stopped reaction.Reaction solution is under agitation slowly poured in 60 DEG C of water, has solid to separate out, and filter, solid with methylene chloride recrystallization, obtains title compound, light green solid.
1H?NMR(400MHz,DMSO-d 6)δppm:10.24(s,1H),8.49-8.52(m,1H),7.80-7.92(m,4H),7.53-7.63(m,1H),7.31-7.34(m,1H)。
Step 2, (2S)-2-(((9-oxo-9H-Thiaxanthene-2-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 2-hydroxyl-9-oxo-9H-Thiaxanthene, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.
1H?NMR(400MHz,DMSO-d 6)δppm:11.49(s,1H),8.41-8.44(d,1H),8.24(s,1H),7.83-7.91(m,3H),7.56-7.76(m,3H),6.19-6.22(t,1H),5.94-5.99(m,2H),5.86(s,1H),4.75-4.82(m,1H),4.23-4.29(m,2H),3.78-3.82(m,3H),1.11-1.24(m,6H),1.05-1.11(m,6H)。
LC-MS?m/z:[M+H] +＝664。
Embodiment 9 (2S)-2-(((9-oxo-9H-xanthene-3-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,3-hydroxyl-9-oxo-9H-xanthene
The preparation of 1.13-methoxyl group-9-oxo-9H-xanthene
In reaction flask; 2-hydroxyl-4 methoxybenzaldehyde (1.5g) is dissolved in DMF (40mL); slowly 1 is added under room temperature; 2-dibromobenzene (4.6g) and salt of wormwood (2.76g); then 120 DEG C are heated to; stir 12h under nitrogen protection; TLC display reaction terminates; be cooled to room temperature, with ethyl acetate (150mL) dilution, organic phase washed with water, the 10%LiCl aqueous solution and saturated common salt water washing; dry; removing organic phase, crude product obtains title compound through silicagel column (PE:EA=10:1) purifying, white solid product.
1H?NMR(400MHz,DMSO-d 6)δppm:8.16-8.19(q,1H),8.09-8.12(d,1H),7.83-7.88(m,1H),7.62-7.65(m,1H),7.45-7.50(m,1H),7.17-7.20(d,1H),7.05-7.08(dd,1H),3.94(s,3H)。
The preparation of 1.23-hydroxyl-9-oxo-9H-xanthene
In reaction flask, compound 3-methoxyl group-9-oxo-9H-xanthene (300mg) obtained for above-mentioned steps 1.1 is dissolved in methylene dichloride (5mL), under dry ice acetone bath and nitrogen protection, drips BBr 3(983mg), stir until reaction terminates, go out with 1mL shrend, add 10mL methylene dichloride, organic phase washed with water, saturated common salt water washing, dry methylene chloride, pressure reducing and steaming organic phase obtains thick product, title compound is obtained, white solid product after silica column purification (PE:EA=5:1).
1H?NMR(400MHz,DMSO-d 6)δppm:11.00(s,1H),8.15-8.17(t,1H),8.04-8.06(d,1H),7.29(m,1H),7.60-7.63(m,1H),7.45(m,1H),6.88-6.93(m,2H)。
Step 2, (2S)-2-(((9-oxo-9H-xanthene-3-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 3-hydroxyl-9-oxo-9H-xanthene, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.
1H?NMR(400MHz,DMSO-d 6)δppm:11.51(s,1H),8.18-8.23(m,2H),7.87-7.90(m,1H),7.66-7.68(d,1H),7.49-7.55(m,3H),7.33-7.35(m,1H),6.33-6.40(m,1H),5.91-6.07(m,2H),5.57-5.59(d,1H),4.82-4.86(m,1H),4.35-4.44(m,2H),3.88-4.03(m,3H),1.14-1.29(m,6H),1.10-1.15(m,6H)。
LC-MS?m/z:[M+H] +＝648。
Embodiment 10 (2S)-2-(((9H-carbazole-2-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
With 2-hydroxyl-9H-carbazole, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.Isomer I is obtained, white solid and isomer II, white solid after HPLC purifying.
HPLC condition: instrument: Shimadzu Corporation's preparative HPLC (Preparative liquid chromatography Shimadzu); Chromatographic column: Benetnoch tM-C18,20x250mm, 10 μm; Determined wavelength: 254nm; Moving phase H 2o:MeOH=30:70.
Corresponding isomer I:
1H?NMR(300MHz,DMSO-d 6)δ:11.48(s,1H),11.27(s,1H),8.06(d,2H),7.60-7.58(d,1H),7.46-7.45(d,1H),7.36-7.35(d,2H),7.33-7.31(m,1H),7.14-7.13(d,1H),6.08-6.12(m,2H),5.90(d,1H),5.59(d,1H),4.91-4.82(m,1H),4.67-4.32(m,1H),4.32-4.21(m,1H),4.05-4.01(m,1H),3.88-3.79(m,2H),2.3(s,3H),1.58(d,3H),1.30-1.20(m,9H)。
ESI-MS?m/z:[M+Na] +＝641.3。
Corresponding isomer II:
1H?NMR(300MHz,DMSO-d 6)δppm:11.48(s,1H),11.26(s,1H),8.01(d,2H),7.61-7.59(d,1H),7.47-7.45(d,1H),7.36-7.34(d,2H),7.33-7.31(m,1H),7.14-7.13(d,1H),6.08-6.12(m,2H),5.91(d,1H),5.59(d,1H),4.91-4.82(m,1H),4.67-4.32(m,1H),4.32-4.21(m,1H),4.05-4.01(m,1H),3.88-3.79(m,2H),2.3(s,3H),1.30-1.20(m,9H),1.58(d,3H)。
ESI-MS?m/z:[M+Na] +＝641.3。
Embodiment 11 (2S)-2-(((2; 3; 4; 9-tetrahydrochysene-1H-carbazole-6-base oxygen base) (((2R; 3R, 4R, 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,6-hydroxyl-2,3,4,9-1H-tetrahydro carbazole
In reaction flask, add methoxyphenyl hydrazine hydrochloride (5g, 29mmol), acetic acid (30mL), pimelinketone (2.8g, 29mmol), is heated to 70 DEG C, Quan Rong, continue stirring after 5 minutes, a large amount of yellow solid is separated out, and TLC monitoring is to reacting completely, drop to room temperature, add HBr/H 2o solution 30mL, reheats 80 DEG C, reacts 4 hours, TLC monitoring disappears to raw material, stopped reaction, drops to room temperature, adds ethyl acetate and water extraction, organic phase saturated sodium bicarbonate solution washs 2 times, drying, concentrated, through preparative chromatography column purification (PET:EA=20%), obtain title compound, yellow solid.
Step 2, (2S)-2-(((2; 3; 4; 9-tetrahydrochysene-1H-carbazole-6-base oxygen base) (((2R; 3R, 4R, 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
With 6-hydroxyl-2,3,4,9-1H-tetrahydro carbazole, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.
1H?NMR(500MHz,DMSO-d 6)δppm:11.47(s,1H),10.64(s,1H),7.51(d,1H),7.17-7.11(m,2H),6.84(d,1H),6.03-5.99(m,1H),5.86-5.80(m,2H),5.50-5.44(m,1H),4.88-4.83(m,1H),4.36-4.33(m,1H),4.22-4.19(m,1H),4.05-4.00(m,1H),3.84-3.75(m,2H),2.68-2.54(m,4H),1.81-1.78(m,4H),1.26-1.21(m,6H),1.17-1.14(m,6H)。
ESI-MS?m/z:[M+H] +＝623.0。
Embodiment 12 (2S)-2-(((9H-carbazole-3-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,3-hydroxyl-9H-carbazole
1.13-the preparation of formyl radical-9-tert-butoxycarbonyl-9H-carbazole
In reaction flask; N-Boc-3-bromine carbazole (3g) is dissolved in THF (10mL); be cooled to-78 DEG C under nitrogen protection; add n-BuLi (6.5mL), reaction solution stirs 1h at-78 DEG C, then adds DMF (952mg) wherein; after adding; stop cooling, make temperature of reaction naturally rise to room temperature, with saturated NH 4cl solution (1mL) cancellation, with EA (20mL) dilution, organic phase washed with water, saturated common salt are washed, dry organic phase, concentrated except desolventizing, thick product obtains title compound, white solid through silica gel chromatography (PE:EA=10:1).
The preparation of 1.23-formyl radical oxygen base-9-tert-butoxycarbonyl-9H-carbazole
In reaction flask; 3-formyl radical-9-tert-butoxycarbonyl-9H-carbazole (600mg) is dissolved in THF (10mL), then adds metachloroperbenzoic acid (m-CPBA, 688mg); reaction solution is at stirring at room temperature 5h; with methylene dichloride (20mL) dilution, organic phase washed with water, saturated sodium bicarbonate aqueous solution is washed; dry organic phase; concentrated except desolventizing, obtain title compound, yellow solid.
The preparation of 1.33-hydroxyl-9-tert-butoxycarbonyl-9H-carbazole
In reaction flask, 3-formyl radical oxygen base-9-tert-butoxycarbonyl-9H-carbazole (670mg) is dissolved in MeOH/H 2in O (7.5mL (volume ratio 2:1)), then add K 2cO 3(890mg), reaction solution stirs 12h in room temperature, dilute by ethyl acetate (20mL), organic phase washed with water, saturated aqueous common salt are washed, dry organic phase, concentrated except desolventizing, thick product obtains title compound, faint yellow solid through silica gel column chromatography (PET:EA=20:1) purifying.
The preparation of 1.43-hydroxyl-9H-carbazole
3-hydroxyl-9-tert-butoxycarbonyl-9H-carbazole (240mg) is dissolved in methylene dichloride (4mL), trifluoroacetic acid (TFA is dripped at 0 DEG C, 2mL), stir 2h, reaction terminates, reaction solution is under agitation poured in appropriate frozen water, regulate pH to neutral with saturated sodium hydrogen carbonate solution, with dichloromethane extraction (10mL × 2), organic phase washed with water, saturated common salt water washing, dry methylene chloride, pressure reducing and steaming organic phase obtains thick product, title compound is obtained, faint yellow solid through silica column purification (PET:EA=3:1).
Step 2, (2S)-2-(((9H-carbazole-3-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 3-hydroxyl-9H-carbazole, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.
1H?NMR(400MHz,DMSO-d 6)δppm:11.53(s,1H),11.28(s,1H),7.59-8.06(m,2H),7.57-7.60(m,1H),7.40-7.47(m,3H),7.16-7.28(m,1H),7.12-7.14(m,1H),6.03-6.06(m,2H),5.52-5.55(m,1H),4.82-4.86(m,2H),3.86(m,1H),3.85(m,2H),1.21-1.27(m,6H),1.11-1.14(m,6H)。
LC-MS?m/z:[M+H] +＝619。
Embodiment 13 (2S)-2-(((9-methyl-9H-carbazole-3-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,3-hydroxyl-9-methyl-9H-carbazole
The preparation of 1.13-methoxyl group-9-methyl-9H-carbazole
In reaction flask, at 0 DEG C, 3-hydroxyl-9H-carbazole (400mg) is dissolved in dry DMF (10mL), adds methyl iodide (0.55mL), keep 0 DEG C to continue to stir 0.5h.Add sodium hydride (352mg) again, stir after 5 minutes, rise to stirring at room temperature and terminate to TLC display reaction.With ethyl acetate (50mL) dilution, organic phase washed with water, 10%LiCl solution and saturated common salt water washing, dry, removing organic phase, crude product obtains title compound, pale solid through silicagel column (PE:EA=10:1) purifying.
