EA039977B1 - DIHYDROPYRIMIDINE COMPOUNDS AND METHOD FOR THEIR PRODUCTION AND APPLICATION - Google Patents

Description

Field of invention

The invention relates to a dihydropyrimidine compound having antiviral activity, a pharmaceutical composition containing it, a method for its preparation and its use in the prevention or treatment of viral diseases, including, but not limited to, viral hepatitis type A, viral hepatitis type B, viral hepatitis type C, influenza , herpes and acquired immunodeficiency syndrome (AIDS).

State of the art invention

A virus is made up of a nucleic acid molecule (DNA or RNA) and proteins, or is made up of only proteins (such as a prion). Viruses can cause various infectious diseases.

Common diseases caused by the virus include, but are not limited to, viral hepatitis type A, viral hepatitis type B, viral hepatitis C, influenza, herpes, and acquired immunodeficiency syndrome (AIDS).

Currently, antiviral drugs used in clinical practice act by inhibiting viral attachment and undressing, viral gene replication and maturation or release, or by affecting the host's immune system. These antiviral drugs mainly include reverse transcriptase inhibitors, protein capsid assembly inhibitors and the like.

Hepatitis B virus (HBV) is a common hepatophilic DNA viral pathogen. The virus can result in acute hepatitis, chronic hepatitis, liver fibrosis, liver cirrhosis, liver cancer, and the like.

Drugs for the treatment of type B hepatitis include interferon and nucleoside analogs (such as lamivudine and adenofir dipivoxil). Among them, interferon interacts with the cell surface receptor, ensuring the production of antiviral proteins by cells, thereby inhibiting the replication of the hepatitis B virus. Its disadvantages are a relatively low rate of effective reaction and the need for long-term injection. Nucleoside analogs act primarily by inhibiting the replication of the viral polymerase (reverse transcriptase). Their disadvantage is that these drugs require long-term use, which often results in virus mutation and leads to drug resistance.

In addition, type B viral hepatitis can be treated with non-nucleoside analogues. The heteroaryl dihydropyrimidine compound (Bay41-4109) discovered by Deres et al. can prevent HBV virus replication by inhibiting assembly of the viral capsid protein (Science, 2003, 299, 893-896). The specific mechanism of action is as follows: The dihydropyrimidine compound causes disrupted assembly of capsid proteins, resulting in unstable capsid proteins and accelerated degradation of capsid proteins (Biochem. Pharmacol., 2003, 66, 2273-2279).

The heteroaryl dihydropyrimidine compound HAP1 discovered by Zlotnick et al. (Proc. Natl. Acad. Sci., 2005, 102, 8138-8143), and a heteroaryl dihydropyrimidine compound (GLS4) reported by SUNSHINE LAKE PHARMA CO., LTD. (Antimicrob. Agents Chemother., 2013, 57, 5344-5354; WO 2015078391, US 2016206616 and WO 2015144093) also have anti-HBV activity.

Although the compounds above exhibit some degree of viral suppression, their antiviral activity is still not satisfactory. Moreover, some compounds also exhibit significant toxic effects (eg, GLS4 exhibits significant hERG cardiotoxicity).

The essence of the invention

Through intensive research, a dihydropyrimidine compound has been discovered. The dihydropyrimidine of the present invention is surprisingly more effective in inhibiting HBV DNA replication than the disclosed dihydropyrimidine modulators of HBV protein capsid assembly (e.g., at the cellular level, the antiviral activity of the preferred compounds of the present invention is 10 times that of the preferred compound in WO 2015144093 (compound of example 9) ). The compound of the present invention does not have the cardiotoxicity exhibited by the described dihydropyrimidine compounds (eg hERG inhibitory activity of GLS4 and the preferred compound in WO 2015144093 (compound of example 9)). Moreover, compared to the described dihydropyrimidine compounds (such as the compound of example 5 in WO 201403748), the compound of the present invention has a significantly reduced inductive effect on the CYP450 isoform 3A4. In addition, the compound of the present invention showed better pharmacokinetic properties (such as better exposure, blood drug concentration and bioavailability) in pharmacokinetic tests in rats, beagles and cynomolgus monkeys. However, the preferred compounds of the present invention have excellent liver targeting properties, and the degree of drug exposure in the liver can reach approximately 10-fold compared to plasma, indicating that the compounds have the ability to reach the liver, thereby contributing to an increase in efficacy in treatment of liver diseases.

One aspect of the present invention relates to a compound, or a stereoisomer or tautomer thereof, wherein the compound has the following structure:

where R ¹ represents C _1-6 alkyl;

R ² is H;

Q is -(CR ^a R ^a ')g- or -O-;

each R ^a and R ^a ' represent H;

R ⁴ and R ⁵ are H or halogen;

R ⁶ are H;

each R ^b , at each occurrence, is independently selected from the group consisting of H, C _1-6 haloalkyl, and C _1-6 alkyl;

each R ^c , at each occurrence, is independently selected from the group consisting of Cl, Br, C _1-6 haloalkyl, and C 1-6 alkyl;

R ⁶ is attached to the ring carbon atom(s) designated * or ** in the general formula;

g is 1;

i is equal to 1;

m is 0; and t is 1.

Another aspect of the present invention relates to a pharmaceutical composition having antiviral activity, comprising a prophylactically or therapeutically effective amount of a compound of the present invention, or a stereoisomer or tautomer thereof, and one or more pharmaceutically acceptable carriers, and the pharmaceutical composition is preferably a solid, liquid or transdermal formulation.

Another aspect of the present invention relates to a process for the preparation of a pharmaceutical composition comprising admixture of a compound of the present invention, or a stereoisomer or tautomer thereof, and one or more pharmaceutically acceptable carriers.

Another aspect of the present invention relates to the use of a compound of the present invention or a stereoisomer or tautomer or pharmaceutical composition of the present invention for the preparation of a medicament for the prevention or treatment of a viral disease.

The viral disease is selected from the group consisting of type A viral hepatitis, type B viral hepatitis, type C viral hepatitis, influenza, herpes, and acquired immunodeficiency syndrome (AIDS).

Brief description of the drawings

The figure shows the effect of compound 10-227, control compound 3 and rifampicin on the induction of CYP450 isoform 3A4.

Definitions.

Unless otherwise defined in the context, all technical and scientific terms used in the present invention have the same meanings as commonly known to a person skilled in the art. The methods used in the present invention refer to methods as is generally clear in the art, including variations on these methods or substitutions for equivalent methods that will be clear to a person skilled in the art. While it is believed that the following terms may be understood by a person skilled in the art, the following definitions are nonetheless used to better illustrate the present invention.

The terms contains, includes, has, or refers to, as well as other variations used in the present invention, are inclusive or non-limiting and do not exclude additional, non-listed elements or method steps.

As used in the present invention, the term alkylene refers to a saturated divalent hydrocarbyl, preferably refers to a saturated divalent hydrocarbyl containing 1, 2, 3, 4, 5 or 6 carbon atoms, such as methylene, ethylene, propylene or butylene.

As used in the present invention, the term alkenylene refers to a divalent hydrocarbyl containing one or more double bonds, preferably containing 2, 3, 4, 5 or 6 carbon atoms, such as vinylene, propenylene or allylidene. When the compound of the present invention

- 2 039977 the tenium contains an alkenylene group, the compound may exist as a pure E (on opposite sides) form, a pure Z (on one side) form, or any mixture thereof.

As used in the present invention, the term alkyl is defined as a linear or branched saturated aliphatic hydrocarbon. In some embodiments, the implementation of the alkyl contains 1-12, for example 1-6 carbon atoms. For example, as used in the present invention, the term C1.6 alkyl refers to a linear or branched group containing 1-6 carbon atoms (such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert -butyl, n-pentyl or nhexyl), which is optionally substituted with one or more (for example, 1-3) suitable substituents, such as halogen (in this case, the group can be called haloalkyl) (for example, CF3, C2F5, CHF ₂ , CH2F, CH2CF3, CH ₂ Cl or -CH ₂ CH ₂ Cf ₃ etc.). The term C _1-4 alkyl refers to a straight or branched aliphatic hydrocarbon chain containing 1-4 carbon atoms (ie methyl, ethyl, npropyl, isopropyl, n-butyl, isobutyl, sec-butyl or tert-butyl).

As used in the present invention, the term cycloalkyl refers to a saturated or unsaturated, non-aromatic monocyclic or polycyclic (e.g., bicyclic) hydrocarbon ring (e.g., monocyclic, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, or cyclononyl, or bicyclic, including spiro, fused or bridged ring system (such as bicyclo[1.1.1]pentyl, bicyclo[2.2.1]heptyl, bicyclo[3.2.1]octyl or bicyclo[5.2.0]nonyl, or decahydronaphthalene, etc.) ) which is optionally substituted with one or more (eg 1-3) suitable substituents. Cycloalkyl contains 3-15 carbon atoms. For example, the term _{C3_6} cycloalkyl refers to a saturated or unsaturated, non-aromatic monocyclic or polycyclic (e.g., bicyclic) hydrocarbon ring containing 3-6 carbon atoms forming a ring (e.g., cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl) that is optionally substituted with one or more (eg 1-3) suitable substituents, eg cyclopropyl substituted with methyl.

As used in the present invention, the term aryl refers to an all-carbon monocyclic or fused polycyclic aromatic group containing a conjugated π electron system. For example, as used in the present invention, the term C ₆ - ₁₄ aryl refers to an aromatic group containing 6-14 carbon atoms, such as phenyl or naphthyl. Aryl is optionally substituted with one or more (such as 1-3) suitable substituents (eg halogen, -OH, -CN, -NO ₂ , C1-6 alkyl, etc.).

The term aralkyl preferably means alkyl substituted with aryl, where aryl and alkyl are as defined in the present invention. Typically, an aryl group may contain 614 carbon atoms and an alkyl group may contain 1-6 carbon atoms. Examples of the aralkyl group include, but are not limited to, benzyl, phenylethyl, phenylpropyl, phenylbutyl.

As used in the present invention, the term heteroaryl refers to a monovalent monocyclic, bicyclic or tricyclic aromatic ring system containing 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14 ring atoms, in particular 1 or 2 or 3 or 4 or 5 or 6 or 9 or 10 carbon atoms, and containing at least one heteroatom (such as O, N, or S) which may be the same or different. Moreover, in some cases, it may be benzo-condensed. In particular, heteroaryl is selected from the group consisting of thienyl, furyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, triazolyl, thiadiazolyl, etc., and their benzo derivatives; or pyridinyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, etc., and their benzo derivatives.

As used in the present invention, the term halo or halogen is defined to include F, Cl, Br, or I.

As used in the present invention, the term alkylthio means an alkyl group, as defined above, connected to the main molecular moiety through a sulfur atom. Representative examples of C1-6 alkylthio include, but are not limited to, methylthio, ethylthio, t-butylthio, and hexylthio.

