CN115960148B

CN115960148B - Novel cytidine derivative, and pharmaceutical composition and application thereof

Info

Publication number: CN115960148B
Application number: CN202211039144.9A
Authority: CN
Inventors: 张哲峰; 孟月垒
Original assignee: Nanjing Zhihe Medical Technology Co ltd
Current assignee: Nanjing Zhihe Medical Technology Co ltd
Priority date: 2021-08-27
Filing date: 2022-08-29
Publication date: 2023-12-12
Anticipated expiration: 2042-08-29
Also published as: CN115960148A; WO2023025319A1; CN117886870A

Abstract

The invention discloses a novel cytidine derivative, a pharmaceutical composition and application thereof, wherein the cytidine derivative is shown as a formula (I) ₀ ) Shown; the compound can be used for preparing medicines for resisting viral infection.

Description

Novel cytidine derivative, and pharmaceutical composition and application thereof

Technical Field

The invention relates to the technical field of pharmaceutical chemistry, in particular to a novel cytidine derivative, a pharmaceutical composition and application thereof.

Background

Influenza is an acute respiratory tract infection due to infection with influenza virus. Is a particularly important disease in high risk groups such as infants, the elderly and the like, and has high complication rate of pneumonia in the elderly.

Because influenza viruses have high variability, development of anti-influenza virus drugs is far from serious because of concerns about emergence or side effects on drug-resistant strains, epidemic of pathogenic or lethal novel influenza viruses, and the like, and therefore, development of novel-structure antiviral drugs is still desired in the art.

Disclosure of Invention

The present inventors have developed a novel cytidine derivative having an antiviral effect and low cytotoxicity.

One aspect of the present invention provides a method as described in (I ₀ ) Novel cytidine derivatives, tautomers, stereoisomers, isotopic derivatives, and pharmaceutically acceptable salts thereof, as shown:

formula (I) ₀ ) In,

R ₁ and R is ₂ Independently selected from hydrogen, Alternatively, R ₁ And R is ₂ Adjacent thereto oxygen forms an acetal or ketal;

wherein n is ₁ Selected from 0, 1, 2 or 3;

n ₂ selected from 1, 2 or 3;

R _a and R is _b Each independently selected from the group consisting of hydroxy, substituted or unsubstituted with one or more groups a: C1-C8 alkyl, C1-C8 alkoxy, C2-C8 alkenyl, C3-C8 cycloalkyl, C6-C18 aryl, aryloxy, arylalkyl, alkylaryl;

R _c and R is _d Independently selected from hydrogen, substituted with one or more groups A orUnsubstituted C1-C8 alkyl;

R ₃ and R is ₄ The same or different are each independently selected from hydrogen orR ₃ And R is ₄ Not both are hydrogen;

wherein n is _a Selected from 0, 1, 2, 3, 4, or 5;

n _b selected from 1, 2, 3, 4, or 5;

n ₃ selected from 0, 1, 2, 3, 4, or 5;

n ₄ selected from 0, 1, 2, 3, or 4;

R ₅ and R is ₆ The same or different, independently selected from hydrogen, C1-C8 alkyl substituted or unsubstituted with one or more groups A; or R is ₅ 、R ₆ Cycloalkyl groups attached to the carbon;

R ₇ is hydrogen, halogen, amino, C1-C8 alkyl substituted or unsubstituted with one or more groups A;

Z is selected from

Wherein n is ₅ Each independently 0, 1, 2, 3, 4, or 5;

R ₈ selected from H, hydroxy, nitro, halogen, the following substituted or unsubstituted with one or more groups a: amino, C1-C8 alkyl, C6-C18 aryl, C1-C8 alkoxy, aminoalkyl, C1-C8 alkylaryl, arylcarbonyl, C1-C8 alkylcarbonyloxy;

R _e and R is _f Each independently selected from hydrogen, the following substituted or unsubstituted with one or more groups a: C1-C8 alkyl, C3-C8 cycloalkyl, heterocycloalkyl, C6-C18 aryl, heteroaryl, non-aromatic heterocyclyl;

R _X1 、R _X2 、R _X3 、R _X4 、R _X5 、R _X6 、R _X7 and R is _X8 Each independently selected from hydrogen, deuterium;

the group A is: hydroxy, carboxyl, amino, halogen, cyano, aldehyde, nitro, trifluoromethyl, C3-C8 cycloalkyl, C1-C8 alkoxy, chlorobenzoyl.

In some embodiments, the invention provides a compound of formula (I ₀ -1) novel cytidine derivatives, tautomers, stereoisomers, isotopic derivatives, and pharmaceutically acceptable salts thereof:

formula (I) ₀ The substituents in 1) are as defined above.

In some embodiments, the invention provides a compound of formula (I ₀ -2) novel cytidine derivatives, tautomers, stereoisomers, isotopic derivatives, and pharmaceutically acceptable salts thereof:

Formula (I) ₀ -the substituents in 2) are as defined above.

In some embodiments, the invention provides a compound of formula (I ₀ -3) novel cytidine derivatives, tautomers, stereoisomers, isotopic derivatives and pharmaceutically acceptable salts thereof:

formula (I) ₀ -3) the substituents are as defined above. In some embodiments, the invention provides a compound of formula (I ₀ -4) novel cytidine derivatives, tautomers, stereoisomers, isotopic derivatives and pharmaceutically acceptable salts thereof:

formula (I) ₀ -4) substituents in the formulaThe definition is as described above.

One aspect of the present invention provides a method as described in (I ₀ -1) novel cytidine derivatives, tautomers, stereoisomers, and pharmaceutically acceptable salts thereof:

formula (I) ₀ In the step of-1), the step of,

R ₁ and R is ₂ The same or different are each independently selected from hydrogen, Alternatively, R ₁ And R is ₂ Together with the oxygen adjacent thereto to form an acetal or ketal;

wherein n is ₁ And n ₂ Selected from 0, 1, 2 or 3;

R _a and R is _b Each independently selected from the group consisting of hydroxy, alkyl substituted or unsubstituted with group A, alkoxy substituted or unsubstituted with group A, alkenyl substituted or unsubstituted with group A, cycloalkyl substituted or unsubstituted with group A, aryl substituted or unsubstituted with group A, aryloxy substituted or unsubstituted with group A, arylalkyl substituted or unsubstituted with group A, alkylaryl substituted or unsubstituted with group A;

R _c And R is _d Independently selected from hydrogen, C1-C8 alkyl substituted or unsubstituted with group A;

R ₃ and R is ₄ The same or different are independently selected from hydrogen andR ₃ and R is ₄ Not both are hydrogen;

wherein n is _a 、n _b And n ₃ Each independently selected from 0, or 1, or 2, or 3, or 4, or 5;

n ₄ selected from 0, 1, 2, 3 or4；

R ₅ And R is ₆ The same or different, independently selected from hydrogen, C1-C8 alkyl substituted or unsubstituted with group A; or R is ₅ 、R ₆ C is cycloalkyl connected with the catalyst;

R ₇ selected from hydrogen, halogen, amino, C1-C8 alkyl substituted or unsubstituted with group A;

z is selected from

Wherein n is ₅ Each independently is 0, or 1, or 2, or 3, or 4, or 5;

R ₈ selected from H, hydroxy, halogen, C1-C8 alkyl substituted or unsubstituted by group A, aryl substituted or unsubstituted by group A, C1-C8 alkyloxy substituted or unsubstituted by group A, aminoalkyl substituted or unsubstituted by group A, C1-C8 alkylaryl substituted or unsubstituted by group A, arylcarbonyl substituted or unsubstituted by group A, C1-C8 alkylcarbonyloxy substituted or unsubstituted by group A;

In some embodiments, the invention provides a compound of formula (I ₀ -1-1) novel cytidine derivatives, tautomers, stereoisomers, isotopic derivatives and pharmaceutically acceptable salts thereof:

formula (I) ₀ The substituents in 1-1) are as defined above.

In some embodiments, the invention provides a compound of formula (I ₀ -1-2) novel cytidine derivatives, tautomers, stereoisomers, isotopic derivatives and pharmaceutically acceptable salts thereof:

formula (I) ₀ The substituents in 1-2) are as defined above.

In some embodiments, the invention provides a compound of formula (I ₀ -1-3) novel cytidine derivatives, tautomers, stereoisomers, isotopic derivatives and pharmaceutically acceptable salts thereof:

formula (I) ₀ The substituents in 1-3) are as defined above.

formula (I) ₀ In the step of-1), the step of,

R ₁ and R is ₂ Independently selected from hydrogen, Alternatively, R ₁ And R is ₂ Together with the oxygen adjacent thereto to form an acetal or ketal;

wherein n is ₁ And n ₂ Selected from 0, 1, 2 or 3;

R _a and R is _b Are each independently selected from hydroxy, alkyl substituted or unsubstituted with group A, alkoxy substituted or unsubstituted with group A, alkenyl substituted or unsubstituted with group A, cycloalkyl substituted or unsubstituted with group A, aryl substituted or unsubstituted with group A, aryloxy substituted or unsubstituted with group A, arylalkyl substituted or unsubstituted with group A, taken by group A Substituted or unsubstituted alkylaryl groups;

R ₃ and R is ₄ Identical or different, each independently selected from hydrogen, orR ₃ And R is ₄ Not both hydrogen;

wherein n is _a And n _b Each independently selected from 1, or 2, or 3, or 4, or 5;

n ₃ selected from 0, or 1, or 2, or 3, or 4, or 5;

n ₄ selected from 0, 1, 2, 3 or 4;

R ₅ and R is ₆ The same or different, independently selected from hydrogen, C1-C8 alkyl substituted or unsubstituted with group A; or R is ₅ 、R ₆ Cycloalkyl groups attached to the carbon;

R ₇ is hydrogen, halogen, amino, C1-C8 alkyl substituted or unsubstituted by the group A;

z is selected fromWherein,

n ₅ each independently is 0, or 1, or 2, or 3, or 4, or 5;

R _e And R is _f Each independently selected from hydrogen, the following substituted or unsubstituted with one or more groups a: C1-C8 alkyl, C3-C8 cycloalkyl, heterocycloalkyl, C6-C18 aryl, heteroarylA radical, a non-aromatic heterocyclic radical; wherein R is _e And R is _f Cannot be hydrogen at the same time;

formula (I) ₀ The substituents in 1-1) are as defined above.

formula (I) ₀ The substituents in 1-2) are as defined above.

formula (I) ₀ The substituents in 1-3) are as defined above.