1H?NMR(400MHz,DMSO-d 6)δppm:8.04-8.06(d,1H),7.58-7.59(d,1H),7.11-7.48(m,5H),3.94(m,3H),3.83(m,3H)。
LC-MS?m/z:[M+H] +＝212。
The preparation of 1.23-hydroxyl-9-methyl-9H-carbazole
In reaction flask, 3-methoxyl group-9-methyl-9H-carbazole (115mg) is dissolved in methylene dichloride (5mL), under dry ice acetone bath and nitrogen protection, slowly drips BBr 3(0.1mL), drip and finish, rise to stirring at room temperature and terminate to TLC display reaction.With water (1mL) cancellation, add methylene dichloride (10mL), organic phase washed with water, saturated common salt are washed, dry methylene chloride, pressure reducing and steaming organic phase obtains thick product, be separated (PET:EA=5:1) through column chromatography purification and obtain title compound, white solid product.
1H?NMR(400MHz,DMSO-d 6)δppm:9.01(s,1H),8.01-8.03(d,1H),7.37-7.51(m,4H),7.09-7.13(m,1H),6.94-6.97(m,1H),3.80(s,3H)。
Step 2, (2S)-2-(((9-methyl-9H-carbazole-3-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 3-hydroxyl-9-methyl-9H-carbazole, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.
1H?NMR(400MHz,DMSO-d 6)δppm:11.53(s,1H),7.95-8.13(m,2H),7.46-7.60(m,4H),7.18-7.37(m,2H),5.92-6.08(m,3H),5.51-5.55(m,1H),4.83-4.88(m,1H),4.24-2.32(m,2H),4.03-4.06(m,1H),3.81-3.87(m,5H),1.10-1.27(m,12H)。
LC-MS?m/z:[M+H] +＝633。
Embodiment 14 (2S)-2-(((10-methyl-9-oxo-9; 10-acridan-2-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,2-hydroxyl-10-methyl-9-oxo-9,10-acridan
1.12-(4-Methoxyphenylamino) benzoic preparation
In reaction flask; 2-fluorobenzoic acid (1.68g) and P-nethoxyaniline (1.23g) are dissolved in THF (l5mL), slowly add Lithamide (460mg) under room temperature, under nitrogen protection; be heated to 50 DEG C and stir 4h, reaction terminates.Be cooled to room temperature, with water (1mL) cancellation, reaction mixture ethyl acetate (50mL) and concentrated hydrochloric acid (2mL) dilution, organic phase washed with water and saturated common salt water washing, thick product is obtained after concentrated, add Virahol (10mL), and be heated to 65 DEG C and dissolve completely to solid, cool to room temperature, add water (10mL) again, cool under ice bath, filter and obtain title compound, yellow solid.
1H?NMR(400MHz,DMSO-d 6)δppm:12.98(s,1H),9.50(s,1H),7.90-7.93(m,1H),7.37-7.39(m,1H),7.35(m,2H),6.97-7.02(m,3H),6.71-6.74(m,1H),3.81(s,3H)。
LC-MS?m/z:[M+H] +＝244。
The preparation of 1.22-methoxyl group-9-oxo-9,10-acridan
In reaction flask, 2-(4-anisole amido) phenylformic acid (800mg) is suspended in POCl 3(10mL), in, reflux is until reacted (LC-MS detection), pressure reducing and steaming POCl 3; Add ethanol in concentrated solution: hydrochloric acid=8:1 (V:V) solution (10mL) of 10%, reflux 1h, cool to room temperature, adds water (10mL), has yellow solid to separate out, and filters, obtains title compound, yellow solid.
1H?NMR(400MHz,DMSO-d 6)δppm:11.77(s,1H),8.22-8.24(d,1H),7.40-7.73(m,5H),7.22-7.26(m,1H),3.86(s,3H)。
LC-MS?m/z:[M+H] +＝226。
The preparation of 1.32-hydroxyl-9-oxo-9,10-acridan
In reaction flask, be added to by 2-methoxyl group-9-oxo-9,10-acridan (670mg) in the 40%HBr aqueous solution (13mL), then reflux 24h, reaction terminates, cool to room temperature, filters, obtains title compound, yellow solid.
1H?NMR(400MHz,DMSO-d 6)δppm:11.64(s,1H),8.18-8.20(m,1H),7.68-7.69(m,1H),7.45-7.56(m,3H),7.17-7.27(m,2H)。
LC-MS?m/z:[M+H] +＝212。
The preparation of 1.42-methoxyl group-10-methyl-9-oxo-9,10-acridan
In reaction flask, by 2-hydroxyl-9-oxo-9,10-acridan (211mg) is dissolved in DMF (3mL), at 0 DEG C, add NaH (160mg), then stir 30min, then add MeI (0.18mL), continue reaction 30min to room temperature, reaction terminates.With water (1mL) cancellation, mixture ethyl acetate (10mL) is diluted, organic phase washed with water, the 10%LiCl aqueous solution and saturated common salt water washing, dry, concentrate after filtering and obtain thick product, title compound is obtained, yellow solid through silica gel column chromatography (PE:EA=3:1) purifying.
LC-MS?m/z:[M+H] +＝240。
The preparation of 1.52-hydroxyl-10-methyl-9-oxo-9,10-acridan
In reaction flask, 2-methoxyl group-10-methyl-9-oxo-9,10-acridan (156mg) is dissolved in methylene dichloride (3mL), under dry ice acetone bath and nitrogen protection, slowly drips BBr 3(0.125mL), drip and finish, rise to stirring at room temperature and terminate to TLC display reaction.With water (1mL) cancellation, add methylene dichloride (10mL), organic phase washed with water, saturated common salt are washed, dry methylene chloride, pressure reducing and steaming organic phase obtains thick product, title compound is obtained, yellow solid product through column chromatography for separation (PE:EA=5:1) purifying.
1H?NMR(400MHz,DMSO-d 6)δppm:9.69(s,1H),8.29-8.31(d,1H),7.67-7.79(m,4H),7.26-7.34(m,2H),3.91(s,3H)。
LC-MS?m/z:[M+H] +＝212。
Step 2, (2S)-2-(((10-methyl-9-oxo-9; 10-acridan-2-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 2-hydroxyl-10-methyl-9-oxo-9,10-acridan, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-are raw material, with the method for embodiment 1, obtained title compound (white solid).
1H?NMR(400MHz,DMSO-d 6)δppm:11.53(s,1H),8.32-8.34(d,1H),8.13-8.14(d,1H),7.92(m,3H),7.85(m,1H),7.71(m,1H),7.35(m,1H),6.16(t,1H),5.86-6.04(m,2H),5.53-5.55(d,1H),4.81-4.85(m,1H),4.41(m,1H),4.28(m,1H),4.01(m,1H),3.96(s,3H),3.80(m,2H),1.22-1.28(m,6H),1.10-1.134(m,6H)。
LC-MS?m/z:[M+H] +＝661。
Embodiment 15 (2S)-2-(((the fluoro-9H-carbazole of 1--4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of the fluoro-4-hydroxyl of step 1,1--9H-carbazole
The preparation of the fluoro-2-methoxyphenylboronic acid of 1.15-
The bromo-4-fluoroanisole (5g) of 2-is thrown in reaction flask; then anhydrous THF (8OmL) is added; n-Butyl Lithium (20mL) is slowly dripped under ice bath, nitrogen protection; drip off; reaction 1h, more slowly add trimethyl borate (5.5mL), stirring at room temperature 3h; TCL monitors, stopped reaction.The reaction solution hydrochloric acid of 18% is adjusted to pH value to 1, then adds ethyl acetate (200mL), saturated sodium-chloride water solution, organic layer washes 3 times, merges organic phase, anhydrous sodium sulfate drying, concentrate and obtain title compound, white solid, thick product can be directly used in next step reaction.
The preparation of 1.25-fluorine 2-methoxyl group-2'-nitrobiphenyl
By fluoro-for 5-2-methoxyphenylboronic acid (500mg); the bromo-2-oil of mirbane (591mg) of 1-; [1; 1 '-bis-(diphenylphosphine) ferrocene] palladium chloride dichloromethane complex (215mg), salt of wormwood (811mg) is thrown in reaction flask, then adds 1; 4-dioxane 20mL and water 4mL; be heated to 90 DEG C under nitrogen protection and stir 1.5h, TCL monitoring, stopped reaction.Reaction solution adds ethyl acetate (100mL), saturated sodium-chloride water solution (50mL), extract with separating funnel, organic layer washes 3 times, merge organic phase, anhydrous sodium sulphate is come dry, concentrates and obtains crude product, crude product obtains title compound, faint yellow solid through column chromatography purification (sherwood oil: ethyl acetate=100:1-50:1).
1H?NMR(400MHz,DMSO-d 6)δppm:7.99-8.01(d,1H),7.76-7.79(m,1H),7.63-7.65(m,1H),7.50-7.53(m,1H),7.23-7.28(m,2H),7.03-7.07(m,1H),3.60(s,3H)。
The preparation of the fluoro-4-methoxyl group of 1.31--9H-carbazole
By 5-fluorine 2-methoxyl group-2 '-nitrobiphenyl (1g), drop in reaction flask, then add triphenyl phosphorus (2.1g), 1,2-dichlorobenzene (10mL), stirred under nitrogen atmosphere is heated to 190 DEG C and spends the night.TLC monitors, stopped reaction.Obtain crude product by concentrated for reaction solution, crude product obtains title compound, white solid through column chromatography (sherwood oil: ethyl acetate=80:1 to 30:1) purifying.
LC-MS?m/z:[M+H] +＝216。
The preparation of the fluoro-4-hydroxyl of 1.41--9H-carbazole
In reaction flask, fluoro-for 1-4-methoxyl group-9H-carbazole (250mg) is dissolved in methylene dichloride (5mL), under dry ice acetone bath and nitrogen protection, slowly drips BBr 3(0.22mL), drip and finish, rise to stirring at room temperature and terminate to TLC display reaction.With water (1mL) cancellation, add methylene dichloride (20mL), organic phase washed with water, saturated common salt are washed, dry methylene chloride, pressure reducing and steaming organic phase obtains thick product, title compound is obtained, white solid after column chromatography for separation (PET:EA=5:1) purifying.
LC-MS?m/z:[M+H] +＝202。
Step 2,2S)-2-(((the fluoro-9H-carbazole of 1--4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 1-fluoro-4-hydroxyl-9H-carbazole, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound, white solid.
1H?NMR(400MHz,DMSO-d 6)δppm:11.96-11.96(m,1H),11.60(s,1H),8.26-8.29(d,1H),7.51-7.61(m,3H),7.15-7.33(m,3H),6.32-6.38(m,1H),5.96-5.98(m,2H),5.34(m,1H),4.90-4.93(m,1H),4.38-4.54(m,2H),4.11-4.14(m,1H),3.91-3.92(m,2H),1.05-1.14(m,12H)。
LC-MS?m/z:[M+H] +＝637。
Embodiment 16 (2S)-2-(((the fluoro-9-methyl of 1--9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of the fluoro-4-hydroxyl of step 1,1--9-methyl-9H-carbazole
The preparation of the fluoro-4-methoxyl group of 1.11--9-methyl-9H-carbazole
By fluoro-for 1-4-methoxyl group-9H-carbazole (200mg), throw in reaction flask, ice bath slowly adds sodium hydride (45mg) under stirring, add, continue reaction 30 minutes, and then add methyl iodide (0.15mL), under room temperature, react 1h.TLC monitors, stopped reaction.Reaction solution adds water cancellation, and then add ethyl acetate (80mL), saturated sodium-chloride water solution 50mL, layering, organic layer washes 3 times, merges organic phase, anhydrous sodium sulfate drying, concentratedly obtains title compound, white solid.
The preparation of the fluoro-4-hydroxyl of 1.21--9-methyl-9H-carbazole
In reaction flask, fluoro-for 1-4-methoxyl group-9-methyl-9H-carbazole (190mg) is dissolved in methylene dichloride (10mL), under dry ice acetone bath and nitrogen protection, drips BBr 3(0.2mL), after stirring 2h at-78 DEG C, room temperature continues stirring reaction 2h again, and reaction terminates, and adds methylene dichloride (10mL), with the sodium hydrogen carbonate solution cancellation that 1mL is saturated, the water washing of organic phase washed with water saturated common salt, dry organic layer, concentrating under reduced pressure organic phase obtains thick product, title compound is obtained, white solid through silica column purification (PE:EA=5:1).