As used in the present invention, the term nitrogen-containing heterocyclic system refers to a saturated or unsaturated monocyclic or bicyclic group containing 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or 13 carbon atoms and at least at least one nitrogen atom in the ring, which may additionally optionally contain one or more (for example, one, two, three or four) ring members selected from the group consisting of N, O, C=O, S, S=O and S (=O) ₂ -. The nitrogen-containing heterocyclic system is attached to the rest of the molecule through a nitrogen atom. In particular, a 3-14 membered nitrogen-containing heterocyclic system is a group containing 3-14 carbon atoms and heteroatoms (wherein at least one is nitrogen) in the ring, including, but not limited to, a 3-membered nitrogen-containing heterocyclic system (such as aziridinyl), 4-membered nitrogen-containing heterocyclic system (such as azetidinyl), 5-membered nitrogen-containing heterocyclic system (such as pyrrolyl, pyrrolidinyl, pyrrolinyl, pyrrolidone, imidazolyl, imidazolidinyl, imidazolinyl, pyrazolyl, pyrazolinyl), 6-membered nitrogen-containing heterocyclic system system (such as piperidinyl, morpholinyl, thiomorpholinyl, piperazinyl), a 7-membered nitrogen-containing heterocyclic system, and the like.

- 3 039977

The term "substituted" means that one or more (e.g., one, two, three, or four) hydrogens at said atom are substituted by a selected group, provided that the normal valence of said atom under the circumstances is not exceeded and that the substitution results in a stable compound. The number of substituents chosen is only valid if these combinations result in stable compounds.

Where a substituent is described as being optionally substituted, the substituent may be either (1) unsubstituted or (2) substituted. Where a carbon of a substituent is described as being optionally substituted with one or more of the list of substituents, one or more of the hydrogens on that carbon may be substituted separately and/or together with an independently selected optional substituent. Where the nitrogen of a substituent is described as being optionally substituted with one or more of their list of substituents, each one or more of the hydrogens at that nitrogen may be replaced with an independently selected optional substituent.

If the substituents are described as being independently selected, each substituent may be identical to or different from the other substituent(s).

As used in the present invention, the term one or more means one or more than one (eg, 2, 3, 4, 5, or 10), as appropriate.

As used in the present invention, unless otherwise indicated, the point of attachment of a substituent may be at any suitable substituent position.

When a bond to a substituent is shown crossing the ring and the position of attachment is not indicated, then that substituent may be connected to any of the ring-forming atoms on that ring that can be substituted.

The present invention also includes all pharmaceutically acceptable isotopically labeled compounds which are identical to the compounds of the present invention except that one or more atoms are replaced by an atom having the same atomic number but an atomic mass or mass number different from the atomic mass or mass number that predominate in nature. Examples of isotopes suitable for inclusion in the compound of the present invention include, but are not limited to, hydrogen isotopes such as deuterium (d, ² H), tritium (t, ³ H); carbon, such as ^PS , ¹³ C and ¹⁴ C; chlorine such as ³⁶ Cl; fluorine such as ¹⁸ F; iodine, such as ¹²³ I and ¹²⁵ I; nitrogen, such as ¹³ N and 15N; oxygen such as ¹⁵ O, ¹⁷ O and ¹⁸ O; phosphorus such as ³² P; and sulfur, such as ³⁵ S. Certain isotopically labeled compounds of the present invention are useful in drug and/or substrate tissue distribution studies (eg, assays). Pharmaceutically acceptable solvates according to the present invention include solvates in which the crystallization solvent can be isotopically displaced, for example, D2O, acetone-d ₆ or DMSO-d ₅ .

The term stereoisomer refers to isomers with at least one asymmetric center. A compound containing one or more (eg, one, two, three or four) asymmetric centers can give rise to a racemic mixture, one enantiomer, a diastereomeric mixture, and a single diastereomer. Some individual molecules may exist as geometric isomers (cis/trans). Likewise, a compound of the present invention may exist as a mixture of two or more structurally distinct forms in rapid equilibrium (generally referred to as a tautomer). Typical examples of the tautomer include keto-enol tautomer, phenol-keto tautomer, nitroso-oxime tautomer, imine-enamine tautomer, and the like. For example, a dihydropyrimidine group may exist as the following tautomers in solution equilibrium:

It is clear that all these isomers and mixtures thereof in any ratio (such as 60, 65, 70, 75, 80, 85, 90, 95, 96, 97, 98 and 99%) are included within the scope of the present invention.

The carbon-carbon bonds of the compound of the present invention can be depicted in the present invention using a solid line ( )' a solid wedge ( , a dotted wedge ' ¹¹¹ ' or a wavy line

The use of a solid line to depict bonds to asymmetric carbons is intended to indicate that all possible stereoisomers (eg, specific enantiomers, racemic mixtures, etc.) at a given carbon are included. The use of a wavy line to depict bonds to an alkenyl group is intended to indicate that all possible stereoisomers are included (e.g., a particular cis-trans isomer, a mixture of cis and trans isomers in any ratio, or racemic mixtures, etc.) this chemical bond. The use of either a solid wedge or a dotted wedge to depict bonds to asymmetric carbons indicates that the stereoisomer shown is present. When present in racemic compounds, solid and dotted wedges are used to determine relative stereochemistry, not absolute stereochemistry. Unless otherwise indicated, the compound of the present invention may exist as stereoisomers,

- 4 039977 which include cis and trans isomers, optical isomers such as R and S enantiomers, diastereomers, geometric isomers, rotational isomers, conformational isomers, atropisomers and mixtures thereof. The compound of the present invention may exhibit more than one type of isomerism, and be composed of a mixture of them (such as racemates and diastereomeric pairs).

As used in the invention, the term approximately refers to a range within ±10%, preferably within ±5%, and more preferably within ±2% of the specified value.

Detailed description of the invention

Compound and method for its preparation.

In embodiments, the present invention relates to a compound or a stereoisomer or tautomer thereof, wherein the compound has the following structure:

where R ¹ represents C1-6 alkyl;

R ² is H;

Q is -(CR ^a R ^a ')g- or -O-;

each R ^a and R ^a ' represent H;

R ⁴ and R ⁵ are H or halogen;

R ⁶ are H;

each R ^b , at each occurrence, is independently selected from the group consisting of H, C _1-6 haloalkyl, and C ₁ . ₆ alkyl;

each R ^c , at each occurrence, is independently selected from the group consisting of Cl, Br, C _1-6 haloalkyl, and C _1-6 alkyl;

R ⁶ is attached to the ring carbon atom(s) designated * or ** in the general formula;

g is 1;

i is equal to 1;

m is 0; and t is 1.

In preferred embodiments, the present invention relates to a compound or a stereoisomer or tautomer thereof, wherein each R ^b , at each occurrence, is independently selected from the group consisting of H and C _1-6 alkyl;

each R ^c at each occurrence is independently selected from the group consisting of Cl, Br and C ₁ . ₆ alkyl.

In preferred embodiments, the present invention relates to a compound or a stereoisomer or tautomer thereof, wherein the compound has the following structure:

- 5 039977

In some embodiments, the present invention relates to a compound or a stereoisomer or tautomer thereof, wherein the compound is selected from the group consisting of:

- 6 039977

Pharmaceutical composition and use.

The present invention relates to a pharmaceutical composition having antiviral activity, comprising a prophylactically or therapeutically effective amount of a compound of the present invention, or a stereoisomer or tautomer thereof, and one or more pharmaceutically acceptable carriers or excipients.

The present invention further relates to a process for preparing a pharmaceutical composition comprising admixing a compound of the present invention, or a stereoisomer or tautomer thereof, and one or more pharmaceutically acceptable carriers or excipients.

The present invention relates to the use of a compound of the present invention, or a stereoisomer or tautomer thereof, or a pharmaceutical composition of the present invention, for the preparation of a medicament for the prevention or treatment of a viral disease.

The compound of the present invention exerts its antiviral effects by inhibiting the assembly of the protein capsid. As such, the compound of the present invention can be used to treat any viral disease involving protein capsid assembly when the virus infects the host, including, but not limited to, hepatitis type A virus (HAV), hepatitis type B virus (HBV), hepatitis type C virus. (HCV), influenza virus, herpes virus (HSV) and human immunodeficiency virus (HIV).

Thus, viral diseases that can be prevented and treated with the compound of the present invention include, but are not limited to, viral hepatitis type A, viral hepatitis type B, viral hepatitis type C, influenza, herpes, and acquired immunodeficiency syndrome (AIDS), and

- 7 039977 also related symptoms or diseases resulting from diseases above (eg inflammation, liver fibrosis, cirrhosis and liver cancer, etc.).

The term "pharmaceutically acceptable carrier" in the present invention refers to a diluent, excipient, excipient or medium with which a therapeutic agent is administered and is medically suitable for contact with tissues of human beings and animals without excessive toxicity, irritation, allergic reaction, or other problem or complication, according to an acceptable risk/benefit ratio.

A pharmaceutically acceptable carrier that can be used in the pharmaceutical composition of the present invention includes, but is not limited to, sterile liquids such as water and oils, including petroleum, animal, vegetable or synthetic oils such as peanut oil, soybean oil, mineral oil, sesame oil and the like. Water is an exemplary carrier when the pharmaceutical composition is administered intravenously. Physiological saline solutions, as well as aqueous solutions of dextrose and glycerol, can also be used as liquid carriers, in particular for injectable solutions. Suitable pharmaceutical excipients include starch, glucose, lactose, sucrose, gelatin, maltose, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, skimmed milk powder, glycerin, propylene glycol, water, ethanol, and the like. The pharmaceutical composition, if necessary, may also contain small amounts of wetting agent or emulsifier, or pH buffering agents. Oral formulations may include standard carriers such as mannitol, lactose, starch, magnesium stearate, sodium saccharin, cellulose, pharmaceutical grade magnesium carbonate, and the like. Examples of suitable pharmaceutical carriers are described, for example, in Remington's Pharmaceutical Sciences (1990).

The pharmaceutical composition of the present invention may act systemically and/or locally. In this regard, it can be administered by a suitable route, such as by injection (intravenous, intra-arterial, subcutaneous, intraperitoneal, intramuscular injection, including infusion) or transdermal administration, or administered orally, buccally, nasally, transmucosally, topically, as an ophthalmic formulation, or by inhalation . For these routes of administration, the administration can be carried out with a suitable dosage form.

These dosage forms include, but are not limited to, tablets, capsules, lozenges, lozenges, powders, sprays, creams, ointments, suppositories, gels, pastes, lotions, ointments, aqueous suspensions, injectables, elixirs, and syrups.

As used herein, the term effective amount refers to an amount of a compound administered that will alleviate, to some extent, one or more of the symptoms of the disease being treated.

Dosing regimens can be adjusted to provide the optimal response required. For example, a single administration may be performed, several divided doses may be administered over time, or the dose may be proportionally reduced or increased depending on the therapeutic situation. It should be noted that dosage values may vary depending on the type and severity of the disease and may include single or multiple doses. In addition, it is clear that for any particular subject, specific dosing regimens should be adjusted over time depending on individual needs.