One aspect of the present invention provides a method as described in (I ₀ -5) novel cytidine derivatives, tautomers, stereoisomers, and pharmaceutically acceptable salts thereof:

formula (I) ₀ In the step-5), the step of,

R ₁ and R is ₂ Independently selected from the group consisting of hydroxy, In particular, when R _x6 And R is _x7 When both are hydrogen or deuterium, R ₁ 、R ₂ R is independently selected from C1-8 alkoxy ₁ And R is ₂ May form acetals or ketals with carbon atoms to which they are attached;

wherein n is as above ₁ And n ₂ Each independently selected from 0, 1, 2 or 3;

R _a and R is _b Each independently selected from hydroxy, substituted or unsubstituted with one or more groups a: alkyl, alkoxy, alkenyl, cycloalkyl, aryl, aryloxy, arylalkyl, alkylaryl;

R _c and R is _d Each independently selected from hydrogen, C1-C8 alkyl substituted or unsubstituted with one or more groups A;

wherein n is _a 、n _b And n ₃ Independently selected from 0, or 1, or 2, or 3, or 4, or 5, respectively;

n ₄ selected from 0, 1, 2, 3 or 4;

R ₇ Is hydrogen, halogen, amino, or a group ofOr a plurality of groups A are substituted or unsubstituted C1-C8 alkyl groups;

z is selected fromWherein,

n ₅ each independently is 0, or 1, or 2, or 3, or 4, or 5;

R ₈ selected from H, hydroxy, halogen, the following substituted or unsubstituted with one or more groups a: amino, C1-C8 alkyl, aryl, alkyloxy, aminoalkyl substituted or unsubstituted with one or more groups A, C1-C8 alkylaryl, arylcarbonyl, C1-C8 alkylcarbonyloxy;

R _X1 、R _X2 、R _X3 、R _X4 、R _X5 、R _X6 、R _X7 and R is _X8 Independently selected from hydrogen, deuterium, in particular R _X1 、R _X2 、R _X3 、R _X4 、R _X5 、R _X6 、R _X7 And R is _X8 And cannot be hydrogen at the same time.

In some embodiments, the above formula (I ₀ ) And/or (I) ₀ -2)-(I ₀ -4) in R _X1 、R _X2 、R _X3 、 R _X4 Each independently selected from hydrogen, deuterium;

in some embodiments, the above formula (I ₀ In-1), R _X1 、R _X2 、R _X3 、R _X4 、R _X5 、R _X6 、 R _X7 And R is _X8 Are all hydrogen.

In some embodiments, the above formula (I ₀ -2)-(I ₀ -4) in R _X5 、R _X6 、R _X7 And R is _X8 Are all hydrogen.

In some embodiments, the above formula (I ₀ )-(I ₀ -4) in R ₁ And R is ₂ Are all hydrogen;

in some embodiments, the above formula (I ₀ )-(I ₀ In-1), R ₁ Is hydrogen, R ₂ Selected from the group consisting ofWherein n is ₁ Selected from 0, 1, 2 or 3;

in some more specific embodiments, n ₁ Selected from 0 or 1;

n ₂ selected from 1, 2 or 3;

in some more specific embodiments, n ₂ 1 is shown in the specification;

R _a and R is _b Each independently selected from the group consisting of hydroxy, substituted or unsubstituted with one or more groups a: C1-C8 alkyl, C1-C8 alkoxy, aryloxy, alkylaryl;

in some more specific embodiments, R is as defined above _a And R is _b Each independently is preferably selected from the group consisting of hydroxy, substituted or unsubstituted with one or more groups a: C1-C8 alkyl, C1-C8 alkyloxy;

in some embodiments, the above formula (I ₀ )-(I ₀ In-1), R ₂ Is hydrogen, R ₁ Selected from the group consisting ofWherein n is as above ₁ 、n ₂ 、R _a And R is _b Respectively as defined above;

in some embodiments, the above formula (I ₀ )-(I ₀ In-1), R ₁ And R is ₂ Are not hydrogen, are each independently selected fromWherein n is as above ₁ 、n ₂ 、R _a And R is _b Respectively as defined above.

In some embodiments, the above formula (I ₀ )-(I ₀ -2) in R ₄ Is hydrogen, R ₃ Is that

In some embodiments, n _a Selected from 0, 1, 2, or 3;

In some more specific embodiments, n _a Is 0;

in some more specific embodiments, n _a 1 is shown in the specification;

in some embodiments, n _b Selected from 0, 1, 2, or 3;

in some more specific embodiments, n _b 1 is shown in the specification;

in some embodiments, n ₃ Selected from 0, 1, 2, 3;

in some more specific embodiments, n ₃ Is 0;

in some more specific embodiments, n ₃ Is 2;

in some more specific embodiments, n ₃ 3;

in some embodiments, n ₄ Selected from 0, 1, 2;

in some more specific embodiments, n ₄ Is 0;

in some more specific embodiments, n ₄ Is 2;

in some embodiments, R as described above _e Is hydrogen, R _f Is the following group, optionally substituted with one or more groups A: C1-C8 alkyl, C3-C8 cycloalkyl, heterocycloalkyl, C6-C18 aryl, heteroaryl, non-aromatic heterocyclyl;

in some more specific embodiments, R is as defined above _e Is hydrogen, R _f Is methyl;

in some embodiments, R as described above _e And R is _f Are C1-C8 alkyl which is substituted or unsubstituted by the radical A;

in some more specific embodiments, R is as defined above _e And R is _f Are all C1-C8 alkyl groups;

in some embodiments, R as described above ₅ And R is ₆ The same or different, independently selected from hydrogen, C1-C8 alkyl substituted or unsubstituted with one or more groups A; or R is ₅ 、R ₆ C is cycloalkyl connected with the catalyst;

in some more specific embodiments, R is as defined above ₅ And R is ₆ Are all C1-C4 alkyl groups; preferably, R ₅ And R is ₆ Homomethyl;

in some embodiments, R as described above ₅ Is methyl, R ₆ Selected from C1-C4 alkyl substituted or unsubstituted with one or more groups A; preferably, R ₅ Is methyl, R ₆ Is ethyl;

in some embodiments, R as described above ₇ Is hydrogen, halogen, amino, C1-C8 alkyl substituted or unsubstituted with one or more groups A;

in some more specific embodiments, R is as defined above ₇ Selected from hydrogen, or methyl;

in some embodiments, Z above is

In some embodiments, n is as described above ₅ Selected from 0, 1, 2, 3, 4, or 5;

in some more specific embodiments, n is as described above ₅ Preferably selected from 0, 1, or 2;

in some embodiments, R as described above ₈ Selected from hydrogen, hydroxy, nitro, halogen, the following groups substituted or unsubstituted with one or more groups a: amino, C1-C8 alkyl, C6-C18 aryl, C1-C8 alkoxy, C1-C8 alkylaryl, C1-C8 alkylcarbonyloxy;

In some more specific embodiments, R is as defined above ₈ Preferably selected from hydrogen, nitro, chlorine, bromine, the following groups substituted or unsubstituted with one or more groups A: amino, C1-C3 alkyl, C6-C18 aryl, C1-C3 alkoxy, C1-C3 alkylcarbonyloxy;

in some more specific casesIn an embodiment, Z is selected from the group consisting of 4-chlorobenzoyl, hydrogen, chlorine, 4-chlorobenzyl,

In some embodiments, the above formula (I ₀ )-(I ₀ -1) and/or (I) ₀ -3) in R ₃ And R is ₄ Are allTherein, Z, R ₅ 、R ₆ 、R ₇ 、R _e 、R _f 、n _a 、 n _b 、n ₃ And n ₄ As defined above.

In some embodiments, the above formula (I ₀ )-(I ₀ -1) and/or (I) ₀ -4) in R ₃ Is hydrogen, R ₄ Is thatTherein, Z, R ₅ 、R ₆ 、R ₇ 、R _e 、R _f 、n _a 、 n _b 、n ₃ And n ₄ As defined above.

In some embodiments, the present invention provides the above-described novel cytidine derivatives, tautomers, stereoisomers, isotopic derivatives, and pharmaceutically acceptable salts thereof, selected from the group consisting of:

in another aspect, in some embodiments, the present invention provides pharmaceutical compositions comprising the novel cytidine derivatives, tautomers, stereoisomers, isotopic derivatives, and pharmaceutically acceptable salts thereof, described above.

In some embodiments, the invention discloses a pharmaceutical composition, which is composed of the compound, isomer or pharmaceutically acceptable salt thereof of the invention as an active ingredient or a main active ingredient, and a pharmaceutically acceptable carrier.

In yet another aspect, the present invention provides, in some embodiments, the above-described novel cytidine derivatives, tautomers, stereoisomers, isotopic derivatives, and pharmaceutically acceptable salts thereof, useful for treating and/or preventing diseases associated with antiviral disorders.