LC-MS?m/z:[M+H] +＝216。
Step 2, (2S)-2-(((the fluoro-9-methyl of 1--9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 1-fluoro-4-hydroxyl-9-methyl-9H-carbazole, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.
Be separated further through HPLC and obtain isomer I (white solid) and isomer II (white solid).
HPLC condition:
Instrument: SHIMADZU LC-6A; Chromatographic column: Shim-pack PREP-ODS (H) (250*20mm, 10um); Flow velocity: 12mL/min; Determined wavelength: 220/254nm; Moving phase: A phase: H 2o (0.1%FA) B phase: CH 3oH.
Isomer I: 1h NMR (400MHz, DMSO-d 6) δ ppm:11.52 (s, 1H), 8.24-8.26 (d, 1H), 7.66-7.68 (d, 1H), 7.45-7.59 (m, 2H), 7.12-7.30 (m, 3H), 6.28-6.35 (m, 1H), 5.91-6.03 (m, 2H), 5.21 (s, 1H), 4.83-4.87 (t, 1H), 4.31-4.44 (m, 2H), 4.05 (m, 4H), 3.84-3.89 (m, 2H), 1.18-1.23 (m, 6H), 1.12-1.24 (m, 6H).
LC-MS?m/z:[M+H] +＝651。
Isomer II: 1h NMR (400MHz, DMSO-d 6) δ ppm:11.53 (s, 1H), 8.23-8.25 (d, 1H), 7.65-7.67 (d, 1H), 7.5-7.56 (m, 2H), 7.12-7.18 (m, 3H), 6.33-6.37 (m, 1H), 5.97-6.07 (m, 2H), 5.21-5.27 (m, 1H), 4.73-4.76 (t, 1H), 4.35-4.36 (m, 2H), 4.04-4.09 (m, 4H), 3.85-3.89 (m, 2H), 1.20-1.23 (m, 6H), 0.99-1.14 (m, 6H).
LC-MS?m/z:[M+H] +＝651。
Embodiment 17 (2S)-2-(((9-oxo-9H-xanthene-2-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,2-hydroxyl-9-oxo-9H-xanthene
The preparation of 1.12-methoxyl group-9-oxo-9H-xanthene
In reaction flask, 2-nitrobenzaldehyde (1.5g) is dissolved in toluene (10mL), p methoxy phenol (1.24g) is slowly added under room temperature, cupric chloride (66mg), triphenylphosphine (196mg) and potassiumphosphate (4.6g), reaction solution exposes in atmosphere, be heated to 110 DEG C and stir 24h, TLC display reaction terminates, be cooled to room temperature, filter, filtrate is diluted by ethyl acetate (150mL), organic phase washed with water and saturated common salt water washing, dry, removing organic phase, crude product obtains title compound through silicagel column (PE:EA=10:1) purifying, faint yellow solid product.
1H?NMR(400MHz,DMSO-d 6)δppm:8.17-8.20(m,1H),7.83-7.90(m,1H),7.60-7.65(m,2H),7.53-7.55(d,1H),7.44-7.48(m,2H),3.87(s,3H)。
The preparation of 1.22-hydroxyl-9-oxo-9H-xanthene
In reaction flask, 2-methoxyl group-9-oxo-9H-xanthene (200mg) is dissolved in methylene dichloride (5mL), under dry ice acetone bath and nitrogen protection, drips BBr 3(437mg), 2h is stirred at-78 DEG C, continue to stir 2h in room temperature again, reaction terminates, and adds the saturated sodium hydrogen carbonate solution of 1mL and methylene dichloride (10mL), organic phase washed with water, saturated common salt water washing, dry organic layer, concentrating under reduced pressure organic phase obtains thick product, obtains title compound, faint yellow solid product after silica column purification (PET:EA=5:1).
1H?NMR(400MHz,DMSO-d 6)δppm:10.00(s,1H),8.17-8.19(m,1H),7.83-7.88(m,1H),7.55-7.66(m,2H),7.44-7.49(m,2H),7.31-7.35(m,1H)。
Step 2, (2S)-2-(((9-oxo-9H-xanthene-2-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 2-hydroxyl-9-oxo-9H-xanthene, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound (white solid).
1H?NMR(400MHz,DMSO-d 6)δppm:11.54(s,1H),8.24-8.27(d,1H),7.93-8.05(m,2H),7.73-7.82(m,3H),7.54-7.84(m,2H),6.27-6.28(m,1H),5.94-6.13(m,2H),5.60-5.62(m,1H),4.86-4.89(t,1H),4.31-4.49(m,2H),4.07(m,1H),3.89-3.93(m,2H),1.28-1.34(m,6H),1.15-1.20(m,6H)。
LC-MS?m/z:[M+H] +＝648。
Embodiment 18 (2S)-2-(((9-methyl-2; 3; 4; 9-tetrahydrochysene-1H-carbazole-6-base oxygen base) (((2R; 3R, 4R, 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,6-hydroxyl-9-methyl-2,3,4,9-1H-tetrahydro carbazole
The preparation of 1.16-methoxyl group-9-methyl-2,3,4,9-1H-tetrahydro carbazole
In reaction flask, add 6-methoxyl group-2,3,4,9-1H-tetrahydro carbazole (2.5g, 12mmol) and THF (20mL), be cooled to-10 DEG C after dissolving, slowly add NaH (1.5g, 36mmol), finish, stir after 30 minutes, then add methyl iodide 2.5mL, finish, to room temperature reaction 30 minutes, reheat 50 DEG C, react 1.5 hours, reaction terminates, and is cooled to room temperature, add water cancellation, extraction into ethyl acetate, dry, concentrated, preparative chromatography purifying, PET:EA=10%, obtains title compound, white solid.
The preparation of 1.26-hydroxyl-9-methyl-2,3,4,9-1H-tetrahydro carbazole
In reaction flask, add the compound 6-methoxyl group-9-methyl-2,3 that above-mentioned steps 1.1 is obtained, 4,9-1H-tetrahydro carbazole (0.7g, 3.2mmol), the 40%HBr aqueous solution and each 10mL of acetic acid, be heated to 110 DEG C, reacts 6 hours, raw material primitive reaction is complete, and cool to room temperature, adds water and extraction into ethyl acetate, concentrated, preparative chromatography purifying, PET:EA=10%, obtain title compound, yellow solid.
Step 2, (2S)-2-(((9-methyl-2; 3; 4; 9-tetrahydrochysene-1H-carbazole-6-base oxygen base) (((2R; 3R, 4R, 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
Be raw material with 6-hydroxyl-9-methyl-2,3,4,9-1H-tetrahydro carbazole, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-, with the method for embodiment 1, obtained title compound.
1HNMR(500MHz,DMSO)δppm:11.48(s,1H),7.49-6.90(m,4H),6.04-5.97(m,2H),5.90-5.81(m,1H),5.49-5.41(m,1H),4.88-4.83(m,1H),4.38-4.33(m,1H),4.22-4.20(m,1H),4.02-3.99(m,1H),3.82-3.79(m,2H),3.57(s,3H),2.69(m,4H),1.84-1.77(m,4H),1.27-1.22(m,6H),1.19-1.13(m,6H)。
ESI-MS?m/z:[M+Na] +＝659.2。
Embodiment 19 (2S)-2-(((9-methyl-9H-carbazole-2-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,2-hydroxyl-9-methyl-9H-carbazole
The preparation of 1.12-methoxyl group-9-methyl-9H-carbazole
By 2-hydroxyl-9H-carbazole (915mg, 5mmol), THF (15mL) adds in reaction flask, under 0 DEG C of condition, add NaH (130mg, 20mmol) in batches, reaction 30min, add THF (5ml) solution of methyl iodide (568mg, 20mmol) again, spend the night (about 10h) in 30 DEG C of reactions.TLC monitoring reaction is complete, and add water cancellation, and extraction into ethyl acetate (50ml × 3), silica column purification, obtains title compound.
The preparation of 1.22-hydroxyl-9-methyl-9H-carbazole
Join in reaction flask by 2-methoxyl group-9-methyl-9H-carbazole (730mg), add the 40%HBr aqueous solution and glacial acetic acid, after 105 DEG C of reaction 24h, TLC monitoring reaction is complete.Reaction solution is added 20ml water, with dichloromethane extraction 3 times, silica column purification.Obtain title compound.
Step 2, (2S)-2-(((9-methyl-9H-carbazole-2-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 2-hydroxyl-9-methyl-9H-carbazole, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.
1HNMR(300MHz,DMSO-d6)δppm:11.45(s,1H),8.10-8.11(d,1H),7.95-7.97(d,1H),7.60-7.61(m,1H),7.59-7.60(m,1H),7.56-7.58(m,1H),7.44-7.45(m,1H),7.36-7.38(m,1H),7.19-7.22(m,1H),7.07-7.09(m,1H),6.05-6.07(m,2H),5.91(m,1H),5.26(s,1H),4.83-4.89(m,1H),4.31-4.48(m,1H),3.92-4.14(m,1H),3.84-3.93(m,5H),1.25-1.26(m,6H),1.11-1.14(m,6H)。
ESI-MS?m/z:[M+H] +＝633.3。
Embodiment 20 (2S)-2-(((dibenzo [b; d] furans-3-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,3-hydroxyl-dibenzo [b, d] furans
The preparation of 1.13-amino-dibenzo [b, d] furans
By 3-nitro diphenylene-oxide (4g), reduced iron powder (5.26g), ammonium chloride (5g) is thrown in reaction flask, adds ethanol (150mL), in 90 DEG C of backflows, monitors raw material reaction completely, stopped reaction to TLC.Cooling, suction filtration, after filtrate reduced in volume, through column chromatographic isolation and purification, obtains title compound.
1H?NMR(400MHz,DMSO-d 6)δppm:7.84-7.82(m,1H),7.71-7.69(d,1H),7.52-7.50(m,1H),7.30-7.25(m,2H),6.75-6.75(d,1H),6.64-6.62(m,1H),5.57(s,2H)。
LC-MS?m/z:[M+H] +＝184。
The preparation of the bromo-dibenzo of 1.23-[b, d] furans
By 3-amino-dibenzo [b, d] furans (1g, 5.46mmol) be dissolved in acetonitrile (10mL), nitrous acid special butyl ester (731mg is slowly dripped under condition of ice bath, 7.10mmol), and then slowly add cupric bromide (1.46g, 6.55mmol), finish, reaction solution to 50 DEG C stirring reaction 1 hour, reaction terminates, raw material reaction is complete, add water (1mL) cancellation, filter, filtrate adds methylene dichloride (20mL), wash with water (40mL × 2), anhydrous sodium sulfate drying, purification by silica gel column chromatography obtains title compound, white solid.
1H?NMR(400MHz,DMSO-d 6)δppm:8.16(m,1H),8.11-8.13(d,1H),8.02(s,1H),7.71-7.73(d,1H),7.56-7.58(m,2H),7.42-7.45(m,1H)。
The preparation of 1.33-formyl radical-dibenzo [b, d] furans
Bromo-for 3-dibenzo [b, d] furans (247mg, 1mmol) is dissolved in anhydrous tetrahydro furan (3mL), n-Butyl Lithium (0.6mL) is slowly dripped in-78 DEG C, finish, stir 30 minutes at continuing low temperature-78 DEG C, and then slowly drip DMF (0.1mL), drip off, rise to stirring at room temperature 10 minutes, reaction terminates, and drips aqueous ammonium chloride solution (0.2mL) cancellation, silica column purification obtains title compound, yellow solid.
