WO2024067812A1 - Aromatic acetylene derivative, preparation method therefor, and use thereof - Google Patents

Abstract

The present invention relates to an aromatic acetylene derivative, a preparation method therefor, and a medical use of a pharmaceutical composition containing said derivative. Specifically, the present invention relates to an aromatic acetylene derivative represented by general formula (I), a preparation method therefor, a pharmaceutically acceptable salt thereof, and a use thereof as therapeutic agents, particularly LPXC inhibitors, the definitions of the substituents in the general formula (I) being the same as those in the description.

Description

Aromatic acetylene derivatives and preparation method and use thereof Technical Field

The present invention relates to an aromatic acetylene derivative, a preparation method thereof, a pharmaceutical composition containing the derivative, and the use of the aromatic acetylene derivative as a therapeutic agent, in particular as an LPXC inhibitor.

Background technique

The 1930s to 1960s were the golden age of antibiotic development. Since then, antibiotics have been widely used around the world, but bacterial resistance has also emerged. Drug-resistant bacteria have become a major threat to human health. Multidrug-resistant Gram-negative bacteria are one of the main pathogens of infection. Currently, there is a serious shortage of drugs used in clinical treatment of multidrug-resistant Gram-negative bacterial infections, and drugs with high toxicity are still used. In recent years, although bacterial resistance has been a hot topic in the international medical community, the pace of research and development has been slow, and there are very few compounds entering clinical research at home and abroad. Therefore, finding a new type of Gram-negative antibacterial drug is an important issue that needs to be solved urgently.

UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase (LPXC) is a Zn ⁺ -dependent metalloenzyme. It is the first rate-limiting enzyme in the synthesis of lipid A, which is an important component of the outer membrane of Gram-negative bacteria. It can anchor lipopolysaccharide on the outer membrane of the cell to maintain the integrity of its own cells. At the same time, it acts as a hydrophobic external barrier to prevent external factors such as antibiotics from entering the cell and protect bacteria from being invaded. In addition, lipid A is also the active ingredient of bacterial endotoxins, which enters the blood through the intestinal mucosa, activates the body's immune response, and even causes severe septic shock, which is also the reason why Gram-negative bacteria have pathogenic infections. Therefore, by inhibiting LPXC, the biosynthesis of Gram-negative lipid A can be inhibited, thereby effectively controlling Gram-negative bacterial infections.

At present, further understanding of the structure and characteristics of LPXC is mostly obtained through the separation, purification and analytical identification of LPXC crystals from Escherichia coli, Pseudomonas aeruginosa and hyperthermophilic bacteria. The structures of LPXC from these three different sources are highly similar, all containing two domains, and the active region is located at the junction of the two domains. Each domain contains an α-helix and a β-fold, and the β-fold sandwiches the α-helix to form a "β-α-α-β" sandwich structure. Although the amino acid sequences of the two domains are slightly different, they have the same spatial structure. In addition, each domain has a corresponding insertion region composed of β-folds to form different functional regions. Studies have shown that LPXC has a high homology in Gram-negative bacteria and has no common sequences with various enzyme systems in mammals. From a biological point of view, due to its unique advantages of broad spectrum and low toxicity, inhibiting LPXC as a target will be an ideal direction for studying antibacterial drugs.

There are no new LPXC inhibitors on the market. The small molecule compound RC-01 developed by Toyama Chemical Co., Ltd. has entered the clinical phase I, and other pharmaceutical companies such as Novartis and Taiwan Taisho Pharmaceutical are all in the preclinical stage for LPXC research. Therefore, the research and application of LPXC inhibitors have made some progress, but there is still huge room for improvement, and it is still necessary to continue to research and develop new LPXC inhibitors.

Summary of the invention

In view of the above technical problems, the present invention provides an aromatic acetylene derivative represented by general formula (I) or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof:

in:

W is selected from O, S(O) _r or N;

Ring C is selected from a 5- to 10-membered spiro heterocyclic group or a monocyclic heterocyclic group, wherein the monocyclic heterocyclic group contains a double bond;

X, Y, Z, and Q are each independently selected from CR ^A or N atoms, and at most two atoms among X, Y, Z, and Q are N atoms at the same time;

^RA is selected from hydrogen, halogen, hydroxy, cyano, alkyl or alkoxy; wherein the alkyl or alkoxy is optionally further substituted with one or more substituents selected from halogen, hydroxy, cyano, alkyl or alkoxy;

R ¹ is selected from the group consisting of absence, hydrogen, hydroxy, cyano, halogen, alkyl, alkoxy, cycloalkyl, heterocyclyl, aryl, heteroaryl, -C(O)R ³ , -C(O)NR ⁶ R ⁷ or -S(O) _r R ⁴ ; wherein the alkoxy, alkyl, cycloalkyl, heterocyclyl, aryl or heteroaryl is optionally further substituted by one or more R ^B ;

R ³ and R ⁴ are each independently selected from a hydrogen atom, a hydroxyl group, an alkyl group, a cycloalkyl group, an aryl group, a heteroaryl group or a heterocyclic group; wherein the alkyl group, the cycloalkyl group, the aryl group, the heteroaryl group or the heterocyclic group is optionally further substituted by one or more substituents selected from hydroxyl, halogen, nitro, cyano, alkyl, alkoxy, haloalkyl, haloalkoxy, cycloalkyl, heterocyclic group, aryl, heteroaryl, =O, -C(O)R ⁸ , -C(O)OR ⁸ , -OC(O)R ⁸ , -NR ⁹ R ¹⁰ , -C(O)NR ⁹ R ¹⁰ , -SO ₂ NR ⁹ R ¹⁰ or -NR ⁹ C(O)R ¹⁰ ;

R ^{and B} are each independently selected from cyano, halogen, alkyl, hydroxyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -OR ⁵ , -C(O)R ⁵ , -C(O)OR ⁵ , -NHC(O)R ⁵ , -NHC(O)OR ⁵ , -NR ⁶ R ⁷ , -C(O)NR ⁶ R ⁷ , -CH ₂ NHC(O)OR ⁵ , -CH ₂ NR ⁶ R ⁷ or -S(O) _r R ⁵ ; wherein the alkyl, cycloalkyl, heterocyclyl, aryl or heteroaryl is optionally further substituted by one or more selected from hydroxyl, halogen, nitro, cyano, alkyl, alkoxy, haloalkyl, haloalkoxy, cycloalkyl, heterocyclyl, aryl, heteroaryl, =O, -C(O)R ⁸ , -C(O)OR ⁸ , -OC(O)R ⁸ , -NR ⁹ R ¹⁰ , -C(O)NR ⁹ R ¹⁰ , _-SO2NR9R10 or ^-NR9C ( ^{O ) R10} ;

Alternatively, two ^RBs are attached to the same carbon atom to form a -C(=O)-;

^R2 are the same or different and are independently selected from hydroxy, cyano, halogen, alkyl or alkoxy; wherein the alkyl or alkoxy is optionally further substituted with one or more substituents selected from halogen, hydroxy, cyano, alkyl or alkoxy;

R ⁵ is independently selected from hydrogen, alkyl, cycloalkyl, heterocyclic, aryl or heteroaryl, wherein the alkyl, cycloalkyl, heterocyclic, aryl or heteroaryl is optionally further substituted by one or more selected from hydroxy, halogen, nitro, cyano, alkyl, alkoxy, haloalkyl, haloalkoxy, cycloalkyl, heterocyclic, aryl, heteroaryl, =O, -C(O)R ⁸ , -C(O)OR ⁸ , -OC(O)R ⁸ , -NR ⁹ R ¹⁰ , -C(O)NR ⁹ R ¹⁰ , -SO ₂ NR ⁹ R ¹⁰ or -NR ⁹ C(O)R ¹⁰ ;

^R6 and ^R7 are each independently selected from a hydrogen atom, a hydroxyl group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein the alkyl group, the alkoxy group, the cycloalkyl group, the heterocyclic group, the aryl group or the heteroaryl group is optionally further substituted with one or more substituents selected from hydroxyl group, a halogen, a nitro group, a cyano group, an alkyl group, an alkoxy group, ^a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, =O, -C(O) ^R8 , -C(O)OR8, -OC(O) ^R8 , ^-NR9R10 , ^-C (O ⁾ _NR9R10 ^{, -SO2NR9R10} or ^-NR9C ( ^O ) ^R10 ;

Alternatively, ^R6 and ^R7 together with the atoms to which they are attached form a 4- to 8-membered heterocyclic group, wherein the 4- to 8-membered heterocyclic group contains one or more N, O or S(O)r, and the ^4- to 8-membered heterocyclic group is optionally further substituted by one or more substituents selected from hydroxy, halogen, nitro, cyano, alkyl, alkoxy ^, cycloalkyl, heterocyclic group, aryl, heteroaryl, =O, -C(O) ^R8 , -C(O) ^OR8 , -OC(O) ^R8 , ^-NR9R10, -C(O ⁾ _NR9R10 ^{, -SO2NR9R10} or ^-NR9C (O) ^R10 ;

R ⁸ , R ⁹ and R ¹⁰ are each independently selected from a hydrogen atom, an alkyl group, an amino group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein the alkyl group, the cycloalkyl group, the heterocyclic group, the aryl group or the heteroaryl group is optionally further substituted with one or more substituents selected from a hydroxyl group, a halogen group, a nitro group, an amino group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, a carboxyl group or a carboxylate group;

m is 0, 1 or 2; m is preferably 0; and

r is independently 0, 1 or 2.

A preferred embodiment of the present invention provides a compound of general formula (I) or its stereoisomer, tautomer or pharmaceutically acceptable salt, which is a compound of general formula (II) or its stereoisomer, tautomer or pharmaceutically acceptable salt:

Ring C, R ¹ , R ² , W and m are as defined in the general formula (I).

A preferred embodiment of the present invention provides a compound of formula (I) or (II) or its stereoisomers, tautomers or pharmaceutically acceptable salts, wherein W is N.

A preferred embodiment of the present invention provides a compound of general formula (I) or (II) or a stereoisomer, a tautomer or a pharmaceutically acceptable salt thereof, wherein ring C is selected from:

A preferred embodiment of the present invention provides a compound of general formula (I) or (II) or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof, wherein:

R ¹ is selected from hydrogen atom, hydroxyl group, alkyl group, cycloalkyl group, heterocyclic group, -C(O)R ³ , -C(O)NR ⁶ R ⁷ or -S(O) _r R ⁴ ; wherein the alkyl group or cycloalkyl group is optionally further substituted by one or more R ^B ;

R ^{and B} are each independently selected from cyano, hydroxy, heterocyclyl, aryl, heteroaryl, -OR ⁵ , -C(O)OR ⁵ , -NR ⁶ R ⁷ , -C(O)NR ⁶ R ⁷ or -S(O) _r R ⁵ ; wherein the heterocyclyl or heteroaryl is optionally further substituted by one or more substituents selected from hydroxy, cyano, alkoxy, haloalkyl, =O, -C(O)OR ⁸ , -C(O)NR ⁹ R ¹⁰ ;

Alternatively, two ^RBs are attached to the same carbon atom to form a -C(=O)-;

R ³ is selected from hydrogen atom, alkyl, cycloalkyl, heteroaryl or heterocyclic group; wherein the alkyl, cycloalkyl, heteroaryl or heterocyclic group is optionally further substituted by one or more substituents selected from hydroxyl, alkoxy, cyano, heteroaryl, =O, -NR ⁹ R ¹⁰ , -C(O)OR ⁸ or -C(O)NR ⁹ R ¹⁰ ;

R ⁴ is a hydrogen atom or a hydroxyl group;

R ⁵ is each independently selected from a hydrogen atom, an alkyl group or a hydroxyalkyl group;

R ⁶ and R ⁷ are each independently selected from a hydrogen atom or an alkyl group, wherein the alkyl group is optionally substituted by one or more substituents selected from hydroxyl, cyano, halogen or alkoxy;

R ⁸ , R ⁹ , and R ¹⁰ are each independently selected from a hydrogen atom or an alkyl group; the alkyl group is preferably a methyl group;

r is 2.

In a preferred embodiment of the present invention, the compound described by general formula (I) is selected from:

or a stereoisomer, a tautomer or a pharmaceutically acceptable salt thereof.

Note: If there is a discrepancy between a drawn structure and the name given for that structure, the drawn structure will be given greater weight.

Furthermore, the present invention provides a pharmaceutical composition comprising an effective dose of a compound of general formula (I) or (II) or its stereoisomers, tautomers or pharmaceutically acceptable salts, and a pharmaceutically acceptable carrier, excipient or a combination thereof.

The present invention provides a use of a compound of general formula (I) or (II) or its stereoisomer, tautomer or pharmaceutically acceptable salt, or a pharmaceutical composition thereof (such as the pharmaceutical composition described in the aforementioned technical solution) in the preparation of an LPXC inhibitor.

The present invention also provides a use of a compound of formula (I) or (II) or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof (e.g., the pharmaceutical composition described in the aforementioned technical solution) in the preparation of a medicament for treating a disease mediated by LPXC, wherein the disease mediated by LPXC is preferably a bacterial infection caused by Gram-negative bacteria; wherein the disease mediated by LPXC is more preferably selected from bacterial infections caused by Gram-negative bacteria such as Escherichia coli, Pseudomonas aeruginosa, Proteus, Shigella dysenteriae, Klebsiella pneumoniae, Brucella, Salmonella typhi, Acinetobacter, Yersinia, Legionella pneumophila, Bordetella pertussis, Shigella, Pasteurella, Vibrio cholerae, and Neisseria meningitidis.

The present invention further provides a use of a compound of general formula (I) or (II) or its stereoisomers, tautomers or pharmaceutically acceptable salts, or a pharmaceutical composition thereof (such as the pharmaceutical composition described in the aforementioned technical solution) in the preparation of a drug for treating bacterial infections caused by Gram-negative bacteria.

The present invention provides a compound described in general formula (I) or (II) or its stereoisomer, tautomer or pharmaceutically acceptable salt, or a pharmaceutical composition thereof (such as the pharmaceutical composition described in the above technical solution) for use in the preparation of a drug for antibacterial infection. The invention relates to a method for treating a bacterial infection in a human body, wherein the bacterial infection is caused by gram-negative bacteria such as Escherichia coli, Pseudomonas aeruginosa, Proteus, Shigella dysenteriae, Klebsiella pneumoniae, Brucella, Salmonella typhi, Acinetobacter, Yersinia, Legionella pneumophila, Bordetella pertussis, Shigella, Pasteurella, Vibrio cholerae, and Neisseria meningitidis.

The present invention provides a method for preventing and/or treating a disease mediated by LPXC, comprising administering to a patient a therapeutically effective dose of a compound of formula (I) or (II) or a stereoisomer, a tautomer or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof (e.g., a pharmaceutical composition described in the aforementioned technical solution), wherein the disease mediated by LPXC is preferably a bacterial infection caused by Gram-negative bacteria; wherein the disease mediated by LPXC is more preferably selected from bacterial infections caused by Gram-negative bacteria such as Escherichia coli, Pseudomonas aeruginosa, Proteus, Shigella dysenteriae, Klebsiella pneumoniae, Brucella, Salmonella typhi, Acinetobacter, Yersinia, Legionella pneumophila, Bordetella pertussis, Shigella, Pasteurella, Vibrio cholerae, and Neisseria meningitidis.

The present invention also provides a method for preventing and/or treating bacterial infections caused by Gram-negative bacteria, comprising administering to a patient a therapeutically effective dose of a compound of general formula (I) or (II) or its stereoisomers, tautomers or pharmaceutically acceptable salts, or a pharmaceutical composition thereof (e.g., the pharmaceutical composition described in the aforementioned technical solution).

The present invention further provides a method for preventing and/or treating bacterial infections caused by Gram-negative bacteria such as Escherichia coli, Pseudomonas aeruginosa, Proteus, Shigella dysenteriae, Klebsiella pneumoniae, Brucella, Salmonella typhi, Acinetobacter, Yersinia, Legionella pneumophila, Bordetella pertussis, Shigella, Pasteurella, Vibrio cholerae, and Neisseria meningitidis, comprising administering to a patient a therapeutically effective dose of a compound of formula (I) or (II) or its stereoisomers, tautomers, or pharmaceutically acceptable salts thereof, or a pharmaceutical composition thereof (e.g., the pharmaceutical composition described in the aforementioned technical solution).

Detailed description of the invention

Unless otherwise stated, some of the terms used in the specification and claims of the present invention are defined as follows:

"Alkyl" when used as a group or a part of a group refers to a straight or branched aliphatic hydrocarbon group including _C1 - _C20 . Preferably, it is _C1 - _C10 alkyl, and more preferably _C1 - _C6 alkyl. Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, etc. Alkyl can be substituted or unsubstituted.

"Cycloalkyl" refers to a non-aromatic cyclic alkyl group in which one or more of the atoms forming the ring are carbon atoms, including monocyclic, polycyclic, condensed, bridged and spirocyclic rings, preferably having a 5- to 7-membered monocyclic ring or a 7- to 10-membered bicyclic or tricyclic ring. Examples of "cycloalkyl" include, but are not limited to, cyclopropyl, cyclopentyl, and cyclobutyl. Cycloalkyl groups may be substituted or unsubstituted.

"Spirocycloalkyl" refers to a polycyclic group with 5 to 18 members, two or more ring structures, and one carbon atom (called spiro atom) shared between the single rings, containing one or more double bonds in the ring, but no ring has a completely conjugated π electron aromatic system. Preferably, it is 6 to 14 members, and more preferably 7 to 10 members. According to the number of spiro atoms shared between the rings, spirocycloalkyl is divided into single spiro, double spiro or multi-spiro cycloalkyl, preferably single spiro and double spiro cycloalkyl, preferably 4-membered/5-membered, 4-membered/6-membered, 5-membered/5-membered or 5-membered/6-membered. Non-limiting examples of "spirocycloalkyl" include, but are not limited to, spiro[4.5]decyl, spiro[4.4]nonyl, spiro[3.5]nonyl, spiro[2.4]heptyl.

"Fused cycloalkyl" refers to a 5 to 18-membered, all-carbon polycyclic group containing two or more cyclic structures sharing a pair of carbon atoms, one or more rings may contain one or more double bonds, but no ring has a completely conjugated π electron aromatic system, preferably 6 to 12 members, more preferably 7 to 10 members. According to the number of constituent rings, it can be divided into bicyclic, tricyclic, tetracyclic or polycyclic fused cycloalkyl, preferably bicyclic or tricyclic, more preferably 5-membered/5-membered or 5-membered/6-membered bicyclic alkyl. Non-limiting examples of "fused cycloalkyl" include, but are not limited to: bicyclo[3.1.0]hexyl, bicyclo[3.2.0]hept-1-enyl, bicyclo[3.2.0]heptyl, decahydronaphthyl or tetradecahydrophenanthrenyl.

"Bridged cycloalkyl" refers to a 5-18 membered, all-carbon polycyclic group containing two or more cyclic structures, sharing two carbon atoms that are not directly connected to each other, one or more rings may contain one or more double bonds, but no ring has a completely conjugated π electron aromatic system, preferably 6-12 members, more preferably 7-10 members. Preferably 6-14 members, more preferably 7-10 members. According to the number of constituent rings, it can be divided into bicyclic, tricyclic, tetracyclic or polycyclic bridged cycloalkyl, preferably bicyclic, tricyclic or tetracyclic, more preferably bicyclic or tricyclic. Non-limiting examples of "bridged cycloalkyl" include, but are not limited to: (1s, 4s)-bicyclo[2.2.1]heptyl, bicyclo[3.2.1]octyl, (1s, 5s)-bicycloo[3.3.1]nonyl, bicyclo[2.2.2]octyl, (1r, 5r)-bicyclo[3.3.2]decyl.