In some embodiments, the present invention provides the use of the above pharmaceutical composition for the preparation of an antiviral drug; among these, the viruses described above include, but are not limited to: arenaviridae, filoviridae, and coronaviridae, and the like, including, but not limited to, adenovirus, rhinovirus, influenza virus, lassa virus, respiratory syncytial virus, severe acute respiratory syndrome virus, parainfluenza virus, coronavirus, and the like.

In some embodiments, the present invention provides the use of the above pharmaceutical composition for the preparation of an antiviral drug; among these, the influenza viruses and coronaviruses described above include, but are not limited to: influenza A virus, influenza B virus, SARS virus, MERS virus, covd-19 virus, etc.

In some embodiments, the novel cytidine derivatives of the present invention can be formulated as pharmaceutical compositions for administration to a patient in a variety of suitably selected modes of administration, including systemic, e.g., oral or parenteral, intravenous, intramuscular, transdermal, subcutaneous, and the like.

Compared with the compound A disclosed by CN111372592A, the compound disclosed by the invention has better stability.

Compared with the compound A disclosed in CN111372592A, the compound disclosed by the invention has better anti-influenza virus activity, lower cytotoxicity and higher selection index.

Compared with the compound A, the compound disclosed by the invention has better anti-novel coronavirus activity.

The bioavailability of the compound disclosed by the invention is approximately 1.4 times that of the compound A, and the compound has a better safety range.

Compared with the combined administration of the compound A and the compound C, the compound disclosed by the invention has more excellent treatment effect and prevention effect in the aspect of resisting influenza virus, and the two metabolic components play a certain synergistic effect after the compound enters a body through analysis of the excellent treatment effect and prevention effect. The compounds of the invention can be used as antiviral drugs with novel structures.

The structure of compound a is as follows:

the structure of compound C is as follows:

definition:

the following terms and phrases used herein are intended to have the following meanings unless otherwise indicated. A particular term or phrase, unless otherwise specifically defined, should not be construed as being ambiguous or otherwise clear, but rather should be construed in a generic sense. When trade names are presented herein, it is intended to refer to their corresponding commercial products or active ingredients thereof.

Certain compounds of the invention may exist in unsolvated forms or solvated forms such as, for example, hydrated, ethanolic forms. In general, solvated forms, which are equivalent to unsolvated forms, are intended to be encompassed within the scope of the present invention.

The term "pharmaceutically acceptable" is intended to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.

The term "pharmaceutically acceptable salt" refers to salts of the compounds of the present invention prepared from the compounds of the present invention which have the specified substituents found herein with relatively non-toxic acids or bases. When the compounds of the present invention contain relatively acidic functional groups, base addition salts may be obtained by contacting neutral forms of such compounds with a sufficient amount of a base in pure solution or in a suitable inert solvent. Pharmaceutically acceptable base addition salts include aluminum, sodium, potassium, calcium, manganese, iron, ammonium, organic ammonia, or magnesium salts or similar salts. When the compounds of the present invention contain relatively basic functional groups, the acid addition salts may be obtained by contacting the neutral form of such compounds with a sufficient amount of an acid in pure solution or in a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include inorganic acid salts including, for example, hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, bicarbonate, phosphoric acid, monohydrogen phosphate, dihydrogen phosphate, sulfuric acid, hydrogen sulfate, hydroiodic acid, phosphorous acid, and the like; and organic acid salts including acids such as acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid, and methanesulfonic acid; also included are salts of amino acids (e.g., arginine, etc.), and salts of organic acids such as glucuronic acid. Certain specific compounds of the invention contain basic and acidic functionalities that can be converted to either base or acid addition salts.

The term "alkyl" means a saturated aliphatic radical, including straight and branched chain groups, alkyl groups which may be substituted or unsubstituted. When substituted alkyl, the substituent is preferably one or more, more preferably 1 to 3, most preferably 1 or 2 substituents.

The term "alkenyl" means an aliphatic radical having an unsaturated carbon-carbon double bond, and includes straight and branched chain alkyl groups which may be substituted or unsubstituted. The carbon-carbon double bond may be one or more.

The term "cycloalkyl" means a single or fused ring of all carbons ("fused" ring means that each ring in the system shares an adjacent pair of carbon atoms with the other rings in the system), wherein one or more of the rings does not have a fully attached pi-electron system, examples of cycloalkyl (without limitation) are cyclopropane, cyclobutane, cyclopentane, cyclopentene, cyclohexane, adamantane, cyclohexadiene, cycloheptane, and cycloheptatriene. Cycloalkyl groups may be substituted and unsubstituted.

The term "aryl" means an all-carbon monocyclic or fused multicyclic group of 1 to 12 carbon atoms having a fully conjugated pi-electron system. Non-limiting examples of aryl groups are phenyl, naphthyl and anthracenyl. Aryl groups may be substituted or unsubstituted. When substituted, the substituents are preferably one or more, more preferably one, two or three, and even more preferably one or two.

The term "arylalkyl" denotes an aryl-substituted hydrocarbon group.

The term "heteroaryl" means a multi-atom monocyclic or fused ring radical containing one, two, three or four ring heteroatoms selected from N, O or S, the remaining ring atoms being C, additionally having a fully conjugated pi-electron system. Non-limiting examples of unsubstituted heteroaryl groups are pyrrole, furan, thiophene, imidazole, oxazole, thiazole, pyrazole, pyrimidine, quinoline, isoquinoline, purine, tetrazole, triazine, and carbazole.

The term "alkoxy" refers to a group in which an alkyl group is attached to oxygen, where the alkyl group may be straight chain, branched or cyclic alkyl.

The term "hydroxy" denotes an-OH group.

The term "amino" means-NH ₂ A group.

The term "carboxyl" refers to a-COOH group.

The term "halogen" means fluorine, chlorine, bromine or iodine.

The term "pharmaceutically acceptable carrier" refers to any formulation or carrier medium representative of a carrier capable of delivering an effective amount of the active agents of the present invention, which does not interfere with the biological activity of the active agents and which does not have toxic or side effects to the host or patient, including water, oils, vegetables and minerals, cream bases, lotion bases, ointment bases, and the like. Such matrices include suspending agents, viscosity enhancers, transdermal enhancers, and the like.

The term "stereoisomer" refers to a compound that has the same chemical constitution but differs in the arrangement of atoms or groups in space.

The numerical ranges stated herein, e.g., "C1-C8", means that the group can contain 1 carbon atom, 2 carbon atoms, 3 carbon atoms, etc., up to and including 8 carbon atoms.

Drawings

FIG. 1 shows the results of therapeutic administration tests against influenza virus (H1N 1).

FIG. 2 shows the results of a prophylactic administration test against influenza virus (H1N 1).

Detailed Description

A number of exemplary methods for preparing the compounds of the present application are provided in the examples below. The present application is described in detail below by way of examples, but is not meant to be limiting in any way. The present application has been described in detail herein, and specific embodiments thereof are also disclosed, it will be apparent to those skilled in the art that various changes and modifications can be made to the specific embodiments of the application without departing from the spirit and scope of the application. Certain compounds of the application can be used as intermediates for preparing other compounds of the application, the structure of all of which is determined by liquid or nuclear magnetism.

The starting materials in the examples of the present application were all purchased commercially.

Example 1: synthesis of Compound PY-01

The reaction formula:

the preparation method comprises the following steps:

step 1: preparation of Compound PY-0102:

tetrahydrofuran (300 ml), PY-01-SM2 (31.8 g,100 mmol) and 4-dimethylaminopyridine (DMAP, 18.33g,150 mols) are sequentially added into a reaction bottle, 1-ethyl- (3-dimethylaminopropyl) carbodiimide (EDC, 18.63g,120 mmol) and a compound PY-01-SM1 (28.43 g,100 mmol) are added after dissolution, the temperature is raised to 70 ℃, the reaction is stirred, the TLC monitors the reaction completion, the system is cooled, evaporated to dryness, 200ml of ethyl acetate is added, 100ml of water is added, an organic phase is separated, water is washed twice again, dried, evaporated to dryness under reduced pressure, and the residue is purified by a column to obtain a product PY-0102 (44.34 g), and the yield is 75.8%. ESI-MS (+): m/z= 585.16.

Step 2: preparation of Compound PY-0101:

the reaction flask was charged with the compound PY-0102 (44.00 g,75.2 mmol), N-dimethylformamide (250 ml), DIPEA (19.44 g,150.4 mmol), pyBroP (38.55 g, 82.7 mmol) was added after dissolution, hydroxylamine hydrochloride (62.71 g,90.24 mmol) was added after stirring the system at room temperature for 30min, the reaction was completed at 40-50℃for 4-6 h, the temperature was reduced, water was added, ethyl acetate was added for extraction 3 times, the organic phases were combined, washed twice, evaporated to dryness under reduced pressure, and the residue was recrystallized from methyl tert-butyl ether/N-heptane to give the product PY-0101 (39.00 g) in 86.4% yield. ESI-MS (+): m/z= 600.17.

Step 3: preparation of Compound PY-01:

PY-0101 (38.00 g,63.3 mmol) and formic acid (500 ml) were added to the flask, and the reaction was carried out at room temperature for 20 hours. After the completion of the reaction, the residue was recrystallized from isopropyl alcohol/methyl tert-butyl ether to give compound PY-01 (29.21) in 82.4% yield. ESI-MS (+): m/z=560.2. 1H NMR (DM SO-D6,500 MHz): delta 10.02 (s, 1H), 9.49 (s, 1H), 7.72-7.75 (m, 4H), 7.63-7.64 (d, 2H), 6.94-6.96 (d, 2H), 6.78-6.80 (d, 1H), 5.71-5.72 (d, 1H), 5.46-5.48 (d, 1H), 5.32-5.33 (d, 1H), 5.20-5.21 (d, 1H), 4.28-4.36 (m, 2H), 3.97 (s, 1H), 3.89-3.92 (m, 1H), 3.78-3.79 (d, 1H), 1.64-1.66 (d, 6H).