1H?NMR(400MHz,DMSO-d 6)δppm:10.15(s,1H),8.38-8.40(d,1H),8.300(m,1H),8.24(s,1H),7.98-7.98(m,1H),7.79-7.82(d,1H),7.65-7.66(m,1H),7.47-7.49(m,1H)。
The preparation of 1.43-methanoyl-dibenzo [b, d] furans
By 3-formyl radical-dibenzo [b; d] furans (160mg; 0.81mmol) be dissolved in anhydrous methylene chloride (4mL); then metachloroperbenzoic acid (m-CPBA) is slowly added under room temperature condition; finish, reaction solution at room temperature stirs and spends the night to reaction end.Then add water (2mL), saturated sodium bicarbonate aqueous solution (4mL), methylene dichloride (20mL), extraction, filtration drying, concentrate to obtain title compound, yellow solid.
The preparation of 1.53-hydroxyl-dibenzo [b, d] furans
3-methanoyl-dibenzo [b, d] furans (130mg, 0.6mmol) is dissolved in methyl alcohol (3mL), then 5% aqueous sodium hydroxide solution (3mL) is slowly added, finish, in room temperature reaction 30 minutes, reaction terminated, with the salt acid for adjusting pH value to 2 of 2N, with dichloromethane extraction, washing, anhydrous sodium sulfate drying, column chromatography (sherwood oil: ethyl acetate=50:1) obtains title compound, white solid.
1H?NMR(400MHz,DMSO-d 6)δppm:9.94(s,1H),7.95-7.98(m,1H),7.88-7.90(m,1H),7.58-7.60(d,1H),7.32-7.34(m,2H),7.01-7.02(d,1H),6.83-6.86(m,1H)。
Step 2, (2S)-2-(((dibenzo [b; d] furans-3-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 3-hydroxyl-dibenzo [b, d] furans, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.
1H?NMR(400MHz,DMSO-d 6)δppm:11.51(s,1H),8.11-8.15(m,2H),7.68-7.71(d,1H),7.60-7.61(m,2H),7.51(m,1H),7.39(m,1H),7.28(m,1H),6.17-6.18(m,1H),5.89(m,1H),5.88(m,1H),5.56-5.59(d,1H),4.81-4.85(m,1H),4.40(m,1H),4.28(m,1H),4.03(m,1H),3.85-3.88(m,2H),1.23-1.29(m,6H),1.09-1.12(m,6H)。
LC-MS?m/z:[M+H] +＝620。
Embodiment 21 (2S)-2-(((spiral shell [cyclopropane-1; 9 '-fluorenes]-2 '-Ji oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,2 '-hydroxyl spiral shell [cyclopropane-1,9 '-fluorenes]
The preparation of the bromo-9-of 1.12-(2-bromotrifluoromethane)-9H-fluorenes
In reaction flask, 2-bromine fluorenes (2g) is dissolved in THF (10mL), slowly adds LiHMDS (LHMDS) (9.8mL) in-78 DEG C, add and stir 1h, then to 0 DEG C.Said mixture is slowly joined 1, in THF (10mL) solution of 2-ethylene dibromide (10.5g), reaction mixture continues to react 1h at 0 DEG C, TLC display reaction terminates, with methyl alcohol (1mL) cancellation, except desolventizing, crude product obtains title compound through silica gel column chromatography (PE:EA=100:1) purifying, yellow solid product.
1H?NMR(400MHz,DMSO-d 6)δppm:7.83-7.91(m,3H),7.57-7.65(m,2H),7.37-7.41(m,2H),4.16(m,1H),3.40-3.44(m,2H),2.43-2.50(m,2H)。
The preparation of 1.22 '-bromine spiral shell [cyclopropane-1,9 '-fluorenes]
In reaction flask, bromo-for 2-9-(2-bromotrifluoromethane)-9H-fluorenes (1.1g) is dissolved in DMF (5mL), slowly adds NaH (248mg) in 0 DEG C, then to room temperature, react 12h.TLC display reaction terminates, and uses saturated NH 4the cancellation of Cl (1mL) solution, add EA (20mL) dilution, and use the LiCl solution (5mL × 3) of 10% successively, saturated aqueous common salt (5mL) washes, merge organic phase, anhydrous sodium sulfate drying, filters, decompression removing ethyl acetate, crude product obtains title compound, yellow solid through silica gel column chromatography (PE:EA=50:1) purifying.
1H?NMR(400MHz,DMSO-d 6)δppm:7.87-7.95(m,2H),7.51-7.53(m,1H),7.46(s,1H),7.31-7.38(m,2H),7.19-7.21(m,1H),1.80-1.86(m,2H),1.74-1.79(m,2H)。
The preparation of 1.32 '-formyl radical spiral shell [cyclopropane-1,9 '-fluorenes]
In reaction flask, 2 '-bromine spiral shell [cyclopropane-1,9 '-fluorenes] (500mg) is dissolved in THF (5mL), under-78 DEG C with nitrogen protection, slowly adds n-Butyl Lithium (0.86mL), under maintaining this temperature, react 1h; Then add DMF (0.5mL), stop cooling, be naturally warming up to room temperature, and TLC display reaction terminates, and uses saturated NH 4the cancellation of Cl (1mL) solution, add EA (20mL) dilution, and use the LiCl solution (5mL × 2) of 10% successively, saturated aqueous common salt (5mL) washs, and merges organic phase, anhydrous sodium sulfate drying, filter, removal of solvent under reduced pressure, crude product silica gel column chromatography (PET:EA=30:1) purifying obtains title compound, yellow solid.
1H?NMR(400MHz,DMSO-d 6)δppm:10.03(s,1H),8.16-8.18(d,1H),8.06-8.09(m,1H),7.92-7.94(m,1H),7.766(s,1H),7.41-7.43(m,2H),7.27-7.30(m,1H),1.90-1.94(m,2H),1.81-1.84(m,2H)。
The preparation of 1.42 '-methanoyl spiral shell [cyclopropane-1,9 '-fluorenes]
In reaction flask; by 2 '-formyl radical spiral shell [cyclopropane-1; 9 '-fluorenes] (210mg) be dissolved in DCM (methylene dichloride) (10mL); slowly add under room temperature m-CPBA (328mg) (metachloroperbenzoic acid); then at 25 DEG C, react 3h, terminate to TLC display reaction.Add DCM (20mL) dilution again, and successively with saturated NaHCO 3solution (5mL × 2), saturated aqueous common salt (5mL) washs, and merges organic phase, anhydrous sodium sulfate drying, filters, and decompression removing DCM, obtains title compound, yellow solid, is directly used in next step reaction.
The preparation of 1.52 '-hydroxyl spiral shell [cyclopropane-1,9 '-fluorenes]
In reaction flask, 2 '-methanoyl spiral shell [cyclopropane-1,9 '-fluorenes] (260mg) is dissolved in MeOH/H 2in O (3mL/lmL), under room temperature, slowly add K 2cO 3(303mg), then at 50 DEG C, react 1.5h, terminate to TLC display reaction.Decompression removing methyl alcohol, with EA (20mL) dilution, and uses H successively 2o (5mL × 2), saturated aqueous common salt (5mL) is washed, and merges organic phase, anhydrous sodium sulfate drying, filters, and decompression removing EA, crude product obtains title compound, faint yellow solid through silica gel column chromatography (PE:EA=15:1) purifying.
1H?NMR(400MHz,DMSO-d 6)δppm:9.48(s,1H),7.67-7.74(m,2H),7.08-7.29(m,3H),6.74-6.77(m,1H),6.52-6.53(m,1H),1.67-1.71(m,2H),1.60-1.63(m,2H)。
Step 2, (2S)-2-(((spiral shell [cyclopropane-1; 9 '-fluorenes]-2 '-Ji oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 2 '-hydroxyl spiral shell [cyclopropane-1,9 '-fluorenes], phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained title compound.
1H?NMR(400MHz,DMSO-d 6)δppm:11.44(s,1H),7.88-7.91(s,2H),7.57-7.59(m,1H),7.17-7.36(m,4H),7.07(s,1H),6.07-6.13(m,2H),5.93(br,1H),5.56-5.58(m,1H),4.83-4.87(m,1H),4.37-4.41(m,1H),4.25-4.27(m,1H),4.0-4.04(m,1H),3.80-3.87(m,2H),1.69-1.78(m,4H),1.23-1.28(m,6H),1.13-1.15(m,6H)。
LC-MS?m/z:[M+H] +＝644。
Embodiment 22 (2S)-2-(((pyrido [1; 2-a] benzoglyoxaline-7-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,7-Hydroxy-pyridine also [1,2a] benzoglyoxaline
The preparation of 1.11-bromine 4-methoxyl group-2-oil of mirbane
4-methoxyl group-2-N-methyl-p-nitroaniline (1.68g) is dissolved in acetonitrile (30mL); then at ice bath temperature, nitrite tert-butyl (1.34g) is slowly dripped; again cupric bromide (2.67g) is slowly joined in reaction flask; reaction solution is heated to 80 DEG C of reactions 2 hours under nitrogen protection; reaction terminates; concentrating under reduced pressure, obtains title compound through silica gel chromatography (PE:EA=20:1), yellow solid.
The preparation of the bromo-5-anisidine of 1.22-
1-bromine 4-methoxyl group-2-oil of mirbane (2.1g) is dissolved in methyl alcohol (10mL), then iron powder (1.5g) is added, concentrated hydrochloric acid (2mL) is added again in reaction solution, after reaction solution backflow 2h, cool to room temperature, add sodium carbonate solid until do not have bubble to emerge, extract with EA, EA layer saturated common salt is washed, drying, filters, removal of solvent under reduced pressure, thick product is separated (PE:EA=20:1) purifying through silica gel column chromatography and obtains title compound, flaxen oily matter.
LC-MS?m/z:[M+H] +＝202。
The preparation of 1.3N-(the bromo-5-p-methoxy-phenyl of 2-) pyridyl-2-amine
Bromo-for 2-5-anisidine (1.2g) is dissolved in anhydrous dioxane (6mL), add 2-bromopyridine (0.68mL), three (dibenzalacetone) two palladium (Pd2 (dba) 3, 100mg), 4, the two diphenylphosphine-9 of 5-, 9-dimethyl xanthene (Xant-phos, 50mg) with cesium carbonate (2.87g), finish, reaction solution is heated to 100 DEG C of stirring reactions 3 hours under nitrogen protection, reaction terminates, removal of solvent under reduced pressure, direct silica gel column chromatography is separated (PET:EA=30:1) and obtains title compound, faint yellow oily product.
LC-MS?m/z:[M+H]+＝280。
The preparation of 1.47-methoxv-pyridine also [1,2a] benzoglyoxaline
N-(the bromo-5-p-methoxy-phenyl of 2-) pyridyl-2-amine (1g) is dissolved in anhydrous acetonitrile (10mL); then salt of wormwood (590mg) is added; N; N'-dimethyl-ethylenediamine (315mg) and cuprous iodide (68mg), reaction solution is heated to 80 DEG C of reactions under nitrogen protection and spends the night, and reaction terminates; filter; by filtrate reduced in volume, the direct silica gel column chromatography of concentrated solution is separated and obtains title compound, yellow solid.
1H?NMR(400MHz,DMSO-d 6)δppm:9.06-9.08(d,1H),8.23-8.25(d,1H),7.66-7.68(d,1H),7.56-7.58(m,1H),7.31-7.32(d,1H),7.01-7.05(m,1H),3.91(s,3H)。
LC-MS?m/z:[M+H] +＝199。
The preparation of 1.57-Hydroxy-pyridine also [1,2a] benzoglyoxaline
By 7-methoxv-pyridine also [1,2a] benzoglyoxaline (210mg) is dissolved in methylene dichloride (10mL), be cooled to-78 DEG C, then boron tribromide (550mg) is slowly added, finish, react 1 hour at ambient temperature, after reaction terminates, use frozen water cancellation, then add methyl alcohol (5mL), wash with saturated sodium bicarbonate aqueous solution, extract by ethyl acetate (30mL × 3), anhydrous sodium sulfate drying, concentrates and obtains title compound, faint yellow solid.