"Heterocyclyl", "heterocycloalkyl", "heterocycle" or "heterocyclic" are used interchangeably in this application and refer to non-aromatic heterocyclic groups, wherein one or more atoms forming the ring are selected from nitrogen, oxygen or S(O)r (wherein r is selected from 0, 1 or 2) heteroatoms, including monocyclic, polycyclic, fused, bridged and spirocyclic groups, and the heterocyclic group may be substituted or unsubstituted.

Examples of "monocyclic heterocyclyl" include, but are not limited to, morpholinyl, oxetanyl, thiomorpholinyl, tetrahydrofuranyl, tetrahydropyranyl, 1,1-dioxo-thiomorpholinyl, piperidinyl, 2-oxo-piperidinyl, pyrrolidinyl, 2-oxo-pyrrolidinyl, piperazin-2-one, 8-oxa-3-aza-bicyclo[3.2.1]octyl, piperazinyl, hexahydropyrimidine, 1,4-dioxane, or

"Spiro heterocyclic group" refers to 5 to 18 yuan, two or more cyclic structures, and a polycyclic group that shares one atom between monocyclic rings, containing one or more double bonds in the ring, but no ring has a completely conjugated π-electron aromatic system, wherein one or more ring atoms are selected from nitrogen, oxygen or S (O) _r (wherein r is selected from 0, 1 or 2) heteroatoms, and the remaining ring atoms are carbon. Preferably 5 to 14 yuan, more preferably 6 to 10 yuan, most preferably 7 to 10 yuan. According to the number of shared spiral atoms between rings, spiro cycloalkyl is divided into single spiral heterocyclic group, double spiral heterocyclic group or multi-spiro heterocyclic group, preferably single spiral heterocyclic group and double spiral heterocyclic group. More preferably 4 yuan/4 yuan, 4 yuan/5 yuan, 4 yuan/6 yuan, 5 yuan/5 yuan or 5 yuan/6 yuan single spiral heterocyclic group. Non-limiting examples of "spiroheterocyclyl" include, but are not limited to, 1,7-dioxaspiro[4.5]decyl, 2-oxa-7-azaspiro[4.4]nonyl, 7-oxaspiro[3.5]nonyl, 5-oxaspiro[2.4]heptyl,

"Fused heterocyclic group" refers to an all-carbon polycyclic group containing two or more cyclic structures that share a pair of atoms with each other, one or more rings may contain one or more double bonds, but no ring has a completely conjugated π electron aromatic system, wherein one or more ring atoms are selected from nitrogen, oxygen or S(O) _r (wherein r is selected from 0, 1 or 2) heteroatoms, and the remaining ring atoms are carbon. Preferably, it is 6 to 14 members, more preferably 7 to 10 members. According to the number of constituent rings, it can be divided into a bicyclic, tricyclic, tetracyclic or polycyclic fused heterocyclic group, preferably a bicyclic or tricyclic, more preferably a 5-membered/5-membered or 5-membered/6-membered bicyclic fused heterocyclic group. Non-limiting examples of "fused heterocyclic group" include, but are not limited to: octahydropyrrolo[3,4-c]pyrrolyl, octahydro-1H-isoindolyl, 3-azabicyclo[3.1.0]hexyl, octahydrobenzo[b][1,4]dioxine.

"Bridged heterocyclic group" refers to a 5-14-membered, 5-18-membered, polycyclic group containing two or more cyclic structures, sharing two atoms that are not directly connected to each other, one or more rings may contain one or more double bonds, but no ring has a completely conjugated π electron aromatic system, wherein one or more ring atoms are selected from nitrogen, oxygen or S(O) _r (wherein r is selected from 0, 1 or 2) heteroatoms, and the remaining ring atoms are carbon. Preferably it is 6 to 14 members, more preferably 7 to 10 members. According to the number of constituent rings, it can be divided into a bicyclic, tricyclic, tetracyclic or polycyclic bridged heterocyclic group, preferably a bicyclic, tricyclic or tetracyclic, more preferably a bicyclic or tricyclic. Non-limiting examples of "bridged heterocyclic group" include, but are not limited to: 2-azabicyclo[2.2.1]heptyl, 2-azabicyclo[2.2.2]octyl, 2-azabicyclo[3.3.2]decyl.

"Aryl" refers to a carbocyclic aromatic system containing one or two rings, wherein the rings may be connected together in a fused manner. The term "aryl" includes monocyclic or bicyclic aromatic groups, such as phenyl, naphthyl, and tetrahydronaphthyl aromatic groups. Preferably, the aryl group is a C ₆ -C ₁₀ aromatic group, more preferably, the aryl group is phenyl and naphthyl, and most preferably, naphthyl. The aryl group may be substituted or unsubstituted.

"Heteroaryl" refers to an aromatic 5- to 6-membered monocyclic or 8- to 10-membered bicyclic ring which may contain 1 to 4 atoms selected from nitrogen, oxygen and/or sulfur. Examples of "heteroaryl" include, but are not limited to, furanyl, pyridyl, 2-oxo-1,2-dihydropyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, thienyl, isoxazolyl, oxazolyl, oxadiazolyl, imidazolyl, pyrrolyl, pyrazolyl, triazolyl, tetrazolyl, thiazolyl, isothiazolyl, 1,2,3-thiadiazolyl, benzodioxolyl, benzothienyl, benzimidazolyl, indolyl, isoindolyl, 1,3-dioxo-isoindolyl, quinolyl, indazolyl, benzisothiazolyl, benzoxazolyl, benzisoxazolyl, isothiazolyl, 1H-1,2,4-triazolyl, 4H-1,2,4 -triazolyl, pyridyl, pyrimidinyl, pyrazin-2(1H)-onyl, pyrimidin-4(3H)-onyl, pyridazin-3(2H)-onyl, 1H-indolyl, 1H-benzo[d]imidazolyl, 1H-pyrrolo[2,3-c]pyridinyl, 3H-imidazo[4,5-c]pyridinyl, isoquinolyl, quinazolinyl, 2H-isoindolyl, furano[3,2-b]pyridinyl, furano[2,3-c]pyridinyl, thieno[2,3-c]pyridinyl, benzofuranyl, benzo[b]thienyl, 1H-pyrrolo[3,2-b]pyridinyl, 2H-pyrrolo[3,4-c]pyridinyl. The heteroaryl group may be substituted or unsubstituted.

"Alkoxy" refers to a group of (alkyl-O-), wherein alkyl is as defined herein. C ₁ -C ₆ alkoxy is preferred, and examples thereof include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, and the like.

"Nitro" refers to a _-NO2 radical.

"Hydroxy" refers to an -OH group.

"Halogen" refers to fluorine, chlorine, bromine and iodine.

"Amino" refers to _-NH2 .

"Cyano" refers to -CN.

"Benzyl" refers to _-CH2 -phenyl.

"Carboxy" refers to -C(O)OH.

"Carboxylate" refers to -C(O)O-alkyl or -C(O)O-cycloalkyl, wherein alkyl and cycloalkyl are as defined above.

"Hydroxyalkyl" or "hydroxyalkyl" refers to an alkyl group substituted with a hydroxy group wherein alkyl is as defined above.

"Aminoalkyl" refers to an alkyl group substituted with an amino group, wherein alkyl is as defined above.

"Haloalkyl" refers to an alkyl group substituted with a halogen, wherein alkyl is as defined above.

"Haloalkoxy" refers to an alkoxy group substituted with a halogen group, wherein alkoxy is as defined above.

"DMSO" refers to dimethyl sulfoxide.

"BOC" refers to tert-butoxycarbonyl.

"Bn" refers to benzyl.

"THP" refers to 2-tetrahydropyranyl.

"TFA" refers to trifluoroacetic acid.

"Ts" refers to p-toluenesulfonyl.

"Leaving group", or leaving group, is an atom or functional group that leaves a larger molecule in a chemical reaction. It is a term used in nucleophilic substitution reactions and elimination reactions. In nucleophilic substitution reactions, the reactant attacked by the nucleophile is called the substrate, and the atom or group of atoms that breaks away from the substrate molecule with a pair of electrons is called the leaving group. Groups that are easy to accept electrons and have a strong ability to bear negative charges are good leaving groups. The smaller the pKa of the conjugate acid of the leaving group, the easier it is for the leaving group to leave other molecules. The reason is that when the pKa of its conjugate acid is smaller, the corresponding leaving group does not need to combine with other atoms, and the tendency to exist as an anion (or electrically neutral leaving group) is enhanced. Common leaving groups include but are not limited to halogens, mesyl, -OTs or -OH.

"Substituted" means that one or more hydrogen atoms, preferably up to 5, more preferably 1 to 3 hydrogen atoms in the group are replaced independently of each other by a corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and the skilled person can determine (by experiment or theory) possible or impossible substitutions without undue effort. For example, amino or hydroxy groups with free hydrogens may be unstable when combined with carbon atoms with unsaturated (e.g. olefinic) bonds.

The term "substituted" or "substituted" as used herein, unless otherwise specified, means that a group may be substituted by one or more groups selected from the following: alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxyl, nitro, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, cycloalkyloxy, heterocycloalkyloxy, cycloalkylthio, heterocycloalkylthio, amino, haloalkyl, hydroxyalkyl, carboxyl, carboxylate, =O, -OR ⁵ , -C(O)R ⁵ , -C(O)OR ⁵ , -NHC(O)R ⁵ , -NHC(O)OR ⁵ , -NR ⁶ R ⁷ , -C(O)NR ⁶ R ⁷ , -CH ₂ NHC(O)OR ⁵ , -CH ₂ NR ⁶ R ⁷ or -S(O)rR ⁵ ;

R ⁶ is selected from a hydrogen atom, an alkyl group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein the alkyl group, the cycloalkyl group, the heterocyclic group, the aryl group or the heteroaryl group is optionally further substituted by one or more substituents selected from hydroxyl, halogen, nitro, cyano, alkyl, alkoxy, haloalkyl, haloalkoxy, cycloalkyl, heterocyclic group, aryl, heteroaryl, =O, -C(O)R ⁸ , -C(O)OR ⁸ , -OC(O)R ⁸ , -NR ⁹ R ¹⁰ , -C(O)NR ⁹ R ¹⁰ , -SO ₂ NR ⁹ R ¹⁰ or -NR ⁹ C(O)R ¹⁰ ;

^R6 and ^R7 are each independently selected from a hydrogen atom, a hydroxyl group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein the alkyl group, the alkoxy group, the cycloalkyl group, the heterocyclic group, the aryl group or the heteroaryl group is optionally further substituted with one or more substituents selected from hydroxyl group, a halogen, a nitro group, a cyano group, an alkyl group, an alkoxy group, ^a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, =O, -C(O) ^R8 , -C(O)OR8, -OC(O) ^R8 , ^-NR9R10 , ^-C (O ⁾ _NR9R10 ^{, -SO2NR9R10} or ^-NR9C ( ^O ) ^R10 ;

Alternatively, ^R6 and ^R7 together with the atoms to which they are attached form a 4- to 8-membered heterocyclic group, wherein the 4- to 8-membered heterocyclic group contains one or more N, O or S(O)r, and the ^4- to 8-membered heterocyclic group is optionally further substituted by one or more substituents selected from hydroxy, halogen, nitro, cyano, alkyl, alkoxy ^, cycloalkyl, heterocyclic group, aryl, heteroaryl, =O, -C(O) ^R8 , -C(O) ^OR8 , -OC(O) ^R8 , ^-NR9R10 , -C(O ⁾ _NR9R10 ^{, -SO2NR9R10} or ^-NR9C (O) ^R10 ;

R ⁸ , R ⁹ and R ¹⁰ are each independently selected from a hydrogen atom, an alkyl group, an amino group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein the alkyl group, the cycloalkyl group, the heterocyclic group, the aryl group or the heteroaryl group is optionally further substituted with one or more substituents selected from a hydroxyl group, a halogen group, a nitro group, an amino group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, a carboxyl group or a carboxylate group;

r is selected from 0, 1 or 2.

The compounds of the present invention may contain asymmetric centers or chiral centers and therefore exist in different stereoisomeric forms. It is contemplated that all stereoisomeric forms of the compounds of the present invention, including but not limited to diastereomers, enantiomers and atropisomers and geometric (conformation) isomers and mixtures thereof, such as racemic mixtures, are within the scope of the present invention.

Unless otherwise indicated, structures depicted herein also encompass all isomeric (e.g., diastereoisomers, enantiomers, and atropisomers and geometric (conformational) isomeric forms of such structures; for example, R and S configurations at various asymmetric centers, (Z) and (E) double bond isomers, and (Z) and (E) conformational isomers. Therefore, individual stereoisomers as well as enantiomeric mixtures, diastereomeric mixtures, and geometric (conformational) isomeric mixtures of the compounds of the invention are within the scope of the invention.

"Pharmaceutically acceptable salts" refer to salts of the above compounds that can retain their original biological activity and are suitable for medical use. Pharmaceutically acceptable salts of the compounds represented by general formula (I) may be metal salts or amine salts formed with suitable acids.

"Pharmaceutical composition" means a mixture containing one or more compounds described herein or their physiologically pharmaceutically acceptable salts or prodrugs and other chemical components, as well as other components such as physiologically pharmaceutically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration to an organism, facilitate the absorption of the active ingredient, and thus exert biological activity.

Synthesis method of the compound of the present invention

In order to achieve the purpose of the present invention, the present invention adopts the following technical solution:

The present invention provides a method for preparing a compound of general formula (I) or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof, the method comprising:

When W is N:

method 1:

The compound of general formula (I-a) undergoes a coupling reaction with the compound of general formula (I-b) in the presence of a catalyst, and optionally further undergoes a deprotection reaction to obtain a compound of general formula (I-c). The compound of general formula (I-c) optionally further undergoes a substitution reaction with the compound of general formula (I-d) to obtain a compound of general formula (I).

in:

X ₁ and X ₂ are selected from halogen;

PG is an amino protecting group, preferably tert-butyloxycarbonyl;

PG' is a hydroxyl protecting group, preferably 2-tetrahydropyranyl;

Ring C, X, Y, Z, Q, R ¹ , R ² and m are as described in the general formula (I).

Method 2:

The compound of general formula (I-a) undergoes a coupling reaction with the compound of general formula (II-b) in the presence of a catalyst to obtain a compound of general formula (II-c). The compound of general formula (II-c) further undergoes a deprotection reaction to obtain a compound of general formula (I).

in:

_X1 is selected from halogen;

PG' is a hydroxyl protecting group, preferably 2-tetrahydropyranyl;

Ring C, X, Y, Z, Q, R ¹ , R ² and m are as described in the general formula (I).

Detailed ways

The present invention is further described below with reference to the embodiments, but these embodiments are not intended to limit the scope of the present invention.

Example

The examples provide the preparation of representative compounds represented by formula (I) and related structural identification data. It must be noted that the following examples are used to illustrate the present invention rather than to limit the present invention. ¹ H NMR spectra were obtained using a Bruker instrument (400 MHz), chemical shifts are expressed in ppm. Tetramethylsilane internal standard (0.00 ppm) was used. ¹ H NMR notation: s = singlet, d = doublet, t = triplet, m = multiplet, br = broadened, dd = double of doublets, dt = double of triplets, dq = double of quartets. Coupling constants, when provided, are in Hz.

The mass spectrum is obtained by LC/MS, and the ionization method can be ESI or APCI.

The thin layer chromatography silica gel plate uses Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plate. The silica gel plate used in thin layer chromatography (TLC) adopts a specification of 0.15mm-0.2mm, and the specification used for thin layer chromatography separation and purification products is 0.4mm-0.5mm.

Column chromatography generally uses Yantai Huanghai silica gel 200-300 mesh silica gel as the carrier.

In the following examples, unless otherwise specified, all temperatures are in degrees Celsius. Unless otherwise specified, various starting materials and reagents are commercially available or synthesized according to known methods. Commercially available raw materials and reagents are used directly without further purification. Unless otherwise specified, commercial manufacturers include but are not limited to Aldrich Chemical Company, ABCR GmbH & Co. KG, Acros Organics, Guangzan Chemical Technology Co., Ltd. and Jingyan Chemical Technology Co., Ltd.

DMSO-d ₆ : deuterated dimethyl sulfoxide.

Argon atmosphere means that the reaction bottle is connected to an argon balloon with a capacity of about 1L.

Unless otherwise specified in the examples, the solution in the reaction refers to an aqueous solution.

The compound is purified using a silica gel column chromatography eluent system and thin layer chromatography, wherein the eluent system is selected from: A: petroleum ether and ethyl acetate system; B: dichloromethane and methanol system; C: dichloromethane and ethyl acetate system; wherein the volume ratio of the solvent varies according to the polarity of the compound, and a small amount of acidic or alkaline reagents, such as acetic acid or triethylamine, may also be added for adjustment.

Example 1

(S)-1-(1-((5-(4-((7-azaspiro[3.5]nonan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

first step

Ethyl 4-(4-iodophenyl)-2,4-dioxobutanoate

Sodium hydride (1.63 g, 40.64 mmol, 60% oil dispersion) was added to toluene (20 mL), 1-(4-iodophenyl)ethan-1-one 1a (5 g, 20.32 mmol) was added, and the mixture was heated to 50°C. A toluene solution (20 mL) of diethyl oxalate 1b (4.45 g, 30.48 mmol) was added dropwise, and the mixture was heated to 50°C for 2 hours. The mixture was cooled, poured into ice water, and acidified with 1M hydrochloric acid. The mixture was extracted with ethyl acetate (100 mL × 2). The combined organic phase was washed with saturated brine (100 mL × 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System A) to obtain ethyl 4-(4-iodophenyl)-2,4-dioxobutanoate 1c (2.8 g) with a yield of 39.81%.

MS m/z(ESI):347.0[M+1].

Step 2

Ethyl 5-(4-iodophenyl)isoxazole-3-carboxylate

Ethyl 4-(4-iodophenyl)-2,4-dioxobutanoate 1c (2.8 g, 8.09 mmol) and hydroxylamine hydrochloride (1.69 g, 24.27 mmol) were added to ethanol (25 mL) and heated under reflux for 1 hour. After the reaction was completed, the mixture was concentrated under reduced pressure and ethyl acetate (100 mL) was added to dissolve the mixture. The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System A) to give ethyl 5-(4-iodophenyl)isoxazole-3-carboxylate 1d (2.3 g) with a yield of 82.86%.

MS m/z(ESI):343.8[M+1].

third step

(5-(4-iodophenyl)isoxazol-3-yl)methanol

Ethyl 5-(4-iodophenyl)isoxazole-3-carboxylate 1d (1.20 g, 3.50 mmol) was added to methanol (25 mL), sodium borohydride (198.46 mg, 5.25 mmol) was added in batches, the temperature was raised to 80°C, and the reaction was carried out for 4 hours. After the reaction was completed, ice water was added to quench the reaction, and ethyl acetate was used for extraction (50 mL×2). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain (5-(4-iodophenyl)isoxazol-3-yl)methanol 1e (0.36 g), with a yield of 34.19%.