The amount of a compound of the present invention administered will depend on the subject being treated, the severity of the disorder or disease, the frequency of administration, the pharmacokinetics of the compound, and the discretion of the attending physician. Typically, an effective dose is in the range of about 0.0001 to about 50 mg per kg of body weight per day, such as about 0.01 to about 10 mg/kg/day, in single or divided doses. For a 70 kg human, this would be an amount from about 0.007 mg to about 3500 mg/day, such as from about 0.7 mg to about 700 mg/day. In some cases, dose levels below the lower end of the above range may be more than sufficient, while in other cases, even larger doses can be used without causing any harmful side effect, provided that these large doses are first divided into several small doses to injections during the day.

The amount or dose of a compound of the present invention in a pharmaceutical composition is from about 0.01 mg to about 1000 mg, corresponding to 0.1-500 mg, preferably 0.5-300 mg, more preferably 1-150 mg, particularly preferably 1-50 mg , for example 1.5, 2, 4, 10, 25 mg, etc.

Unless otherwise indicated, the term treatment, as used herein, means reversing, alleviating, inhibiting a process, or preventing the disorder or disease to which the term is applied, or one or more of the symptoms of that disorder or disease.

As used in the present invention, the term subject includes a human or non-human animal. Examples of a human subject include a human subject suffering from a disease (such as the disease described in the present invention) (called a patient),

- 8 039977 or a normal subject. The term non-human animal as used herein includes all vertebrates, such as non-mammalian vertebrates (e.g., birds, amphibians, reptiles) and mammals, such as non-human primates, livestock, and/or domesticated animals. animals (such as sheep, dogs, cats, cows, pigs and the like).

In other embodiments, the pharmaceutical composition of the present invention may further comprise one or more additional therapeutic agents or prophylactic agents that are drugs for the treatment of viral hepatitis type B, including, but not limited to, lamivudine, telbivudine, entecavir, adefovir dipivoxil, tenofovir , tenofovir disoproxil fumarate and tenofovir alafenamide fumarate.

Examples

The invention is further described with reference to the following examples, which are not intended to limit the scope of the present invention in any way. Any suitable combination of conditions is possible.

Unless otherwise indicated, commercial anhydrous and HPLC grade solvents were used without further purification.

Ή NMR spectra were recorded at room temperature on a Bruker instrument (400 MHz) with TMS as internal standard. Chemical shifts (5) are given in ppm and binding constants (J) are given in hertz (Hz). The splitting multiplicity of the Ή NMR spectra is given using the following abbreviations: s (singlet), d (doublet), t (triplet), q (quartet), m (multiplet), br (broadened).

LC-MS was performed on an Aglient 1200 liquid chromatograph connected to an Aglient 6120 quadrupole mass spectrometer, with detection at 214 and 254 nm. Preparative liquid chromatography was performed on a SHIMADZU CBM-20A and Aglient 1260 preparative liquid chromatograph, C18 OBD 19x150 mm 5 μM preparative column, detection at 214 nm, where mobile phase A is

was water, mobile phase B was acetonitrile (with the addition of 0.5% formic acid slots), and the elution was carried out with a linear gradient in the following way: Time (min) A% IN% 0 90 10 15 40 60 thirty 10 90

Abbreviations as used in the present invention have the following meanings:

Reduction Meaning Reduction Meaning DAST Diethylaminosulfur trifluoride SPLA acetic acid DEA diethylamine IPA isopropanol EA ethyl acetate Meon methanol EtOH ethanol RE Petroleum ether NOT X hexane those thin layer chromatography

-9039977

Example 1 Preparation of ethyl 4-(2-chloro-4-fluorophenyl)-6-((3,3-difluoro-4-hydroxypiperidin-1yl)methyl)-2-(triaZol-2-yl)-1,4- dihydropyrimidine-5-carboxylate (10-88)

Stage 1: preparation of ethyl 4-(2-chloro-4-fluorophenyl)-6-methyl-2-(triazol-2-yl)-1,4-dihydropyrimidine-5-carboxylate (1-1).

At room temperature, ethyl acetoacetate (4.7 g, 36.0 mmol), triazole-2-carboximidamide hydrochloride (5.4 g, 36.0 mmol), 2-chloro-4-fluorobenzaldehyde (5.8 g, 36.0 mmol ) and potassium acetate (6.0 g, 60.0 mmol) were added to 2,2,2-trifluoroethanol (100 ml), heated to boiling, and the reaction was carried out for 16 hours. The reaction solution was cooled to room temperature, the solvent was distilled off at reduced pressure and the title compound (6.0 g) was obtained after working up. ESI-MS (m/z): 380.1 [M+H]+ .

Stage 2: preparation of ethyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triaZol-2-yl)-1,4-dihydropyrimidine-5-carboxylate (1-2).

Compound (1-1) (5.7 g, 15.0 mmol) was dissolved in carbon tetrachloride (60 ml) and heated to 50°C, N-bromosuccinimide (2.7 g, 15.0 mmol) was added in one portion and the reaction was carried out for 30 min. The reaction solution was cooled to room temperature, filtered to remove insolubles, and concentrated to give a crude product, which was purified to give the title compound (6.0 g). ESI-MS (m/z): 458.0 [M+H] ⁺ .

Stage 3: preparation of ethyl 4-(2-chloro-4-fluorophenyl)-6-((3,3-difluoro-4-hydroxypiperidin-1yl)methyl)-2-(triazol-2-yl)-1,4- dihydropyrimidine-5-carboxylate (10-88).

Compound (1-2) (68 mg, 0.14 mmol), 3,3-difluoropiperidin-4-ol hydrochloride (34 mg, 0.2 mmol) and N,N-diisopropylethylamine (50 mg, 0.2 mmol) at room temperature 4 mmol) was added to dichloromethane (3 ml) and the reaction was carried out at room temperature overnight. The reaction solution was concentrated to give a crude product, which was purified to give the title compound (10-88) (27 mg).

The structure was characterized in the following way:

1H NMR (400 MHz, DMSO-de): δ 9.54 (s, 1H), 7.99 (d, J=3.14 Hz, 1H), 7.94 (d, J=3.14 Hz, 1H), 7.44-7.40 (m, 2H), 7.19 (td, J=8.44, 2.64 Hz, 1H), 6.05 (s, 1H), 5.62 (t , J=5.40 Hz, 1H), 4.00-3.92 (m, 4H), 3.76 (s, 1H), 3.06-2.67 (m, 4H), 1.86 ( d, J=3.52 Hz, 1H), 1.74 (d, J=3.08 Hz, 1H), 1.04 (t, J=7.12 Hz, 3H). ESI-MS (m/z): 515.2 [M+H] ⁺ .

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Example 2 Preparation of ethyl 4-(2-bromo-4-fluorophenyl)-6-((3,3-difluoro-4-hydroxypiperidin-1yl)methyl)-2-(triazol-2-yl)-1,4- dihydropyrimidine-5-carboxylate (10-93)

in Example 1 (according to the reaction scheme above, using methods similar to those except that 2-chloro-4-fluorobenzaldehyde was replaced in step 1 with 2-bromo-4-fluorobenzaldehyde), the title compound (27 mg) was obtained.

The structure was characterized in the following way:

Ή NMR (400 MHz, DMSO-d6): δ 9.54 (s, 1H), 7.99 (d, J=3.14 Hz, 1H), 7.94 (d, J=3.14 Hz, 1H), 7.44-7.40 (m, 2H), 7.19 (td, J=8.44, 2.64 Hz, 1H), 6.05 (s, 1H), 5.62 (t , J=5.40 Hz, 1H), 4.00-3.92 (m, 4H), 3.76 (s, 1H), 3.06-2.67 (m, 4H), 1.86 ( d, J=3.52 Hz, 1H), 1.74 (d, J=3.08 Hz, 1H), 1.04 (t, J=7.12 Hz, 3H). ESI-MS (m/z): 559.2 [M+H]+.

Example 3 Preparation of (S)-ethyl 4-(2-chloro-4-fluorophenyl)-6-((3,3-difluoro-4-hydroxypiperidin-1yl)methyl)-2-(triazol-2-yl)- 1,4-dihydropyrimidine-5-carboxylate isomer A and (S)-ethyl 4-(2-chloro-4fluorophenyl)-6-((3,3-difluoro-4-hydroxypiperidin-1-yl)methyl)-2 -(triazol-2-yl)-1,4-dihydropyrimidine5-carboxylate isomer B

OH stage 4 _to isomer A + isomer B

Step 1: Separation of ethyl 4-(2-chloro-4-fluorophenyl)-6-methyl-2-(triazol-2-yl)-1,4-dihydropyrimidine-5-carboxylate (3-1).

Compound (3-1) (10 g) was separated by chiral chromatography using the following separation conditions: CHIRALPAK IE separation column 0.46 cm ID x 15 cm length, mobile phase:

MeOH/DEA=10 0/0.1 (v/v), flow rate: 1.0 ml/min, wavelength: UV 254 nm, temperature: 35°C.

Resolution gave (5)-ethyl 4-(2-chloro-4-fluorophenyl)-6-methyl-2-(triazol-2-yl)-1,4dihydropyrimidine-5-carboxylate (3-2) 4.7 g, e.i.%=99.9%, Rt=2.642 min. ESI-MS (m/z): 380.1 [M+H]+; And

- 11 039977 (R)-ethyl 4-(2-chloro-4-fluorophenyl)-6-methyl-2-(triazol-2-yl)-1,4-dihydropyrimidine-5-carboxylate (3-2') 4 .5 g, e.i.%=99.9%, Rt=4.783 min. ESI-MS (m/z): 380.1 [M+H]+.

Stage 2-stage 3.

(S)-Ethyl 4-(2-chloro-4-fluorophenyl)-6-((3,3-difluoro-4-hydroxypiperidin-1-yl)methyl)-2-(triazol-2yl)-1,4- dihydropyrimidine-5-carboxylate (3-4) was prepared using compound (3-2) as starting compound and using methods similar to those in Step 2 and Step 3 of Example 1. ESI-MS (m/z): 515.2 [M+N] ⁺ .

Stage 4. Separation of the connection (3-4).

Compound (3-4) (340 mg) was separated by chiral chromatography using the following separation conditions: CHIRALPAK IE separation column 0.46 cm ID x 15 cm length, mobile phase: HEX: 1PA=100/0.1 ( v/v), flow rate: 1.0 ml/min, wavelength: UV 254 nm, temperature: 35°C. Of these, the product with R _t =5.961 min was isomer A, ee%=99.3%, 122 mg, the structure was characterized by the following method:

Ή NMR (400 MHz, DMSO-d ₆ ): δ 9.54 (s, 1H), 7.99 (d, J=3.14 Hz, 1H), 7.94 (d, J=3.14 Hz , 1H), 7.44-7.40 (m, 2H), 7.19 (td, J=8.44, 2.64 Hz, 1H), 6.05 (s, 1H), 5.62 ( t, J=5.40 Hz, 1H), 4.00-3.92 (m, 4H), 3.76 (s, 1H), 3.06-2.67 (m, 4H), 1.86 (d, J=3.52 Hz, 1H), 1.74 (d, J=3.08 Hz, 1H), 1.04 (t, J=7.12 Hz, 3H). ESI-MS (m/z): 515.2 [M+H]+; And

Of these, the product with R _t =7.130 min was isomer B, ee%=99.5%, 131 mg, the structure was characterized by the following method:

1H NMR (400 MHz, DMSO-d ₆ ): δ 9.54 (s, 1H), 7.99 (d, J=3.14 Hz, 1H), 7.94 (d, J=3.14 Hz , 1H), 7.44-7.40 (m, 2H), 7.19 (td, J=8.44, 2.64 Hz, 1H), 6.05 (s, 1H), 5.62 ( t, J=5.40 Hz, 1H), 4.00-3.92 (m, 4H), 3.76 (s, 1H), 3.06-2.67 (m, 4H), 1.86 (d, J=3.52 Hz, 1H), 1.74 (d, J=3.08 Hz, 1H), 1.04 (t, J=7.12 Hz, 3H). ESI-MS (m/z): 515.2 [M+H] ⁺ .