1H?NMR(400MHz,DMSO-d 6)δppm:9.55(br,1H),8.95-8.97(d,1H),8.07-8.09(d,1H),7.45-7.57(m,2H),7.05-7.06(d,1H),6.91-6.94(m,1H),6.84-6.86(m,1H)。
LC-MS?m/z:[M+H] +＝185。
Step 2, (2S)-2-(((pyrido [1; 2-a] benzoglyoxaline-7-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 7-Hydroxy-pyridine, also [1,2a] benzoglyoxaline, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-, for raw material, with the method for embodiment 1, obtain title compound.
1H?NMR(400MHz,DMSO-d 6)δppm:11.51(br,1H),9.07-9.09(d,1H),8.30-8.32(d,1H),7.55-7.68(m,4H),7.22-7.25(m,1H),7.01-7.04(m,1H),5.92-6.14(m,3H),5.51-5.57(m,1H),4.82-4.85(m,1H),4.28-4.38(m,2H),4.02-4.04(m,1H),3.80-3.89(m,2H),1.19-1.28(m,6H),1.12-1.15(m,6H)。
LC-MS?m/z:[M+H] +＝620。
Embodiment 23 (2S)-2-(((pyrido [1; 2-a] benzoglyoxaline-8-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,8-Hydroxy-pyridine also [1,2a] benzoglyoxaline
The preparation of 1.12-bromine 4-methoxyl group-1-oil of mirbane
5-methoxyl group-2-N-methyl-p-nitroaniline (3g) is dissolved in acetonitrile (30mL); then at ice bath temperature, nitrite tert-butyl (2.78mL) is slowly dripped; again cupric bromide (4.7g) is slowly joined in reaction flask; reaction solution is heated to 65 DEG C of reactions 2 hours under nitrogen protection; reaction terminates; directly remove solvent under reduced pressure, obtain title compound through column chromatographic isolation and purification.
The preparation of the bromo-4-anisidine of 1.22-
2-bromine 4-methoxyl group-1-oil of mirbane (4.3g) is dissolved in ethanol (15mL), then iron powder (4g) and ammonium chloride (7.8g) is added, finally add water (15mL), reaction solution is placed in 90 DEG C of reactions 3 hours, reaction terminates, by reaction solution cool to room temperature, leach solid, and use washed with dichloromethane filter cake, filtrate dichloromethane extraction, saturated common salt water washing, dry, title compound is obtained, brown oil through column chromatographic isolation and purification.
1H?NMR(400MHz,DMSO-d 6)δppm:6.69-6.96(m,1H),6.67-6.74(m,2H),4.08(s,2H),3.64(s,3H)。
LC-MS?m/z:[M+H] +＝202。
The preparation of 1.3N-(the bromo-4-p-methoxy-phenyl of 2-) pyridyl-2-amine
Bromo-for 2-4-anisidine (1.2g) is dissolved in anhydrous dioxane (6mL); add 2-bromopyridine (0.68mL); three (dibenzalacetone) two palladium (100mg); two diphenylphosphine-9,9-dimethyl xanthene (50mg) of 4,5-and cesium carbonate (2.8) are g); finish; reaction solution is heated to 100 DEG C of stirring reactions 3 hours under nitrogen protection, and reaction terminates, concentrating under reduced pressure.Residue is directly separated through silicagel column and obtains title compound, yellow solid.
1H?NMR(400MHz,DMSO-d 6)δppm:8.13(s,1H),7.99-8.00(m,1H),7.46-7.54(m,1H),7.22(m,1H),6.93-6.94(m,1H),6.63-6.66(m,2H),3.76(s,3H)。
LC-MS?m/z:[M+H] +＝280。
The preparation of 1.48-methoxv-pyridine also [1,2a] benzoglyoxaline
N-(the bromo-4-p-methoxy-phenyl of 2-) pyridyl-2-amine (1g) is dissolved in anhydrous acetonitrile (10mL); then salt of wormwood (590mg) is added; N; N'-dimethyl-ethylenediamine (315mg) and cuprous iodide (68mg), reaction solution is heated to 80 DEG C of reactions under nitrogen protection and spends the night, and reaction terminates; leach solid; filtrate reduced in volume, concentrated solution is directly separated through silicagel column and obtains title compound, yellow solid.
1H?NMR(400MHz,DMSO-d 6)δppm:8.98-9.00(d,1H),7.90(s,1H),7.68-7.71(d,1H),7.58-7.60(m,1H),7.42-7.44(m,1H),7.11-7.14(m,1H),6.92-6.95(m,1H)，3.88(s,3H)。
LC-MS?m/z:[M+H] +＝199。
The preparation of 1.58-Hydroxy-pyridine also [1,2a] benzoglyoxaline
By 8-methoxv-pyridine also [1,2a] benzoglyoxaline (100mg) is dissolved in methylene dichloride (3mL), be cooled to-78 DEG C, then boron tribromide (0.096mL) is slowly added, finish, react 1 hour at ambient temperature, after reaction terminates, use frozen water cancellation, then add methyl alcohol (5mL), wash with saturated sodium bicarbonate aqueous solution, extract by ethyl acetate (30mL × 2), anhydrous sodium sulfate drying, concentrates and obtains title compound, faint yellow solid.
1H?NMR(400MHz,DMSO-d 6)δppm:9.54(s,1H),8.84-8.86(d,1H),7.60-7.62(d,1H),7.54-7.55(m,2H),7.38-7.40(m,1H),7.01-7.04(d,1H),6.85-6.88(m,1H)。
LC-MS?m/z:[M+H] +＝185。
Step 2, (2S)-2-(((pyrido [1; 2-a] benzoglyoxaline-7-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With 8-Hydroxy-pyridine, also [1,2a] benzoglyoxaline, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-, for raw material, with the method for embodiment 1, obtain title compound.
1H?NMR(400MHz,DMSO-d 6)δppm:11.50(s,1H),8.98-8.99(d,1H),8.19(s,1H),7.77-7.80(m,1H),7.64(m,1H),7.54(m,2H),7.40-7.43(m,1H),6.97-6.99(m,1H),6.07-6.13(m,2H),5.91(m,1H),5.51-5.55(m,1H),4.80-4.84(m,1H),4.41-4.44(m,1H),4.31(m,1H),3.88-4.02(m,1H),3.82-3.86(m,2H),1.21-1.26(m,6H),1.08-1.11(m,6H)。
LC-MS?m/z:[M+H] +＝620。
Embodiment 24 (2S)-2-(((9-methyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) ethyl propionate
With the 4-hydroxyl-9-methyl-9H-carbazole of embodiment 2 step 1 preparation, dichloro oxygen phosphorus, ALANINE carbethoxy hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained target compound.
1H?NMR(300MHz,DMSO-d 6)δppm:11.46(s,1H),8.20-8.23(d,1H),7.61-7.63(m,1H),7.40-7.50(m,4H),7.17-7.24(m,2H),6.24-6.32(m,1H),5.90-6.05(m,2H),5.15(s,1H),4.83-4.89(m,1H),4.32(m,1H),4.04(m,1H),3.92-3.93(m,2H),3.84-3.92(m,5H),1.05-1.22(m,6H)，1.01-1.02(m,3H)。
LC-MS?m/z:[M+H] +＝619。
Embodiment 25 (2S)-2-(((9-methyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) propionic acid ring pentyl ester
The synthesis of step 1, ALANINE ring ester hydrochloride
Take ALANINE (8.9g, 100mmol) to drop in 250mL round-bottomed flask, add cyclopentanol 20ml, under condition of ice bath, slowly add thionyl chloride (11.9g, 100mmol), slowly be warming up to room temperature, at room temperature reaction is spent the night, concentrated, adds isopropyl ether, pull an oar under-10 DEG C of conditions, separate out a large amount of white solid, filter, obtain title compound.
Step 2, (2S)-2-(((9-methyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) benzyl propionate
With the 4-hydroxyl-9-methyl-9H-carbazole of embodiment 2 step 1 preparation, phosphorus oxychloride, ALANINE ring ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained target compound.
1H?NMR(300MHz,DMSO-d 6)δppm:11.15(s,1H),8.21-8.24(d,1H),7.59-7.62(m,1H),7.41-7.50(m,4H),7.16-7.21(m,2H),6.22-6.26(m,1H),6.04-6.18(m,2H),5.19(s,1H),5.02-5.17(m,1H),4.44-4.45(m,1H),4.42(m,1H),4.01-4.02(m,1H),3.88-3.92(m,5H),1.25-1.28(m,6H)，1.21-122(m,8H)。
LC-MS?m/z:[M+H] +＝659。
Embodiment 26 (2S)-2-(((9-methyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) benzyl propionate
With the 4-hydroxyl-9-methyl-9H-carbazole of embodiment 2 step 1 preparation, phosphorus oxychloride, ALANINE benzyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained target compound.
1H?NMR(300MHz,DMSO-d 6)δppm:11.508(s,1H),8.22-8.25(d,1H),7.59-7.62(m,1H),7.47-7.50(m,3H),7.38-7.41(m,4H),7.13-7.18(m,3H),6.36-6.40(m,1H),5.90-6.04(m,2H),4.83-5.15(m,3H),4.33-4.44(m,2H),4.04(m,2H),3.88(m,3H),3.55(m,2H),1.24-1.26(m,6H)。
LC-MS?m/z:[M+H] +＝681。
Embodiment 27 (2S)-2-(((9-methyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) propionic acid peopentyl ester
The synthesis of step 1, ALANINE peopentyl ester hydrochloride
With ALANINE and neopentyl alcohol for raw material, with the method for embodiment 25 step 1, obtained title compound.
Step 2, (2S)-2-(((9-methyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) propionic acid peopentyl ester
With the 4-hydroxyl-9-methyl-9H-carbazole of embodiment 2 step 1 preparation, phosphorus oxychloride, ALANINE peopentyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained target compound.
1H?NMR(300MHz,DMSO-d 6)δppm:11.48(s,1H),8.22-8.24(d,1H),7.59-7.62(m,1H),7.45-7.48(m,4H),7.15-7.22(m,2H),6.29-6.33(m,1H),5.91-5.98(m,1H),5.20-5.22(s,1H),4.46-4.47(m,1H),4.32(m,1H),4.29-4.31(m,1H),4.06-4.12(m,1H),3.96-3.99(m,1H),3.84-3.92(m,4H),3.80-3.87(m,2H),1.24-1.28(m,6H)，1.22-1.23(m,9H)。
LC-MS?m/z:[M+H] +＝661。
Embodiment 28 (2S)-2-(((9; 9-dimethyl-9H-fluorenes-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The synthesis of step 1,2'-methyl diphenyl-2-methyl-formiate
Take 2-methylphenylboronic acid (1g, 7.35mmol), o-iodobenzoic acid methyl esters (1.75g, 6.7mmol), Pd (dppf) Cl 2(0.47g, 0.67mmol), cesium carbonate (2.8g; 20mmol) throw in the eggplant-shape bottle of 250mL, then add Isosorbide-5-Nitrae-dioxane 50mL and water 5mL; the lower 90 DEG C of reaction 1.5h of nitrogen protection, after reaction terminates, add ethyl acetate (100mL); saturated sodium-chloride water solution (50mL), extracts with separating funnel, washes 3 times; then organic phase is collected; come dry by anhydrous sodium sulphate, filter, concentrate, column chromatography purification obtains title compound.
LC-MS?m/z:[M+H] +＝227。
The synthesis of step 2,4-methyl-9-Fluorenone
Take step 1 gains 2 '-methyl diphenyl-2-carboxylate methyl ester (1g, 4.42mmol), throw in the eggplant-shape bottle of 100mL, then vitriol oil 2ml is added, stirring at room temperature 1.5h, stopped reaction, reaction solution is poured in frozen water, add saturated sodium carbonate solution, regulate pH value to neutral, add ethyl acetate (100mL), extract with separating funnel, wash 3 times, then collect organic phase, come dry by anhydrous sodium sulphate, filter, concentrated, column chromatography purification obtains title compound.