MS m/z(ESI):302.0[M+1].

the fourth step

(5-(4-iodophenyl)isoxazol-3-yl)methanesulfonate

(5-(4-iodophenyl)isoxazol-3-yl)methanol 1e (0.36 g, 1.20 mmol) and triethylamine (241.99 mg, 2.39 mmol, 333.31 μL) were added to dichloromethane (5 mL), cooled to 0°C, and methanesulfonyl chloride 1f (205.45 mg, 1.79 mmol) was added dropwise, and the mixture was heated to room temperature for 4 hours. After the reaction was completed, water was added to quench the reaction, and dichloromethane (50 mL×3) was used for extraction. The organic phases were combined and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: A system) to obtain (5-(4-iodophenyl)isoxazol-3-yl)methanesulfonic acid methyl ester 1g (0.45 g), with a yield of 99.26%.

MS m/z(ESI):379.8[M+1].

the fifth step

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole

2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazole 1h (100 mg, 509.57 μmol) was added to N,N-dimethylformamide (2 mL), cooled to 0°C, sodium hydride (50.96 mg, 764.35 μmol, 60% oil dispersion) was added in batches, the temperature was raised to room temperature for reaction for 1 hour, (5-(4-iodophenyl)isoxazol-3-yl)methanesulfonic acid methyl ester 1g (193.21 mg, 509.57 μmol) was added, and the reaction was continued at room temperature for 4 hours. After the reaction was completed, water was added to quench the reaction, and the mixture was extracted with ethyl acetate (10 mL×3). The organic phases were combined and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System A) to give 5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (0.12 g), yield: 49.13%.

MS m/z(ESI):480.1[M+1].

Step 6

tert-butyl 2-ethynyl-7-azaspiro[3.5]nonane-7-carboxylate

2-Ethynyl-7-azaspiro[3.5]nonane-7-carboxylic acid tert-butyl ester

Under ice bath, dimethyl (1-diazo-2-oxopropyl)phosphonate (1.36 g, 7.11 mmol) was added to a solution of potassium carbonate (1.64 g, 11.8 mmol) and tert-butyl 2-formyl-7-azaspiro[3.5]nonane-7-carboxylate 1j (1.5 g, 5.92 mmol, commercially available) in methanol (15 mL), and the mixture was reacted at room temperature for 24 hours. After the reaction was completed, water (50 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (50 mL×3). The combined organic phase was washed with saturated brine (100 mL), and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System A) to obtain tert-butyl 2-ethynyl-7-azaspiro[3.5]nonane-7-carboxylate 1k (1.3 g), with a yield of 88.1%.

MS m/z(ESI):194.1[M+1-56].

Step 7

tert-butyl

2-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonane-7-carboxylate

tert-Butyl 2-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonane-7-carboxylate

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (1.3 g, 2.71 mmol), tert-butyl 2-ethynyl-7-azaspiro[3.5]nonane-7-carboxylate 1k (1.26 g, 3.53 mmol), bistriphenylphosphine palladium dichloride (190 mg, 271 μmol), cuprous iodide (51.8 mg, 271 μmol) and triethylamine (823 mg, 8.14 mmol, 1.13 mL) were added to N,N-dimethylformamide (13.87 mL), the argon atmosphere was replaced three times, and the reaction was carried out at room temperature overnight. After the reaction was completed, water (30 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (100 mL×3). The combined organic phases were washed with saturated brine (100 mL), and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System B) to give 2-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonane-7-carboxylic acid tert-butyl ester 11 (1.25 g), yield: 76.7%.

MS m/z(ESI):601.1[M+1].

Step 8

(S)-1-(1-((5-(4-((7-azaspiro[3.5]nonan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

2-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonane-7-carboxylic acid tert-butyl ester 11 (1.2 g, 2.00 mmol) was dissolved in dichloromethane (30 mL), trifluoroacetic acid (228 mg, 2.00 mmol) was slowly added, and the reaction was carried out at room temperature for 4 hours. After the reaction was completed, the reaction mixture was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm ID; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (0.85 g) with a yield of 76.2%.

MS m/z(ESI):417.1[M+1].

Example 2

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)butanoic acid

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)butanoic acid

first step

ethyl

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)butanoate

(S)-ethyl 4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)butanoate

(S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (100 mg, 240 μmol), ethyl 4-bromobutyrate 2a (70.3 mg, 360 μmol, commercially available), potassium carbonate (332 mg, 2.40 mmol) and sodium iodide (119 mg, 792 μmol) were added to N,N-dimethylformamide (3 mL) in sequence and heated to 70 °C for 4 hours. After the reaction was completed, water (30 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (50 mL×3). The combined organic phases were washed with saturated brine (100 mL), and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System B) to give (S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)butanoic acid ethyl ester 2b (65 mg), yield: 51.0%.

MS m/z(ESI):531.1[M+1].

Step 2

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)butanoic acid

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)butanoic acid

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)butanoic acid ethyl ester 2b (30 mg, 56.5 μmol) was dissolved in a mixed solvent of methanol (0.5 mL), tetrahydrofuran (0.5 mL) and water (0.5 mL), sodium hydroxide (2.26 mg, 56.5 μmol) was added, and the mixture was heated to 70°C for 4 hours. After the reaction was completed, a small amount of dilute hydrochloric acid was added to adjust the pH to about 4, and the organic solvent was removed by concentration to prepare a liquid phase separation (separation column) AKZONOBEL Kromasil; 250×21.2 mm ID; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)butanoic acid 2 (1.8 mg), yield: 4.9%.

MS m/z(ESI):503.1[M+1].

Example 3

(S)-1-(1-((5-(4-((6-azaspiro[3.4]octan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((6-azaspiro[3.4]octan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

first step

tert-butyl 2-ethynyl-6-azaspiro[3.4]octane-6-carboxylate

2-Ethynyl-6-azaspiro[3.4]octane-6-carboxylic acid tert-butyl ester

Under argon protection, dimethyl (1-diazo-2-oxopropyl)phosphonate (241 mg, 1.25 mmol) was added to a solution of potassium carbonate (289 mg, 2.09 mmol) and tert-butyl 2-formyl-6-azaspiro[3.4]octane-6-carboxylate 3a (250 mg, 1.04 mmol) in methanol (4 mL) under ice bath, and reacted at room temperature for 24 hours. After the reaction was completed, water (20 mL) was added to dilute the reaction, and ethyl acetate was extracted (50 mL×3). The combined organic phase was washed with saturated brine (50 mL), and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: A system) to obtain tert-butyl 2-ethynyl-6-azaspiro[3.4]octane-6-carboxylate 3b (180 mg), with a yield of 73.2%.

MS m/z(ESI):170.1[M-55].

Step 2

tert-butyl

2-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octane-6-carboxylate

tert-Butyl 2-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octane-6-carboxylate

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (0.4 g, 835 μmol), tert-butyl 2-ethynyl-6-azaspiro[3.4]octane-6-carboxylate 3b (365 mg, 1.08 mmol), bistriphenylphosphine palladium dichloride (58.5 mg, 83.5 μmol), cuprous iodide (15.9 mg, 83.5 μmol) and triethylamine (253 mg, 2.50 mmol, 348 μL) were mixed with N,N-dimethylformamide (4.65 mL), argon was replaced three times, and the mixture was reacted at room temperature overnight. After the reaction was completed, water (20 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (30 mL×3). The combined organic phase was washed with saturated brine (50 mL), and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System B) to give 2-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octane-6-carboxylic acid tert-butyl ester 3c (0.4 g) in a yield of 81.7%.

MS m/z(ESI):587.3[M+1].

third step

(S)-1-(1-((5-(4-((6-azaspiro[3.4]octan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((6-azaspiro[3.4]octan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

2-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octane-6-carboxylic acid tert-butyl ester 3c (0.3 g, 511 μmol) was dissolved in dichloromethane (5 mL), trifluoroacetic acid (58 mg, 511 μmol) was slowly added dropwise, and the mixture was reacted at room temperature for 4 hours. After the reaction was completed, the mixture was concentrated. Preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) gave (S)-1-(1-((5-(4-((6-azaspiro[3.4]octan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 3 (120 mg) in a yield of 43.2%.

MS m/z(ESI):403.1[M+1].

Example 4

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octan-6-yl)butanoic acid

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octan-6-yl)butanoic acid

first step

ethyl

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octan-6-yl)butanoate

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octan-6-yl)butanoic acid ethyl ester

(S)-1-(1-((5-(4-((6-azaspiro[3.4]octan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 3 (100 mg, 248 μmol), ethyl 4-bromobutyrate 2a (72.7 mg, 373 μmol), potassium carbonate (343 mg, 2.48 mmol) and sodium iodide (74.5 mg, 497 μmol) were added to N,N-dimethylformamide (3 mL) and heated to 70°C for 4 hours. After the reaction was completed, water (20 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (50 mL×3). The combined organic phases were washed with saturated brine (50 mL), and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System B) to give (S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octan-6-yl)butanoic acid ethyl ester 4a (60 mg) with a yield of 46.7%.

MS m/z(ESI):517.1[M+1].

Step 2

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octan-6-yl)butanoic acid

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octan-6-yl)butanoic acid

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octan-6-yl)butanoic acid ethyl ester 4a (60 mg, 116 μmol) was dissolved in a mixed solvent of methanol (1 mL), tetrahydrofuran (1 mL) and water (1 mL), sodium hydroxide (23 mg, 580 μmol) was added, and the mixture was heated to 70°C for 4 hours. After the reaction was completed, a small amount of dilute hydrochloric acid was added to adjust the pH to about 4. The reaction mixture was concentrated to remove most of the organic solvent and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octan-6-yl)butanoic acid 4 (30 mg) with a yield of 40.7%.

MS m/z(ESI):489.1[M+1].

Example 5

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propanoic acid

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propanoic acid

first step

methyl

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propanoate

(S)-methyl 3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propanoate

(S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (50 mg, 120 μmol), methyl 3-bromopropanoate 5a (30 mg, 180 μmol), potassium carbonate (166 mg, 1.20 mmol) and sodium iodide (59.4 mg, 396 μmol) were added to N,N-dimethylformamide (3 mL) and heated to 70 °C for 4 hours. After the reaction was completed, water (20 mL) was added to dilute the reaction, and the mixture was extracted with ethyl acetate (30 mL×3). The combined organic phases were washed with saturated brine (50 mL), and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System A) to give (S)-methyl 3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)propanoate 5b (40 mg), yield: 66.7%.

MS m/z(ESI):503.1[M+1].

Step 2

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propanoic acid

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propanoic acid

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)propanoic acid methyl ester 5b (80 mg, 159 μmol) was dissolved in a mixed solvent of methanol (1 mL), tetrahydrofuran (1 mL) and water (1 mL), sodium hydroxide (6.4 mg, 159 μmol) was added, and the mixture was heated to 70 °C for 4 hours. After the reaction was completed, a small amount of dilute hydrochloric acid was added to adjust the pH to about 4, and the reaction mixture was concentrated to remove most of the organic solvent. Preparative liquid separation was performed (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)propanoic acid 5 (20 mg) with a yield of 19.8%.

MS m/z(ESI):489.1[M+1].

Example 6

(S)-1-(1-((5-(4-((2-azaspiro[3.3]heptan-6-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((2-azaspiro[3.3]hept-6-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

first step

tert-butyl 6-ethynyl-2-azaspiro[3.3]heptane-2-carboxylate

tert-Butyl 6-ethynyl-2-azaspiro[3.3]heptane-2-carboxylate

Under argon protection, dimethyl (1-diazo-2-oxopropyl)phosphonate (614 mg, 3.20 mmol) was slowly added to a methanol (8 mL) solution of potassium carbonate (736 mg, 5.33 mmol) and tert-butyl 6-formyl-2-azaspiro[3.3]heptane-2-carboxylate 6a (0.6 g, 2.66 mmol) under ice bath, and reacted overnight at room temperature. After the reaction was completed, water (20 mL) was added to dilute the reaction, and ethyl acetate was extracted (30 mL×3). The combined organic phase was washed with saturated brine (50 mL) and concentrated under reduced pressure to obtain the residue. The product was purified by silica gel column chromatography (eluent: System A) to give tert-butyl 6-ethynyl-2-azaspiro[3.3]heptane-2-carboxylate 6b (0.455 g). Yield: 77.4%.

MS m/z(ESI):166.1[M-55].

Step 2

tert-butyl

6-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptane-2-carboxylate

tert-Butyl 6-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptane-2-carboxylate

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (0.6 g, 1.25 mmol), tert-butyl 6-ethynyl-2-azaspiro[3.3]heptane-2-carboxylate 6b (455 mg, 1.44 mmol), bistriphenylphosphine palladium dichloride (87.8 mg, 125 μmol), cuprous iodide (23.9 mg, 125 μmol) and triethylamine (380 mg, 3.76 mmol, 522 μL) were added sequentially to N,N-dimethylformamide (4 mL), argon was replaced, and the reaction was carried out at room temperature overnight. After the reaction was completed, water (20 mL) was added to dilute the reaction, and the mixture was extracted with ethyl acetate (100 mL×3). The combined organic phases were washed with saturated brine (100 mL), and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System B) to give 6-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptane-2-carboxylic acid tert-butyl ester 6c (0.65 g) in a yield of 90.7%.

MS m/z(ESI):573.1[M+1].

third step

(S)-1-(1-((5-(4-((2-azaspiro[3.3]heptan-6-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((2-azaspiro[3.3]hept-6-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

6-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptane-2-carboxylic acid tert-butyl ester 6c (20 mg, 34.92 μmol) was dissolved in dichloromethane (1 mL), trifluoroacetic acid (46 mg, 405 μmol) was slowly added dropwise, and the reaction was carried out at room temperature for 3 hours. After the reaction was completed, the product was concentrated and the residue was separated by preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-1-(1-((5-(4-((2-azaspiro[3.3]hept-6-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 6 (5 mg) in a yield of 28.5%.

MS m/z(ESI):389.1[M+1].

Example 7

(S)-4-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptan-2-yl)butanoic acid

(S)-4-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)butanoic acid

first step

ethyl

(S)-4-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptan-2-yl)butanoate

(S)-4-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)butanoic acid ethyl ester

(S)-1-(1-((5-(4-((2-azaspiro[3.3]hept-6-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 6 (135 mg, 348 μmol), ethyl 4-bromobutyrate 2a (102 mg, 521 μmol, commercially available), potassium carbonate (480 mg, 3.48 mmol) and sodium iodide (172 mg, 1.15 mmol) were dissolved in N,N-dimethylformamide (3 mL) and heated to 70 °C for 3 hours. After the reaction was completed, water (20 mL) was added to dilute the reaction, and the mixture was extracted with ethyl acetate (50 mL×3). The combined organic phases were washed with saturated brine (50 mL), and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System B) to give (S)-4-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)butanoic acid ethyl ester 7a (50 mg) with a yield of 28.6%.

MS m/z(ESI):503.1[M+1].

Step 2

(S)-4-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptan-2-yl)butanoic acid

(S)-4-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)butanoic acid

(S)-4-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)butanoic acid ethyl ester 7a (30 mg, 59.69 μmol) was dissolved in a mixed solvent of methanol (0.5 mL), tetrahydrofuran (0.5 mL) and water (0.5 mL), sodium hydroxide (2.4 mg, 59.7 μmol) was added, and the temperature was raised to 70°C for 3 hours. After the reaction was completed, an appropriate amount of dilute hydrochloric acid was added to adjust the pH to about 4, and the organic solvent was removed by concentration, and then subjected to preparative liquid separation (separation) The column was AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-4-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)butanoic acid 7 (10 mg), yield: 27.0%.

MS m/z(ESI):475.1[M+1].

Example 8

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)butanamide

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)butanamide

(S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)butanoic acid ethyl ester 2b (30 mg, 56.53 μmol) was dissolved in ethanol (1.5 mL), and aqueous ammonia (1 mL) was added dropwise. The mixture was reacted at room temperature overnight. After the reaction was completed, the mixture was concentrated. After preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+ _H2O , mobile phase B: _CH3CN ), (S)-4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)butanamide 8 (5 mg) was obtained. Yield: 13.65%.

MS m/z(ESI):502.1[M+1].

Example 9

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propanamide

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propanamide

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propanoic acid methyl ester 5b (60 mg, 119.38 μmol) was dissolved in ethanol (2 mL), and aqueous ammonia (1 mL) was added dropwise and reacted at room temperature overnight. After the reaction was completed, the reaction mixture was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)propanamide 9 (10 mg) with a yield of 7.18%.

MS m/z(ESI):488.1[M+1].

Example 10

(S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)acetic acid

(S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)acetic acid

first step

methyl

(S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)acetate

(S)-methyl 2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)acetate

(S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (0.1 g, 240.09 μmol), methyl 2-bromoacetate 10a (55.1 mg, 360.13 μmol, commercially available), potassium carbonate (332 The mixture was added with 4-(4-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)acetate (20 mg, 17.05%) to give (S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)acetate 10b (20 mg, 17.05% yield).

MS m/z(ESI):489.3[M+1].

Step 2

(S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)acetic acid

(S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)acetic acid

(S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)acetate 10b (15 mg, 30.70 μmol) was dissolved in a mixed solvent of methanol (0.5 mL), tetrahydrofuran (0.5 mL) and water (0.5 mL), sodium hydroxide (6.14 mg, 153.51 μmol) was added, and the reaction was carried out at 80 ° C for 8 hours. After the reaction was completed, 1 M dilute hydrochloric acid (3 mL) and water (50 ml) were added, and the mixture was extracted with ethyl acetate (50 mL×3). The organic phases were combined and concentrated under reduced pressure. Preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+ _H2O , mobile phase B: _CH3CN ) was performed to obtain (S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)acetic acid 10 (2.90 mg) with a yield of 18.91%.

MS m/z(ESI):475.0[M+1].

Embodiment 11

(S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)ethan-1-ol

(S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)ethan-1-ol

2-Bromoethanol 11a (30 mg, 240.07 μmol, commercially available), (S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (100 mg, 240.07 μmol), sodium iodide (216 mg, 1.44 mmol), and cesium carbonate (78.2 mg, 240.07 μmol) were added to N,N-dimethylformamide (2 mL) respectively. After reacting at 80 ° C for 6 hours, the reaction was basically completed. Water (50 mL) was added to quench, extracted three times with ethyl acetate (50 mL×3), and the organic phases were combined. The reaction mixture was washed with saturated brine (50 mL), concentrated under reduced pressure, and chromatographed on a silica gel column (eluent: System B) to give (S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)ethan-1-ol 11 (10 mg) in a yield of 6.89%.

MS m/z(ESI):461.0[M+1].

Example 12

(S)-2-hydroxy-1-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)ethan-1-one

(S)-2-Hydroxy-1-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)ethan-1-one

2-Hydroxyacetic acid 12a (10 mg, 131.49 μmol, commercially available), (S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (54.8 mg, 131.49 μmol), N,N-diisopropylethylamine (51.0 mg, 394.47 μmol), and O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (100 mg, 262.98 μmol) were added to N,N-dimethylformamide (1 mL) in sequence and reacted at room temperature for 16 hours. After the reaction was substantially completed, water (50 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (50 mL×3) three times. The organic phases were combined, washed with saturated brine (50 mL), and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-2-hydroxy-1-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)ethan-1-one 12 (10 mg) in a yield of 12.28%.