Example 4 Preparation of (R)-ethyl 4-(2-chloro-4-fluorophenyl)-6-((3,3-difluoro-4-hydroxypiperidin-1yl)methyl)-2-(triazol-2-yl)- 1,4-dihydropyrimidine-5-carboxylate isomer A and (R)-ethyl 4-(2-chloro-4fluorophenyl)-6-((3,3-difluoro-4-hydroxypiperidin-1-yl)methyl)-2 -(triazol-2-yl)-1,4-dihydropyrimidine5-carboxylate isomer B

The desired product was obtained using compound (3-2') as the starting compound and using methods similar to those in Example 3. Of these, the product with Rt=8.171 min was isomer A, ee%=99.1%, 137 mg, the structure was characterized in the following way:

1H NMR (400 MHz, DMSO-d ₆ ): δ 9.54 (s, 1H), 7.99 (d, J=3.14 Hz, 1H), 7.94 (d, J=3.14 Hz , 1H), 7.44-7.40 (m, 2H), 7.19 (td, J=8.44, 2.64 Hz, 1H), 6.05 (s, 1H), 5.62 ( t, J=5.40 Hz, 1H), 4.00-3.92 (m, 4H), 3.76 (s, 1H), 3.06-2.67 (m, 4H), 1.86 (d, J=3.52 Hz, 1H), 1.74 (d, J=3.08 Hz, 1H), 1.04 (t, J=7.12 Hz, 3H). ESI-MS (m/z): 515.2 [M+H]+; and of these, the product with Rt=7.088 min was isomer B, ee%=99.4%, 128 mg, the structure was characterized by the following method:

1H NMR (400 MHz, DMSO-d6): δ 9.54 (s, 1H), 7.99 (d, J=3.14 Hz, 1H), 7.94 (d, J=3.14 Hz, 1H), 7.44-7.40 (m, 2H), 7.19 (td, J=8.44, 2.64 Hz, 1H), 6.05 (s, 1H), 5.62 (t , J=5.40 Hz, 1H), 4.00-3.92 (m, 4H), 3.76 (s, 1H), 3.06-2.67 (m, 4H), 1.86 ( d, J=3.52 Hz, 1H), 1.74 (d, J=3.08 Hz, 1H), 1.04 (t, J=7.12 Hz, 3H). ESI-MS (m/z): 515.2 [M+H] ⁺ .

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Example 5 Preparation of 2-((1-((6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6dihydropyrimidin-4-yl)methyl) -3,3-difluoropiperidin-4-yl)oxy)acetic acid (10-95)

Stage 1: obtaining benzyl 3,3-difluoro-4-hydroxypiperidine-1-carboxylate (5-2).

3,3-Difluoropiperidin-4-ol hydrochloride (5-1) (100 mg, 0.73 mmol) was dissolved in dichloromethane (2 ml), triethylamine (147 mg, 1.46 mmol) was added, K-(benzyloxycarbonyloxy) solution succinimide (12 mg, 0.48 mmol) in dichloromethane (2 ml) was added portionwise with ice bath cooling and the reaction was warmed to room temperature and held for 1 hour. The title compound 100 mg was obtained after workup. ESI-MS (m/z): 272.2 [M+H]+.

Stage 2: obtaining benzyl 4-(2-ethoxy-2-oxoethoxy)-3,3-difluoropiperidine-1-carboxylate (5-3).

Compound (5-2) (100 mg, 0.37 mmol) was dissolved in tetrahydrofuran (2 ml), sodium hydride (18 mg, 0.74 mmol) was added under cooling in an ice bath, and the reaction was warmed to room temperature and kept for 1 h. Then, the reaction was placed in an ice bath, a solution of ethyl bromoacetate (94 mg, 0.56 mmol) in tetrahydrofuran was added in portions, and the reaction was warmed to room temperature and stirred for an additional 5 h after completion of the batch addition. The solvent was distilled off under reduced pressure to give the title compound 150 mg. ESI-MS (m/z): 358.2 [M+H] ⁺ .

Step 3: Preparation of 2-((1-((benzyloxy)carbonyl)-3,3-difluoropiperidin-4-yl)oxy)acetic acid (5-4).

At room temperature, compound (5-3) (132 mg, 0.37 mmol) was dissolved in tetrahydrofuran (1 ml), sodium hydroxide (89 mg, 2.22 mmol in 0.3 ml H ₂ O) was added and the reaction was carried out at room temperature overnight. The reaction system was poured into water and the title compound 80 mg was obtained after workup. ESI-MS (m/z): 328.2 [M-H].

Step 4: Obtaining 2-((3,3-difluoropiperidin-4-yl)oxy)acetic acid (5-5).

At room temperature, compound (5-4) (80 mg, 0.24 mmol) was dissolved in methanol (2 ml), palladium-carbon (10%, 10 mg) was added, and the reaction was carried out under a hydrogen atmosphere at room temperature for 3 h. The insoluble material was filtered off and the solvent was distilled off under reduced pressure to give the title compound 40 mg. ESI-MS (m/z): 196.2 [M+H]+.

Step 5: Preparation of 2-((1-((6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6dihydropyrimidin-4-yl)methyl) -3,3-difluoropiperidin-4-yl)oxy)acetic acid (10-95).

The title compound 10 mg was prepared in a manner similar to that described in Step 3 of Example 1, except that 3,3-difluoropiperidin-4-ol hydrochloride was replaced with compound (5-5).

The structure was characterized in the following way:

1H NMR (400 MHz, DMSO-d6) δ 12.66 (s, 1H), 9.54 (d, J=2.4 Hz, 1H), 8.07-7.82 (m, 2H), 7 .53-7.33 (m, 2H), 7.18 (td, J=8.5, 2.7 Hz, 1H), 6.05 (s, 1H), 4.30-4.09 (m , 2H), 4.06-3.89 (m, 4H), 3.84 (s, 1H), 3.10-2.81 (m, 3H), 2.72 (s, 1H), 2, 00 (s, 1H), 1.85 (s, 1H), 1.04 (t, J=7.0 Hz, 3H). ESI-MS (m/z): 573.2 [M+H]+.

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Example 6 Preparation of (R)-ethyl 4-(2-chloro-4-fluorophenyl)-6-((3,3-difluoroazetidin-1-yl)methyl)-2(triazol-2-yl)-1,4 -dihydropyrimidine-5-carboxylate (10-7)

The title compound 4 mg was prepared in a manner analogous to that described in step 3 of Example 1 hydrochloride (3,3-difluoro-piperidin-4-ol was replaced by 3,3-difluoroazetidine hydrochloride).

The structure was characterized in the following way:

¹ H NMR (400 MHz, DMSO-d6) δ 9.46 (s, 1H), 8.06-7.99 (m, 1H), 7.95 (d, J=3.1 Hz, 1H), 7.46-7.36 (m, 2H), 7.19 (td, J=8.5, 2.6 Hz, 1H), 6.03 (s, 1H), 4.15 (s, 2H) , 3.97 (t, J=7.1 Hz, 2H), 3.85 (t, J=12.6 Hz, 4H), 1.05 (t, J=7.1 Hz, 3H). ESI-MS (m/z): 471.1 [M+H]+.

Example 7 Preparation of (R)-1-((6-(2-chloro-4-fluorophenyl)-5-ethoxycarbonyl-2-(triazol-2-yl)-3,6dihydropyrimidin-4-yl)methyl)-3 -fluoroazetidine-3-carboxylic acid (10-11)

Stage 1: obtaining 3-fluoroazetidine-3-carboxylic acid.

1-(tert-Butoxycarbonyl)-3-fluoroazetidine-3-carboxylic acid (30 mg, 0.14 mmol) was dissolved in dichloromethane (2 ml), trifluoroacetic acid (0.5 ml) was added at room temperature and the reaction was carried out for 1 hour. The solvent was distilled off under reduced pressure to give the trifluoroacetate salt of the title compound 33 mg. ESI-MS (m/z): 120.1 [M+H] ⁺ .

Step 2: Preparation of (R)-1-((6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6dihydropyrimidin-4-yl)methyl) -3-fluoroazetidine-3-carboxylic acid (10-11).

The title compound 6 mg was prepared in a manner analogous to that described in step 3 of Example 1 (3,3-difluoro-piperidin-4-ol hydrochloride was replaced by 3-fluoroazetidine-3-carboxylic acid trifluoroacetate).

The structure was characterized in the following way:

¹ H NMR (400 MHz, DMSO-d6) δ 9.47 (s, 1H), 8.01 (d, J=3.08 Hz, 1H), 7.95 (d, J=3.09 Hz, 1H), 7.40 (dd, J=15.82, 6, 61 Hz, 2H), 7.21-7.15 (m, 1n), 6.02 (s, 1H), 4.12 (s , 2H), 3.96 (dd, J=14.10, 7.12 Hz, 2H), 3.91-3.81 (m, 2H), 3.71 (dd, J=20.97, 9 .47 Hz, 2H), 1.05 (t, J=7.06 Hz, 3H). ESI-MS (m/z): 497.0 [M+H] ⁺ .

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Example 8 Preparation of (4R)-ethyl 4-(2-chloro-4-fluorophenyl)-6-((3,3-difluoro-4-methoxypiperidin-1yl)methyl)-2-(triazol-2-yl)- 1,4-dihydropyrimidine-5-carboxylate (10-98)

Stage 1: obtaining tert-butyl 3,3-difluoro-4-methoxypiperidine-1-carboxylate (8-2).

tert-Butyl 3,3-difluoro-4-hydroxypiperidine-1-carboxylate (8-1) (100 mg, 0.42 mmol) and tetrahydrofuran (5 ml) were added to a 25 ml flask and cooled to 0°C under nitrogen protection , sodium hydride (20 mg, 0.5 mmol) was added thereto, and the reaction was carried out for 30 minutes. Then, iodomethane (120 mg, 0.84 mmol) was added thereto, and the reaction was carried out for 16 hours. The reaction solution was slowly poured into water, and a crude product of the title compound 100 mg was obtained after work-up and used directly in the next reaction without purification. ESI-MS (m/z): 252.2 [M+H]+.

Step 2: Preparation of 3,3-difluoro-4-methoxypiperidine trifluoroacetate salt (8-3).