Example 2: synthesis of Compound PY-02

The reaction formula:

the preparation method comprises the following steps:

step 1: preparation of Compound PY-0201

Referring to the preparation of step 1 of example 1, substituting compound PY-0101 for compound PY-01-SM1 produced compound PY-0201 (13.21 g) in 73.5% yield. ESI-MS (+): m/z= 900.22.

Step 2: preparation of Compound PY-02

Referring to the preparation of step 3 of example 1, compound PY-0101 was replaced with compound PY-0201 to prepare compound PY-02 (10.14 g) in 83.2% yield. ESI-MS (+): m/z= 860.19.

Example 3: synthesis of Compound PY-03

The reaction formula:

The preparation method comprises the following steps:

step 1: preparation of Compound PY-0303

The three-necked flask was charged with the compound PY-03-SM3 (25.0 g,102.0 mmol 1) and 500ml of methylene chloride under nitrogen. The resulting solution was cooled to 0deg.C, and DMAP (1.3 g,10.6 mmol) and imidazole (27.9 g,409.0 mmol 1) were added sequentially. TBSCl (61.7 g,40.0 mmol) was added over 10 minutes and the resulting mixture was warmed to ambient temperature and stirred for 18 hours. 300mL of water was added to the system, stirred at room temperature for 2 hours, the mixture was separated, the aqueous phase was extracted 3 times with dichloromethane, the combined organic layers were washed with brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and the residue was purified by column to give compound PY-0303 (43.4 g) in 72.5% yield. ESI-MS (+): m/z= 587.33.

Step 2: preparation of Compound PY-0302

A1L round bottom flask was charged with compound PY-0303 (28.0 g,47.7mo 1) and dichloromethane (700 mL). The solution was cooled to 0 ℃ using an ice bath; DMAP (0.583 g,4.77mmo 1) and N, N-diisopropylethylamine (30.9 g, 235 mmo 1) were added successively. 2,4, 6-triisopropylphenyl-1-sulfonyl chloride (28.9 g,95 mmol) was slowly added to the flask, after which the flask was warmed to ambient temperature and stirred for 18 hours. The system was cooled to 0deg.C and N, N diisopropylethylamine (24.6 g,19 lmol) was added dropwise followed by immediately adding solid hydroxylamine hydrochloride (13.26 g,191 mmol). The mixture was warmed to room temperature and stirred for 3 hours. The reaction was quenched with water (200 mL) and the resulting layers were separated. The aqueous layer was extracted with dichloromethane (200 mL) and the combined organics were washed with brine, dried over sodium sulfate, concentrated under reduced pressure and the resulting residue purified by column to give compound PY-0302 (20.0 g) in 69.5% yield. ESI-MS (+): m/z= 602.34.

Step 3: preparation of Compound PY-0301

Referring to the preparation of step 1 of example 1, substituting compound PY-0302 for compound PY-01-SM1 produced compound PY-0301 (18.6 g) in 76.2% yield. ESI-MS (+): m/z= 902.40.

Step 4: preparation of Compound PY-03

To a three-necked flask, compound PY-0301 (17.5 g,19.4 mol) and tetrahydrofuran (50 mL) were added followed by triethylamine trihydrofluoride (3.1 g,19.4mmo 1), and the mixture was stirred at ambient temperature for 18 hours. The mixture was concentrated under reduced pressure and the residue was dissolved in a minimum amount of MeOH, the solution was slowly added to an erlenmeyer flask containing rapidly stirring dichloromethane (50 mL) and the mixture was stirred at room temperature for 15 min. Filtration and recrystallization from methylene chloride and petroleum ether gave the title compound PY-03 (7.65 g) in 70.5% yield. ESI-MS (+): m/z= 560.25.1H NMR (DMS O-D6,400 MHz): delta 11.34-11.35 (s, 1H), 10.79-10.80 (s, 1H), 7.70-7.74 (m, 4H), 7.61-7.63 (d, 2H), 7.48-7.51 (d, 1H), 6.94-6.96 (d, 2H), 5.72-5.78 (m, 2H), 5.32-5.34 (d, 1H), 5.02-5.08 (d, 2H), 3.96-4.04 (m, 2H), 3.83-3.84 (m, 1H), 3.55 (s, 2H), 1.76 (s, 3H), 1.69 (s, 3H).

Example 4: synthesis of Compound ZJT1

The reaction formula:

The preparation method comprises the following steps:

step 1: preparation of Compound ZJT1-03

500ml of 80% methylamine solution by mass fraction, anisole (36.4 g,0.34 mol), cuprous chloride (35.6 g,0.36 mol) are added into a three-port bottle, the temperature of the solution is controlled to be 35 ℃, p-bromobenzamide (101.9 g, 0.35 mol) is dropwise added, the temperature of the solution is raised to 45 ℃ after the addition, the stirring speed is maintained for reacting for 7 hours, the temperature of the solution is lowered to 30 ℃, the reaction solution is poured into 30% sodium bisulfite solution by mass fraction, the temperature of the solution is maintained to be 0-5 ℃, an organic layer is separated, a water layer is extracted by methylamine solution for 6 times, and after the organic layer is combined, methylamine is distilled out, so that the compound ZJT1-03 is obtained for standby.

Step 2: preparation of Compound ZJT1-02

Hexane 300m1 is added into the standby product of the compound ZJT1-03, the temperature is raised to 60 ℃, cuprous chloride (59.4 g,0.6 mol) is added, reflux reaction is carried out for 2h, hexane is steamed out, the temperature of the solution is reduced to 5 ℃, 200ml of 60% potassium bicarbonate solution by mass fraction is added, stirring is carried out for 2h, filtration and washing with potassium sulfate solution are carried out, thus obtaining the compound ZJT1-02 (79.2 g), and the yield is 84.1%. ESI-MS (+): m/z= 276.98.

Step 3: preparation of Compound ZJT1

Acetone (120 g,2.07 mol) and compound ZJT1-02 (59.6 g,0.21 mol) were added to the three-necked flask and stirred for about 10 minutes, sodium hydroxide (60.0 g,1.5 mol) was added thereto, chloroform (45 ml) was added dropwise thereto under stirring at 20 to 30℃and, after the chloroform dropwise addition was completed, the mixture was kept at 20 to 30℃for 1.5 hours, and then heated to reflux for 3.5 hours. The organic solvent was distilled off under reduced pressure, 100ml of a proper amount of water and 130ml of toluene were added to the residue, 36% hydrochloric acid (about 60 g) was added dropwise, the feed solution was acidified to p=3.5 to 4.5, then 100ml was added, the system was cooled to room temperature and stirred for 2 hours, and the compound ZJT1 (66.5 g) was obtained by filtration and drying in a yield of 87.2%. ESI-MS (-) m/z= 361.01.

Example 5: synthesis of Compound ZJT2

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure for each step of example 4, the starting material was replaced with p-nitrobenzamide to give compound ZJT2 (55.2 g) in 85.8% yield. ESI-MS (-) m/z= 328.09.

Example 6: synthesis of Compound ZJT3

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure for each step of example 4, the starting material was replaced with paraaminobenzamide to give compound ZJT3 (51.9 g) in 86.1% yield. ESI-MS (-) m/z= 298.12.

Example 7: synthesis of Compound ZJT4

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure for each step of example 4, the starting material was replaced with 3-chlorobenzamide to give compound ZJT4 (57.6 g) in 82.9% yield. ESI-MS (-) m/z= 317.07.

Example 8: synthesis of Compound ZJT5

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure for each step of example 4, the starting material was replaced with 3, 4-dichlorobenzamide to give compound ZJT5 (46.1 g) in 79.6% yield. ESI-MS (-) m/z= 351.03.

Example 9: synthesis of Compound ZJT6

The reaction formula:

The preparation method comprises the following steps:

referring to the procedure for each step of example 4, the starting material was replaced with p-methoxybenzamide to give compound ZJT6 (31.3 g) in 87.1% yield. ESI-MS (-) m/z= 313.12.

Example 10: synthesis of Compound ZJT7

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure for each step of example 4, the starting material was replaced with 3,4, 5-trimethoxybenzamide to afford compound ZJT7 (32.6 g) in 84.3% yield. ESI-MS (-) m/z= 373.14.

Example 11: synthesis of Compound PY-ZJT8

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure for each step of example 4, the starting material was replaced with 3- (trifluoromethyl) benzamide to give compound ZJT8 (36.8 g) in 81.1% yield. ESI-MS (-) m/z= 351.09.

Example 12: synthesis of Compound PY-ZJT9

The reaction formula:

the preparation method comprises the following steps:

the compound PY-01-SM2 (21.5 g,67.5 mmol), water (300 mL), methylene chloride (300 mL), tetrabutylammonium bisulfate (2.3 g,6.75 mmol) and sodium bicarbonate (22.7 g,270 mmol) were charged to a three-necked flask. Chloromethylchlorosulfonate (13.5 g,81.7 mmol) was added dropwise at room temperature, and the reaction was stirred at room temperature overnight. The organic and aqueous layers were separated and the aqueous layer was extracted with dichloromethane (300 mL). The combined organic phases were dried, concentrated and purified by column chromatography (0 to 5% ethyl acetate/hexanes gradient elution) to give compound ZJT9 (18.7 g) in 75.4% yield.