MS m/z(ESI):475.1[M+1].

Example 13

(S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)acetamide

(S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)acetamide

2-Bromoacetamide 13a (10 mg, 72.48 μmol, commercially available), (S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (30.2 mg, 72.48 μmol), potassium carbonate (30.1 mg, 217.45 μmol) and sodium iodide (10.9 mg, 72.48 μmol) were added to N,N-dimethylformamide (1 mL) in sequence and reacted at 80°C for 4 hours. After the reaction was substantially completed, water (50 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (50 mL×3) three times. The organic phases were combined, washed with saturated brine (50 mL), and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)acetamide 13 (5 mg) in a yield of 13.84%.

MS m/z(ESI):474.0[M+1].

Embodiment 14

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-3-oxopropanoic acid

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-3-oxopropanoic acid

first step

methyl

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-3-oxopropanoate

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-3-oxopropanoic acid methyl ester

(S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (100 mg, 240.09 μmol), methyl 3-chloro-3-oxopropanoate 14a (32.8 mg, 240.09 μmol, commercially available), and triethylamine (72.9 mg, 720.26 μmol) were added to N,N-dimethylformamide (3 mL) and reacted at room temperature for 8 hours. After the reaction was substantially completed, the product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-methyl 3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)-3-oxopropanoate 14b (40 mg) in a yield of 32.25%.

MS m/z(ESI):517.0[M+1].

Step 2

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-3-oxopropanoic acid

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-3-oxopropanoic acid

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)-3-oxopropanoic acid methyl ester 14b (5 mg, 9.68 μmol) was dissolved in a mixed solvent of methanol (0.25 mL), tetrahydrofuran (0.25 mL) and water (0.25 mL), sodium hydroxide (1.16 mg, 29.04 μmol) was added, and the reaction was carried out at 80 ° C for 4 hours. After the reaction was completed, dilute hydrochloric acid (3 mL) and water (50 mL) were added, and the mixture was extracted with ethyl acetate (50 mL×3). The organic phases were combined, washed with saturated brine (50 mL), and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)-3-oxopropanoic acid 14 (2.70 mg) in a yield of 52.73%.

MS m/z(ESI):503.0[M+1].

Embodiment 15

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-3-oxopropanamide

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-3-oxopropanamide

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)-3-oxopropanoic acid methyl ester 14b (20 mg, 38.72 μmol) was dissolved in methanol (1 mL), followed by dropwise addition of aqueous ammonia (0.8 mL) and reaction at room temperature for 36 hours. After the reaction was basically completed, the product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)-3-oxopropanamide 15 (2.66 mg) with a yield of 10.60%.

MS m/z(ESI):502.0[M+1].

Example 16

(S)-2-amino-1-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)ethan-1-one

(S)-2-Amino-1-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)ethan-1-one

first step

tert-butyl

(S)-(2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-2-oxoethyl)carbamate

(S)-tert-Butyl (2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-2-oxoethyl)carbamate

2-(tert-Butyloxycarbonylamino)acetic acid 16a (31.54 mg, 180.07 μmol, commercially available), N,N-diisopropylethylamine (62.06 mg, 480.18 μmol), and O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (68.47 mg, 180.07 μmol) were dissolved in N,N-dimethylformamide (2 mL). After stirring for 5 min, (S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (50.00 mg, 120.04 μmol) was added and the reaction was allowed to react at room temperature overnight. After the reaction was completed, water (50 mL) was added and extracted with ethyl acetate (50 mL×3). The organic phases were combined and washed with saturated sodium chloride (20 mL). The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated to obtain a crude product, which was then separated by preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-tert-butyl(2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)-2-oxoethyl)carbamate 16b (20 mg) in a yield of 29.04%.

MS m/z(ESI):574.3[M+1].

Step 2

(S)-2-amino-1-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)ethan-1-one

(S)-2-Amino-1-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)ethan-1-one

(S)-tert-Butyl (2-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)-2-oxoethyl)carbamate 16b (20 mg, 34.86 μmol) was dissolved in dichloromethane (2 mL), trifluoroacetic acid (0.3 ml, 2.63 mmol) was added dropwise, and the reaction was carried out at room temperature for 1.5 hours. After the reaction was completed, the reaction mixture was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-2-amino-1-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)ethan-1-one 16 (3 mg) with a yield of 3.91%.

MS m/z(ESI):474.0[M+1].

Embodiment 17

(S)-2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonane-7-carbaldehyde

(S)-2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonane-7-carbaldehyde

(S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (30 mg, 56.55 μmol) was dissolved in formamide (2 mL), triethylamine (28.61 mg, 282.73 μmol) was added dropwise, and the mixture was reacted at room temperature for 6 hours. After the reaction was completed, the reaction mixture was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonane-7-carbaldehyde 17 (3 mg) in a yield of 9.02%.

MS m/z(ESI):445.2[M+1].

Embodiment 18

(S)-2-amino-3-hydroxy-1-(2-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propan-1-one

(S)-2-Amino-3-hydroxy-1-(2-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propan-1-one

first step

tert-butyl

((S)-3-hydroxy-1-(2-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phe nyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-1-oxopropan-2-yl)carbamate

tert-Butyl ((S)-3-hydroxy-1-(2-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-1-oxopropan-2-yl)carbamate

(tert-Butyloxycarbonyl)-L-serine 18a (29.01 mg, 141.37 μmol, commercially available), N,N-diisopropylethylamine (60.90 mg, 471.22 μmol), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (53.75 mg, 141.37 μmol) were dissolved in N,N-dimethylformamide (2 mL). After stirring for 5 minutes, (S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (50 mg, 94.24 μmol) was added and reacted at room temperature overnight. After the reaction was completed, water (50 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate three times (50 ml×3). The organic phases were combined, washed with saturated aqueous sodium chloride solution (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System B) to give tert-butyl ((S)-3-hydroxy-1-(2-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)-1-oxopropan-2-yl)carbamate 18b (20 mg), yield: 35.15%.

MS m/z(ESI):604.3[M+1].

Step 2

(S)-2-amino-3-hydroxy-1-(2-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propan-1-one

(S)-2-Amino-3-hydroxy-1-(2-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)propan-1-one

Tert-butyl ((S)-3-hydroxy-1-(2-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)-1-oxopropan-2-yl)carbamate 18b (20 mg, 33.13 μmol) was dissolved in dichloromethane (3 mL), and trifluoroacetic acid (0.5 g, 4.39 mmol) was added dropwise, and the reaction was carried out at room temperature for 3 hours. After the reaction was completed, the reaction mixture was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-2-amino-3-hydroxy-1-(2-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)propan-1-one 18 (5 mg) with a yield of 24.44%.

MS m/z(ESI):504.0[M+1].

Embodiment 19

(S)-2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonane-7-carboxamide

(S)-2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonane-7-carboxamide

(S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (50 mg, 120.04 μmol), tetrahydrofuran (2 mL), and triethylamine (36.44 mg, 360.13 μmol) were added to the reaction bottle in sequence, and finally trimethylsilyl isocyanate 19a (17.98 mg, 156.06 μmol, commercially available) was added and the reaction was allowed to react at room temperature overnight. After the reaction was completed, the reaction mixture was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonane-7-carboxamide 19 (10 mg) with a yield of 13.80%.

MS(ESI):460.2[M+1].

Embodiment 20

(S)-2-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptan-2-yl)acetamide

(S)-2-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)acetamide

2-Bromoacetamide 13a (30 mg, 217.45 μmol, commercially available), (S)-1-(1-((5-(4-((2-azaspiro[3.3]hept-6-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 6 (84.47 mg, 217.45 μmol), potassium carbonate (90.16 mg, 652.35 μmol) and sodium iodide (32.62 mg, 217.45 μmol) were added to N,N-dimethylformamide (1.5 mL) in sequence and reacted at 80°C for 4 hours. After the reaction was substantially completed, water (50 mL) was added to quench the reaction, and the mixture was extracted three times with ethyl acetate (50 mL×3). The organic phases were combined and concentrated, and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-2-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)acetamide 20 (3 mg) in a yield of 2.34%.

MS m/z(ESI):446.1[M+1].

Embodiment 21

(S)-6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptane-2-carboxamide

(S)-6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptane-2-carboxamide

(S)-1-(1-((5-(4-((2-azaspiro[3.3]hept-6-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 6 (35 mg, 90.10 μmol), tetrahydrofuran (2 mL), and triethylamine (27.35 mg, 270.30 μmol) were added to the reaction bottle in sequence, and trimethylsilyl isocyanate 19a (13.49 mg, 117.13 μmol, commercially available) was added and the reaction was allowed to react at room temperature overnight. After the reaction was completed, the reaction mixture was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptane-2-carboxamide 21 (9 mg) with a yield of 17.40%.

MS m/z(ESI):432.0[M+1].

Embodiment 22

(S)-6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptane-2-carbaldehyde

(S)-6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptane-2-carbaldehyde

(S)-1-(1-((5-(4-((2-azaspiro[3.3]hept-6-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl) Ethanol 6 (35 mg, 90.10 μmol), formamide (2 mL), and triethylamine (27.35 mg, 270.30 μmol) were added to a single-mouth bottle in sequence and reacted at room temperature overnight. After the reaction was completed, the mixture was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptane-2-carbaldehyde 22 (5 mg) with a yield of 9.94%.

MS m/z(ESI):417.2[M+1].

Embodiment 23

(S)-2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octane-6-carbaldehyde

(S)-2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octane-6-carbaldehyde

(S)-1-(1-((5-(4-((6-Azaspiro[3.4]octan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 3 (10 mg, 24.85 μmol) was dissolved in formamide (2 mL), triethylamine (25.14 mg, 248.45 μmol) was added and the reaction was allowed to react at room temperature overnight. After the reaction was completed, the reaction mixture was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octane-6-carbaldehyde 23 (2.40 mg) with a yield of 15.08%.

MS m/z(ESI):431.3[M+1].

Embodiment 24

(S)-2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octane-6-carboxamide

(S)-2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octane-6-carboxamide

(S)-1-(1-((5-(4-((6-azaspiro[3.4]octan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 3 (36.26 mg, 90.10 μmol) was dissolved in tetrahydrofuran (2 mL), and triethylamine (27.35 mg, 270.30 μmol) and trimethylsilyl isocyanate 19a (13.49 mg, 117.13 μmol) were added and reacted at room temperature for 4 hours. After the reaction was completed, the mixture was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octane-6-carboxamide 24 (3 mg), yield: 5.36%. MS m/z (ESI): 446.0 [M+1].

Embodiment 25

(S)-1-(1-((5-(4-((7-((1H-pyrazol-5-yl)methyl)-7-azaspiro[3.5]nonan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((7-((1H-pyrazol-5-yl)methyl)-7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

first step

tert-butyl

(S)-5-((2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)methyl)-1H-pyrazole-1-carboxylate

(S)-tert-Butyl 5-((2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)methyl)-1H-pyrazole-1-carboxylate

(S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl) Ethanol 1 (50 mg, 120.04 μmol), tert-butyl 3-(bromomethyl)pyrazole-1-carboxylate 25a (31.35 mg, 120.04 μmol, commercially available), and potassium carbonate (19.91 mg, 144.05 μmol) were added to N,N-dimethylformamide (1.5 mL) in sequence and reacted at 75 °C for 8 hours. After the reaction was completed, water (50 mL) was added, and ethyl acetate (50 mL×3) was added for extraction three times. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to obtain a crude product, which was purified by silica gel column chromatography (eluent: B system) to obtain the product (S)-5-((2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)methyl)-1H-pyrazole-1-carboxylic acid tert-butyl ester 25b (40 mg), yield: 55.84%.

MS m/z(ESI):497.3[M-100+1].

Step 2

(S)-1-(1-((5-(4-((7-((1H-pyrazol-5-yl)methyl)-7-azaspiro[3.5]nonan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((7-((1H-pyrazol-5-yl)methyl)-7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-tert-Butyl 5-((2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)methyl)-1H-pyrazole-1-carboxylate 25b (40 mg, 67.03 μmol) was dissolved in dichloromethane (1 mL), and trifluoroacetic acid (67.03 μmol, 0.25 mL) was slowly added. The reaction was completed after 3 hours at room temperature. The residue was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-1-(1-((5-(4-((7-((1H-pyrazol-5-yl)methyl)-7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 25 (10 mg), yield: 23.21%.

MS m/z(ESI):497.3[M+1].

Embodiment 26

(S)-2-hydroxy-1-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptan-2-yl)ethan-1-one

(S)-2-Hydroxy-1-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)ethan-1-one

1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (25.91 mg, 135.15 μmol), 1-hydroxybenzotriazole (18.26 mg, 135.15 μmol), N-methylmorpholine (20.05 mg, 198.22 μmol), 2-hydroxyacetic acid 12a (10.28 mg, 135.15 μmol) was dissolved in dichloromethane (2 mL). After reacting for 5 minutes, (S)-1-(1-((5-(4-((2-azaspiro[3.3]hept-6-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 6 (35 mg, 90.10 μmol) was added and reacted at room temperature overnight. After the reaction was completed, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction three times. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to obtain a crude product. The crude product was separated by preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-2-hydroxy-1-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)ethan-1-one 26 (2.80 mg) with a yield of 5.27%.

MS m/z(ESI):447.2[M+1].

Embodiment 27

(S)-1-(1-((5-(4-((7-((1H-imidazol-2-yl)methyl)-7-azaspiro[3.5]nonan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((7-((1H-imidazol-2-yl)methyl)-7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (50 mg, 120.04 μmol), 2-(bromomethyl)-1H-imidazole 27a (26.14 mg, 108.04 μmol), sodium iodide (1.99 mg, 12.00 μmol) and potassium carbonate (36.50 mg, 264.10 μmol) were added to N,N-dimethylformamide (1.5 mL) in sequence and heated to 65 °C for 8 hours. After the reaction was basically completed, the product was filtered and the filtrate was separated by preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-1-(1-((5-(4-((7-((1H-imidazol-2-yl)methyl)-7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 27 (10 mg) in a yield of 12.96%.

MS(ESI):497.1[M+1].

Embodiment 28

(S)-2-hydroxy-1-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octan-6-yl)ethan-1-one

(S)-2-Hydroxy-1-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octan-6-yl)ethan-1-one

2-Hydroxyacetic acid 12a (14.17 mg, 186.34 μmol), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (35.72 mg, 186.34 μmol), 1-hydroxybenzotriazole (25.18 mg, 186.34 μmol), and N-methylmorpholine (27.64 mg, 273.30 μmol) were added to a dichloromethane (2 mL) solution. After reacting for 5 minutes, (S)-1-(1-((5-(4-((6-azaspiro[3.4]octan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazole-2-yl)ethan-1-ol 3 (50 mg, 124.23 μmol) was added and reacted at room temperature for 16 hours. After the reaction was completed, water (50 mL) was added, and the mixture was extracted with dichloromethane (50 mL×3) three times. The organic layers were combined and concentrated, and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+ _H2O , mobile phase B: _CH3CN ) to obtain (S)-2-hydroxy-1-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-6-azaspiro[3.4]octan-6-yl)ethan-1-one 28 (5 mg) with a yield of 6.30%.

MS m/z(ESI):461.2[M+1].

Embodiment 29

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-3-oxopropanenitrile

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)-3-oxopropionitrile

2-Cyanoacetic acid 29a (15.32 mg, 180.07 μmol, commercially available), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (34.52 mg, 180.07 μmol), 1-hydroxybenzotriazole (24.33 mg, 180.07 μmol), and N-methylmorpholine (26.71 mg, 264.10 μmol) were added to N,N-dimethylformamide (1.5 mL) solution, and after 5 minutes of reaction, (S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazole-2-yl)ethan-1-ol 1 (50 mg, 120.04 μmol) was added and reacted at room temperature for 16 hours. After the reaction was completed, water (50 mL) and dichloromethane (50 mL×3) were added. After extraction three times, the organic phases were combined, concentrated, and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)-3-oxopropionitrile 29 (3 mg), yield: 3.97%.

MS m/z(ESI):484.2[M+1].

Embodiment 30

(S)-1-(1-((5-(4-((7-(azetidin-3-yl)-7-azaspiro[3.5]nonan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((7-(azetidin-3-yl)-7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

first step

tert-butyl

(S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)azetidine-1-carboxylate

(S)-tert-butyl 3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]nonan-7-yl)azetidine-1-carboxylate

At room temperature, (S)-1-(1-((5-(4-((7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 1 (26.81 mg, 64.36 μmol) and tert-butyl 3-oxoazetidine-1-carboxylate 30a (16.53 mg, 96.53 μmol, commercially available) were dissolved in a mixed solution of 1,2-dichloroethane (0.5 mL) and methanol (0.5 mL), and acetic acid (20 mg, 64.36 μmol) was added dropwise. After reacting at 50 °C for 6 hours, sodium cyanoborohydride (12.13 mg, 193.07 μmol) was slowly added at room temperature and reacted at room temperature for 16 hours. After the reaction was completed, water (1 mL) was added to quench the reaction, and the mixture was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)azetidine-1-carboxylic acid tert-butyl ester 30b (25 mg), yield: 67.95%. MS m/z (ESI): 572.4 [M+1].

Step 2

(S)-1-(1-((5-(4-((7-(azetidin-3-yl)-7-azaspiro[3.5]nonan-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((7-(azetidin-3-yl)-7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

At room temperature, (S)-tert-butyl 3-(2-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-7-azaspiro[3.5]non-7-yl)azetidine-1-carboxylate 30b (25 mg, 43.73 μmol) was dissolved in dichloromethane (2 mL), and trifluoroacetic acid (0.5 g, 4.39 mmol) was added dropwise, and the reaction was carried out at room temperature for 4 hours. After the reaction was completed, the mixture was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-1-(1-((5-(4-((7-(azetidin-3-yl)-7-azaspiro[3.5]non-2-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 30 (9 mg), yield: 31.63%. MS m/z (ESI): 472.3 [M+1].

Embodiment 31

(S)-2-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptan-2-yl)ethan-1-ol

(S)-2-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)ethan-1-ol

At room temperature, (S)-1-(1-((5-(4-((2-azaspiro[3.3]hept-6-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 6 (25 mg, 49.75 μmol), 2-bromoethanol 11a (6.84 mg, 54.73 μmol, commercially available) and potassium carbonate (27.51 mg, 199.01 μmol) were added to acetonitrile (0.5 mL) and reacted at 60 °C for 12 hours. After the reaction was completed, the reaction was filtered and the filtrate was concentrated. The product was separated by preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-2-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)ethan-1-ol (8 mg) in a yield of 26.48%.

MS m/z(ESI):433.0[M+1].

Embodiment 32

(S)-3-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]heptan-2-yl)-3-oxopropanenitrile

(S)-3-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)-3-oxopropionitrile

At room temperature, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (57.23 mg, 298.52 μmol), 1-hydroxybenzotriazole (40.34 mg, 298.52 μmol), N-methylmorpholine (84.55 mg, 835.85 μmol), and 2-cyanoacetic acid 32a (30.47 mg, 358.22 μmol, commercially available) were dissolved in dichloromethane (2 mL). After reacting for 5 minutes, (S)-1-(1-((5-(4-((2-azaspiro[3.3]hept-6-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 6 (60 mg, 119.41 μmol) was added and the reaction was allowed to proceed overnight at room temperature. After the reaction was completed, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to obtain a crude product. The crude product was subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-3-(6-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-2-azaspiro[3.3]hept-2-yl)-3-oxopropionitrile 32 (20 mg) with a yield of 27.94%.