Compound (8-2) (100 mg, 0.4 mmol) and dichloromethane (6 ml) were added to a 25 ml flask, the reaction was cooled to 0°C, then trifluoroacetic acid (2 ml) was added thereto, and the reaction was carried out for 3 h. The solvent was distilled off under reduced pressure, obtaining a crude product of 120 mg. ESI-MS (m/z): 152.2 [M+H]+.

Stage 3: preparation of (4R)-ethyl 4-(2-chloro-4-fluorophenyl)-6-((3,3-difluoro-4-methoxypiperidin-1yl)methyl)-2-(triazol-2-yl)- 1,4-dihydropyrimidine-5-carboxylate (10-98).

The title compound 50 mg was prepared in a manner analogous to that described in Step 3 of Example 1 (3,3-difluoropiperidin-4-ol hydrochloride was replaced by compound (8-3)).

The structure was characterized in the following way:

¹ H NMR (400 MHz, DMSO-d6) δ 9.54 (s, 1H), 8.00 (dd, J=3.1, 1.7 Hz, 1H), 7.94 (d, J=3 .1 Hz, 1H), 7.477.35 (m, 2H), 7.18 (td, J=8.5, 2.6 Hz, 1H), 6.05 (s, 1H), 4.06-3 .91 (m, 4H), 3.60 (d, J=5.0 Hz, 1H), 3.41 (d, J=1.6 Hz, 3H), 3.12-2.93 (m, 1H), 2.92-2.76(m, 1H), 2.67(s, 1H), 1.94(s, 1H), 1.77(s, 1H), 1.04(t, J =7.1 Hz, 3H). ESI-MS (m/z): 529.2 [M+H]+.

Example 9 Preparation of (R)-ethyl 4-(2-chloro-4-fluorophenyl)-6-(((3R,4R)-3-fluoro-4-hydroxypyrrolidin1-yl)methyl)-2-(triazol-2 -yl)-1,4-dihydropyrimidine-5-carboxylate (10-34)

Stage 1: obtaining (3R,4R)-4-fluoropyrrolidin-3-ol (9-2).

While cooling in an ice bath, (3R,4R)-tert-butyl 3-hydroxy-4-fluoropyrrolidine-1-carboxylate (9-1) (60 mg, 0.3 mmol) was dissolved in dichloromethane (1 ml), trifluoroacetic acid was added (0.3 ml) and the reaction was carried out at room temperature for 1.5 hours. The solvent was distilled off under reduced pressure to give the trifluoroacetate salt of the title compound 60 mg. ESI-MS (m/z): 106.1 [M+H]+.

Stage 2: preparation of (R)-ethyl 4-(2-chloro-4-fluorophenyl)-6-(((3R,4R)-3-fluoro-4-hydroxypyrrolidin-1yl)methyl)-2-(triazole-2 -yl)-1,4-dihydropyrimidine-5-carboxylate (10-34).

The title compound 30 mg was prepared in a manner analogous to that described in Step 3 of Example 1 (3,3-difluoropiperidin-4-ol hydrochloride was replaced by compound (9-2)).

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The structure was characterized in the following way:

¹ H NMR (400 MHz, CDCl ₃ ) δ 7.86 (t, J=2.6 Hz, 1H), 7.59 (s, 2H), 7.49-7.33 (m, 1H), 7 .16 (ddd, J=8.3, 4.3, 2.6 Hz, 1H), 7.02 (d, J=9.0 Hz, 1H), 6.18 (d, J=2.3 Hz, 1H), 5.19 (d, J=50.5 Hz, 1H), 4.93-4.26 (m, 4H), 4.06 (kvd, J=7.1, 1.7 Hz , 2H), 3.53 (s, 2H), 1.10 (td, J=7.1, 4.5 Hz, 3H). ESI-MS (m/z): 483.2 [M+H]+.

Example 10 Preparation of 2-(((3R,4R)-1-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol2-yl)-3, 6-dihydropyrimidin-4-yl)methyl)-4-fluoropyrrolidin-3-yl)oxy)acetic acid (10-36)

Step 1: Preparation of (3R,4R)-tert-butyl 3-(2-ethoxy-2-oxoethoxy)-4-fluoropyrrolidine-1-carboxylate (10-b).

tert-Butyl (3R,4R)-3-hydroxy-4-fluoropyrrolidine-1-carboxylate (10-a) (100 mg, 0.49 mmol) was dissolved in tetrahydrofuran (5 ml), sodium hydride was added with cooling in an ice bath (40 mg, 60% in oil, 0.98 mmol) and the reaction was warmed to room temperature and held for 2 hours. While cooling in an ice bath, ethyl bromoacetate (124 mg, 0.74 mmol) was added and the reaction was carried out at room temperature in during the night. Saturated ammonium chloride (3 ml) was added, followed by dilution with dichloromethane (15 ml), the reaction was washed with saturated brine and dried over anhydrous sodium sulfate. The drying agent was removed by filtration, the solvent was distilled off under reduced pressure to give the title compound 120 mg. ESI-MS (m/z): 292.2 [M+H] ⁺ .

Step 2: Preparation of 2-(((3R,4R)-1-(tert-butoxycarbonyl)-4-fluoropyrrolidin-3-yl)oxy)acetic acid (10-c).

Compound (10-b) (120 mg, 0.41 mmol) was dissolved in a mixed solution of tetrahydrofuran and water (v:v=1:1, 3 ml), lithium hydroxide monohydrate (104 mg, 2.47 mmol) was added and the reaction was carried out at room temperature for 3.5 hours. The pH was adjusted to 5 with an aqueous citric acid solution, dichloromethane (15 ml) was added, and the reaction was washed with saturated brine and dried over anhydrous sodium sulfate. Anhydrous sodium sulfate was removed by filtration, the solvent was distilled off under reduced pressure to give the title compound 100 mg. ESI-MS (m/z): 263.2 [M+H]+.

Step 3: Preparation of 2-(((3R,4R)-4-fluoropyrrolidin-3-yl)oxy)acetic acid (10-d).

At room temperature, compound (10-c) (100 mg, 0.38 mmol) was dissolved in dichloromethane (1 ml), trifluoroacetic acid (0.3 ml) was added and the reaction was carried out at room temperature for 1.5 h. The solvent was distilled off under reduced pressure to give the trifluoroacetate salt of the title compound 60 mg. ESI-MS (m/z): 164.1 [M+H] ⁺ .

Stage 4: preparation of 2-(((3R,4R)-1-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2yl)-3, 6-dihydropyrimidin-4-yl)methyl)-4-fluoropyrrolidin-3-yl)oxy)acetic acid (10-36).

The title compound 13 mg was prepared in a manner similar to that described in Step 3 of Example 1 (3,3-difluoropiperidin-4-ol hydrochloride was replaced by compound (10-d)).

The structure was characterized in the following way:

1H NMR (400 MHz, DMSO-d6) δ 12.66 (s, 1H), 9.47 (d, J=12.6 Hz, 1H), 7.98 (dd, J=5.1, 3, 2 Hz, 1H), 7.93 (dd, J=3.2, 1.9 Hz, 1H), 7.41 (dd, J=9.5, 6.4, 3.0 Hz, 2H), 7.19 (tt, J=8.5, 2.0 Hz, 1H), 6.04 (d, J=1.4 Hz, 1H), 5.15 (d, J=51.7 Hz, 1H ), 4.40-3.77 (m, 9H), 3.12-2.87 (m, 2H), 1.04 (td, J=7.1, 2.1 Hz, 3H). ESI-MS (mz): 541.2 [M+H] ⁺ .

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Example 11 Preparation of (4R)-ethyl 4-(2-chloro-4-fluorophenyl)-6-((3,3-difluoro-4-hydroxypyrrolidin-1yl)methyl)-2-(triazol-2-yl)- 1,4-dihydropyrimidine-5-carboxylate (10-40)

Stage 1: obtaining 2,2-difluorovinyl p-methylbenzenesulfonate (11-2).

2,2,2-Trifluoroethyl p-methylbenzenesulfonate (11-1) (2.57 g, 10.0 mmol) was dissolved in tetrahydrofuran (15 ml), the reaction solution was cooled to -78° C. under nitrogen protection, and n -butyllithium (2.5M in hexane, 8 ml, 20 mmol). The reaction solution was stirred at -78° C. for 40 min, and water (4.5 g, 25 mmol) and tetrahydrofuran (10 ml) were slowly added in portions. The reaction solution was slowly warmed to room temperature. Water (60 ml) was added to the reaction and extracted with ethyl acetate (30 mlx2). The organic phase was collected, washed with saturated sodium chloride solution (50 ml x 2), dried over anhydrous sodium sulfate, filtered to remove anhydrous sodium sulfate, concentrated under reduced pressure, followed by purification by column chromatography to give the title compound 2.4 g. ESI-MS ( m/z): 235.1 [M+H]+.

Stage 2: obtaining 1-benzyl-4,4-difluoropyrrolidin-3-yl p-methylbenzenesulfonate (11-3).

Compound (11-2) (2340 mg, 10.0 mmol) and N-methoxymethyl-N-trimethylsilylbenzylamine (9500 mg, 40.0 mmol) were stirred at 130° C. in an oil bath for 5 min and trifluoroacetic acid ( 110 mg, 1.1 mmol). The reaction was stirred for an additional 1 hour, cooled to room temperature, diluted with ethyl acetate and purified to give the title compound 3.0 g. ESI-MS (m/z): 368.1 [M+H] ⁺ .

Stage 3: obtaining 1-benzyl-4,4-difluoropyrrolidin-3-ol (11-4).

Compound (11-3) (500 mg, 1.35 mmol) was dissolved in methanol (5 ml), magnesium turnings (326 mg, 13.6 mmol) were added and the reaction was stirred at room temperature for 2 h. Ice water ( 20 ml), concentrated hydrochloric acid was added in portions until all solids were dissolved. The reaction was extracted with ethyl acetate, the aqueous phase was collected, adjusted to pH 7 with saturated sodium hydroxide and extracted with ethyl acetate. The organic phase was collected, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered to remove anhydrous sodium sulfate, concentrated under reduced pressure, and the crude product was purified by preparative silica gel chromatography (petroleum ether/ethyl acetate=3/2) to give the title compound 240 mg. ESI-MS (m/z): 214.2 [M+H] ⁺ .

Stage 4: obtaining 4,4-difluoropyrrolidin-3-ol (11-5).

Compound (11-4) (100 mg, 0.47 mmol) was dissolved in methanol (2 ml), palladium-carbon (8 mg, 10% Pd, 0.047 mmol) was added and the reaction was stirred overnight at room temperature under a hydrogen atmosphere . The palladium-carbon was removed by filtration, and the reaction was concentrated under reduced pressure to give the title compound 50 mg. ESI-MS (m/z): 124.1 [M+H] ⁺ .

Stage 5: preparation of (4R)-ethyl 4-(2-chloro-4-fluorophenyl)-6-((3,3-difluoro-4-hydroxypyrrolidin-1yl)methyl)-2-(triazol-2-yl)- 1,4-dihydropyrimidine-5-carboxylate (10-40).