Example 13: synthesis of Compound PY-04

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure of each step of example 1, compound ZJT1 was used as a starting material instead of compound PY-01-SM2, to give compound PY-04 (10.8 g), with a total yield of 51.1%. ESI-MS (+): m/z= 604.09.

Example 14: synthesis of Compound PY-05

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure of each step of example 2, substituting compound ZJT1 for compound PY-01-SM2 as a starting material gave compound PY-05 (12.3 g), with a total yield of 57.6%. ESI-MS (+): m/z= 948.09.

Example 15: synthesis of Compound PY-06

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure for each step of example 1, substituting compound ZJT2 for compound PY-01-SM2 as a starting material gave compound PY-06 (14.1 g), in a total yield of 46.9%. ESI-MS (+): m/z= 571.16.

Example 16: synthesis of Compound PY-07

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure of each step of example 1, substituting compound ZJT3 for compound PY-01-SM2 as a starting material gave compound PY-07 (13.6 g), with a total yield of 48.5%. ESI-MS (+): m/z= 541.19.

Example 17: synthesis of Compound PY-08

The reaction formula:

the preparation method comprises the following steps:

step 1: preparation of Compound PY-0804:

PY-01-SM2 (1.91 g,6 mmol) was added to dichloromethane (20 mL) under nitrogen, cooled to 0deg.C, thionyl chloride (0.86 g,7.2 mmol) was slowly added, and after the addition was completed, the reaction was stirred at room temperature for 2.0h. Concentrating the system to dryness, removing residual sulfoxide chloride with toluene (10 mL×3), and collecting residual PY-0804 without treatment;

step 2: preparation of Compound PY-0803:

dichloromethane (25 mL) was added to the residue (compound PY-0804,6 mmol) obtained in step 1, and then the above mixture was slowly added to a dichloromethane solution (20 mL) in which aluminum trichloride (0.48 g,3.6 mmol) was dissolved, and stirred at room temperature for 20 minutes after the addition. The system was cooled to 0℃and acetaldehyde (0.27 g,6 mmoles) was added dropwise over 10 minutes. The reaction mixture was stirred at room temperature for 1 hour and the system was gradually added to a vigorously stirred ice-water slurry. The organic phases were combined by extraction with dichloromethane 3 times, washed with ice water 2 times, separated out, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by column chromatography to give compound PY-0803 (1.11 g) in 48.7% yield. ESI-MS (+): m/z= 381.06.

Step 3: preparation of Compound PY-0802:

the reaction flask was charged with compound PY-01-SM1 (0.56 g,2.0 mmol), triethylamine (0.31 g, 3 mmol) and compound PY-0803 (0.92 g,2.4 mmol), and the system was heated to 50deg.C and stirred for 24 hours with the addition of N, N-dimethylformamide (50 mL). Cooling to room temperature, adding water, stirring, filtering to obtain crude product, and purifying with silica gel column to obtain compound PY-0802 (0.96 g) with yield of 76.4%, ESI-MS (+): m/z= 628.20.

Step 4: preparation of Compound PY-0801:

the reaction flask was charged with PY-0802 (0.47 g,0.75 mmol), N-dimethylformamide (25 ml), DIPEA (0.20 g,1.50 mmol), pyBroP (0.39 g,0.83 mmol) was added after dissolution, hydroxylamine hydrochloride (0.63 g,0.90 mmol) was added after stirring the system at room temperature for 30min, the reaction was completed at 40-50℃for 4-6 h, TLC detection was completed, the temperature was lowered, water was added, ethyl acetate was extracted 3 times, the organic phases were combined, washed twice, evaporated under reduced pressure, and the residue was purified by column chromatography to give the product compound PY-0801 (0.36 g), yield 74.5%. ESI-MS (+): m/z= 644.19.

Step 5: preparation of Compound PY-08:

the reaction flask was charged with PY-0801 (0.36 g,0.56 mmol) and formic acid (10 mL) and the system was reacted at room temperature for 20 hours. After the completion of the reaction, the residue was purified by column chromatography to give PY-08 (0.23 g) in 68.7% yield as ESI-MS (+): m/z= 604.16.1H NMR (DMSO-D6, 400 MHz): delta 11.32-11.33 (s, 1H), 10.77-10.78 (s, 1H), 7.71-7.76 (m, 4H), 7.59-7.61 (D, 2H), 7.45-7.49 (D, 1H), 6.91-6.94 (D, 2H), 6.61 (m, 4H), 5.70-5.75 (m, 2H), 5.29-5.31 (D, 1H), 5.01-5.06 (D, 2H), 3.94-4.03 (m, 2H), 3.81-3.83 (m, 1H), 3.53 (s, 2H), 1.72 (D, 3H), 1.70 (s, 6H).

Example 18: synthesis of Compound PY-09

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure of step 3 and step 4 of example 3, compound PY-01-SM2 was replaced with compound ZJT5 to give compound PY-09 (9.3 g), with a total yield of 50.1%. ESI-MS (+): m/z= 592.12.

Example 19: synthesis of Compound PY-10

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure of step 3 and step 4 of example 3, compound PY-01-SM2 was replaced with compound ZJT6 to give compound PY-10 (19.4 g), with a total yield of 55.3%. ESI-MS (+): m/z= 554.21.

Example 20: synthesis of Compound PY-11

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure of step 1 and step 2 of example 1, compound PY-1102 was prepared by substituting compound ZJT7 for compound PY-01-SM 2;

referring to the procedure of each step of example 2, compound PY-11 (5.7 g) was prepared in a total yield of 35.3% by substituting compound ZJT7 for compound PY-01-SM2 and compound PY-1102 for compound PY-0101, respectively. ESI-MS (+): m/z= 972.33.

Example 21: synthesis of Compound PY-24

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure of each step of example 1, substituting compound ZJT8 for compound PY-01-SM2 as a starting material gave compound PY-24 (5.1 g), with a total yield of 45.9%. ESI-MS (+): m/z= 594.16.

Example 22: synthesis of Compound PY-26

The reaction formula:

the preparation method comprises the following steps:

step 1: synthesis of Compound PY-2602

Referring to the procedure of step 3 in example 1, compound PY-2602 (6.1 g) was prepared in a yield of 78.3%. ESI-MS (+): m/z= 545.12.

Step 2: compound PY-2601

Into a three-necked flask, compound PY-2602 (5.6 g,10.24 mol) and ethyl acetate (120 mL) were added. Triethylamine (5.2 g,51.12 mol) and DMAP (0.063 g,5.12 mmol) were added to the above system with stirring. Cooled to below 10 ℃ and trifluoroacetic anhydride (6.45 g,30.72 mmol) was slowly added to the system over 5 minutes. The system exothermed during addition. After the addition was completed, the system was stirred at room temperature for 1h, and the reaction was completed by TLC. The reaction was quenched with 40mL of water and stirred at room temperature for 20 minutes. The layers were separated and the organics were washed with water (2X 50 mL), saturated aqueous bicarbonate (50 mL X2), water (50 mL), brine (50 mL X2), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure to give the residue compound PY-2601. No further purification was required for the next step. ESI-MS (+): m/z= 737.09.

Step 3: preparation of Compound PY-26

The procedure of step 2 in reference example 1 was followed to obtain compound PY-26 (2.2 g) in 83.6% yield. ESI-MS (+): m/z= 754.12.

Example 23: synthesis of Compound PY-27

The reaction formula:

the preparation method comprises the following steps:

step 1: synthesis of Compound PY-2702

The reaction flask was charged with the compounds PY-01-SM1 (14.2 g,50.0 mmol), triethylamine (75.9 g, 75.0 mmol) and ZJT9 (36.7 g,100.0 mmol), 500ml of N, N-dimethylformamide was added and the system was heated to 50℃and stirred for 24 hours. Cooling to room temperature, adding water, stirring, filtering to obtain crude product, and purifying with silica gel column to obtain compound PY-2702 (17.2 g), yield 55.8%, ESI-MS (+): m/z= 615.17.

Step 2: compound PY-2701

Referring to the procedure of step 2 in example 1, compound PY-2701 (10.1 g) was produced in a yield of 82.3%. ESI-MS (+): m/z= 630.18.

Step 3: preparation of Compound PY-27

The procedure of step 3 of example 1 was followed to give compound PY-27 (5.3 g) in 79.6% yield. ESI-MS (+): m/z= 590.15.

Example 24: synthesis of Compound PY-28

The reaction formula:

the preparation method comprises the following steps:

step 1: synthesis of Compound PY-2801

Referring to the procedure of step 1 in example 23, compound PY-2801 (12.2 g) was prepared in a yield of 51.4% and ESI-MS (+): m/z= 932.41.

Step 2: compound PY-28

Referring to the procedure of step 4 in example 3, compound PY-28 (5.2 g) was produced in a yield of 68.3%. ESI-MS (+): m/z= 590.15.

Example 25: synthesis of Compound PY-34

The reaction formula:

the preparation method comprises the following steps:

step 1: preparation of Compound PY-3404:

PY-01-SM2 (1.91 g,6 mmol) was added to dichloromethane (20 mL) under nitrogen, cooled to 0deg.C, thionyl chloride (0.86 g,7.2 mmol) was slowly added, and after the addition was completed, the reaction was stirred at room temperature for 2.0h. Concentrating the system to dryness, removing residual sulfoxide chloride with toluene (10 mL×3), and collecting residue of PY-3404 without treatment;

step 2: preparation of Compound PY-3403:

dichloromethane (25 mL) was added to the residue (compound PY-3404,6 mmol) obtained in step 1, and then the above mixture was slowly added to a dichloromethane solution (20 mL) in which aluminum trichloride (0.48 g,3.6 mmol) was dissolved, and after the addition, stirred at room temperature for 20 minutes. The system was cooled to 0℃and acetaldehyde (0.27 g,6 mmoles) was added dropwise over 10 minutes. The reaction mixture was stirred at room temperature for 1 hour and the system was gradually added to a vigorously stirred ice-water slurry. Extraction with dichloromethane 3 times, combining the organic phases, washing with ice water 2 times, separating the organic phase, drying over anhydrous sodium sulfate, filtering, concentrating the filtrate under reduced pressure, purifying the residue by column chromatography to give compound PY-3403 (1.11 g), yield 48.7%. ESI-MS (+): m/z= 381.06.