MS m/z(ESI):456.2[M+1].

Embodiment 33

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-4-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-4-yl)methanone

first step

tert-butyl 4-((trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxylate

tert-Butyl 4-((Trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxylate

Tert-butyl 4-(((trifluoromethyl)sulfonyl)oxy)-3,6-dihydropyridine-1(2H)-carboxylate 33a (1 g, 3.02 mmol, commercially available), ethynyltrimethylsilane 33b (444.68 mg, 4.53 mmol, 639.83 μL, commercially available), bis(triphenylphosphine)palladium(II) dichloride (352.28 mg, 452.75 μmol), triethylamine (305.43 mg, 3.02 mmol), cuprous iodide (57.48 mg, 301 .83μmol) were successively added to a toluene (3mL) solution, and the mixture was stirred for 16 hours at room temperature. The reaction solution was distilled under reduced pressure to remove the organic solvent, water (20mL) was added, and the mixture was extracted with ethyl acetate (20mL×3). The organic phases were combined, dried over anhydrous sodium sulfate, and distilled under reduced pressure. The residue was purified by silica gel column chromatography (eluent: system A) to obtain tert-butyl 4-((trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxylate 33c (820mg) in a yield of 97.22%.

MS m/z(ESI):224.1[M+1-56].

Step 2

tert-butyl 4-ethynyl-3,6-dihydropyridine-1(2H)-carboxylate

tert-Butyl 4-ethynyl-3,6-dihydropyridine-1(2H)-carboxylate

4-((Trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxylic acid tert-butyl ester 33c (800 mg, 2.86 mmol) was added to methanol (2 mL), followed by potassium fluoride (332.65 mg, 5.73 mmol), and the mixture was stirred for 12 hours at room temperature. The reaction solution was distilled under reduced pressure to remove the organic solvent, water (20 mL) was added, and ethyl acetate was extracted (20 mL×3). The organic phases were combined, dried over anhydrous sodium sulfate, and distilled under reduced pressure. The residue was purified by silica gel column chromatography (eluent: A system) to obtain 4-ethynyl-3,6-dihydropyridine-1(2H)-carboxylic acid tert-butyl ester 33d (370 mg), with a yield of 62.36%.

MS m/z(ESI):152.1[M+1-56].

third step

tert-butyl

4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxylate

tert-Butyl 4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxylate

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (800 mg, 1.67 mmol), tert-butyl 4-ethynyl-3,6-dihydropyridine-1(2H)-carboxylate 33d (363.24 mg, 1.75 mmol), allylpalladium(II) chloride dimer (121.84 mg, 333.81 μmol), triethylenediamine (374.44 mg, 3.34 mmol), tri-tert-butylphosphine (10% in toluene solution) (67.54 mg, 333.81 μmol) were added to acetonitrile (2 mL) in sequence. The reaction was carried out at room temperature overnight. After the reaction was completed, water (30 mL) was added to quench the reaction, and ethyl acetate was used for extraction (100 mL×3). The combined organic phase was washed with saturated brine (100 mL), and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: B system) to obtain 4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxylic acid tert-butyl ester 33e (660 mg), yield: 70.78%. MS m/z(ESI):559.2[M+1].

the fourth step

(S)-1-(1-((5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxylic acid tert-butyl ester 33e (660 mg, 1.18 mmol) was added to dichloromethane (1.5 mL), followed by dropwise addition of trifluoroacetic acid (0.5 mL), and stirring was continued at room temperature for 1 hour. After the reaction was completed, saturated _NaHCO3 solution was added under an ice bath until no bubbles were generated (pH>7), and the mixture was extracted with ethyl acetate (20 mL×3). The organic phases were combined, dried over anhydrous sodium sulfate, and distilled under reduced pressure. The residue was purified by silica gel column chromatography (eluent: B system) to give (S)-1-(1-((5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazole-2-yl)ethan-1-ol 33f (320 mg) in a yield of 72.34%.

MS m/z(ESI):375.2[M+1].

the fifth step

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-4-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-4-yl)methanone

Isonicotinic acid 33g (32.88mg, 267.07μmol, commercially available), (S)-1-(1-((5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 33f (50mg, 133.53μmol), N,N-diisopropylethylamine (34.52mg, 267.07μmol), (7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (50.77mg, 133.53μmol) were added to N,N-dimethylformamide (1mL) in sequence and stirred at room temperature for 16 hours. After the reaction was completed, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to obtain a crude product. The crude product was subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-4-yl)methanone 33 (13.2 mg) with a yield of 15.99%.

MS m/z(ESI):480.3[M+1].

Embodiment 34

(S)-4-((4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3, 6-dihydropyridin-1(2H)-yl)methyl)benzoic acid

(S)-4-((4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methyl)benzoic acid

4-Formylbenzoic acid 34a (24.06 mg, 160.24 μmol, commercially available), (S)-1-(1-((5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 33f (50 mg, 133.53 μmol), and acetic acid (8.02 mg, 133.53 μmol) were added to dichloromethane (1 mL) in sequence and stirred at room temperature for 1 hour. Then, sodium triacetoxyborohydride (28.30 mg, 133.53 μmol) was added and stirred at room temperature for 16 hours. After the reaction was completed, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to obtain a crude product. The crude product was subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-4-((4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methyl)benzoic acid 34 (7.35 mg) with a yield of 8.66%.

MS m/z(ESI):509.0[M+1].

Embodiment 35

((1R,5S)-bicyclo[3.1.0]hexan-6-yl)(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

((1R,5S)-Bicyclo[3.1.0]hexan-6-yl)(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

first step

3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((trimethylsilyl)ethynyl)phenyl)isoxazole

3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((trimethylsilyl)ethynyl)phenyl)isoxazole

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (8 g, 16.69 mmol), ethynyltrimethylsilane 33b (3.28 g, 33.38 mmol, 4.72 mL, commercially available), allylpalladium(II) chloride dimer (609.21 mg, 1.67 mmol) and triethylenediamine (3.74 g, 33.38 mmol) and tri-tert-butylphosphine (10% toluene solution) (3.38 g, 1.67 mmol) were added to acetonitrile (20 mL) in sequence, argon was replaced 3 times, and stirring was continued at room temperature for 12 hours. The system was concentrated to dryness under reduced pressure, water (50 mL) was added, and extraction was carried out with ethyl acetate (50 mL×3). The combined organic phase was dried over anhydrous sodium sulfate and then distilled under reduced pressure. The residue was purified by silica gel column chromatography (eluent: System A) to give 3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((trimethylsilyl)ethynyl)phenyl)isoxazole 35a (7.5 g), yield: 99.94%.

MS m/z(ESI):450.3[M+1].

Step 2

5-(4-ethynylphenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole

5-(4-ethynylphenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole

3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((trimethylsilyl)ethynyl)phenyl)isoxazole 35a (7.5 g, 16.68 mmol) and potassium fluoride (1.94 g, 33.36 mmol) were added to methanol in sequence. The product was added into 4-nitropropene (50 mL) and stirred at room temperature for 5 hours until the reaction was complete. Water (50 mL) was added and extracted with ethyl acetate (50 mL×3). The combined organic phase was dried over anhydrous sodium sulfate and then distilled under reduced pressure. The residue was purified by silica gel column chromatography (eluent: system A) to give 5-(4-ethynylphenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 35b (5.8 g) in a yield of 92.21%.

MS m/z(ESI):378.2[M+1].

third step

1,2,3,6-tetrahydropyridin-4-yl trifluoromethanesulfonate

1,2,3,6-Tetrahydropyridin-4-yl trifluoromethanesulfonate

4-(((trifluoromethyl)sulfonyl)oxy)-3,6-dihydropyridine-1(2H)-carboxylic acid tert-butyl ester 33a (1.7 g, 5.13 mmol) was added to dichloromethane (1.5 mL), followed by dropwise addition of trifluoroacetic acid (0.5 mL). The mixture was stirred for 1 hour at room temperature. After the reaction was complete, saturated NaHCO ₃ solution was added to the reaction solution until no bubbles were generated and pH>7. Water (50 mL) was then added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to give 1,2,3,6-tetrahydropyridin-4-yl trifluoromethanesulfonate 35c (1.1 g) with a yield of 92%. The crude product was directly used in the next step.

MS m/z(ESI):232.1[M+1].

the fourth step

1-(2,2,2-trifluoroacetyl)-1,2,3,6-tetrahydropyridin-4-yl trifluoromethanesulfonate

1-(2,2,2-Trifluoroacetyl)-1,2,3,6-tetrahydropyridin-4-yl trifluoromethanesulfonate

1,2,3,6-Tetrahydropyridin-4-yl trifluoromethanesulfonate 35c (900 mg, 3.89 mmol) and triethylamine (393.92 mg, 3.89 mmol) were added to dichloromethane (10 mL) in sequence under an ice bath, and then trifluoroacetic anhydride (820 mg, 3.89 mmol) was added dropwise to the mixed solution, and then the temperature was gradually raised to room temperature and stirred for 20 minutes. After the reaction raw materials were detected to be completely reacted, water (50 mL) was added, and ethyl acetate was extracted (50 mL×3). The combined organic phase was dried over anhydrous sodium sulfate and then distilled under reduced pressure. The residue was purified by silica gel column chromatography (eluent: A system) to obtain 1-(2,2,2-trifluoroacetyl)-1,2,3,6-tetrahydropyridin-4-yl trifluoromethanesulfonate 35d (1.17 g) with a yield of 91%.

MS m/z(ESI):328.0[M+1].

the fifth step

2,2,2-trifluoro-1-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

2,2,2-Trifluoro-1-(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

Under argon atmosphere, 1-(2,2,2-trifluoroacetyl)-1,2,3,6-tetrahydropyridin-4-yl trifluoromethanesulfonate 35d (1.13 g, 3.44 mmol), 5-(4-ethynylphenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 35b (1 g, 2.65 mmol), cuprous iodide (50.46 mg, 264.95 μmol), bis(triphenylphosphine)palladium(II) dichloride (412.30 mg, 529.89 μmol) and triethylamine (268.10 mg, 2.65 mmol) were added to toluene (5 mL) in sequence, and stirring was continued at room temperature for 16 hours. After the reaction was completed, the reaction solution was distilled under reduced pressure to remove the organic solvent, and the residue was purified by silica gel column chromatography (eluent: A system) to obtain 2,2,2-trifluoro-1-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2- (400 mg), yield: 27.22%.

MS m/z(ESI):555.3[M+1].

Step 6

3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole

3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole

2,2,2-Trifluoro-1-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 35e (400 mg, 721.29 μmol) and potassium carbonate (797.52 mg, 5.77 mmol) were added sequentially to a mixture of methanol (10 mL) and water (0.5 mL). After argon was replaced three times in the solution, the reaction was stirred at 70° C. for 2 hours. After the reaction was completed, the reaction solution was distilled under reduced pressure to remove the organic solvent. The crude product was purified by silica gel column chromatography (eluent: B system) to obtain 3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazole-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole 35f (180 mg) in a yield of 54.42%.

MS m/z(ESI):459.3[M+1].

Step 7

bicyclo[3.1.0]hexan-6-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Bicyclo[3.1.0]hexane-6-carboxylic acid 35g (16.51 mg, 130.85 μmol, commercially available), 3-((2-((1S)-1-((tetrahydro 2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole 35f (30 mg, 65.42 μmol), N-methylmorpholine (19.85 mg, 196.27 μmol), 1-hydroxybenzotriazole (17.68 mg, 130.85 μmol), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (25.08 mg, 130.85 μmol) were added to dichloromethane (1 mL) in sequence, and stirring was continued at room temperature for 16 hours. After the reaction was complete, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to give bicyclo[3.1.0]hexan-6-yl(4-((4-(3-((2-((1S)-1-((tetrahydro 2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 35h (37 mg), and the crude product was directly used in the next reaction.

MS m/z(ESI):567.2[M+1].

Step 8

((1R,5S)-bicyclo[3.1.0]hexan-6-yl)(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

((1R,5S)-Bicyclo[3.1.0]hexan-6-yl)(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Bicyclo[3.1.0]hex-6-yl(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 35h (37 mg, 65.29 μmol) was added to dichloromethane (1.5 mL), followed by dropwise addition of trifluoroacetic acid (0.5 mL), and stirring was continued for half an hour at room temperature. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give ((1R,5S)-bicyclo[3.1.0]hexan-6-yl)(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 35 (4.05 mg) in a yield of 9.88%.

MS m/z(ESI):483.2[M+1].

Embodiment 36

(S)-cyclopropyl(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(S)-Cyclopropyl(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

first step

cyclopropyl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Cyclopropyl(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Cyclopropanecarboxylic acid 36a (11.26 mg, 130.85 μmol), 3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole 35f (30 mg, 65.42 μmol), N-methylmorpholine (19.85 mg, 196.27 μmol), 1-hydroxybenzotriazole (17.68 mg, 130.85 μmol), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (25.08 mg, 130.85 μmol) were added to dichloromethane (1 mL) in sequence, and stirring was continued at room temperature for 6 hours. After the reaction was complete, water (50 mL) was added, and dichloromethane (50 mL × 3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to obtain cyclopropyl (4-((4-(3-((2-((1S)-1-((tetrahydro 2H-pyran (2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 36b (34 mg), yield: 98.68%. The crude product was directly used for the next step reaction.

MS m/z(ESI):527.3[M+1].

Step 2

(S)-cyclopropyl(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(S)-Cyclopropyl(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Cyclopropyl(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 36b (34 mg, 64.56 μmol) was added to dichloromethane (1.5 mL), followed by dropwise addition of trifluoroacetic acid (0.5 mL), and stirring was continued for half an hour at room temperature. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-cyclopropyl(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 36 (25.29 mg) in a yield of 69.68%.

MS m/z(ESI):443.2[M+1].

Embodiment 37

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-(1H-1,2,4-triazol-1-yl)ethan-1-one

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-(1H-1,2,4-triazol-1-yl)ethan-1-one

first step

1-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-(1H-1,2,4-triazol-1-yl)ethan-1-one

1-(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-(1H-1,2,4-triazol-1-yl)ethan-1-one

2-(1H-1,2,4-triazol-1-yl)acetic acid 37a (16.63 mg, 130.85 μmol), 3-((2-((1S)-1-((tetrahydro 2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole 35f (30 mg, 65.42 μmol), N-methylmorpholine (19.85 mg, 196.27 μmol), 1-hydroxybenzotriazole (17.68 mg, 130.85 μmol), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (25 .08mg, 130.85μmol) were added to dichloromethane (1mL) in turn, and stirred for 16 hours at room temperature. After the reaction was complete, water (50mL) was added, and dichloromethane (50mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to give 1-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-(1H-1,2,4-triazol-1-yl)ethan-1-one 37b (37mg), yield: 99.63%. It was directly used for the next step reaction.

MS m/z(ESI):568.4[M+1].

Step 2

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-(1H-1,2,4-triazol-1-yl)ethan-1-one

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-(1H-1,2,4-triazol-1-yl)ethan-1-one

1-(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-(1H-1,2,4-triazol-1-yl)ethan-1-one 37b (37 mg, 65.18 μmol) was added to dichloromethane (1.5 mL), followed by dropwise addition of trifluoroacetic acid (0.5 mL), and stirring was continued for half an hour at room temperature. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-(1H-1,2,4-triazol-1-yl)ethan-1-one 37 (27.39 mg) in a yield of 69.62%.

MS m/z(ESI):484.2[M+1].

Embodiment 38

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyrazin-2-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyrazin-2-yl)methanone

first step

pyrazin-2-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Pyrazin-2-yl(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Pyrazine-2-carboxylic acid 38a (16.24 mg, 130.85 μmol), 3-((2-((1S)-1-((tetrahydro 2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole 35f (30 mg, 65.42 μmol), N-methylmorpholine (19.85 mg, 196.27 μmol), 1-hydroxybenzotriazole (17.68 mg, 130.85 μmol), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (25.08 mg, 130.85 μmol) were added to dichloromethane (1 mL) in sequence, and stirring was continued at room temperature for 6 hours. After the reaction was complete, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to give pyrazin-2-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 38b (36 mg) in a yield of 97.45%. The crude product was used directly in the next step.

MS m/z(ESI):565.3[M+1].

Step 2

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyrazin-2-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyrazin-2-yl)methanone

Pyrazin-2-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 38b (36 mg, 63.76 μmol) was added to dichloromethane (1.5 mL), followed by dropwise addition of trifluoroacetic acid (0.5 mL), and stirring was continued for half an hour at room temperature. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyrazin-2-yl)methanone 38 (14.76 mg) with a yield of 38.12%.

MS m/z(ESI):481.2[M+1].

Embodiment 39

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-d ihydropyridin-1(2H)-yl)(pyridazin-4-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridazin-4-yl)methanone

first step

pyridazin-4-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Pyridazin-4-yl(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Pyridazine-4-carboxylic acid 39a (16.24 mg, 130.85 μmol, commercially available), 3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole 35f (30 mg, 65.42 μmol), N-methylmorpholine (19.85 mg, 196.27 μmol), 1-hydroxybenzotriazole (17.68 mg, 130.85 μmol), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (25.08 mg, 130.85 μmol) were added to dichloromethane (1 mL) in sequence, and stirring was continued at room temperature for 6 hours. After the reaction was complete, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to give pyridazine-4-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 39b (36 mg) in a yield of 97.45%. The crude product was used directly in the next step.

MS m/z(ESI):565.3[M+1].

Step 2

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridazin-4-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridazin-4-yl)methanone

Pyridazin-4-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 39b (36 mg, 63.76 μmol) was added to dichloromethane The mixture was added to 1% paraffin (1.5 mL), and then trifluoroacetic acid (0.5 mL) was added dropwise. The mixture was stirred for half an hour at room temperature. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridazin-4-yl)methanone 39 (13.22 mg) with a yield of 33.83%.

MS m/z(ESI):481.2[M+1].

Embodiment 40

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyrimidin-5-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyrimidin-5-yl)methanone

first step

pyrimidin-5-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Pyrimidin-5-yl(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Pyrimidine-5-carboxylic acid 40a (21.65 mg, 174.46 μmol, commercially available), 3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole 35f (40 mg, 87.23 μmol), 1-hydroxybenzotriazole (23.57 mg, 174.46 μmol), N-methylmorpholine (26.47 mg, 261.69 μmol), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33.44 mg, 174.46 μmol) were added to dichloromethane (1.5 mL) in sequence, and stirring was continued at room temperature for 16 hours. After the reaction was complete, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to give pyrimidin-5-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 40b (49 mg) with a yield of 99.48%. It was directly used for the next step reaction.

MS m/z(ESI):565.3[M+1].

Step 2

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyrimidin-5-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyrimidin-5-yl)methanone

Pyrimidin-5-yl(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 40b (49 mg, 86.78 μmol) was added to dichloromethane (1.5 mL), followed by dropwise addition of trifluoroacetic acid (0.5 mL), and stirring was continued at room temperature for 1 hour. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyrimidin-5-yl)methanone 40 (22.3 mg) with a yield of 42.36%.