LC-MS?m/z:[M+H] +＝195。
The synthesis of step 3,4,9,9-trimethylammonium-9-fluorenes
In the single port bottle of 50ml, argon replaces 3 times, measure zinc methide toluene solution (30ml, 30mmol), titanium tetrachloride (3.5ml is added under the condition of-50 DEG C, 60mmol) stir 1 hour, again by step 2 gains 4-methyl-9-Fluorenone (1.94g, 10mmol) be added in 2ml methylene dichloride, in slow instillation single port bottle, after reacting completely, pour cancellation in frozen water into, add ethyl acetate (500mL), extract with separating funnel, wash 3 times, then organic phase is collected, come dry by anhydrous sodium sulphate, filter, concentrated, column chromatography purification obtains title compound.
LC-MS?m/z:[M+H] +＝209。
Step 4,9,9-dimethyl-4-methylol-9H-fluorenes
In the single port bottle of 100mL, add step 3 gains 4,9,9-trimethylammonium-9-fluorenes (1.1g, 5mmol), tetracol phenixin 5ml, NBS (1g, 5.6mmol) and Diisopropyl azodicarboxylate (AIBN, 700mg, 4.2mmol), heated overnight at reflux, add ethyl acetate (500mL), extract with separating funnel, wash 3 times, then collect organic phase, come dry by anhydrous sodium sulphate, concentrate and obtain crude product.Gained crude product is dropped in the single port bottle of 250mL, add salt of wormwood (910mg, 6.6mmol), water 20ml and 1,4-dioxane 50ml, is heated to 90 DEG C, reacts 3 hours, react completely, add ethyl acetate (500mL), extract with separating funnel, wash 3 times, then collect organic phase, come dry by anhydrous sodium sulphate, filter, concentrated, column chromatography purification obtains title compound.
LC-MS?m/z:[M+H] +＝225。
Step 5,9,9-dimethyl-4-formyl radical-9H-fluorenes
Step 4 gains 9,9-dimethyl-4-methylol-9H-fluorenes (400mg, 1.7mmol) are dropped in the single port bottle of 50mL, add methylene dichloride 15ml, under condition of ice bath, add Pyridinium chlorochromate on silica gel (PCC, 1.2g, 6mmol), after reacting completely, add ethyl acetate (200mL), extract with separating funnel, wash 3 times, then collect organic phase, come dry by anhydrous sodium sulphate, filter, concentrated, column chromatography purification obtains title compound.
LC-MS?m/z:[M+H] +＝223。
The synthesis of step 6,4-hydroxyl-9,9-dimethyl-9-fluorenes
By step 5 gains 9, 9-dimethyl-4-formyl radical-9H-fluorenes (200mg, 0.95mmol) drop in the single port bottle of 50mL, add methylene dichloride 15ml, under ice bath, add metachloroperbenzoic acid (mCPBA, 500mg, 3mmol), rise to room temperature, stirred overnight at room temperature, after reacting completely, add ethyl acetate (200mL), extract with separating funnel, organic addition 10% sodium thiosulfate solution washes 3 times, come dry by anhydrous sodium sulphate, filter, after concentrate drying, add 15ml methyl alcohol, KOH (56mg, 1mmol), normal-temperature reaction, after reacting completely, add ethyl acetate (200mL), extract with separating funnel, wash 3 times, then organic phase is collected, come dry by anhydrous sodium sulphate, filter, concentrated, column chromatography purification obtains title compound.
LC-MS?m/z:[M+H] +＝211。
Step 7, (2S)-2-(((9; 9-dimethyl-9H-fluorenes-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
With step 6 gains 4-hydroxyl-9,9-dimethyl-9-fluorenes, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained target compound.
1H?NMR(300MHz,DMSO-d 6)δppm:11.46(s,1H),8.20-8.23(d,1H),7.66-7.69(m,1H),7.38-7.47(m,4H),7.19-7.21(m,2H),6.23-6.27(m,1H),6.02-6.19(m,1H),5.86-5.95(m,1H),5.08(s,1H),4.88-4.90(m,1H),4.45-47(m,1H),4.30-41(m,1H),4.28-4.31(m,1H),3.92-3.94(m,2H),1.05-1.22(m,18H)。
LC-MS?m/z:[M+H] +＝659。
Embodiment 29 (2S)-2-(((9-cyclopropyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The synthesis of step 1,4-methoxyl group-9H-carbazole
Take 4-hydroxycarbazole (9.1g, 50mmol), salt of wormwood (6.9g, 50mmol) drop in the three-necked bottle of 500ml, add 250ml acetone, under condition of ice bath, slowly add methyl-sulfate (4.8ml, 50mmol), under room temperature, react 2h, after reaction terminates, add ethyl acetate (100mL) dilution, organic phase washed with water, saturated common salt is washed, dry, filters, concentrated, column chromatography purification obtains title compound.
LC-MS?m/z:[M+H] +＝198。
The synthesis of step 2,9-cyclopropyl-4-methoxyl group-9H-carbazole
Take step 1 gains 4-methoxyl group-9H-carbazole (3.96g, 20mmol), neutralized verdigris (4.0g, 20mmol), cyclopropylboronic acid (3.44g, 40mmol) with DMAP (7.32g, 60mmol) drop in the eggplant-shape bottle of 500ml, add 200ml toluene and 20ml bis-(trimethyl silicon based) sodium amide (NaHMDS), 95 DEG C of reaction 4h, after reacting completely, add ethyl acetate (500mL) dilution, organic phase washed with water, saturated common salt is washed, dry, filters, concentrated, column chromatography purification obtains title compound.LC-MS?m/z:[M+H] +＝238。
The synthesis of step 3,9-cyclopropyl-4-hydroxyl-9H-carbazole
Taking step 2 gains 9-cyclopropyl-4-methoxyl group-9H-carbazole (1.19g, 5mmol) drops in the eggplant-shape bottle of 250ml, adds 100ml methylene dichloride, under-50 DEG C of conditions, slowly adds BBr 3(7ml, 7.5mmol), finishes, and-20 DEG C of reaction 4h, after reacting completely, slowly add 200ml water, ethyl acetate (500mL) extracts, organic phase washed with water, and saturated common salt is washed, drying, filters, and concentrated, column chromatography purification obtains title compound.
LC-MS?m/z:[M+H] +＝224。
Step 4, (2S)-2-(((9-cyclopropyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With step 3 gains 9-cyclopropyl-4-hydroxyl-9H-carbazole, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained target compound.
1H?NMR(300MHz,DMSO-d 6)δppm:11.45(s,1H),7.97-7.98(d,1H),7.53-7.56(m,2H),7.36-7.38(m,2H),7.27-7.30(m,3H),6.20-6.22(m,1H),6.18-6.20(m,1H),6.00-6.05(m,1H),5.31(s,1H),4.86-4.89(m,1H),4.45-4.47(m,1H),4.32-4.33(m,1H),4.29-4.31(m,1H),3.87-3.89(m,2H),2.42(m,1H),1.42-1.43(m,6H)，1.32(m,2H),1.28(m,2H),1.23-1.24(m,6H)。
LC-MS?m/z:[M+H] +＝659。
Embodiment 30 (2S)-2-(((dibenzo [b; d] furans-1-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,3,4,6,7,8,9-six diphenyl hydrogens also [b, d] furans-1 (2H)-one
In the there-necked flask of 250mL, add hydroresorcinol (11g, 0.1mol), pimelinketone (10g, 0.1mol), dimethylbenzene 100ml, tosic acid 0.45g (0.01mol), adds water trap, 170 DEG C of backflow 5h, after reacting completely, cool to room temperature, concentrated, column chromatography purification obtains title compound.
The preparation of step 2,1-hydroxyl dibenzo [b, d] furans
In the single port bottle of 250mL, add step 1 gains 3,4; 6,7,8; 9-six diphenyl hydrogen is [b, d] furans-1 (2H)-one 5.2g (0.027mmol), cymene 80mL, 10%Pa/C3g also; under nitrogen protection, 170 DEG C are refluxed 18 hours, after reacting completely; cooling; filter, concentrated, column chromatography purification obtains title compound.
Step 3, (2S)-2-(((dibenzo [b; d] furans-1-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) preparation of isopropyl propionate
With step 2 gains 1-hydroxyl dibenzo [b, d] furans, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-be raw material, with the method for embodiment 1, obtained target compound.
1HNMR((300MHz,DMSO-d 6)δppm:11.49(s,1H),8.43(m,1H),8.16(d,1H),7.74(d,1H),7.53-7.38(m,5H),6.34(m,1H),6.05(m,1H),5.89(m,1H),5.37(m,1H),5.19(m,1H),4,84(m,1H),4.45(m,1H),4.04(m,1H),3.91(m,2H),1,26-1.10(m,12H)。
LC-MS?m/z:[M+H] +＝620。
Embodiment 31 (2S)-2-(((5-methyl-6-oxo-5; 6-dihydrophenanthridine-10-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,3-hydroxy-n-(2-iodophenyl) benzamide
M-Salicylic acid (1g, 7.25mmol) is dissolved in 20ML N,N-dimethylacetamide, drip thionyl chloride (1.3g, 10.9mmol) at 0-4 DEG C, finish, after room temperature reaction 0.5h, drip adjacent Iodoaniline (1.6g, 7.25mmol), finish, continue reaction 3h under room temperature, after reacting completely, add water and extraction into ethyl acetate, dry, filter, concentrated, column chromatography purification obtains title compound.
The preparation of step 2,3-methoxyl group-N-(2-iodophenyl)-N-methyl-benzamide
In the single port bottle of 100mL, add step 1 gains 3-hydroxy-n-(2-iodophenyl) benzamide (0.5g, 1.47mmol) and be dissolved in DMF, under ice bath, add NaH (0.2g, 4.42mmol), after stirring 0.5h, add methyl-sulfate 1.33g (10.5mml), finish, room temperature reaction 2h, reacts completely, and add water under ice bath cancellation, ethyl acetate and water extraction, dry, filter, concentrated, column chromatography purification obtains title compound.
The preparation of step 3,10-methoxyl group-5-methyl phenanthridines-6 (5H)-one
In the single port bottle of 100mL, add step 2 gains 3-methoxyl group-N-(2-iodophenyl)-N-methyl-benzamide (0.5g, 1.36mmol), be dissolved in DMF20ml, add Pd (OAc) 2(50mg, 0.22mmol), PPh 3(70mg, 0.267mmol), K 2cO 3(0.4g, 2.9mmol), nitrogen replacement three times, 160 DEG C of backflow 3h, lower the temperature after reacting completely, add water and extraction into ethyl acetate, dry, and filter, concentrated, column chromatography purification obtains title compound.
The preparation of step 4,10-hydroxy-5-methyl base phenanthridines-6 (5H)-one
In the single port bottle of 100mL, add step 3 gains 10-methoxyl group-5-methyl phenanthridines-6 (5H)-one (0.15g, 0.6mmol), HBr/H 2o (40%, 5ml), acetic acid 5mL, 110 DEG C of reactions, after reacting completely, add water and extraction into ethyl acetate, dry, filter, and concentrated, column chromatography purification obtains title compound.
Step 5, (2S)-2-(((5-methyl-6-oxo-5; 6-dihydrophenanthridine-10-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
With step 4 gains 10-hydroxy-5-methyl base phenanthridines-6 (5H)-one, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained target compound.