Step 3: preparation of Compound PY-3402:

into the reaction flask were added compound PY-01-SM1 (0.56 g,2.0 mmol), triethylamine (0.31 g, 3 mmol) and compound PY-3403 (0.92 g,2.4 mmol), and the added N, N-dimethylformamide (50 mL), and the system was heated to 50℃and stirred for 24 hours. Cooling to room temperature, adding water, stirring, filtering to obtain crude product, and purifying with silica gel column to obtain compound PY-3402 (0.96 g), yield 76.4%, ESI-MS (+): m/z= 628.20.

Step 4: preparation of Compound PY-3401:

into the reaction flask were added PY-3402 (0.47 g,0.75 mmol), N-dimethylformamide (25 ml) and DIPEA (0.20 g,1.50 mmol), after dissolution, tripyrrolidinylphosphonium bromide hexafluorophosphate (PyBroP, 0.39g,0.83 mmol) was added, the system was stirred at room temperature for 30min, hydroxylamine hydrochloride (0.63 g,0.90 mmol) was added, the reaction was completed at 40-50℃for 4-6 h, TLC detection was completed, the temperature was lowered, water was added, ethyl acetate extraction was carried out 3 times, the organic phase was combined, washed twice with water, evaporated under reduced pressure, and the residue was purified by column to give the product compound PY-3401 (0.36 g), yield 74.5%. ESI-MS (+): m/z= 644.19.

Step 5: preparation of Compound PY-34:

the reaction flask was charged with PY-3401 (0.36 g,0.56 mmol) and formic acid (10 mL) and the system was reacted at room temperature for 20 hours. After the completion of the reaction, the residue was purified by column chromatography to give compound PY-34 (0.23 g) in a yield of 68.7% and ESI-MS (+): m/z= 604.16.1H NMR (DMSO-D6, 400 MHz): delta 11.32-11.33 (s, 1H), 10.77-10.78 (s, 1H), 7.71-7.76 (m, 4H), 7.59-7.61 (D, 2H), 7.45-7.49 (D, 1H), 6.91-6.94 (D, 2H), 6.61 (m, 4H), 5.70-5.75 (m, 2H), 5.29-5.31 (D, 1H), 5.01-5.06 (D, 2H), 3.94-4.03 (m, 2H), 3.81-3.83 (m, 1H), 3.53 (s, 2H), 1.72 (D, 3H), 1.70 (s, 6H).

Example 26: synthesis of Compound PY-35

The reaction formula:

the preparation method comprises the following steps:

step 1: preparation of Compound PY-3501

Referring to the preparation of step 3 of example 25, compound PY-3401 was substituted for compound PY-01-SM1 to prepare compound PY-3501 (1.26 g) in 69.5% yield. ESI-MS (+): m/z= 988.27.

Step 2: preparation of Compound PY-35

With reference to the preparation of step 5 of example 25, compound PY-3401 was replaced with compound PY-3501 to prepare compound PY-35 (0.19 g) in 63.2% yield. ESI-MS (+): m/z= 948.24.

Example 27: synthesis of Compound PY-36

The reaction formula:

the preparation method comprises the following steps:

step 1: preparation of Compound PY-3603

The compound PY-03-SM3 (12.5 g,51.0 mmol) and methylene chloride (250 mL) were added to a three-necked flask under nitrogen. The resulting solution was cooled to 0℃and 4-dimethylaminopyridine (DMAP, 0.70g,5.7 mmol) and imidazole (14.0 g,205.6 mmol) were added sequentially. T-butyldimethylchlorosilane (TBSC 1, 30.9g,205.0 mmol) was added over 10 minutes and the resulting mixture was warmed to ambient temperature and stirred for 18 hours. Water was added to the system, stirred at room temperature for 2 hours, the mixture was separated, the aqueous phase was extracted 3 times with dichloromethane, the combined organic layers were washed with brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and the residue was purified by column to give compound PY-3603 (21.1 g) in 70.6% yield. ESI-MS (+): m/z= 587.33.

Step 2: preparation of Compound PY-3602

The compound PY-3603 (14.0 g,23.9 mmol) and methylene chloride (350 mL) were added to a three-necked flask. The solution was cooled to 0 ℃ using an ice bath. DMAP (0.29 g,0.24 mmol) and N, N-diisopropylethylamine (15.5 g,120 mmol) were added sequentially. 2,4, 6-triisopropylbenzene-1-sulfonyl chloride (14.5 g,47.5 mmol) was slowly added to the flask, after which the flask was warmed to ambient temperature and stirred for 18 hours. The system was cooled to 0deg.C and N, N diisopropylethylamine (12.3 g,95.5 mmol) was added dropwise followed immediately by solid hydroxylamine hydrochloride (6.63 g,95.5 mmol). The mixture was warmed to room temperature and stirred for 3 hours. The reaction was quenched with water and the resulting layers were separated. The aqueous layer was extracted with dichloromethane and the combined organics were washed with brine, dried over sodium sulfate, concentrated under reduced pressure and the resulting residue was purified by column to give compound PY-3602 (9.41 g) in 65.4% yield. ESI-MS (+): m/z= 602.34.

Step 3: preparation of Compound PY-3601

Referring to the preparation of step 3 of example 25, compound PY-3601 (10.1 g) was prepared in 71.3% yield by substituting compound PY-3602 for compound PY-01-SM 1. ESI-MS (+): m/z= 946.42.

Step 4: preparation of Compound PY-36

To a three-necked flask, compound PY-0301 (9.5 g,1.0 mmol) and tetrahydrofuran (100 mL) were added followed by triethylamine trihydrofluoride (1.61 g,1.0 mmol) and the mixture was stirred at ambient temperature for 18 hours. The mixture was concentrated under reduced pressure and the residue was dissolved in a minimum amount of methanol, the solution was slowly added to the dichloromethane containing the rapid stirring, and the mixture was stirred at room temperature for 15 minutes. Filtration and recrystallization from methylene chloride and petroleum ether gave the title compound PY-36 (4.05 g) in 67.1% yield. ESI-MS (+): m/z= 604.16.

Example 28: synthesis of Compound ZJT10

The reaction formula:

the preparation method comprises the following steps:

step 1: preparation of Compound ZJT 10-02:

pyrimidine-5-D (11.31 g,0.1 mol), BSA (40.7 g,0.2 mol) and anhydrous acetonitrile (250 mL) were added to a reaction flask under nitrogen, heated to 80℃and stirred for 30min; cooling to room temperature, adding tetra-acetyl ribose (63.3 g,0.2 mol), dropwise adding TMSOTF (44.5 g,0.2 mol), heating to 80 ℃ after the addition, reacting for 2 hours, cooling to room temperature, and evaporating acetonitrile under reduced pressure; ethyl acetate was added to the residue, which was washed with saturated aqueous sodium hydrogencarbonate solution for 2 times, the organic phase was separated, washed with saturated brine for 2 times, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and the residue was subjected to column chromatography to give the compound ZJT10-02 (31.6 g) in 85.2% yield. ESI-MS (+): m/z= 372.13.

Step 2: preparation of Compound ZJT 10-01:

compound ZJT10-02 (31.0 g,0.084 mol) and 7M ammonia/methanol solution (500 mL,3.5 mol) were added to a reaction flask, stirred at room temperature for 36 hours, the solvent was distilled off, and the residue was subjected to column chromatography to give compound ZJT10-01 (15.5 g), yield 75.2%. ESI-MS (+): m/z= 246.10.

Step 3: preparation of Compound ZJT10

Compound ZJT10-01 (15.0 g,0.061 mol) was dissolved in acetone (300 mL), 2-dimethoxypropane (31.8 g,0.305 mol) was slowly added, after the addition was completed, after stirring for 10 minutes, concentrated sulfuric acid (2.0 g, 0.020mol) was slowly added, and stirring was performed for 30 minutes; saturated sodium bicarbonate was added to the system to quench, the solvent was distilled off under reduced pressure, and the residue was subjected to column chromatography to give Compound ZJT10 (11.5 g), yield 66.1%. ESI-MS (+): m/z= 286.20.

Example 29: synthesis of Compound ZJT11

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure of example 1, uracil-2D was used in place of pyrimidine-5-D as a starting material to give compound ZJT11 (10.2 g) in a total yield of 40.1%. ESI-MS (+): m/z= 287.22.

Example 30: synthesis of Compound ZJT12

The reaction formula:

the preparation method comprises the following steps:

step 1: preparation of Compound ZJT12-01

PY-01-SM1 (6.3 g,22.0 mmol) and methylene chloride (200 mL) were added to the flask at room temperature under nitrogen. Pyridinium dichromate (16.6 g,44.1 mmol), acetic anhydride (22.5 g,220 mmol) and t-butanol (16.3 g,220 mmol) were then added sequentially to the system with stirring. The above system was stirred at room temperature for 24 hours, washed with water, the aqueous phase was separated, extracted 2 times with methylene chloride, the organic phases were combined, washed 2 times with saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated, and the residue was subjected to column chromatography to give compound ZJT12-01 (5.5 g), yield 70.5%. ESI-MS (+): m/z= 355.15.