MS m/z(ESI):481.2[M+1]. ¹ H NMR(400MHz,DMSO-d ₆ )δ9.30(s,1H),8.94(s,2H),7.88(d,J＝8.2Hz,2H),7.79(d,J＝2.0Hz,1H),7.72(d,J＝2.1Hz,1H),7.62(d,J＝8.1Hz,2H),7.17(s,1H),6.34(s,1H),5.72(s,2H),5.26(q,J＝6.6Hz,1H),4.27(s,1H),4.13(s,2H),3.81(s,1H),2.41(s,2H),1.49(d,J＝6.6Hz,3H).

Embodiment 41

(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydrofuran-3-yl)methanone

(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydrofuran-3-yl)methanone

first step

(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydrofuran-3-yl)methanone

(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydrofuran-3-yl)methanone

Tetrahydrofuran-3-carboxylic acid 41a (20.26 mg, 174.46 μmol, commercially available), 3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole 35f (40 mg, 87.23 μmol), 1-hydroxybenzotriazole (23.57 mg, 174.46 μmol), N-methylmorpholine (26.47 mg, 261.69 μmol), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33.44 mg, 174.46 μmol) were added to dichloromethane (1.5 mL) in sequence, and stirring was continued at room temperature for 16 hours. After the reaction was completed, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to give (4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydrofuran-3-yl)methanone 41b (48 mg) with a yield of 98.85%. The crude product was directly used in the next step.

MS m/z(ESI):557.3[M+1].

Step 2

(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydrofuran-3-yl)methanone

(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydrofuran-3-yl)methanone

(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydrofuran-3-yl)methanone 41b (48 mg, 86.23 μmol) was added to dichloromethane (1.5 mL), followed by dropwise addition of trifluoroacetic acid (0.5 mL), and stirring was continued at room temperature for 1 hour. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydrofuran-3-yl)methanone 41 (20.51 mg) with a yield of 36.90%.

MS m/z(ESI):473.3[M+1].

Embodiment 42

(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(morpholin-3-yl)methanone

(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(morpholin-3-yl)methanone

first step

morpholin-3-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Morpholin-3-yl(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Morpholine-3-carboxylic acid 42a (22.88 mg, 174.46 μmol, commercially available), 3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole 35f (40 mg, 87.23 μmol), 1-hydroxybenzotriazole (23.57 mg, 174.46 μmol), N-methylmorpholine (26.47 mg, 261.69 μmol), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33.44 mg, 174.46 μmol) were added to dichloromethane (1.5 mL) in sequence, and stirring was continued for 16 hours at room temperature. After the reaction was complete, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to give morpholin-3-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 42b (45 mg) in a yield of 90.24%. The crude product was used directly in the next step.

MS m/z(ESI):572.3[M+1].

Step 2

(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(morpholin-3-yl)methanone

(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(morpholin-3-yl)methanone

Morpholin-3-yl(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 42b (45 mg, 78.72 μmol) was added to dichloromethane (1.5 mL), followed by dropwise addition of trifluoroacetic acid (0.5 mL), and stirring was continued at room temperature for 1 hour. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(morpholin-3-yl)methanone 42 (10.35 mg) with a yield of 20.33%.

MS m/z(ESI):488.3[M+1].

Embodiment 43

(S)-1-(1-((5-(4-((1-(2-(1H-1,2,4-triazol-1-yl)ethyl)-1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((1-(2-(1H-1,2,4-triazol-1-yl)ethyl)-1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

first step

5-(4-((1-(2-(1H-1,2,4-triazol-1-yl)ethyl)-1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole

5-(4-((1-(2-(1H-1,2,4-triazol-1-yl)ethyl)-1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)-3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole

Ethyl 2-(1H-1,2,4-triazol-1-yl)methanesulfonate 43a (25.02 mg, 130.85 μmol), 3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole 35f (30 mg, 65.42 μmol) and potassium carbonate (9.04 mg, 65.42 μmol) were added to acetonitrile (2 mL) in sequence and stirred at 60 °C for 16 hours. After the reaction was complete, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to give 5-(4-((1-(2-(1H-1,2,4-triazol-1-yl)ethyl)-1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)-3-((2-((1S)-1-((tetrahydro 2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 43b (35 mg), yield: 96.63%. The crude product was directly used for the next step reaction.

MS m/z(ESI):554.4[M+1].

Step 2

(S)-1-(1-((5-(4-((1-(2-(1H-1,2,4-triazol-1-yl)ethyl)-1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

(S)-1-(1-((5-(4-((1-(2-(1H-1,2,4-triazol-1-yl)ethyl)-1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol

5-(4-((1-(2-(1H-1,2,4-triazol-1-yl)ethyl)-1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)-3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 43b (35 mg, 63.22 μmol) was added to dichloromethane (1.5 mL), followed by dropwise addition of trifluoroacetic acid (0.5 mL), and stirring was continued for 1 hour at room temperature. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-1-(1-((5-(4-((1-(2-(1H-1,2,4-triazol-1-yl)ethyl)-1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazol-3-yl)methyl)-1H-imidazol-2-yl)ethan-1-ol 43 (10.69 mg), yield: 28.98%.

MS m/z(ESI):470.3[M+1].

Embodiment 44

(S)-3-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propanenitrile

(S)-3-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propionitrile

first step

3-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propanenitrile

3-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propionitrile

Acrylonitrile 44a (13.89 mg, 261.69 μmol, commercially available), 3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole 35f (60 mg, 130.85 μmol) and potassium carbonate (36.17 mg, 261.69 μmol) were added to acetonitrile (1 mL) and water (0.5 mL) in sequence and stirred continuously for 16 h at room temperature. After the reaction was complete, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to give 3-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propionitrile 44b (66 mg), yield: 98.59%. The crude product was directly used for the next step reaction.

MS m/z(ESI):512.3[M+1].

Step 2

(S)-3-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propanenitrile

(S)-3-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propionitrile

3-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propionitrile 44b (66 mg, 129.00 μmol) was added to dichloromethane (1.5 mL), followed by dropwise addition of trifluoroacetic acid (0.5 mL), and stirring was continued at room temperature for 1 hour. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-3-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propionitrile 44 (32.38 mg), yield: 46.35%. MS m/z (ESI): 428.2 [M+1].

Embodiment 45

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-3-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-3-yl)methanone

first step

pyridin-3-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Pyridin-3-yl(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Nicotinic acid 45a (21.48 mg, 174.46 μmol, commercially available), 3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)-5-(4-((1,2,3,6-tetrahydropyridin-4-yl)ethynyl)phenyl)isoxazole 35f (40 mg, 87.23 μmol), 1-hydroxybenzotriazole (23.57 mg, 174.46 μmol), N-methylmorpholine (26.47 mg, 261.69 μmol), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33.44 mg, 174.46 μmol) were added to dichloromethane (1.5 mL) in sequence, and stirring was continued at room temperature for 16 hours. After the reaction was complete, water (50 mL) was added, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to give pyridin-3-yl (4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine -1(2H)-yl)methanone 45b (49 mg), yield: 99.66%. The crude product was directly used for the next step reaction.

MS m/z(ESI):564.3[M+1].

Step 2

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-3-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-3-yl)methanone

Pyridin-3-yl(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 45b (49 mg, 86.93 μmol) was added to dichloromethane (1.5 mL), followed by dropwise addition of trifluoroacetic acid (0.5 mL), and stirring was continued at room temperature for 1 hour. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-3-yl)methanone 45 (37.08 mg) with a yield of 70.42%.

MS m/z(ESI):480.2[M+1].

According to the synthesis method of Examples 1-45 of the present invention, Examples 46-56 were synthesized, and the structures and characterization data are shown in the following table:

Embodiment 57

(S)-4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde

(S)-4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde

first step

4-((trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde

4-((Trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde

4-((Trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (250 mg, 1.16 mmol, commercially available) and triethylamine (468.92 mg, 4.63 mmol, 642.35 μL) were added to formamide (5.13 mL) in sequence and stirred at room temperature for 12 hours. Ethyl acetate (30 mL) and water (15 mL) were added, the organic phase was separated, the aqueous phase was extracted with ethyl acetate (30 mL×2), the combined organic phase was washed with saturated sodium chloride solution (30 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated to dryness under reduced pressure to obtain 4-((trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde 57b (220 mg), which was directly used in the next step.

MS m/z(ESI):208.0[M+1].

Step 2

4-ethynyl-3,6-dihydropyridine-1(2H)-carbaldehyde

4-Ethynyl-3,6-dihydropyridine-1(2H)-carbaldehyde

4-((Trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde 57b (220 mg, 1.06 mmol) and potassium fluoride (308.23 mg, 5.31 mmol) were added to methanol (5 mL) in sequence and stirred for 3 hours at room temperature. The system was concentrated to dryness under reduced pressure, acetonitrile (3 mL) was added to dissolve, filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain 4-ethynyl-3,6-dihydropyridine-1(2H)-carbaldehyde 57c (140 mg), which was directly used in the next step reaction.

MS m/z(ESI):136.0[M+1].

third step

4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde

4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (250 mg, 495.50 μmol), 4-ethynyl-3,6-dihydropyridine-1(2H)-carbaldehyde 57c (118.19 mg, 743.26 μmol), triethylenediamine (111.16 mg, 991.01 μmol) and allylpalladium(II) chloride dimer (36.17 mg, 99.10 μmol) were added to acetonitrile (2.0 mL) in sequence, argon was replaced 3 times, and stirring was continued at room temperature for 3 hours. Ethyl acetate (30 mL) and water (15 mL) were added, and the liquids were separated. The aqueous phase was extracted with ethyl acetate (30 mL×2), and the combined organic phase was washed with saturated sodium chloride solution (30 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated to dryness under reduced pressure. The residue was separated and purified by silica gel chromatography (eluent: A system) to obtain 4-((4-(3-((2-((1S)-1-((tetrahydro 2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde 57d (240 mg), with a yield of 89.59%. MS m/z(ESI):487.3[M+1].

the fourth step

(S)-4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde

(S)-4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde

4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde 57d (220 mg, 452.15 μmol) and trifluoroacetic acid (0.5 mL) were added to dichloromethane (2 mL) and stirred at room temperature for 1 hour. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carbaldehyde 57 (59.7 mg) with a yield of 25.05%.

MS m/z(ESI):402.9[M+1]. ¹ H NMR(400MHz,DMSO-d ₆ )δ8.09(s,1H),7.91–7.85(m,2H),7.79(d,J＝2.0Hz,1H),7.71(d,J＝2.0Hz,1H),7.61(d,J＝8.2Hz,2H),7.18(s,1H),6.32–6.21(m,1H),5.73(s,2H),5.27(q,J＝6.6Hz,1H),4.04(dq,J＝20.9,3.0Hz,2H),3.55(t,J＝5.7Hz,2H),2.36(dq,J＝5.8,2.9Hz,1H),2.26(h,J＝2.7Hz,1H),1.50(d,J＝6.6Hz,3H).

Embodiment 58

(S)-4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide

(S)-4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide

first step

4-((trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide

4-((Trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide

4-((Trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (50 mg, 278.81 μmol) and triethylamine (84.64 mg, 836.43 μmol, 115.94 μL) were added to tetrahydrofuran (1.95 mL) in sequence, and then isocyanatotrimethylsilane 58a (41.76 mg, 362.45 μmol, 49.13 μL, commercially available) was added and reacted at room temperature for 16 hours. After the reaction was completed, 4-((trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide 58b (50 mg) was concentrated and used directly in the next step.

MS m/z(ESI):223.0[M+1].

Step 2

4-ethynyl-3,6-dihydropyridine-1(2H)-carboxamide

4-Ethynyl-3,6-dihydropyridine-1(2H)-carboxamide

4-((Trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide 58b (50 mg, 224.86 μmol) and potassium fluoride (65.32 mg, 1.12 mmol) were added to methanol (2 mL) in sequence and stirred at room temperature for 12 hours. The system was concentrated to dryness under reduced pressure, ethyl acetate (30 mL) and water (15 mL) were added, the organic phase was separated, the aqueous phase was extracted with ethyl acetate (30 mL×2), the combined organic phase was washed with saturated sodium chloride solution (30 mL×2), and concentrated under reduced pressure to give 4-ethynyl-3,6-dihydropyridine-1(2H)-carboxamide 58c (20 mg), which was directly used in the next step.

MS m/z(ESI):151.1[M+1].

third step

4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide

4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide

Under argon atmosphere, 5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (50 mg, 104.32 μmol), 4-ethynyl-3,6-dihydropyridine-1(2H)-carboxamide 58c (23.50 mg, 156.47 μmol), triethylenediamine (23.40 mg, 208.63 μmol), allylpalladium(II) chloride dimer (7.62 mg, 20.86 μmol), and tri-tert-butylphosphine (10% in toluene solution) (42.21 mg, 20.86 μmol, 10% purity) were added to acetonitrile (2 mL) in sequence. After argon replacement, stirring was continued at room temperature for 4 hours. Ethyl acetate (30 mL) and water (15 mL) were added, and the liquids were separated. The aqueous phase was extracted with ethyl acetate (30 mL×2). The combined organic phase was washed with saturated sodium chloride solution (30 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated to dryness under reduced pressure. The residue was separated and purified by silica gel chromatography (eluent: System A) to give 4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide 58d (40 mg) in a yield of 76.45%.

MS m/z(ESI):502.3[M+1].

the fourth step

(S)-4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide

(S)-4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide

4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide 58d (40 mg, 79.75 μmol) and trifluoroacetic acid (0.5 mL) were added sequentially to dichloromethane (2 mL), and stirring was continued at room temperature for 1 hour. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-carboxamide 58 (13.5 mg) with a yield of 30.26%.

MS m/z(ESI):418.2[M+1].

Embodiment 59

(S)-2-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide

(S)-2-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide

first step

2-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide

2-(4-((Trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide

2-Iodoacetamide 59a (123.77 mg, 669.14 μmol, commercially available), 4-((trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (100 mg, 557.62 μmol) and cesium carbonate (363.37 mg, 1.12 mmol) were added to acetonitrile (2 mL) in sequence and stirred at room temperature for 12 hours. The reaction solution was distilled under reduced pressure to remove the organic solvent, and then ethyl acetate (50 mL) and water (30 mL) were added. The liquids were separated, and the aqueous phase was extracted with ethyl acetate (30 mL×2). The organic phases were combined, dried over anhydrous sodium sulfate, and then the organic solvent was distilled under reduced pressure to obtain 2-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide 59b (130 mg), which was directly used in the next step reaction.

MS m/z(ESI):237.2[M+1].

Step 2

2-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)acetamide

2-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)acetamide

2-(4-((Trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide 59b (130 mg, 549.95 μmol) and potassium fluoride (95.86 mg, 1.65 mmol) were added to methanol (2 mL) in sequence and stirred for 16 hours at room temperature. The reaction solution was distilled under reduced pressure to remove the organic solvent to obtain 2-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)acetamide 59c (80 mg), which was directly used in the next step reaction.

MS m/z(ESI):165.2[M+1].

third step

2-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide

2-(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (150 mg, 312.95 μmol), 2-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)acetamide 59c (56.53 mg, 344.24 μmol), triethylenediamine (70.21 mg, 625.90 μmol), allylpalladium(II) chloride dimer (22.85 mg, 62.59 μmol) and tri-tert-butylphosphine (10% in toluene solution) (12.66 mg, 62.59 μmol) were added to acetonitrile (2 mL) in sequence. After argon replacement, stirring was continued at room temperature for 4 hours. The reaction solution was distilled under reduced pressure to remove the organic solvent, and the residue was separated and purified by silica gel column chromatography (eluent: System A) to give 2-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide 59d (108 mg) in a yield of 66.93%.

MS m/z(ESI):516.3[M+1].

the fourth step

(S)-2-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide

(S)-2-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide

2-(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide 59d (50 mg, 96.97 μmol) and trifluoroacetic acid (0.5 mL) were added sequentially to dichloromethane (1.5 mL), and stirring was continued at room temperature for 3 hours. The crude product was concentrated and subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-2-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)acetamide 59 (26.0 mg) with a yield of 43.60%. MS m/z (ESI): 431.9 [M+1].

Embodiment 60

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(morpholino)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(morpholino)methanone

first step

morpholino(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Morpholino(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

4-((Trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (62.50 mg, 278.81 μmol), morpholine 60a (48.58 mg, 557.62 μmol, 49.07 μL), triphosgene (99.28 mg, 334.57 μmol) and triethylamine (169.28 mg, 1.67 mmol, 235.11 μL) were added to tetrahydrofuran (1.72 mL) in sequence and stirred at room temperature for 12 hours. Ethyl acetate (30 mL) and water (15 mL) were added, the organic phase was separated, the aqueous phase was extracted with ethyl acetate (30 mL×2), and the combined organic phase was quenched with saturated sodium chloride. The mixture was washed with 1% paraformaldehyde (30 mL x 2), dried over anhydrous sodium sulfate, filtered, and concentrated to dryness under reduced pressure. The residue was separated and purified by silica gel column chromatography (eluent: system A) to give morpholino (4-((trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-yl)methanone 60b (40 mg) with a yield of 49.06%.

MS m/z(ESI):293.0[M+1].

Step 2

(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(morpholino)methanone

(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(morpholino)methanone

Morpholino (4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 60b (40 mg, 136.78 μmol) and potassium fluoride (39.73 mg, 683.88 μmol) were added to methanol (1 mL) in sequence and stirred at room temperature for 4 hours. The system was concentrated to dryness under reduced pressure, acetonitrile (10 mL) was added to the residue, filtered, and the filtrate was concentrated to dryness to obtain (4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(morpholino)methanone 60c (20 mg), which was directly used in the next step reaction.

MS m/z(ESI):221.0[M+1].

third step

morpholino(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Morpholino(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (25 mg, 49.55 μmol), (4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(morpholino)methanone 60c (18.19 mg, 74.33 μmol), triethylenediamine (11.12 mg, 99.10 μmol) and allylpalladium(II) chloride dimer (3.62 mg, 9.91 μmol) were added to acetonitrile (2 mL) in sequence, argon was replaced 3 times, and the mixture was stirred at room temperature for 12 hours. The system was concentrated to dryness under reduced pressure, and the residue was separated and purified by silica gel column chromatography (eluent: system A) to give morpholino(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 60d (20 mg) in a yield of 70.61%.

MS m/z(ESI):572.0[M+1].

the fourth step

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(morpholino)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(morpholino)methanone

Morpholino(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 60d (20 mg, 34.99 μmol) and trifluoroacetic acid (0.3 mL) were added to dichloromethane (1 mL) in sequence, and the mixture was stirred for 1 hour at room temperature. The system was concentrated to dryness under reduced pressure, and the residue was separated by preparative liquid phase separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to obtain (S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl) methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(morpholino)methanone 60 (2.5 mg), yield 11.11%.

MS m/z(ESI):487.9[M+1].