1H?NMR(500MHz,DMSO-d 6)δppm:11.47(s,1H),9.04(d,1H)，8.30(d,1H),7.8-7.3(m,6H),6.39-6.30(m,1H),5.80-6.06(m,2H),5.20(m,1H),4.80(m,2H),4.40-4.25(m,2H),3.91(m,2H),3.78(s,3H),1.05-1.25(m,12H)。
LC-MS?m/z:[M+H]+＝661。
Embodiment 32 (2S)-2-(((9-methyl-9H-carbazole-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) methyl propionate
With embodiment 2 step 1 gains 4-hydroxyl-9-methyl-9H-carbazole, phosphorus oxychloride, ALANINE methyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained target compound.
1H?NMR(300MHz,DMSO-d 6)δppm:11.47(s,1H),8.23(d,1H),7.61-7.63(d,1H),7.40-7.50(m,4H),7.17-7.24(m,2H),6.24-6.32(m,1H),5.90-6.05(m,2H),5.15(s,1H),4.83-4.89(m,1H),4.32(m,1H),4.04(m,1H),3.92-3.93(m,5H),3.60(s,3H),1.05-1.25(m,6H)。
LC-MS?m/z:[M+H] +＝605。
Embodiment 33 (2S)-2-(((9H-fluorenes-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1,4-hydroxyl-9H-fluorenes-9-ketone
With embodiment 28 step 2 gains 4-methyl-9-Fluorenone for raw material, with embodiment 28 step 3,4, the method for 5 and 6 obtains title compound.
The preparation of step 2,4-hydroxyl-9H-fluorenes
In the single port bottle of 100mL, add step 1 gains 4-hydroxyl-9H-fluorenes-9-ketone (200mg, 1.02mmol), be dissolved in methyl alcohol 20ml, add 10% palladium carbon 20mg, hydrogen is divided into lower room temperature reaction to spend the night, after reacting completely, filter, concentrated, obtain title compound.
Step 3, (2S)-2-(((9H-fluorenes-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) methyl propionate
With step 2 gains 4-hydroxyl-9H-fluorenes, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained target compound.
1H?NMR(300MHz,DMSO-d 6)δppm:11.46(s,1H),8.23(d,1H),7.61-7.20(m,7H),6.24-6.32(m,1H),5.85-6.05(m,2H),5.20(s,1H),4.83(m,1H),4.30(m,1H),4.14(m,1H),3.90(m,1H),3.80(d,2H),3.62(m,2H),1.05-1.25(m,12H)。
LC-MS?m/z:[M+H]+＝618.2。
Embodiment 34 (2S)-2-(((9-oxo-9H-fluorenes-4-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
With embodiment 33 step 1 gains 4-hydroxyl-9H-fluorenes-9-ketone, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained target compound.
1H?NMR(300MHz,DMSO-d 6)δppm:11.46(s,1H),8.23(d,1H),7.61-7.20(m,7H),6.24-6.32(m,1H),5.85-6.05(m,2H),5.20(s,1H),4.83(m,1H),4.30(m,1H),4.14(m,1H),3.90(m,1H),3.80(d,2H),1.05-1.25(m,12H)。
LC-MS?m/z:[M+H] +＝632。
Embodiment 35 (2S)-2-(((5-methyl-6-oxo-5; 6-dihydrophenanthridine-1-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
The preparation of step 1, N-(3-hydroxy phenyl)-2-iodobenzamide
In the single port bottle of 500ml, add Metha Amino Phenon 5g (45.9mmol), be dissolved in 80ml THF, add o-iodobenzoic acid 14g (56.5mmol), EDCI13g (67.8mmol) and DMAP1.3g (10.6mmol), stirred overnight at room temperature, after reacting completely, concentrated, add ethyl acetate and water extraction, dry, filter, concentrated, column chromatography purification obtains title compound.
The preparation of step 2,1-hydroxy-5-methyl base phenanthridines-6 (5H)-one
With step 1 gains N-(3-hydroxy phenyl)-2-iodobenzamide for raw material, according to embodiment 31 step 2,3 and 4 method obtain title compound.
Step 3, (2S)-2-(((5-methyl-6-oxo-5; 6-dihydrophenanthridine-1-base oxygen base) (((2R; 3R; 4R; 5R)-5-(2; 4-dioxo-3,4-dihydro-pyrimidin-1 (2H)-Ji)-4-fluoro-3-hydroxy-4-methyl tetrahydrofuran (THF)-2-base) methoxyl group) phosphoryl) amino) isopropyl propionate
With step 2 gains 1-hydroxy-5-methyl base phenanthridines-6 (5H)-one, phosphorus oxychloride, ALANINE isopropyl ester hydrochloride, Pentafluorophenol and the fluoro-2'-methyluridine of (2'R)-2'-deoxidation-2'-for raw material, with the method for embodiment 1, obtained target compound.
1H?NMR(300MHz,DMSO-d 6)δppm:11.50(s,1H),9.0(d,1H)8.23(d,1H),7.8-7.20(m,5H),6.40-6.30(m,1H),5.82-6.06(m,2H),5.20(m,1H),4.80(m,2H),4.40-4.25(m,2H),4.12(m,1H),3.91(m,1H),3.80(m,1H),3.65(s,3H),1.05-1.25(m,12H)。
LC-MS?m/z:[M+H]+＝661。
Experimental example 1 pharmacology activity research:
1. experiment material
1.1 reagent:
The list of table 1. reagent
Reagent name Supplier
DMEM cell culture medium Invitrogen
Foetal calf serum (FBS) Gibco
L-glutaminate Invitrogen
Pen .-Strep solution Invitrogen
DPBS/Modified Hyclone
Trypsinase/EDTA Invitrogen
Dimethyl sulfoxide (DMSO) (DMSO) Sigma
Bright-Glo Promega
Growth of Cells fluorometric titration detection reagent Promega
1.2Huh71b clone:
Huh71b clone is provided by Shanghai Yaoming Kangde New Medicine Development Co., Ltd, for comprising the Huh7 clone of the HCV1b replicon with stable luciferase (Luc) report.HCV nonstructural protein gene, neo (G418 resistance) and luciferase reporter gene are cloned into pBR vector construction by gene recombination technology by it.Then the carrier carrying HCV replicon is transfected into huh7 cell, by G418 resistance screening, HCV replicon Absorbable organic halogens copies and associated protein and luciferase are expressed at huh7 cell inner stablity.This cell model is used for the screening of HCV-Ab IgG Compound ira vitro.The activity of the HCV-Ab IgG of compound is measured by the expression level of inspection luciferase.See Lohmann V, et al.1999.Replication of subgenomic hepatitis C virus RNAs in a hepatoma cell line.Science.285 (5424): 110-113.
1.3 positive control drugs:
The contrast medicine structure used in experimental example of the present invention is: it is the compound of WO2008/121634 (PCT/US2008/058183) embodiment 25, i.e. (S)-2-{ [(2R, 3R, 4R, 5R)-5-(2, 4-dioxo-3, 4-dihydro-2H-pyrimidine-1-base)-4-fluoro-3-hydroxy-4-methyl-tetrahydro-furan-2-ylmethoxy]-phenoxy group-phosphoryl amino }-isopropyl propionate ((S)-2-{ [(2R, 3R, 4R, 5R)-5-(2, 4-dioxo-3, 4-dihydro-2H-pyrimidin-1-yl)-4-fluoro-3-hydroxy-4-me thyl-tetrahydro-furan-2-ylmethoxy]-phenoxy-phosphorylamino}-propionic acid isopropyl ester).
This compound is with reference to J.Org.chem, and the method described in 2011,76,8311-8319 is obtained and composed and Mass Spectrometric Identification by hydrogen.
2. experimental procedure:
2.1 compounds prepare: with full-automatic microwell plate pretreatment system (the LabCyte company of POD810, the U.S.) compound of the present invention above embodiment prepared joins in orifice plate, the initial final concentration of compound is 10 μMs, each compound does duplicate hole, 3 times of dilutions, 10 points, DMSO final concentration 0.5%;
2.2 cells prepare: plant Huh71b cell respectively to 96 orifice plates, 125 μ l systems, 8 × 10 3individual cells/well, 37 DEG C, 5%CO 2incubator culturing cell 72 hours;
2.3 cytoactives detect: every hole adds 30 μ l Growth of Cells fluorometric titration detection reagent, 37 DEG C, 5%CO 2incubator culturing cell 1 hour, spectrophotometer detects fluorescent signal value, and the data obtained is used for Compound Cytotoxicity and calculates;
2.4Bright-Glo detects: every hole adds 100 μ l luciferase luminous substrate Bright-Glo, with chemiluminescence detection system EnVision (PerkinElmer company in 5 minutes, the U.S.) detect fluorescent signal value, the data obtained is used for compound potencies and calculates.
2.5 data processing: use following formula the data obtained to be converted to cell viability per-cent (Viability%):
Viability % = CPD ZPE × 100
CPD: the fluorescent signal value of compound well
ZPE (Zero percent effect): invalid effect contrast fluorescent signal value
Following formula is used to be suppression percentage ratio (Inhibition%) by original data processing:
Inhibition % = CPD - HPE ZPE - HPE × 100
CPD: the fluorescent signal value of compound well
HPE (Hundred percent effect): 100% useful effect contrast fluorescent signal value
ZPE (Zero percent effect): invalid effect contrast fluorescent signal value
Suppression percentage ratio importing GraphPad Prism is processed further and draws homologous thread and EC 50value.Data are in table 2.
Table 2
Compound number EC 50(nM) Compound number EC 50(nM)
Positive control 80 Embodiment 1 113.6
Embodiment 2 32 Embodiment 3 60.46
Embodiment 4 75.69 Embodiment 5 539.2
Embodiment 6 363.1 Embodiment 7 31
Embodiment 8 316 Embodiment 9 >3330
Embodiment 10-I 291.1 Embodiment 10-II 1425
Embodiment 11 1475 Embodiment 12 638.4
Embodiment 13 165.3 Embodiment 14 622.1
Embodiment 15 62.53 ? ?
Embodiment 16-I 42.52 Embodiment 16-II 260.1
Embodiment 17 192.9 Embodiment 18 577.1
Embodiment 19 185.5 Embodiment 20 45
Embodiment 21 63 Embodiment 24 33
Embodiment 25 37 Embodiment 26 68
Embodiment 27 150 Embodiment 28 58
Embodiment 29 36 Embodiment 32 46
Embodiment 33 68 ? ?
Experimental example 2 compound of the present invention detects the antiviral activity of HCV infection model (HCVcc) 2a
1 experiment material
1.1 compound
Compound of the present invention prepared by above embodiment 2,7 and 25 and control compound, after being mixed with 10mM mother liquor, be diluted to 10 μMs with the DMEM complete culture solution containing 0.5%DMSO with DMSO, then 3 times of dilutions, totally 10 concentration successively.
1.2 cell
Huh7.5.1 cell, is provided by the bright Kant of medicine (Shanghai) new drug development company limited.
1.3 viral
HCVcc reporter virus, namely the transfection HCV total length mutant strain of luciferase and GFP, can produce the virus with JFH-1 wild-type with identical infection ability, provided by the bright Kant of medicine (Shanghai) new drug development company limited.
1.4 reagent
DMEM cell culture fluid (DMEM medium), purchased from American Invitrogen company;
Foetal calf serum (Fetal bovine serum, FBS), purchased from American Sigma company;
L-glutaminate (L (+)-Glutamine), purchased from American Invitrogen company;
Pen .-Strep (Pen-Strep), purchased from American Invitrogen company;
Phosphate buffered saline buffer (Phosphate buffered saline, PBS), purchased from American Hyclone company;
Pancreatin (Trypsin), purchased from American Invitrogen company;
Dimethyl sulfoxide (DMSO) (Dimethyl sulfoxide, DMSO), purchased from American Sigma company;
Cell pyrolysis liquid (lysis buffer), purchased from American Promega company;
Renillaluciferase detection reagent, purchased from American Promega company;
Alamar Blue detection reagent, purchased from American Invitrogen company.
1.4 instrument
The multi-functional microplate reader of EnVision, purchased from American Perkin-Elmer company.