Step 2: preparation of Compound ZJT12

Compound ZJT12-01 (5.4 g,15.2 mmol) is added to ethanol-D1 (200 mL) at room temperature under nitrogen, and sodium borodeuteride-D4 (2.5 g,60.8 mmol) is added in one portion with stirring. After the addition was completed, the mixture was stirred at room temperature for 1 hour, heated to 55℃for 7 hours, and then stirred at room temperature overnight. The system was cooled to 0deg.C, quenched with acetic acid-D1, evaporated to dryness under reduced pressure to give the residue by column chromatography to give Compound ZJT12 (3.0 g) in a yield of 68.2%. ESI-MS (+): m/z= 287.15

Example 31: synthesis of Compound PY-49

The reaction formula:

the preparation method comprises the following steps:

step 1: preparation of Compound PY-4902

Tetrahydrofuran (30 mL), PY-01-SM2 (3.18 g,10 mmol) and 4-dimethylaminopyridine (DMAP, 1.83g,15 mmol) are sequentially added into a reaction bottle, 1-ethyl- (3-dimethylaminopropyl) carbodiimide (EDC, 1.86g,12 mmol) and ZJT10 (2.85 g,10 mmol) are added after dissolution, the temperature is raised to 70 ℃, the reaction is stirred, TLC monitors the reaction completion, the system is cooled, evaporated to dryness, ethyl acetate and water are added into the remainder, an organic phase is separated, washed twice again, dried, evaporated to dryness under reduced pressure, and the remainder is purified by a column to obtain a product PY-4902 (4.2 g), and the yield is 71.7%. ESI-MS (+): m/z= 586.20.

Step 2: preparation of Compound PY-4901

The reaction flask was charged with the compound PY-4902 (4.1 g,7.0 mmol)), N-dimethylformamide (25 mL), N-diisopropylethylamine (DIPEA, 1.81g,14.0 mmol), after dissolution, tripyrrolidinylphosphonium hexafluorophosphate (PyBroP, 3.59g,7.7 mmol) was added, the system was stirred at room temperature for 30min, hydroxylamine hydrochloride (0.59 g,8.4 mmol) was added, the reaction was completed by TLC detection at 40-50℃for 4-6 h, the temperature was lowered, water was added, ethyl acetate was extracted 3 times, the organic phases were combined, water was used for twice, evaporated under reduced pressure, and the residue was chromatographed to give the product PY-4901 (2.93 g), yield 69.6%. ESI-MS (+): m/z= 601.17.

Step 6: preparation of Compound PY-49

PY-4901 (2.9 g,4.83 mmol) and formic acid (50 mL) were added to the reaction flask, and the system was reacted at room temperature for 20 hours. After the completion of the reaction, the residue was recrystallized from isopropyl alcohol/methyl tert-butyl ether to give compound PY-49 (2.17 g) in 80.1% yield, ESI-MS (+): m/z= 561.14.1H NMR (DM SO-D6,500 MHz): delta 10.05 (s, 1H), 9.44 (s, 1H), 7.74-7.77 (m, 4H), 7.65-7.66 (d, 2H), 6.96-6.98 (d, 2H), 6.79-6.81 (d, 1H), 5.73-5.75 (d, 1H), 5.33-5.34 (d, 1H), 5.22-5.23 (d, 1H), 4.29-4.37 (m, 2H), 3.98 (s, 1H), 3.91-3.95 (m, 1H), 3.79-3.80 (d, 1H), 1.65-1.67 (d, 6H).

Example 32: preparation of Compound PY-50

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure of example 31, ZJT12 was used as a starting material to replace ZJT10 to give compound PY-50 (1.21), in a total yield of 42.2%. ESI-MS (+): m/z= 562.18.

Example 33: preparation of Compound PY-51

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure of example 31, ZJT12 was used as a starting material to replace ZJT10, yielding compound PY-51 (1.52) in a total yield of 38.5%. ESI-MS (+): m/z= 562.20.

Example 34: preparation of Compound PY-52

The reaction formula:

the preparation method comprises the following steps:

step 1: preparation of Compound PY-5203

Under the protection of nitrogen, the compound ZJT10-01 (2.5 g,10.2 mmol) and methylene chloride (50 mL) are added into a reaction bottle, and cooled to 0 ℃; DMAP (0.13 g,1.02 mmol) and imidazole (2.8 g,40.9 mmol) were then added sequentially. T-butyldimethylchlorosilane (TBSCl, 6.17g,4.0 mmol) was added over 10 minutes and the resulting mixture was warmed to ambient temperature and stirred for 18 hours. Water (30 mL) was added to the system, stirred at room temperature for 2 hours, the solution was separated, the aqueous phase was extracted 3 times with dichloromethane, the combined organic layers were washed with brine, dried over sodium sulfate, filtered and concentrated under reduced pressure, and the residue was purified by column to give compound PY-5203 (4.14 g) in 69.0% yield. ESI-MS (+): m/z= 588.33.

Step 2: preparation of Compound PY-5202

The compound PY-5203 (4.00 g,6.8 mmol) and methylene chloride (100 mL) were added to a reaction flask, stirred and cooled to 0℃and then DMAP (0.083 and g,0.68 mmol) and N, N-diisopropylethylamine (4.41 g,34.1 mmol) were added sequentially to the above system, 2,4, 6-triisopropylbenzene-1-sulfonyl chloride (4.13 g,13.6 mmol) was slowly added to the flask with stirring, and after the addition, stirred at room temperature for 18 hours. The system was cooled again to 0deg.C and N, N-diisopropylethylamine (3.51 g,27.2 mmol) was added dropwise followed immediately by solid hydroxylamine hydrochloride (1.9 g,27.3 mmol). The mixture was warmed to room temperature and stirred for 3 hours. The reaction was quenched with water, separated, the aqueous layer was extracted 2 times with dichloromethane, the organic phases were combined, washed with brine, dried over sodium sulfate, concentrated under reduced pressure, and the obtained residue was purified by column to give compound PY-5202 (2.70 g) in 65.9% yield. ESI-MS (+): m/z= 603.34.

Step 3: preparation of Compound PY-5201

Referring to the preparation of step 1 of example 31, substituting compound PY-5202 for compound ZJT10 produced is compound PY-5201 (2.22 g), 66.5% yield. ESI-MS (+): m/z= 903.43.

Step 4: preparation of Compound PY-52

To a three-necked flask, the compound PY-5201 (1.75 g,1.94 mol) and tetrahydrofuran (20 mL) were added followed by triethylamine trihydrofluoride (0.31 g,1.94mmo 1), and the mixture was stirred at ambient temperature for 18 hours. The mixture was concentrated under reduced pressure, and the residue was subjected to column chromatography to give compound PY-52 (0.53 g) in a yield of 48.7%. ESI-MS (+): m/z= 561.16.

Example 35: preparation of Compound PY-54

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure of example 34, ZJT11-01 was used as a starting material in place of ZJT10-01 to give compound PY-54 (0.46), with a total yield of 16.2%. ESI-MS (+): m/z= 562.20.

Example 36: preparation of Compound PY-55

The reaction formula:

the preparation method comprises the following steps:

referring to the procedure of step 1 and step 3 of example 31, using compound PY-4901 as a starting material, compound PY-55 (0.25 g) was prepared in a total yield of 50.2%. ESI-MS (+): m/z= 861.22.

Example 37: preparation of Compound PY-56

The reaction formula:

the preparation method comprises the following steps:

step 1: synthesis of Compound PY-5604

PY-01-SM2 (3.82 g,12 mmol) was added to dichloromethane (50 mL) under nitrogen, cooled to 0deg.C, thionyl chloride (1.72 g,14.4 mmol) was slowly added, and after the addition was completed, the reaction was stirred at room temperature for 2.0h. Concentrating the system to dryness, taking toluene as the remainder to remove the residual sulfoxide chloride for 3 times, wherein the remainder is a compound PY-5604, and performing no treatment for later use;

step 2: synthesis of Compound PY-5603

Dichloromethane (50 mL) was added to the residue obtained in step 1 (compound PY-5604, 12 mmol), and then the above mixture was slowly added to a dichloromethane solution (40 mL) in which aluminum trichloride (0.96 g,7.2 mmol) was dissolved, and after the addition, stirred at room temperature for 20 minutes. The system was cooled to 0deg.C and acetaldehyde (0.54 g,12 mmol) was added dropwise over 10 minutes. The reaction mixture was stirred at room temperature for 1 hour and the system was gradually added to a vigorously stirred ice-water slurry. Extraction with dichloromethane 3 times, combining the organic phases, washing with ice water 2 times, separating the organic phase, drying over anhydrous sodium sulfate, filtering, concentrating the filtrate under reduced pressure, purifying the residue by column chromatography to give compound PY-5603 (2.07 g), yield 45.2%. ESI-MS (+): m/z= 381.06.

Step 3-step 5: synthesis of Compound PY-56

Referring to the procedure of each step of example 31, PY-5603 was used in place of PY-01-SM2 as a starting material to give Compound PY-56 (1.12), in a total yield of 35.3% in three steps. ESI-MS (+): m/z= 605.20.

The following examples were synthesized in the same manner as in the above examples, using commercially available compounds or intermediate compounds appropriately synthesized from the commercially available compounds.