Embodiment 61

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-2-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-2-yl)methanone

first step

pyridin-2-yl(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Pyridin-2-yl(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

4-((Trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (50 mg, 278.81 μmol), picolinic acid 61a (51.49 mg, 418.22 μmol, commercially available), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (127.21 mg, 334.57 μmol), and N,N-diisopropylethylamine (108.10 mg, 836.43 μmol, 145.69 μL) were added to dichloromethane (2.0 mL) in sequence, and stirring was continued at room temperature for 4 hours. Mass spectrometry indicated that the reaction of the starting material was complete and the system was concentrated to dryness under reduced pressure to obtain pyridin-2-yl(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 61b (55 mg), which was directly used in the next step.

MS m/z(ESI):285.1[M+1].

Step 2

(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(pyridin-2-yl)methanone

(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(pyridin-2-yl)methanone

Pyridin-2-yl (4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 61b (55 mg, 193.37 μmol) and potassium fluoride (56.17 mg, 966.85 μmol) were added to methanol (2 mL) in sequence and stirred for 3 hours at room temperature. The system was concentrated to dryness under reduced pressure, acetonitrile (3 mL) was added to dissolve, filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain (4-ethynyl-3,6-dihydropyridin-1(2H)-yl) (pyridin-2-yl)methanone 61c (30 mg), which was directly used in the next step reaction.

MS m/z(ESI):213.0[M+1].

third step

pyridin-2-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

Pyridin-2-yl(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (30 mg, 59.46 μmol), (4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(pyridin-2-yl)methanone 61c (21.03 mg, 89.19 μmol), triethylenediamine (13.34 mg, 118.92 μmol) and allylpalladium(II) chloride dimer (4.34 mg, 11.89 μmol) were added to acetonitrile (2 mL) in sequence, argon was replaced 3 times, and the mixture was stirred at room temperature for 12 hours. The system was concentrated to dryness under reduced pressure, and the residue was separated and purified by silica gel column chromatography (eluent: system A) to give pyridin-2-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 61d (25 mg) in a yield of 74.59%.

MS m/z(ESI):563.9[M+1].

the fourth step

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-2-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-2-yl)methanone

Pyridin-2-yl(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 61d (25 mg, 44.35 μmol) and trifluoroacetic acid (0.5 mL) were added sequentially to dichloromethane (2 mL), and stirring was continued at room temperature for 1 hour. The system was concentrated to dryness under reduced pressure, and the residue was subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(pyridin-2-yl)methanone 61 (15.2 mg) with a yield of 56.00%.

MS m/z(ESI):480.2[M+1].

Embodiment 62

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

first step

1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

1-(4-((Trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

4-((Trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (50 mg, 278.81 μmol), acetic anhydride (42.70 mg, 418.22 μmol, 39.53 μL) and triethylamine (84.64 mg, 836.43 μmol, 116.26 μL) were added to dichloromethane (1 mL) in sequence and stirred at room temperature for 12 hours. The system was concentrated to dryness under reduced pressure, and the residue was separated and purified by silica gel column chromatography (eluent: A system) to obtain 1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 62a (50 mg) with a yield of 81.01%.

MS m/z(ESI):222.0[M+1].

Step 2

1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

1-(4-((Trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 62a (50 mg, 225.87 μmol) and potassium fluoride (65.61 mg, 1.13 mmol) were added to methanol (1 mL) in sequence and stirred for 3 hours at room temperature. The system was concentrated to dryness under reduced pressure, acetonitrile (3 mL) was added to dissolve, filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain 1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 62b (30 mg), which was directly used in the next step reaction.

MS m/z(ESI):150.0[M+1].

third step

1-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

1-(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (30 mg, 59.46 μmol), 1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 62b (14.78 mg, 89.19 μmol), triethylenediamine (13.34 mg, 118.92 μmol) and allylpalladium(II) chloride dimer (4.34 mg, 11.89 μmol) were added to acetonitrile (2 mL) in sequence, argon was replaced 3 times, and the mixture was stirred at room temperature for 12 hours. The system was concentrated to dryness under reduced pressure, and the residue was separated and purified by silica gel column chromatography (eluent: System A) to give 1-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 62c (25 mg) in a yield of 83.99%.

MS m/z(ESI):500.9[M+1].

the fourth step

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

1-(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 62c (25 mg, 49.94 μmol) and trifluoroacetic acid (0.5 mL) were added sequentially to dichloromethane (2 mL), and stirring was continued at room temperature for 1 hour. The system was concentrated to dryness under reduced pressure, and the residue was separated by preparative liquid phase separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 62 (6.8 mg), yield 24.38%. MS m/z (ESI): 417.2 [M+1].

Embodiment 63

(S)-2-hydroxy-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

(S)-2-Hydroxy-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

first step

2-hydroxy-1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

2-Hydroxy-1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

4-((Trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (50 mg, 278.81 μmol), 2-hydroxyacetic acid 12a (42.41 mg, 557.62 μmol), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (127.21 mg, 334.57 μmol), and N,N-diisopropylethylamine (108.10 mg, 836.43 μmol, 145.69 μL) were added to dichloromethane (2.0 mL) in sequence and stirred for 4 hours at room temperature. Mass spectrometry indicated that the reaction of the starting materials was complete, and the system was concentrated to dryness under reduced pressure to obtain 2-hydroxy-1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 63a (40 mg), which was directly used in the next step.

MS m/z(ESI):237.9[M+1].

Step 2

1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)-2-hydroxyethan-1-one

1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)-2-hydroxyethan-1-one

2-Hydroxy-1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 63a (40 mg, 168.51 μmol) and potassium fluoride (48.95 mg, 842.57 μmol) were added to methanol (1 mL) in sequence and stirred for 3 hours at room temperature. The system was concentrated to dryness under reduced pressure, acetonitrile (3 mL) was added to dissolve, filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain 1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)-2-hydroxyethan-1-one 63b (20 mg), which was directly used in the next step reaction.

MS m/z(ESI):166.0[M+1].

third step

2-hydroxy-1-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

2-Hydroxy-1-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (30 mg, 59.46 μmol), 1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)-2-hydroxyethan-1-one 63b (16.37 mg, 89.19 μmol), triethylenediamine (13.34 mg, 118.92 μmol) and allylpalladium (II) chloride dimer (4.34 mg, 11.89 μmol) were added to acetonitrile (2 mL) in sequence, argon was replaced 3 times, and the mixture was stirred at room temperature for 12 hours. The system was concentrated to dryness under reduced pressure, and the residue was separated and purified by silica gel column chromatography (eluent: A system) to obtain 2-hydroxy-1-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 63c (25 mg) with a yield of 81.39%.

MS m/z(ESI):516.9[M+1].

the fourth step

(S)-2-hydroxy-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

(S)-2-Hydroxy-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

2-Hydroxy-1-(4-((4-(3-((2-((1S)-1-((tetrahydro2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 63c (25 mg, 48.39 μmol) and trifluoroacetic acid (0.5 mL) were added to dichloromethane (2 mL) in sequence, and stirring was continued at room temperature for 1 hour. The system was concentrated to dryness under reduced pressure, and the residue was subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-2-hydroxy-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 63 (1.3 mg) with a yield of 4.62%.

MS m/z(ESI):433.2[M+1]. ¹ H NMR(400MHz,DMSO-d ₆ )δ7.89–7.86 (m, 2H), 7.76–7.75 (d, J = 4.0 Hz, 1H), 7.67–7.66 (d, J = 4.0 Hz, 1H), 7.62–7.60 (m, 3H), 7.17–7.16 (d, J = 4.0 Hz, 1H), 6.27–6.24 (m, 1H), 5.73 (s, 2H), 5.24 (q, J = 6.6 Hz, 1H), 4.41–4.04 (m, 4H), 3.64–3.61 (m, 1H), 3.50–3.47 (m, 1H), 2.35–2.28 (m, 2H), 1.50 (d, J = 6.6 Hz, 3H).

Embodiment 64

(1,4-dioxan-2-yl)(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(1,4-dioxan-2-yl)(4-((3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

first step

(1,4-dioxan-2-yl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(1,4-dioxane-2-yl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

4-((Trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (50 mg, 278.81 μmol), 1,4-dioxane-2-carboxylic acid 64a (55.25 mg, 418.22 μmol), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (127.21 mg, 334.57 μmol), and N,N-diisopropylethylamine (108.10 mg, 836.43 μmol, 145.69 μL) were added to dichloromethane (2.0 mL) in sequence and stirred for 4 hours at room temperature. Mass spectrometry indicated that the reaction of the starting materials was complete, and the system was concentrated to dryness under reduced pressure to obtain (1,4-dioxane-2-yl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-yl)methanone 64b (45 mg), which was directly used in the next step.

MS m/z(ESI):294.0[M+1].

Step 2

(1,4-dioxan-2-yl)(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)methanone

(1,4-dioxane-2-yl)(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)methanone

(1,4-dioxane-2-yl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridine-1(2H)-yl)methanone 64b (45 mg, 153.36 μmol) and potassium fluoride (8.91 mg, 153.36 μmol) were added to methanol (2 mL) in sequence and stirred for 3 hours at room temperature. The system was concentrated to dryness under reduced pressure, acetonitrile (3 mL) was added to dissolve, filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain (1,4-dioxane-2-yl)(4-ethynyl-3,6-dihydropyridine-1(2H)-yl)methanone 64c (25 mg), which was directly used in the next step reaction.

MS m/z(ESI):222.1[M+1].

third step

(1,4-dioxan-2-yl)(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(1,4-dioxan-2-yl)(4-((3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (30 mg, 59.46 μmol), (1,4-dioxane-2-yl)(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)methanone 64c (21.93 mg, 89.19 μmol), triethylenediamine (13.34 mg, 118.92 μmol) and allylpalladium(II) chloride dimer (4.34 mg, 11.89 μmol) were added to acetonitrile (2 mL) in sequence, argon was replaced 3 times, and the mixture was stirred at room temperature for 12 hours. The system was concentrated to dryness under reduced pressure, and the residue was separated and purified by silica gel column chromatography (eluent: system A) to give (1,4-dioxane-2-yl)(4-((3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridine-1(2H)-yl)methanone 64d (25 mg) in a yield of 73.42%.

MS m/z(ESI):573.0[M+1].

the fourth step

(1,4-dioxan-2-yl)(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(1,4-dioxan-2-yl)(4-((3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(1,4-Dioxane-2-yl)(4-((3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 64d (25 mg, 43.66 μmol) and trifluoroacetic acid (0.5 mL) were added sequentially to dichloromethane (2 mL), and stirring was continued at room temperature for 1 hour. The system was concentrated to dryness under reduced pressure, and the residue was subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (1,4-dioxane-2-yl)(4-((3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 64 (3.8 mg) with a yield of 14.33%.

MS m/z(ESI):488.9[M+1].

Embodiment 65

(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-2-yl)methanone

(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-2-yl)methanone

first step

(tetrahydro-2H-pyran-2-yl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(Tetrahydro-2H-pyran-2-yl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

4-((Trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (50 mg, 278.81 μmol), tetrahydro-2H-pyran-2-carboxylic acid 65a (54.43 mg, 418.22 μmol, 47.74 μL, commercially available), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (127.21 mg, 334.57 μmol) and N,N-diisopropylethylamine (108.10 mg, 836.43 μmol, 145.69 μL) were added to dichloromethane (1.95 mL) in sequence and stirred for 4 hours at room temperature. Mass spectrometry indicated that the reaction of the starting material was complete, and the system was concentrated to dryness under reduced pressure to obtain (tetrahydro-2H-pyran-2-yl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 65b (60 mg), which was directly used in the next step reaction.

MS m/z(ESI):292.0[M+1].

Step 2

(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-2-yl)methanone

(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-2-yl)methanone

(Tetrahydro-2H-pyran-2-yl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 65b (60 mg, 205.86 μmol) and potassium fluoride (59.80 mg, 1.03 mmol) were added to methanol (2 mL) in sequence and stirred for 3 hours at room temperature. The system was concentrated to dryness under reduced pressure, acetonitrile (3 mL) was added to dissolve, filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain (4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-2-yl)methanone 65c (30 mg), which was used directly in the next step. MS m/z(ESI):220.1[M+1].

third step

(tetrahydro-2H-pyran-2-yl)(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(Tetrahydro-2H-pyran-2-yl)(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (30 mg, 59.46 μmol), (4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-2-yl)methanone 65c (21.73 mg, 89.19 μmol), triethylenediamine (13.34 mg, 118.92 μmol) and allylpalladium (II) chloride dimer (4.34 mg, 11.89 μmol) were added to acetonitrile (2 mL) in sequence, argon was replaced 3 times, and the mixture was stirred at room temperature for 12 hours. The system was concentrated to dryness under reduced pressure, and the residue was separated and purified by silica gel column chromatography (eluent: A system) to obtain (tetrahydro-2H-pyran-2-yl)(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 65d (20 mg) with a yield of 58.94%.

MS m/z(ESI):570.9[M+1].

the fourth step

(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-2-yl)methanone

(4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-2-yl)methanone

(Tetrahydro-2H-pyran-2-yl)(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 65d (20 mg, 35.05 μmol) and trifluoroacetic acid (0.5 mL) were added successively to dichloromethane (2 mL), and stirring was continued at room temperature for 1 hour. The system was concentrated to dryness under reduced pressure, and the residue was subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (4-((4-(3-((2-((S)-1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-2-yl)methanone 65 (4.9 mg) with a yield of 22.44%.

MS m/z(ESI):486.9[M+1].

Embodiment 66

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-4-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro2H-pyran-4-yl)methanone

first step

(tetrahydro-2H-pyran-4-yl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(Tetrahydro-2H-pyran-4-yl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

4-((Trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (50 mg, 278.81 μmol), tetrahydro-2H-pyran-4-carboxylic acid 66a (54.43 mg, 418.22 μmol), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (127.21 mg, 334.57 μmol), and N,N-diisopropylethylamine (108.10 mg, 836.43 μmol, 145.69 μL) were added to dichloromethane (1.85 mL) in sequence, and stirring was continued for 4 hours at room temperature. Mass spectrometry indicated that the reaction of the starting materials was complete, and the system was concentrated to dryness under reduced pressure to obtain (tetrahydro-2H-pyran-4-yl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 66b (50 mg), which was directly used in the next step.

MS m/z(ESI):292.0[M+1].

Step 2

(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-4-yl)methanone

(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-4-yl)methanone

(Tetrahydro-2H-pyran-4-yl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 66b (50 mg, 171.55 μmol) and potassium fluoride (49.84 mg, 857.75 μmol) were added to methanol (2 mL) in sequence and stirred for 3 hours at room temperature. The system was concentrated to dryness under reduced pressure, acetonitrile (3 mL) was added to dissolve, filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain (4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-4-yl)methanone 66c (25 mg), which was directly used in the next step. MS m/z(ESI):220.0[M+1].

third step

(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-4-yl)methanone

(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-4-yl)methanone

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (30 mg, 59.46 μmol), (4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-4-yl)methanone 66c (23.01 mg, 89.19 μmol), triethylenediamine (13.34 mg, 118.92 μmol) and allylpalladium(II) chloride dimer (4.34 mg, 11.89 μmol) were added to acetonitrile (2.0 mL) in sequence, argon was replaced 3 times, and stirring was continued at room temperature for 3 hours. Ethyl acetate (30 mL) and water (15 mL) were added, the organic phase was separated, the aqueous phase was extracted with ethyl acetate (30 mL × 2), the combined organic phase was washed with saturated sodium chloride solution (30 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to dryness under reduced pressure. The residue was separated and purified by silica gel column chromatography (eluent: A system) to obtain (4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy) ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-4-yl)methanone 66d (25 mg), yield 73.68%.

MS m/z(ESI):571.0[M+1].

the fourth step

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-4-yl)methanone

(S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro2H-pyran-4-yl)methanone

(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro-2H-pyran-4-yl)methanone 66d (20 mg, 35.05 μmol) and trifluoroacetic acid (0.5 mL) were added sequentially to dichloromethane (1 mL), and stirring was continued at room temperature for 1 hour. The system was concentrated to dryness under reduced pressure, and the residue was subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)(tetrahydro 2H-pyran-4-yl)methanone 66 (15.8 mg) with a yield of 71.39%.

MS m/z(ESI):486.9[M+1].

Embodiment 67

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-3-methoxypropan-1-one

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-3-methoxypropan-1-one

first step

3-methoxy-1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propan-1-one

3-Methoxy-1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propan-1-one

4-((Trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (50 mg, 278.81 μmol), 3-methoxypropionic acid 67a (43.54 mg, 418.22 μmol, 39.29 μL, commercially available), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (127.21 mg, 334.57 μmol), and N,N-diisopropylethylamine (108.10 mg, 836.43 μmol, 145.69 μL) were added to dichloromethane (2 mL) in sequence, and stirring was continued for 4 hours at room temperature. Mass spectrometry indicated that the reaction of the starting material was complete and the system was concentrated to dryness under reduced pressure to obtain 3-methoxy-1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propan-1-one 67b (50 mg), which was directly used in the next step.

MS m/z(ESI):266.0[M+1].

Step 2

1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)-3-methoxypropan-1-one

1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)-3-methoxypropan-1-one

3-Methoxy-1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propan-1-one 67b (50 mg, 188.38 μmol) and potassium fluoride (54.72 mg, 941.89 μmol) were added to methanol (2 mL) in sequence and stirred for 3 hours at room temperature. The system was concentrated to dryness under reduced pressure, acetonitrile (3 mL) was added to dissolve, filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain 1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)-3-methoxypropan-1-one 67c (20 mg), which was directly used in the next step reaction.

MS m/z(ESI):194.0[M+1].

third step

3-methoxy-1-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propan-1-one

3-methoxy-1-(4-(4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propan-1-one

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (30 mg, 59.46 μmol), 1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)-3-methoxypropan-1-one 67c (20.28 mg, 89.19 μmol), triethylenediamine (13.34 mg, 118.92 μmol) and allylpalladium(II) chloride dimer (4.34 mg, 11.89 μmol) were added to acetonitrile (2.0 mL) in sequence, argon was replaced 3 times, and stirring was continued at room temperature for 3 hours. Ethyl acetate (30 mL) and water (15 mL) were added, the organic phase was separated, the aqueous phase was extracted with ethyl acetate (30 mL×2), the combined organic phase was washed with saturated sodium chloride solution (30 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated to dryness under reduced pressure. The residue was separated and purified by silica gel column chromatography (eluent: System A) to give 3-methoxy-1-(4-(4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propan-1-one 67d (25 mg) in a yield of 77.20%.

MS m/z(ESI):545.0[M+1].

the fourth step

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-3-methoxypropan-1-one

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-3-methoxypropan-1-one

3-Methoxy-1-(4-(4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)propan-1-one 67d (20 mg, 36.72 μmol) and trifluoroacetic acid (0.5 mL) were added to dichloromethane (1 mL) in sequence, and stirring was continued at room temperature for 1 hour. The system was concentrated to dryness under reduced pressure, and the residue was subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-3-methoxypropan-1-one 67 (14.0 mg) with a yield of 64.70%.

MS m/z(ESI):461.0[M+1].