The title compound 33 mg was prepared in a manner analogous to that described in Step 3 of Example 1 (3,3-difluoropiperidin-4-ol hydrochloride was replaced by compound (11-5)).

The structure was characterized in the following way:

¹ H NMR (400 MHz, CDCl ₃ ) δ 9.36 (s, 1H), 7.84 (d, J=3.1 Hz, 1H), 7.48 (s, 1H), 7.31 (s , 1H), 7.14 (dt, J=8.5, 2.5 Hz, 1H), 6.94 (s, 1H), 6.20 (d, J=6.9 Hz, 1H), 4 .45-3.87 (m, 6H), 3.20 (s, 2H), 2.85 (s, 1H), 1.13 (td, J=7.1, 4.7 Hz, 3H). ESI-MS (m/z): 501.0 [M+H] ⁺ .

Example 12 Preparation of 2-((1-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6dihydropyrimidin-4- yl)methyl)-4,4-difluoropyrrolidin-3-yl)oxy)acetic acid (10-42)

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Stage 1: preparation of ethyl 2-((1-benzyl-4,4-difluoropyrrolidin-3-yl)oxy) acetate (12-2).

1-Benzyl-4,4-difluoropyrrolidin-3-ol (12-1) (100 mg, 0.47 mmol) was dissolved in tetrahydrofuran (5 ml), sodium hydride (40 mg 60% in oil, 0.94 mmol) was added with ice bath cooling and the reaction was warmed to room temperature and held for 2 hours. Ethyl bromoacetate (119 mg, 0.71 mmol) was added with ice bath cooling and the reaction was carried out at room temperature until the reaction was complete. Saturated ammonium chloride (3 ml) was added to the reaction, diluted with dichloromethane (15 ml), washed with saturated brine, and dried over anhydrous sodium sulfate. Anhydrous sodium sulfate was removed by filtration, and the solvent was distilled off under reduced pressure to give the title compound 123 mg, which was used directly in the next reaction without purification. ESI-MS (m/z): 300.1 [M+H]+.

Stage 2: obtaining 2-((1-benzyl-4,4-difluoropyrrolidin-3-yl)oxy)acetic acid (12-3).

Compound (12-2) (123 mg, 0.41 mmol) was dissolved in a mixed solution of tetrahydrofuran and water (v:v=1:1, 3 ml), lithium hydroxide monohydrate (104 mg, 2.47 mmol) was added and the reaction was carried out at room temperature for 3.5 hours. The reaction was adjusted to pH 5 with an aqueous citric acid solution, dichloromethane (15 ml) was added, washed with saturated brine, and dried over anhydrous sodium sulfate. The drying agent was removed by filtration, the solvent was distilled off under reduced pressure, and the crude product was purified by preparative chromatography to give the title compound 50 mg. ESI-MS (m/z): 272.1 [M+H] ⁺ .

Step 3: Obtaining 2-((4,4-difluoropyrrolidin-3-yl)oxy)acetic acid (12-4).

Compound (12-3) (50 mg, 0.18 mmol) was dissolved in methanol (2 ml), palladium-carbon (5 mg, 10% Pd, 0.018 mmol) was added and the reaction was stirred overnight at room temperature under a hydrogen atmosphere . The reaction was filtered to remove palladium-carbon and concentrated under reduced pressure to give the title compound 30 mg. ESI-MS (m/z): 182.1 [M+H] ⁺ .

Step 4: Preparation of 2-((1-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6dihydropyrimidin-4- yl)methyl)-4,4-difluoropyrrolidin-3-yl)oxy)acetic acid.

The title compound 25 mg was prepared in a manner similar to that described in Step 3 of Example 1 (3,3-difluoropiperidin-4-ol hydrochloride was replaced by compound (12-4)).

The structure was characterized in the following way:

¹ H NMR (400 MHz, CDCl ₃ ) δ 7.87 (dd, J=3.1, 1.3 Hz, 1H), 7.49 (s, 1H), 7.33 (t, J=7, 4 Hz, 1H), 7.14 (dd, J=8.5, 2.6 Hz, 1H), 6.96 (t, J=8.3 Hz, 1H), 6.19 (d, J= 2.3 Hz, 1H), 4.44 (d, J=16.3 Hz, 1H), 4.26 (dd, J=16.3, 2.3 Hz, 4H), 4.08-4, 01 (m, 2H), 3.43 (s, 4H), 1.12 (t, J=7.1 Hz, 3H). ESI-MS (m/z): 559.2 [M+H] ⁺ .

- 18 039977

Example 13 Preparation of (E)-3-(4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)3,6- dihydropyrimidin-4-yl)methyl)mofolin-3-yl)acrylic acid (10-182)

Stage 1: obtaining tert-butyl 3-(hydroxymethyl)morpholine-4-carboxylate (13-2).

Mofolin-3-ylmethanol hydrochloride (1.0 g, 6.5 mmol), triethylamine (1.64 g, 16 mmol) and dichloromethane (10 ml) were added to a 50 ml three-necked flask, stirred under nitrogen protection and cooled to 0° C, followed by the addition of di-tert-butyl dicarbonate (2.1 g, 10 mmol). After the addition, the reaction was warmed to room temperature and kept for 3 hours. The reaction solution was slowly poured into water and extracted with dichloromethane. The organic phase was collected, washed with saturated brine and dried over anhydrous sodium sulfate. Anhydrous sodium sulfate was removed by filtration, the solvent was distilled off under reduced pressure, and the title compound, 1.32 g, was obtained after purification. ESI-MS (m/z): 162.0 [M+H]+.

Stage 2: obtaining tert-butyl 3-formylmorpholine-4-carboxylate (13-3).

Compound (13-2) (500 mg, 2.3 mmol) and dichloromethane (10 ml) were added to a 50 ml three-neck flask, Dess-Martin reagent (1.3 g, 3.0 mmol) was added thereto at room temperature and the reaction was stirred for 3 hours. The reaction solution was slowly poured into aqueous sodium bicarbonate and extracted with dichloromethane. The organic phase was collected, washed with saturated brine, dried over anhydrous sodium sulfate, filtered to remove anhydrous sodium sulfate, and the solvent was distilled off under reduced pressure to give a crude product of 520 mg. ESI-MS (m/z): 160.0 [M+H] ⁺ .

Step 3: Preparation of (E)-tert-butyl 3-(3-ethoxy-3-oxoprop-1-en-1-yl)morpholine-4-carboxylate (13-4).

Triethylphosphonoacetate (600 mg, 3.3 mmol) and tetrahydrofuran (10 ml) were added to a 50 ml three-necked flask, stirred under nitrogen protection and cooled to 0° C., followed by addition of sodium hydride (320 mg, 6.6 mmol). After the addition, the reaction was carried out for 10 minutes and then a solution of compound (13-3) (520 mg, 3.3 mmol) in tetrahydrofuran (5 ml) was added in portions. After adding portionwise, the reaction was warmed to room temperature and kept for 16 hours. The reaction solution was slowly poured into water and extracted with ethyl acetate. The organic phase was collected, washed with saturated brine and dried over anhydrous sodium sulfate. The drying agent was removed by filtration, and the solvent was distilled off under reduced pressure to give a crude product of 620 mg. ESI-MS (m/z): 186.0 [M+H] ⁺ .

Step 4: Preparation of (E)-3-(4-(tert-butoxycarbonyl)mofolin-3-yl)acrylic acid (13-5).

At room temperature, compound (13-4) (620 mg, 2.2 mmol), anhydrous ethanol (10 ml) and water (10 ml) were added to a 50 ml flask, stirred, followed by the addition of sodium hydroxide (260 mg, 6 .6 mmol) and the reaction was carried out at room temperature for 4 hours. Then, the reaction was stopped by adding water (20 ml) thereto, subjected to rotary evaporation, removing ethanol and extracted with methyl tert-butyl ether. The aqueous phase was adjusted to pH 2-3 with 1N aqueous hydrochloric acid and then extracted with ethyl acetate. The organic phase was collected, washed with saturated brine and dried over anhydrous sodium sulfate. Anhydrous sodium sulfate was removed by filtration, and the solvent was distilled off under reduced pressure to give a crude product of 460 mg, which was used directly in the next reaction without purification. ESI-MS (m/z): 158.0 [M+H] ⁺ .

Step 5: Preparation of (E)-3-(mofolin-3-yl)acrylic acid (13-6).

A solution of 4N hydrochloric acid in dioxane (5 ml) was added to a 25 ml flask, cooled to 0°C, followed by portionwise addition of a solution of compound (13-5) (460 mg, 1.8 mmol) in ethyl acetate (5 ml) and the reaction was carried out at room temperature for 2 hours. The solvent was distilled off under reduced pressure to give the hydrochloride salt of the title compound 300 mg. ESI-MS

- 19 039977 (m/z): 158.0 [M+H] ⁺ .

Step 6: Preparation of (E)-3-(4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)3,6- dihydropyrimidin-4-yl)methyl)mofolin-3-yl)acrylic acid (10-182).

The title compound 47 mg was prepared in a manner analogous to that described in Step 3 of Example 1 (3,3-difluoropiperidin-4-ol hydrochloride was replaced by compound (13-6)).

The structure was characterized in the following way:

1H NMR (400 MHz, DMSO-d ₆ ) δ 12.47 (s, 1H), 9.69 (d, J=28.7 Hz, 1H), 8.11-7.91 (m, 2H), 7.53-7.27 (m, 2H), 7.23-7.10 (m, 1H), 6.66 (ddd, J=35.0, 15.8, 8.8 Hz, 1H), 6.11 (dd, J=28.6, 15.7 Hz, 1H), 6.03 (d, J=14.0 Hz, 1H), 4.02-3.85 (m, 4H), 3 .84-3.70 (m, 2H), 3.70-3.57 (m, 1H), 3.46-3.35 (m, 2H), 2.84 (dd, J=38.9, 12.6 Hz, 1H), 2.48-2.41 (m, 1H), 1.03 (t, J=7.1 Hz, 3H). ESI-MS (m/z): 535.1 [M+H] ⁺ .

Example 14 Preparation of (E)-3-(4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)3,6- dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-180)

According to the reaction scheme above, using methods similar to those in Example 13 and using mofolin-2-ylmethanol hydrochloride as the starting compound, the title compound (22 mg) was obtained.

The structure was characterized in the following way:

1H NMR (400 MHz, DMSO-d ₆ ) δ 12.40 (s, 1H), 9.64 (d, J=2.4 Hz, 1H), 8.02 (d, J=2.92 Hz, 2H), 7.95 (d, J=3.12 Hz, 2H), 7.21-7.16 (m, 1H), 6.83-6.72 (m, 1H), 6.05 (d , 1H), 6.00-5.91 (m, 1H), 4.27-4.22 (m, 1H), 4.003.85 (m, 5H), 3.70-3.61 (m, 1H ), 3.07-2.94 (m, 1H), 2.84-2.66 (m, 1H), 2.42-2.67 (m, 1H), 2.17-2.04 (m , 1H), 1.04 (t, J=7.0 Hz, 3H). ESI-MS (m/z): 535.1 [M+H]+.