2 experimental techniques
1) Huh7.5.1 cell prepares: collect the Huh7.5.1 cell of logarithmic phase, after resuspended with DMEM complete culture solution, to be inoculated in 96 orifice plates (7 × 10 3individual cells/well), be placed in 37 DEG C, 5%CO 2overnight incubation in incubator;
2) virus infection: after HCVcc reporter virus DMEM complete culture solution is resuspended, adds in 100 μ l viral supernatants (MOI=0.2) to above-mentioned 96 orifice plates;
3) compound prepares: in the Huh7.5.1 cell that HCVcc reporter virus infects, add embodiment 2, embodiment 7, the compound of embodiment 25 and above-mentioned control compound, each concentration of each compound establishes duplicate hole; Set up invalid effect control group (Zero percent effect simultaneously, ZPE) and 100% useful effect control group (Hundred percent effect, HPE): the complete culture solution of ZPE group containing 0.5%DMSO replaces compound, and HPE group hole is the cell of virus-free infection;
4) cell cultures: 96 orifice plates are placed in 37 DEG C, 5%CO 272hr is cultivated in incubator;
5) HCV-Ab IgG virus activity detects: cultivate after terminating, discard every hole supernatant, 20 μ l cell pyrolysis liquids and luciferase detection reagent is added in every hole, relative luminous intensity (RLU) is read by the multi-functional microplate reader of EnVision, raw data is used for compound anti-HCV activity and calculates, and calculation formula is:
Inhibition％＝(RLU ZPE-RLU CPD)/(RLU ZPE-RLU HPE)×100
Wherein RLU cPDfor the fluorescent signal value in test compounds hole, RLU zPEfor the fluorescent signal value of invalid effect control wells, RLU hPEit is the fluorescent signal value of 100% useful effect control wells.
6) cell viability detects: in 96 orifice plates, plant the Huh7.5.1 cell into equal amts, and add the compound treatment 72hr of the present invention of above-mentioned concentration gradient; Set up DMSO Vehicle controls group simultaneously.After cultivation terminates, in every hole, add 10%Alamar Blue detection reagent, 37 DEG C, 5%CO 2after 2hr cultivated by incubator, read relative intensity of fluorescence (RFU) by the multi-functional microplate reader of EnVision, use the data obtained to be used for Compound Cytotoxicity and calculate, calculation formula is:
Viability％＝RFU CPD/RFU DMSO×100
7) data processing: Inhibition%, Viability% are imported respectively GraphPad Prism software and carry out data processing, draws the medium effective concentration EC of compound to HCVcc 50with half cytotoxic concentration CC 50, experimental result is in table 3.
Table 3
Compound number EC 50(nM) CC 50(nM) Compound number EC 50(nM) CC 50(nM)
Positive control 12.18 >10000 Embodiment 2 2.074 >10000
Embodiment 7 1.604 >10000 Embodiment 25 0.615 >10000
As can be seen from Table 3, for HCV cell in vitro infection model, the compound of the embodiment of the present invention 2,7 and 25 has excellent antiviral activity, simultaneously little to cytotoxicity.
In addition, experiment also shows, use HCV cell in vitro infection model, other compounds of preparing of embodiments of the invention as embodiment 1,5,6,8,9,10-I, 10-II, 11,12,13,14,15,16-I, 16-II, 17,18,19,20,21,22,23,24,26,27,30,31,32,33,34, the compound of 35 etc. has low medium effective concentration EC to HCVcc GT2a virus 50and high half cytotoxic concentration CC 50, demonstrate good inhibit activities and little cytotoxicity.
Above experimental result shows, compound of the present invention has the ability suppressing HCV virus efficiently, compared with positive control drug, and EC 50have quite or more excellent effect, for the prospect that treatment HCV infection has had.
Although be below described in detail the present invention, it will be appreciated by those skilled in the art that and can carry out various amendment and change to the present invention under prerequisite without departing from the spirit and scope of the present invention.Interest field of the present invention is not limited to done detailed description above, and should belong to claims.

Claims (11)

1. three ring fused heterocycle type nucleoside phosphoramidate compounds, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, as shown in general formula (I),
P* represents chiral phosphorus atom,
Wherein,
(1) R 1be selected from H and alkyl, described alkyl is optionally replaced by one or more alkyl, alkoxyl group, alkylamino, halogen, hydroxyl, amino, nitro, cyano group, alkyl acyl, aminoacyl, alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl;
(2) R 2be selected from H, alkyl, aminoacyl, aryl and heteroaryl, described alkyl is optionally replaced by one or more alkyl, alkoxyl group, halogen, hydroxyl, amino, single alkylamino, two alkylamino, aryl or heteroaryl;
(3) R 3for not existing, or be selected from alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, alkylamino, alkyl sulphonyl, alkyl sulfonyl amino, sulfamoyl alkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, amido, ester group ,-CN, cycloalkyl, Heterocyclylalkyl, aryl, heteroaryl, aralkyl;
(4) D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-,-C (R c1r c2the R that)-, is wherein said a1, R a2be hydrogen, alkyl, halogen, haloalkyl, thiazolinyl or haloalkenyl group separately, or R a1, R a2form cycloalkyl together with the C that they connect, described m is 0 or 1, described R bfor not existing, hydrogen, alkyl, alkyl sulphonyl or alkyl-carbonyl, described R c1, R c2be hydrogen, halogen, alkyl, haloalkyl, thiazolinyl or haloalkenyl group separately, or work as R c1, R c2when being alkyl, C can with R c1, R c2form cycloalkyl;
(5) D 1be selected from oxygen, sulphur ,-SO-,-SO 2-,-CO-,-N (R d)-,-(CH) n-, wherein said R dfor not existing, hydrogen, alkyl or haloalkyl, described n is 0,1 or 2, described-(CH) n-optional alkoxy, alkylamino, halogen, hydroxyl, amino, nitro, cyano group, alkyl acyl, aminoacyl, alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl replaces;
(6) G, G 1be-N-or-CH-separately;
(7) for singly-bound or double bond, wherein two be singly-bound, or one for singly-bound, and another is double bond; With
(8) ring T is selected from:
A, aromatic ring or hetero-aromatic ring, described aromatic ring or hetero-aromatic ring can by R 4replace, wherein R 4for alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, alkyl acyl, aminoacyl, alkylamino acyl group, amido ,-CN, aryl or heteroaryl; With
B, naphthenic hydrocarbon or heterocycloalkane, described naphthenic hydrocarbon or heterocycloalkane can by R 4replace, wherein R 4for alkyl, thiazolinyl, alkynyl, alkoxyl group, halogen, haloalkyl, halogenated alkoxy, hydroxyl, nitro, amino, single alkylamino, two alkylamino, alkyl acyl, aminoacyl, alkylamino acyl group, amido ,-CN, aryl or heteroaryl.
2. compound according to claim 1, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein
R 1be selected from H and C 1-6alkyl, described alkyl is optionally by one or more C 1-6alkyl, C 1-6alkoxyl group, C 1-6alkylamino, halogen, hydroxyl, amino, nitro, cyano group, C 1-6alkyl acyl, aminoacyl, C 1-6alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl replace;
R 2be selected from H, C 1-6alkyl, aminoacyl, aryl and heteroaryl, described alkyl is optionally by one or more C 1-6alkyl, C 1-6alkoxyl group, halogen, hydroxyl, amino, single C 1-6alkylamino, two C 1-6alkylamino, aryl or heteroaryl replace; With
R 3for not existing, or be selected from C 1-6alkyl, C 2-6thiazolinyl, C 2-6alkynyl, C 1-6alkoxyl group, fluorine, chlorine, bromine, halo C 1-6alkyl, C 1-6alkylamino, C 1-6alkyl sulphonyl, C 1-6alkyl sulfonyl is amino, sulfamyl C 1-6alkyl, halo C 1-6alkoxyl group, hydroxyl, nitro, amino, single C 1-6alkylamino, two C 1-6alkylamino ,-CN, amido, ester group, C 38cycloalkyl, C 3-8heterocyclylalkyl, aryl and heteroaryl.
3., according to the compound of claim 1 or 2, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-and-C (R c1r c2the R that)-, is described a1, R a2be hydrogen, C separately 1-6alkyl, halogen, halo C 1-6alkyl, C 2-6thiazolinyl or halo C 2-6thiazolinyl, or R a1, R a2c is formed together with the C that they connect 3-8cycloalkyl, described m is 0 or 1, described R bfor not existing, hydrogen, C 1-6alkyl, C 1-6alkyl sulphonyl or C 1-6alkyl-carbonyl, described R c1, R c2be hydrogen, halogen, C separately 1-6alkyl, C 2-6thiazolinyl or halo C 2-6thiazolinyl, or work as R c1, R c2all be selected from C 1-3during alkyl, C can with R c1, R c2form C 3-7cycloalkyl; Preferred D is selected from-(CR a1r a2) mo-,-(CR a1r a2) ms-,-SO-,-SO 2-,-CO-,-N (R b)-,-N (R b)-CO-and-C (R c1r c2the R that)-, is described a1, R a2be hydrogen, methyl or ethyl separately, described m is 0 or 1, described R bfor not existing, hydrogen, methyl, ethyl, propyl group, sec.-propyl, cyclopropyl, methyl sulphonyl or methyl carbonyl, described R c1, R c2be hydrogen, fluorine, methyl, ethyl, C separately 2-4thiazolinyl or halo C 2-4thiazolinyl, or work as R c1, R c2when being methyl, C can with R c1, R c2form cyclopropyl.
4. according to the compound of any one of claims 1 to 3, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein D 1be selected from oxygen, sulphur ,-SO-,-SO 2-,-CO-,-N (R d)-and-(CH) n-, described R dfor not existing, hydrogen, C 1-6alkyl or halo C 1-6alkyl, described n is 0,1 or 2, described-(CH) n-optionally by C 1-6alkoxyl group, C 1-6alkylamino, halogen, hydroxyl, amino, nitro, cyano group, C 1-6alkyl acyl, aminoacyl, C 1-6alkylamino acyl group, alkylsulfonyl, sulfinyl, sulfydryl, aryl or heteroaryl replace.
5., according to the compound of any one of claims 1 to 3, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein ring T is selected from least containing heteroatomic five yuan of fragrant heterocycles and hexa-atomic fragrant heterocycle, phenyl ring, C 3-8naphthenic hydrocarbon and C 3-8heterocycloalkane, preferred ring T is selected from least containing the heteroatomic five yuan of hetero-aromatic rings of N, O or S and hexa-atomic fragrant heterocycle, phenyl ring, pentamethylene, hexanaphthene, suberane.
6. according to the compound of any one of claim 1-5, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, wherein D 1for not existing, then D, G, G 1the phenyl ring be connected with them forms benzo five-membered or six-ring, as shown in general formula (II),
7. compound according to claim 1, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization, described compound is compound and the non-corresponding isomer thereof of following concrete structure:
8. pharmaceutical composition, it comprises the compound of any one of claim 1-7, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization and pharmaceutically acceptable carrier.
9. the pharmaceutical composition of the compound of any one of claim 1-7, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization or claim 8, it is used for the treatment of flaviviridae, especially infection with hepatitis C virus.
10. be used for the treatment of and/or prevention of flavivirus coe virus, especially the method for infection with hepatitis C virus, comprises the pharmaceutical composition giving the compound of any one of the claim 1-7 treating significant quantity, its steric isomer, prodrug, pharmacy acceptable salt, hydrate, solvate or crystallization or claim 8 to its individuality of needs.
The compound of any one of 11. claim 1-7 or the pharmaceutical composition of claim 8 for the preparation for the treatment of and/or preventing flaviviridae, the application in the medicine of especially infection with hepatitis C virus.
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CN114539201B (en) * 2022-03-02 2024-06-28 苏州昊帆生物股份有限公司 Preparation method of Sibirer linking agent

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