Example 38: stability test

The compounds PY-01, PY-02, PY-03, PY-12, PY-15, PY-18, PY-21, PY-34, PY-49, and Compound A were each prepared by preparing a sample with a 0.5% sodium carboxymethyl cellulose solution and a 1.0% methyl cellulose solution, placing the above samples (mouth seal) in a stability box at 25.0deg.C, storing at 37% humidity under light-proof conditions, and detecting the stability on days 0, 3, and 7 by HPLC (with DAD detector). The results are shown in Table I.

Table one stability test results

The results show that the stability of the series of compounds of the invention is better than that of the control compound A.

Example 39: in vitro anti-influenza virus activity and cytotoxicity assay

Toxicity assay of test compounds against MDCK:

taking MDCK cells in logarithmic growth phase, inoculating into 24-hole cell culture plate at 37deg.C with 5% CO ₂ After 24 hours of culture in an incubator, the culture medium is firstly acted for 2 hours by medicines with different dilution concentrations, then the plate is washed for 3 times by Hanks liquid, and 1ml of DMEM maintaining liquid containing 2 percent of new born calf serum is added into each hole after spin-drying to maintain growth, the temperature is 37 ℃ and the CO is 5 percent ₂ Is cultured in an incubator of (a). The degree of cytopathic effect (CPE) was observed over 7 days, and the half-Toxic Concentration (TC) of the sample on cells was calculated by the Reed-Muench method ₅₀ )。

Determination of anti-influenza Activity of test Compounds:

the MDCK cells in logarithmic growth phase were inoculated into 24-well plates with about cells (1X 10) ⁴ And, respectively), 37 ℃,5% CO ₂ Culturing in an incubator. After 24H, the cells were first infected with a virus solution (H1N 1, A/WSN/33), washed 2 times with Hanks 'solution, then treated with a drug dilution, washed 3 times with Hanks' solution, and finally washed 3 times with a DMEM maintenance solution containing 2% neonatal bovine serum at 37℃with 5% CO ₂ Culturing in an incubator. Respectively after culturing1. The cytopathic effect (CPE) was observed under an inverted microscope at days 2, 3, 4, 5, 6, and 7, and the half-maximal Inhibitory Concentration (IC) of the sample on the virus was calculated by the Reed-Muench method ₅₀ ) Then, a Selection Index (SI) is calculated, the SI being calculated by si=tc ₅₀ /IC ₅₀ . The results are shown in Table II.

Data on anti-influenza virus activity in vitro

Experimental results show that compared with the disclosed compound A, the detected compound sample has better inhibition activity on influenza virus (H1N 1), lower cytotoxicity and higher selection index.

Example 40: in vitro anti-novel coronavirus Activity (EC ₅₀ ) Measurement

Vero E6 cells were seeded at a density into microplates and at 5% CO ₂ Culturing overnight in an incubator at 37 ℃. The next day, a compound diluted in a double ratio (8 concentration points, three multiplex wells) and SARS-CoV-2 virus (B.1.1.7 (Alpha)) were added. Cell controls (cells, no compound treatment or virus infection), virus controls (cells infected with virus, no compound treatment) were set. Cells were cultured in an incubator for 3 or 4 days.

The antiviral activity of a compound is represented by the inhibition (%) of the compound at different concentrations against the cytopathic effect caused by the virus. Nonlinear fitting analysis of inhibition ratios of compounds using GraphPad Prism, EC of compounds was calculated ₅₀ . The results are shown in Table III.

Data on novel coronavirus activity against exterior three-body

Sample numbering	EC ₅₀ (μmol)
		Compound A	1.62
PY-01	0.35
		PY-02	0.58
PY-03	0.66
		PY-12	0.79
PY-15	0.57
		PY-18	0.66
PY-21	0.77
		PY-34	0.82
PY-49	0.33

Experimental results show that compared with the disclosed compound A, the detected compound sample has better inhibitory activity on SARS-CoV-2 virus (B.1.1.7 (Alpha)).

Example 41: evaluation of pharmacokinetic profile of Compound PY-01 in rats

12 SD rats, male, 180g-220g. The feeding conditions are as follows: 20-26 ℃, humidity: 40-70%, light illumination: dark = 12h:12h; rats were fed habitually for 3 days, during which time they had free access to drinking water. The cells were randomly divided into 4 groups of 3 cells. The 2 test pieces (PY-01 and Compound A) were administered by oral gavage and by tail vein injection in a single dose. The animals were fed after 4 hours of administration without water withdrawal, starting to fasted 16-17 hours at 5 pm on the day prior to administration.

Blood was collected 0.25h, 0.5h, 1h, 2h, 3h, 4h, 8h, 24h, or 0.083h, 0.25h, 0.5h, 1h, 2h, 4h, 8h, 24h, before, after single intravenous administration.

At each blood sampling time point, about 300 mu L of venous blood is taken from the eyeballs of the rats, added into a centrifugal tube with heparin sodium precooled by ice water, placed in an ice bath, and centrifuged (4000 rpm,10 min) after standing, split-packed in a unit volume of 50 mu L, placed in a sterile EP tube and preserved at-80 ℃ for later use. The concentration of compound B in the plasma was measured as soon as possible. The drug concentration in plasma was determined using HPLC (High Performance Liquid Chromatography) and LC/MS and the area under the concentration-time curve (AUC) in plasma was calculated using a nonlinear least squares procedure to calculate the bioavailability of the drug in rats.

Absolute bioavailability: F=AUClast-po/AUClast-iv×100%. The results are shown in Table IV.

The structure of compound B is as follows:

table four pharmacokinetic results

The SD rat pharmacokinetics experiment result shows that the compound PY-01 has longer half-life period, and the in vivo exposure amount is larger after oral administration, and the bioavailability is nearly 1.4 times that of the compound A. Compound B has a lower Cmax after administration of compound PY-01 than compound a, indicating a better safety margin for the compounds of the invention.

Example 42: therapeutic administration test against influenza virus in vivo

30C 57BL/6J mice, 6-8 weeks old, SPF-rated, female, were randomly assigned to 6 groups, numbered group 1, group 2, group 3, group 4, group 5, and group 6, respectively, 5 each. After nasal drip inoculation of 100pfu of influenza virus (H1N 1, A/WSN/33) virus at a semi-lethal dose for 24H, group 1 (vehicle group) was given a sodium carboxymethyl cellulose solution at 0.5% by lavage; group 2 was given a control (equimolar, 17.33mg/kg of Compound A and 19.01mg/kg of Compound C, combination); groups 3-5 were respectively gastrected with test drug (29.46 mg/kg of PY-01, 29.46mg/kg of PY-03, 29.46mg/kg of PY-49); group 6 began administration of 29.46mg/kg of PY-01 48 hours after nasal inoculation of the virus. Each group was given equimolar, twice daily, group 1-group 5 or vehicle for 4 days, group 6 for 3 days. Animals were sacrificed on day 5 post virus inoculation, lung tissue samples were harvested, and virus titer determinations were performed after homogenization treatment. The results are shown in FIG. 1.

The results show that the compound PY-01, the compound PY-03 and the compound PY-49 all show excellent anti-influenza virus (H1N 1) effect after 24 hours of virus infection and stronger anti-influenza virus (H1N 1) effect than the combined administration. The compound PY-01 can also significantly reduce the titer of the virus 48 hours after infection with the virus.

Example 43: in vivo anti-influenza virus (H1N 1) prophylactic administration test

BALB/c mice were 30, 6-8 week old, SPF-rated, female, and randomly assigned to 6 groups, numbered group 1, group 2, group 3, group 4, group 5, and group 6, 5 each. Each group was started on day-1 and 100PFU influenza virus (H1N 1, A/Puerto Rico/8/1934) was inoculated on day 0 by nasal drip, and virus was inoculated 2 hours after the day of virus inoculation. Wherein, group 1 (vehicle group) was intragastrically administered with 0.5% sodium carboxymethyl cellulose solution; group 2 was given a control (17.33 mg/kg of Compound A and 19.01mg/kg of Compound C, combination); groups 3-5 were respectively given different doses of test drug by gavage (high, medium, low dose groups of PY-01: 88.37mg/kg, 29.46mg/kg, 9.82 mg/kg); group 6 was given a high dose (88.37 mg/kg) of test drug PY-01 by gavage. Each group was equimolar administered once daily, group 1-5 or vehicle for 5 days, group 6 for 2 days. Animals were sacrificed on day 4 post virus inoculation, lung tissue samples were harvested, and virus titers were determined after homogenization treatment. The results are shown in FIG. 2.

The results show that: each group showed lower viral titers than the vehicle group; the virus titer was lower in the PY-01 high and medium dose groups compared to the combination group on the same days of administration; group 6 (high dose of PY-01 was administered 2 days before virus inoculation and no virus inoculation) had slightly higher virus titers than group 3 (high dose of PY-01 was administered 2 days before virus inoculation and no virus inoculation), but was still lower than the other groups. This demonstrates that the compounds of the present invention have a remarkable prophylactic effect against influenza virus.

While the invention has been described with reference to the preferred embodiments, it is not intended to limit the invention thereto, and it is to be understood that other modifications and improvements may be made by those skilled in the art without departing from the spirit and scope of the invention, which is therefore defined by the appended claims.

Claims

1. Novel cytidine derivatives, tautomers, stereoisomers, isotopic derivatives and pharmaceutically acceptable salts thereof, selected from the group consisting of the following compounds:

2. a pharmaceutical composition comprising the novel cytidine derivative, tautomer, stereoisomer, isotopic derivative, and pharmaceutically acceptable salts thereof as defined in claim 1.

3. Use of a novel cytidine derivative, tautomer, stereoisomer, isotopic derivative, and pharmaceutically acceptable salts thereof, as claimed in claim 1, or a pharmaceutical composition as claimed in claim 2, for the preparation of an anti-influenza a virus medicament.