Example 15 2-((4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6-dihydropyrimidin-4 -yl)methyl)mofolin-3 -yl)methoxy)acetic acid (10-162)

Stage 1: preparation of tert-butyl 3-((2-(tert-butoxy)-2-oxoethoxy)methyl)morpholine-4-carboxylate (15-2) tert-Butyl 3-(hydroxymethyl)morpholine-4-carboxylate (200 mg, 0.99 mmol) and tetrahydrofuran (6 ml) were added to a 50 ml three-necked flask, sodium hydride (47.3 mg, 1.2 mmol) was added thereto at room temperature, stirred for 30 min, followed by the addition of tert -butyl bromoacetate (192 mg, 0.99 mmol) and the reaction was carried out at room temperature for 3 hours. The reaction solution was slowly poured into 10 ml of water, adjusted to pH 2-3 with 1N aqueous hydrochloric acid and extracted with ethyl acetate. The organic phase was collected, washed with brine

- 20 039977 solution, dried over anhydrous sodium sulfate, filtered to remove anhydrous sodium sulfate and the solvent was distilled off under reduced pressure, obtaining a crude product of 300 mg. ESI-MS (m/z): 232.0 [M+H]+.

Stage 2: preparation of trifluoroacetate salt of 2-(mofolin-3-ylmethoxy)acetic acid (15-3).

Compound (15-2) (300 mg, 0.9 mmol) and dichloromethane (6 ml) were added to a 25 ml flask, cooled to 0°C, then trifluoroacetic acid (2 ml) was added thereto, and the reaction was carried out at room temperature in within 3 hours. The solvent was distilled off under reduced pressure to give a crude product of 250 mg. ESI-MS (m/z): 176.0 [M+H] ⁺ .

Step 3: Preparation of 2-((4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6dihydropyrimidin-4- yl)methyl)mofolin-3 -yl)methoxy)acetic acid (10-162).

The title compound 73 mg was prepared in a manner analogous to that described in Step 3 of Example 1 (3,3-difluoropiperidin-4-ol hydrochloride was replaced by compound (15-3)).

The structure was characterized in the following way:

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 12.70 (s, 1H), 9.79 (d, J=22.9 Hz, 1H), 8.01 (t, J=2.8 Hz , 1H), 7.93 (t, J=3.0 Hz, 1H), 7.50-7.37 (m, 2H), 7.18 (kvd, J=8.3, 2.6 Hz, 1H), 6.04 (d, J=1.4 Hz, 1H), 4.32 (dd, J=20.0, 17.6 Hz, 1H), 4.09-3.91 (m, 4H ), 3.87-3.63 (m, 3H), 3.62-3.50 (m, 3H), 3.46 (dd, J=11.2, 8.2 Hz, 1H), 2, 87-2.66 (m, 2H), 2.48-2.39 (m, 1H), 1.05 (td, J=7.0, 1.5 Hz, 3H). ESI-MS (m/z): 553.1 [M+H] ⁺ .

Example 16 Preparation of 2-((1-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6dihydropyrimidin-4- yl)methyl)-5,5-difluoropiperidin-3-yl)methoxy)acetic acid (10-136)

Stage 1: obtaining 1-tert-butyl 3-methyl 5-oxo-piperidine-1,3-dicarboxylate (16-2).

At room temperature, 1-tert-butyl 3-methyl 5-hydroxypiperidine-1,3-dicarboxylate (16-1) (1.0 g, 3.86 mmol) was dissolved in dichloromethane (20 ml), the reaction was cooled to 0° After completion of dissolution, Dess-Martin reagent (3.27 g, 7.71 mmol) was added with stirring, and the reaction was stirred overnight at room temperature after the addition. A large amount of a white solid precipitated into the reaction solution, which was filtered, and the filtrate was washed successively with water (50 ml) and saturated aqueous sodium carbonate (50 ml). The organic phase was dried over anhydrous sodium sulfate and the solvent was distilled off under reduced pressure to give the title compound 1.0 g. ESI-MS (m/z): 202.1 [M+H-56]+.

Stage 2: obtaining 1-tert-butyl 3-methyl 5,5-difluoropiperidine-1,3-dicarboxylate (16-3).

Compound (16-2) (1.0 g, 3.89 mmol) was dissolved in dichloromethane (20 ml), the reaction was cooled to -70°C after dissolving with stirring, DAST (1.9 g, 11.67 mmol) and the reaction was warmed to room temperature and kept for 4.5 h after addition in portions. After complete reaction of the starting compound monitored by LC-MS, the reaction solution was quenched with saturated aqueous sodium bicarbonate (20 ml), extracted with dichloromethane, dried and purified to give the title compound 550 mg. ESI-MS (m/z): 224.1 [M+H-56]+.

Stage 3: obtaining tert-butyl 3,3-difluoro-5-(hydroxymethyl)piperidine-1-carboxylate (16-4).

At room temperature, compound (16-3) (500 mg, 1.79 mmol) was added to methanol (8 ml), the reaction was cooled to 0° C. and sodium borohydride (272 mg, 7.16 mmol) was slowly added in portions. After the addition, the reaction was carried out at room temperature overnight, incomplete reaction of the starting compound was monitored by LC-MS, sodium borohydride (136 mg, 3.58 mmol) was added and the reaction was continued overnight until complete reaction of the starting compound. The reaction was quenched by adding water (20 ml) and extracted with ethyl acetate. The organic phase was combined, washed with water, dried over anhydrous sodium sulfate, and the solvent was distilled off under reduced pressure to give the title compound 500 mg. ESI-MS (m/z): 196.1 [M+H-56]+.

Stage 4: preparation of tert-butyl 5-((2-(tert-butoxy)-2-oxoethoxy)methyl)-3,3-difluoropiperidine-1

- 21 039977 carboxylate (16-5).

At room temperature, compound (16-4) (150 mg, 0.60 mmol) was added to tetrahydrofuran (4 ml), the reaction was cooled to 0° C. and sodium hydride (48 mg, 1.2 mmol) was added slowly. After the addition, the reaction was stirred at 0°C for 30 min and then a solution of tert-butyl bromoacetate (140 mg, 0.72 mmol) in tetrahydrofuran (1.0 ml) was added. The reaction was carried out at room temperature overnight, incomplete reaction of the starting compound was monitored by LC-MS, tert-butyl bromoacetate (35 mg, 0.18 mmol) was added and the reaction was continued overnight until complete reaction of the starting compound. The reaction solution was quenched by adding a saturated aqueous ammonium chloride solution (5 ml), diluted with water (20 ml) and extracted with ethyl acetate. The organic phase was combined, washed with water, dried and purified to give the title compound 120 mg. ESI-MS (m/z): 254.1 [M+H-112]+.

Step 5: Obtaining 2-((5,5-difluoropiperidin-3-yl)methoxy)acetic acid (16-6).

At room temperature, compound (16-5) (60 mg, 0.16 mmol) was added to dichloromethane (3 ml), the reaction was cooled to 0° C. and trifluoroacetic acid (1.0 ml) was added. After the addition, the reaction was carried out at room temperature for 1 h, the incomplete reaction of the starting compound was monitored by LC-MS, then the reaction was continued overnight until the complete reaction of the starting compound. The solvent was distilled off under reduced pressure to give the trifluoroacetate salt of the title compound 60 mg. ESI-MS (m/z): 210.1 [M+H] ⁺ .

Step 6: Preparation of 2-((1-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6dihydropyrimidin-4- yl)methyl)-5,5-difluoropiperidin-3-yl)methoxy)acetic acid (10-136).

The title compound 19 mg was prepared in a manner analogous to that described in Step 3 of Example 1 (3,3-difluoropiperidin-4-ol hydrochloride was replaced by compound (16-5)).

The structure was characterized in the following way:

1H NMR (400 MHz, chloroform-d) δ 7.93 (dd, J=9.67, 3.06 Hz, 1H), 7.54 (s, 1H), 7.46-7.31 (m, 1H), 7.13 (ddd, J=8.55, 2.63, 1.35 Hz, 1H), 6.96 (d, J=8.50 Hz, 1H), 6.24 (d, J =3.93 Hz, 1H), 4.28 (d, J=16.74 Hz, 1H), 4.14 (d, J=16.81 Hz, 1H), 3.94 (t, J=16 .64 Hz, 2H), 3.80 (t, J=8.68 Hz, 1H), 3.65 (t, J=8.64 Hz, 1H), 3.56-3.43 (m, 1H ), 2.94 (d, J=53.06 Hz, 4H), 2.65 (s, 1H), 2.45 (s, 1H), 2.13 (dd, J=26.84, 13, 90 Hz, 1H), 1.92 (d, J=9.36 Hz, 1H), 1.13 (td, J=7.10, 3.76 Hz, 3H). ESI-MS (m/z): 587.2 [M+H] ⁺ .

Example 17 Preparation of (E)-3-(1-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)3,6- dihydropyrimidin-4-yl)methyl)-5,5-difluoropiperidin-3-yl)acrylic acid (10-116)

Stage 1: obtaining tert-butyl 3,3-difluoro-5-formylpiperidine-1-carboxylate (17-2)

At room temperature, tert-butyl 3,3-difluoro-5-(hydroxymethyl)piperidine-1-carboxylate (17-1) (50 mg, 0.2 mmol) was dissolved in dichloromethane (2.0 ml), the reaction was cooled to 0 °C after complete dissolution, the Dess-Martin reagent (102 mg, 0.24 mmol) was added with stirring, and the reaction was carried out at room temperature for 3 hours after the addition. A large amount of a white precipitate precipitated into the reaction solution, which was filtered, and the filtrate was evaporated under reduced pressure to give the title compound 50 mg.

Step 2: Preparation of (E)-tert-butyl 5-(3-ethoxy-3-oxoprop-1-en-1-yl)-3,3-difluoropiperidine-1-carboxylate (17-3).

At room temperature, compound (17-2) (50 mg, 0.2 mmol) was dissolved in dichloromethane (2.0 ml), (carbethoxymethylene)triphenylphosphorane (70 mg, 0.2 mmol) was added with stirring after complete dissolution, and the reaction was carried out at room temperature overnight after addition. The reaction solution was purified to give the title compound 30 mg. ESI-MS (m/z): 264.1 [M+H-56]+.

Step 3: Preparation of (E)-3-[1-(tert-butoxycarbonyl)-5,5-difluoropiperidin-3-yl)acrylic acid

- 22 039977 you (17-4).

At room temperature, compound (17-3) (30 mg, 0.1 mmol) was dissolved in tetrahydrofuran (4.0 ml) and water (2 ml), lithium hydroxide (19 mg, 0.4-7 mmol) was added with stirring after complete dissolution and the reaction was carried out at room temperature for 4 hours after the addition. The reaction solution was diluted with water, adjusted to pH 3-4 with 1N hydrochloric acid, and extracted with ethyl acetate. The organic phase was combined, washed with water and dried to give the title compound 30 mg.