Embodiment 68

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-methoxyethan-1-one

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-methoxyethan-1-one

first step

2-methoxy-1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

2-Methoxy-1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

4-((Trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (50 mg, 278.81 μmol), 2-methoxyacetic acid 68a (37.67 mg, 418.22 μmol, commercially available), O-(7-azabenzotriazole-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (127.21 mg, 334.57 μmol), and N,N-diisopropylethylamine (108.10 mg, 836.43 μmol, 145.69 μL) were added to dichloromethane (2 mL) in sequence and stirred for 4 hours at room temperature. The reaction of the raw materials was complete when detected by mass spectrometry, and the system was concentrated under reduced pressure. The residue was reduced to dryness to afford 2-methoxy-1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 68b (50 mg), which was used directly in the next step.

MS m/z(ESI):252.0[M+1].

Step 2

1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)-2-methoxyethan-1-one

1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)-2-methoxyethan-1-one

2-Methoxy-1-(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 68b (50 mg, 198.89 μmol) and potassium fluoride (57.78 mg, 994.45 μmol) were added to methanol (2 mL) in sequence and stirred for 3 hours at room temperature. The system was concentrated to dryness under reduced pressure, acetonitrile (3 mL) was added to dissolve, filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain 1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)-2-methoxyethan-1-one 68c (20 mg), which was directly used in the next step reaction.

MS m/z(ESI):180.0[M+1].

third step

2-methoxy-1-(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

2-Methoxy-1-(4-(4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (30 mg, 59.46 μmol), 1-(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)-2-methoxyethan-1-one 68c (18.81 mg, 89.19 μmol), triethylenediamine (13.34 mg, 118.92 μmol) and allylpalladium(II) chloride dimer (4.34 mg, 11.89 μmol) were added to acetonitrile (2.0 mL) in sequence, argon was replaced 3 times, and stirring was continued at room temperature for 3 hours. Ethyl acetate (30 mL) and water (15 mL) were added, the organic phase was separated, the aqueous phase was extracted with ethyl acetate (30 mL×2), the combined organic phase was washed with saturated sodium chloride solution (30 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated to dryness under reduced pressure. The residue was separated and purified by silica gel column chromatography (eluent: System A) to give 2-methoxy-1-(4-(4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 68d (25 mg) in a yield of 79.24%.

MS m/z(ESI):531.0[M+1].

the fourth step

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-methoxyethan-1-one

(S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-methoxyethan-1-one

2-Methoxy-1-(4-(4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)ethan-1-one 68d (20 mg, 37.69 μmol) and trifluoroacetic acid (0.5 mL) were added to dichloromethane (1 mL) in sequence and stirred at room temperature for 1 hour. The system was concentrated to dryness under reduced pressure, and the residue was separated by preparative liquid phase separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; Mobile phase A: 0.05% TFA+ _H2O , mobile phase B: _CH3CN ) to give (S)-1-(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)-2-methoxyethan-1-one 68 (13.0 mg) with a yield of 58.89%.

MS m/z(ESI):446.9[M+1].

Embodiment 69

(S)-(1-hydroxycyclopropyl)(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(S)-(1-Hydroxycyclopropyl)(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

first step

(1-hydroxycyclopropyl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(1-Hydroxycyclopropyl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

4-((Trimethylsilyl)ethynyl)-1,2,3,6-tetrahydropyridine 57a (50 mg, 278.81 μmol), 1-hydroxycyclopropane-1-carboxylic acid 69a (42.69 mg, 418.22 μmol, commercially available), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (127.21 mg, 334.57 μmol), and N,N-diisopropylethylamine (108.10 mg, 836.43 μmol, 145.69 μL) were added to dichloromethane (2 mL) in sequence, and stirring was continued for 4 hours at room temperature. Mass spectrometry indicated that the reaction of the starting material was complete and the system was concentrated to dryness under reduced pressure to obtain (1-hydroxycyclopropyl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 69b (50 mg), which was directly used in the next step.

MS m/z(ESI):263.9[M+1].

Step 2

(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(1-hydroxycyclopropyl)methanone

(4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(1-hydroxycyclopropyl)methanone

(1-Hydroxycyclopropyl)(4-((trimethylsilyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 69b (50 mg, 189.82 μmol) and potassium fluoride (55.14 mg, 949.10 μmol) were added to methanol (2 mL) in sequence and stirred for 3 hours at room temperature. The system was concentrated to dryness under reduced pressure, acetonitrile (3 mL) was added to dissolve, filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain (4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(1-hydroxycyclopropyl)methanone 69c (30 mg), which was directly used in the next step reaction.

MS m/z(ESI):191.9[M+1].

third step

(1-hydroxycyclopropyl)(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(1-Hydroxycyclopropyl)(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

5-(4-iodophenyl)-3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazole 1i (30 mg, 59.46 μmol), (4-ethynyl-3,6-dihydropyridin-1(2H)-yl)(1-hydroxycyclopropyl)methanone 69c (20.07 mg, 89.19 μmol), triethylenediamine (13.34 mg, 118.92 μmol) and allylpalladium(II) chloride dimer (4.34 mg, 11.89 μmol) were added to acetonitrile (2.0 mL) in sequence, argon was replaced 3 times, and stirring was continued at room temperature for 3 hours. Ethyl acetate (30 mL) and water (15 mL) were added, the liquids were separated, the aqueous phase was extracted with ethyl acetate (30 mL×2), the combined organic phase was washed with saturated sodium chloride solution (30 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated to dryness under reduced pressure. The residue was separated and purified by silica gel column chromatography (eluent: System A) to give (1-hydroxycyclopropyl)(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 69d (25 mg) in a yield of 77.48%.

MS m/z(ESI):543.0[M+1].

the fourth step

(S)-(1-hydroxycyclopropyl)(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(S)-(1-Hydroxycyclopropyl)(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone

(1-Hydroxycyclopropyl)(4-((4-(3-((2-((1S)-1-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 69d (20 mg, 36.86 μmol) and trifluoroacetic acid (0.5 mL) were added sequentially to dichloromethane (1 mL), and stirring was continued at room temperature for 1 hour. The system was concentrated to dryness under reduced pressure, and the residue was subjected to preparative liquid separation (separation column AKZONOBEL Kromasil; 250×21.2 mm; 5 μm, 20 mL/min; mobile phase A: 0.05% TFA+H ₂ O, mobile phase B: CH ₃ CN) to give (S)-(1-hydroxycyclopropyl)(4-((4-(3-((2-(1-hydroxyethyl)-1H-imidazol-1-yl)methyl)isoxazol-5-yl)phenyl)ethynyl)-3,6-dihydropyridin-1(2H)-yl)methanone 69 (2.2 mg) with a yield of 9.83%.

MS m/z(ESI):458.9[M+1].

Biological evaluation

Test Example 1: Determination of the Inhibition of LpxC Enzyme Activity by the Compounds of the Invention

The following method is used to determine the degree of inhibition of the compounds of the present invention on the enzymatic activity of recombinant Pseudomonas aeruginosa LpxC under in vitro conditions.

The experimental process is briefly described as follows: the test compound was first dissolved in DMSO to prepare a 10mM stock solution. The reaction was carried out in a 96-well microplate. First, 20μL of recombinant Pseudomonas aeruginosa LpxC (purchased from Signalway Antibody, catalog number AP74647-2) was added to the wells, with a final concentration of 5nM; 5μL of the test compound was added, and the compound was diluted 4 times, with 8 concentration points, ranging from 0.61-10000nM; 5μL of LpxC substrate UDP-3-O-(R-3-hydroxydecanoyl)-GlcNAc (purchased from Biosynth Carbosynth, catalog number mu75071) was added, with a final substrate concentration of 10μM, and incubated at 25°C for 120 minutes. Then add 20 μL of 2.0 mg/mL fluorescent amine (purchased from Sigmaaldrich, item number F9015, solvent is 1:1 dimethylformamide/acetonitrile) to the reaction system, mix well, and react for 10 minutes; finally, add 50 μL of 200 mM sodium phosphate buffer (pH 8.0) to terminate the reaction, and read using a microplate reader (BMG), with excitation wavelength and emission wavelength of 390 and 495 nm, respectively. By comparing with the fluorescence intensity ratio of the control group (0.1% DMSO), the percentage inhibition rate of the compound at each concentration was calculated, and the nonlinear regression analysis of the compound concentration logarithm-inhibition rate was performed by GraphPad Prism 5 software to obtain the IC ₅₀ value of the compound, as shown in Table 1 below.

Table 1 Inhibition results of the compounds of the present invention on the enzymatic activity of LpxC

Conclusion: The preferred compounds of the present invention have an IC ₅₀ <100 nM for inhibiting the activity of the recombinant Pseudomonas aeruginosa LpxC enzyme, and have a significant inhibitory effect on the activity of the LpxC enzyme.

Test Example 2: Evaluation of Antibacterial Activity of the Compounds of the Invention

In vitro minimum inhibitory concentration (MIC) determination was performed according to the CLSI guidelines using the broth microdilution method.

The experimental process is briefly described as follows: the test compound is dissolved in DMSO to prepare a 12.8 mg/mL stock solution, and then DMSO is used to prepare 11 two-fold dilutions of 100× high concentration working solutions (the final concentration of the system is 64μg/mL-0.06μg/mL). The strains (K.Pneumoniae ATCC13883, K.Pneumoniae ATCC51504 and E.coli ATCC 25922) frozen in glycerol at -80°C are inoculated into solid agar medium and placed in an incubator at 35°C for 18-24 hours to complete the strain preparation work, and then an appropriate amount of solid plate culture is collected and resuspended in physiological saline, mixed, and the turbidity of the bacterial suspension is adjusted to an appropriate turbidity with a turbidity meter, containing about 1×10 ⁸ cfu/mL of bacteria, and then the bacterial suspension with adjusted turbidity is diluted with a test medium (type: CAMHIIB, purchased from BD) to a bacterial concentration of 5×10 ⁵ cfu/mL to complete the preparation of the inoculum. 198 μL of the inoculum solution was inoculated into a 96-well plate, followed by the addition of 2 μL of a 100× high concentration working solution of the compound, and then the 96-well plate was cultured at 35° C. for 18 to 24 h. After culture, the test plate was observed by naked eye, and the minimum drug concentration that completely inhibited bacterial growth was the minimum inhibitory concentration (MIC) of the compound, as shown in Table 2.

Table 2 Antibacterial activity test results of the compounds of the present invention

Conclusion: The in vitro minimum inhibitory degree of the preferred compounds of the present invention against K. Pneumoniae ATCC13883, K. Pneumoniae ATCC51504 and Escherichia coli (E. coli ATCC 25922) is less than 50 μg/mL; they have good inhibitory effects on both K. Pneumoniae and Escherichia coli.

Claims (10)

A compound represented by general formula (I) or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof:
in: W is selected from O, S(O) _r or N; Ring C is selected from a 5- to 10-membered spiro heterocyclic group or a monocyclic heterocyclic group, wherein the monocyclic heterocyclic group contains a double bond; X, Y, Z, and Q are each independently selected from CR ^A or N atoms, and at most two atoms among X, Y, Z, and Q are N atoms at the same time; ^RA is selected from hydrogen, halogen, hydroxy, cyano, alkyl or alkoxy; wherein the alkyl or alkoxy is optionally further substituted with one or more substituents selected from halogen, hydroxy, cyano, alkyl or alkoxy; R ¹ is selected from the group consisting of absence, hydrogen, hydroxy, cyano, halogen, alkyl, alkoxy, cycloalkyl, heterocyclyl, aryl, heteroaryl, -C(O)R ³ , -C(O)NR ⁶ R ⁷ or -S(O) _r R ⁴ ; wherein the alkoxy, alkyl, cycloalkyl, heterocyclyl, aryl or heteroaryl is optionally further substituted by one or more R ^B ; R ³ and R ⁴ are each independently selected from a hydrogen atom, a hydroxyl group, an alkyl group, a cycloalkyl group, an aryl group, a heteroaryl group or a heterocyclic group; wherein the alkyl group, the cycloalkyl group, the aryl group, the heteroaryl group or the heterocyclic group is optionally further substituted by one or more substituents selected from hydroxyl, halogen, nitro, cyano, alkyl, alkoxy, haloalkyl, haloalkoxy, cycloalkyl, heterocyclic group, aryl, heteroaryl, =O, -C(O)R ⁸ , -C(O)OR ⁸ , -OC(O)R ⁸ , -NR ⁹ R ¹⁰ , -C(O)NR ⁹ R ¹⁰ , -SO ₂ NR ⁹ R ¹⁰ or -NR ⁹ C(O)R ¹⁰ ; R ^{and B} are each independently selected from cyano, halogen, alkyl, hydroxyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -OR ⁵ , -C(O)R ⁵ , -C(O)OR ⁵ , -NHC(O)R ⁵ , -NHC(O)OR ⁵ , -NR ⁶ R ⁷ , -C(O)NR ⁶ R ⁷ , -CH ₂ NHC(O)OR ⁵ , -CH ₂ NR ⁶ R ⁷ or -S(O) _r R ⁵ ; wherein the alkyl, cycloalkyl, heterocyclyl, aryl or heteroaryl is optionally further substituted by one or more selected from hydroxyl, halogen, nitro, cyano, alkyl, alkoxy, haloalkyl, haloalkoxy, cycloalkyl, heterocyclyl, aryl, heteroaryl, =O, -C(O)R ⁸ , -C(O)OR ⁸ , -OC(O)R ⁸ , -NR ⁹ R ¹⁰ , -C(O)NR ⁹ R ¹⁰ , _-SO2NR9R10 or ^-NR9C ( ^{O ) R10} ; Alternatively, two ^RBs are attached to the same carbon atom to form a -C(=O)-; ^R2 are the same or different and are independently selected from hydroxy, cyano, halogen, alkyl or alkoxy; wherein the alkyl or alkoxy is optionally further substituted with one or more substituents selected from halogen, hydroxy, cyano, alkyl or alkoxy; R ⁵ is independently selected from hydrogen atom, alkyl, cycloalkyl, heterocyclic, aryl or heteroaryl, wherein the alkyl, The cycloalkyl, heterocyclyl, aryl or heteroaryl group is optionally further substituted with one or more substituents selected from hydroxy, halogen, nitro, cyano, alkyl, alkoxy, haloalkyl, haloalkoxy, cycloalkyl, heterocyclyl, aryl ^, heteroaryl, =O, -C(O) ^R8 , ^-C (O ^{) OR8} , -OC(O) ^R8 , ^-NR9R10 , -C(O) ^NR9R10 , _-SO2NR9R10 or ^{-NR9C (} O) ^R10 ; ^R6 and ^R7 are each independently selected from a hydrogen atom, a hydroxyl group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein the alkyl group, the alkoxy group, the cycloalkyl group, the heterocyclic group, the aryl group or the heteroaryl group is optionally further substituted with one or more substituents selected from hydroxyl group, a halogen, a nitro group, a cyano group, an alkyl group, an alkoxy group, ^a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, =O, -C(O) ^R8 , -C(O)OR8, -OC(O) ^R8 , ^-NR9R10 , ^-C (O ⁾ _NR9R10 ^{, -SO2NR9R10} or ^-NR9C ( ^O ) ^R10 ; Alternatively, ^R6 and ^R7 together with the atoms to which they are attached form a 4- to 8-membered heterocyclic group, wherein the 4- to 8-membered heterocyclic group contains one or more N, O or S(O)r, and the ^4- to 8-membered heterocyclic group is optionally further substituted by one or more substituents selected from hydroxy, halogen, nitro, cyano, alkyl, alkoxy ^, cycloalkyl, heterocyclic group, aryl, heteroaryl, =O, -C(O) ^R8 , -C(O) ^OR8 , -OC(O) ^R8 , ^-NR9R10 , -C(O ⁾ _NR9R10 ^{, -SO2NR9R10} or ^-NR9C (O) ^R10 ; R ⁸ , R ⁹ and R ¹⁰ are each independently selected from a hydrogen atom, an alkyl group, an amino group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein the alkyl group, the cycloalkyl group, the heterocyclic group, the aryl group or the heteroaryl group is optionally further substituted with one or more substituents selected from a hydroxyl group, a halogen group, a nitro group, an amino group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, a carboxyl group or a carboxylate group; m is 0, 1 or 2; m is preferably 0; and r is independently 0, 1 or 2. The compound according to claim 1 or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, which is a compound or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof described by general formula (II):
Ring C, R ¹ , R ² , W and m are as defined in claim 1 . The compound according to claim 1 or 2, or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein W is N. The compound according to claim 1 or 2, or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein ring C is selected from:
The compound according to claim 1 or 2, or its stereoisomer, tautomer, or pharmaceutically acceptable salt thereof, wherein: R ¹ is selected from hydrogen atom, hydroxyl group, alkyl group, cycloalkyl group, heterocyclic group, -C(O)R ³ , -C(O)NR ⁶ R ⁷ or -S(O) _r R ⁴ ; wherein the alkyl group or cycloalkyl group is optionally further substituted by one or more R ^B ; R ^{and B} are each independently selected from cyano, hydroxy, heterocyclyl, aryl, heteroaryl, -OR ⁵ , -C(O)OR ⁵ , -NR ⁶ R ⁷ , -C(O)NR ⁶ R ⁷ or -S(O) _r R ⁵ ; wherein the heterocyclyl or heteroaryl is optionally further substituted by one or more substituents selected from hydroxy, cyano, alkoxy, haloalkyl, =O, -C(O)OR ⁸ or -C(O)NR ⁹ R ¹⁰ ; Alternatively, two ^RBs are attached to the same carbon atom to form a -C(=O)-; R ³ is selected from hydrogen atom, alkyl, cycloalkyl, heteroaryl or heterocyclic group; wherein the alkyl, cycloalkyl, heteroaryl or heterocyclic group is optionally further substituted by one or more substituents selected from hydroxyl, alkoxy, cyano, heteroaryl, =O, -NR ⁹ R ¹⁰ , -C(O)OR ⁸ or -C(O)NR ⁹ R ¹⁰ ; R ⁴ is a hydrogen atom or a hydroxyl group; R ⁵ is each independently selected from a hydrogen atom, an alkyl group or a hydroxyalkyl group; R ⁶ and R ⁷ are each independently selected from a hydrogen atom or an alkyl group, wherein the alkyl group is optionally substituted by one or more substituents selected from hydroxyl, cyano, halogen or alkoxy; R ⁸ , R ⁹ , and R ¹⁰ are each independently selected from a hydrogen atom or an alkyl group; the alkyl group is preferably a methyl group; r is 2. The compound according to any one of claims 1 to 5 or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein the compound is:






A pharmaceutical composition comprising an effective dose of the compound according to any one of claims 1 to 6 or its stereoisomer, tautomer or pharmaceutically acceptable salt, and a pharmaceutically acceptable carrier, excipient or a combination thereof. Use of the compound according to any one of claims 1 to 6 or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 7 in the preparation of an LPXC inhibitor. Use of the compound according to any one of claims 1 to 6 or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 7 in the preparation of a medicament for treating a disease mediated by LPXC, wherein the disease mediated by LPXC is selected from bacterial infections caused by Gram-negative bacteria. The use according to claim 9, wherein the Gram-negative bacteria is selected from Escherichia coli, Pseudomonas aeruginosa, Proteus, Shigella dysenteriae, Klebsiella pneumoniae, Brucella, Salmonella typhi, Acinetobacter, Yersinia, Legionella pneumophila, Bordetella pertussis, Shigella, Pasteurella, Vibrio cholerae or Neisseria meningitidis.