Step 4: Preparation of (E)-3-(5,5-difluoropiperidin-3-yl)acrylic acid trifluoroacetate salt (17-5).

At room temperature, compound (17-4) (30 mg, 0.1 mmol) was added to dichloromethane (1.5 ml), cooled to 0° C., followed by addition of trifluoroacetic acid (0.5 ml). After the addition, the reaction was carried out at room temperature for 1.5 hours. The solvent in the reaction solution was distilled off under reduced pressure to give the title compound 30 mg. ESI-MS (m/z): 192.1 [M+H]+.

Step 5: Preparation of (E)-3-(1-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)3,6- dihydropyrimidin-4-yl)methyl)-5,5-difluoropiperidin-3-yl)acrylic acid (10-116).

The title compound 10 mg was prepared in a manner analogous to that described in Step 3 of Example 1 (3,3-difluoropiperidin-4-ol hydrochloride was replaced by compound (17-5)).

The structure was characterized in the following way:

¹ H NMR (400 MHz, chloroform-d) δ 7.90 (s, 1H), 7.36 (s, 1H), 7.14 (dt, J=8.53, 2.80 Hz, 1H), 7.02-6.86 (m, 2H), 6.25 (d, J=8.50 Hz, 1H), 5.90 (dd, J=23.97, 15.72 Hz, 1H), 4 .20 (d, J=15.93 Hz, 1H), 4.04 (q, J=5.70, 5.05 Hz, 3H), 3.11 (d, J=62.19 Hz, 3H) , 2.71 (s, 2H), 2.36 (s, 2H), 1.14 (td, J=7.06, 5.31 Hz, 3H). ESI-MS (m/z): 569.2 [M+H] ⁺ .

Example 18 Preparation of 2-((1-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6dihydropyrimidin-4- yl)methyl)-4,4-difluoropyrrolidin-2-yl)methoxy)acetic acid (10-56)

Stage 1: preparation of tert-butyl 2-((2-ethoxy-2-oxoethoxy)methyl)-4,4-difluoropyrrolidine-1-carboxylate (18-2).

tert-Butyl 4,4-difluoro-2-(hydroxymethyl)pyrrolidine-1-carboxylate (18-1) (80 mg, 0.34 mmol) was dissolved in tetrahydrofuran (5 ml), sodium hydride (27 mg 60% in oil, 0.68 mmol) while cooling in an ice bath and the reaction was warmed to room temperature and held for 2 hours. While cooling in an ice bath, ethyl bromoacetate (85 mg, 0.51 mmol) was added and the reaction was carried out at room temperature for 4 hours Saturated ammonium chloride (3 ml) was added to the reaction, diluted with dichloromethane (15 ml), washed with saturated brine and dried over anhydrous sodium sulfate. The title compound 100 mg was obtained after workup and used directly in the next reaction without purification.

Stage 2: obtaining 2-((1-(tert-butoxycarbonyl)-4,4-difluoropyrrolidin-2-yl)methoxy)acetic acid (18-3).

Compound (18-2) (100 mg, 0.31 mmol) was dissolved in a mixed solution of tetrahydrofuran and water (v:v=1:1, 3 ml), lithium hydroxide monohydrate (45 mg, 1.86 mmol) was added and the reaction was carried out at room temperature for 3.5 hours. The reaction was adjusted to pH 5 with an aqueous citric acid solution, dichloromethane (15 ml) was added, washed with saturated brine and dried over anhydrous sodium sulfate and the title compound 80 mg was obtained after working up .

Step 3: Obtaining 2-((4,4-difluoropyrrolidin-2-yl)methoxy)acetic acid (18-4).

At room temperature, compound (18-3) (80 mg, 0.27 mmol) was dissolved in dichloromethane (1 ml), trifluoroacetic acid (0.3 ml) was added and the reaction was carried out at room temperature for 1.5 hours. The solvent was distilled off under reduced pressure to give the trifluoroacetate salt of App.

- 23 039977 title compound 60 mg.

Step 4: Preparation of 2-((1-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6dihydropyrimidin-4- yl)methyl)-4,4-difluoropyrrolidin-2-yl)methoxy)acetic acid (10-56).

The title compound 25 mg was prepared in a manner analogous to that described in Step 3 of Example 1 (3,3-difluoropiperidin-4-ol hydrochloride was replaced by compound (18-4)).

The structure was characterized in the following way:

1H NMR (400 MHz, DMSO-d6) δ 12.52 (s, 1H), 9.54 (s, 1H), 7.96 (dd, J=27.4, 3.2 Hz, 2H), 7 .51-7.27 (m, 2H), 7.19 (td, J=8.5, 2.8 Hz, 1H), 6.01 (d, J=12.5 Hz, 1H), 4, 21 (d, J=2.7 Hz, 1H), 4.01-3.84 (m, 3H), 3.66-3.44 (m, 3H), 3.34-3.26 (m, 4H), 3.14-2.96 (m, 1H), 2.33-2.07 (m, 1H), 1.08-0.98 (m, 3H). ESI-MS (m/z): 573.2 [M+H]+.

Example 19 Preparation of 2-((4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6dihydropyrimidin-4- yl)methyl)mofolin-2-yl)methoxy)acetic acid (10-160)

Stage 1: obtaining tert-butyl 2-(hydroxymethyl)morpholine-4-carboxylate (19-2).

Mofolin-2-ylmethanol hydrochloride (19-1) (500 mg, 3.3 mmol), triethylamine (0.82 g, 8 mmol) and dichloromethane (10 ml) were added to a 50 ml three-necked flask, stirred under nitrogen protection, cooled to 0°C and then added di-tert-butyl dicarbonate (1.1 g, 5 mmol). After the addition, the reaction was carried out at room temperature for 3 hours. The reaction solution was slowly poured into water and extracted with dichloromethane (30 ml×3). The organic phase was collected, washed with saturated brine and dried over anhydrous sodium sulfate. Anhydrous sodium sulfate was removed by filtration, the solvent was distilled off under reduced pressure, and the title compound 0.6 g was obtained after purification. ESI-MS (m/z): 162.0 [M+H] ⁺ .

Stage 2-stage 4.

The title compound 70 mg was obtained from compound (19-2) using methods similar to those in Step 1-Step 3 of Example 15.

The structure was characterized in the following way:

1H NMR (400 MHz, DMSO-d6) δ 12.60 (s, 1H), 9.66 (d, J=2.7 Hz, 1H), 8.07-7.90 (m, 2H), 7 .49-7.35 (m, 2H), 7.18 (td, J=8.5, 2.5 Hz, 1H), 6.05 (s, 1H), 4.04 (s, 1H), 4.01-3.80 (m, 6H), 3.68 (s, 1H), 3.63-3.43 (m, 3H), 2.78 (ddd, J=56.5, 46.6 , 11.4 Hz, 2H), 2.41-2.25 (m, 1H), 2.15 (dt, J=31.9, 10.6 Hz, 1H), 1.05 (t, J= 7.0 Hz, 3H). ESI-MS (m/z): 553.1 [M+H] ⁺ .

- 24 039977

Example 20 Preparation of N-((4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2-yl)-3,6dihydropyrimidin-4- yl)methyl)mofolin-2-yl)methyl)-N-methylglycine (10-168)

Stage 1: preparation of tert-butyl 2-(((2-methoxy-2-oxoethyl)(methyl)amino)methyl)morpholine-4carboxylate (20-2).

tert-Butyl 2-formylmorpholine-4-carboxylate (20-1) (117 mg, 0.5 mmol) and sarcosine methyl ester hydrochloride (84 mg, 0.6 mmol) were dissolved in methanol (3 ml), glacial acetic acid was added (0.2 ml) while cooling in an ice bath, then sodium cyanoborohydride (76 mg, 1.2 mmol) was added in portions and the reaction was warmed to room temperature and kept for 2 hours. Ethyl acetate (20 ml) was added to the reaction, stirred in for 10 min, filtered to remove insolubles and the filtrate was concentrated to give the title compound 98 mg. ESI-MS (m/z): 303.2 [M+H]+.

Step 2: Preparation of N-((4-(tert-butoxycarbonyl)mofolin-2-yl)methyl)-N-methylglycine (20-3).

Compound (20-2) (98 mg, 0.3 mmol) was dissolved in a mixed solvent of methanol and water (v:v=1:1, 4 ml), lithium hydroxide monohydrate (84 mg, 2 mmol) was added, and reaction stirred overnight. The pH was adjusted to 2 with 1N hydrochloric acid, and the solvent was distilled off under reduced pressure to give the title compound 80 mg, which was used directly in the next reaction without purification. ESI-MS (m/z): 289.2 [M+H] ⁺ .

Step 3: Obtaining N-methyl-N-((mofolin-2-ylmethyl)glycine (20-4).

At room temperature, compound (20-3) (80 mg, 0.28 mmol) was dissolved in dichloromethane (3 ml), trifluoroacetic acid (1 ml) was added, and the reaction was carried out at room temperature for 3 hours. The solvent was distilled off under reduced pressure, giving the trifluoroacetate salt of the title compound 94 mg. ESI-MS (m/z): 189.2 [M+H] ⁺ .

Step 4: Preparation of N-((4-(((R)-6-(2-chloro-4-fluorophenyl)-5-ethoxycarbonyl-2-(triazol-2-yl)-3,6dihydropyrimidin-4-yl) methyl)mofolin-2-yl)methyl)-N-methylglycine (10-168).

The title compound 9 mg was prepared in a manner analogous to that described in Step 3 of Example 1 (3,3-difluoropiperidin-4-ol hydrochloride was replaced by compound (20-4)).

The structure was characterized in the following way:

¹ H NMR (400 MHz, DMSO-d6) δ 9.66 (d, 1H), 8.03-8.01 (m, 1H), 7.95-7.94 (m, 1H), 7.44 -7.39(m, 2H), 7.22-7.17(m, 1H), 6.05(d, 1H), 3.99-3.82(m, 5H), 3.70-3 .68 (m, 1H), 3.63-3.54 (m, 1H), 3.25 (d, 1H), 3.16 (d, 1H), 2.92-2.56 (m, 4H ), 2.34 (d, 3H), 2.35-2.24 (m, 1H), 2.14-1.98 (m, 1H), 1.04 (t, J=7.1 Hz, 3H). ESI-MS (m/z): 566.1 [M+H] ⁺ .

- 25 039977

Example 21 Preparation of 2-((1-(((S)-5-(ethoxycarbonyl)-6-(4-fluorophenyl)-6-methyl-2-(triazol-2-yl)3,6-dihydropyrimidin-4 -yl)methyl)-3,3-difluoropiperidin-4-yl)oxy)acetic acid (10-224)

Step 1: Preparation of (E)-ethyl 2-acetyl-3-(4-fluorophenyl)but-2-enoate.

At room temperature, ethyl acetoacetate (3.12 g, 24.0 mmol), 4-fluorophenylethynyl (2.88 g, 24.0 mmol) and indium triflate (216 mg, 0.384 mmol) were added to o-xylene (15 ml), the reaction was heated to 120°C and maintained at this temperature for 2 h, LC-MS showed the completion of the reaction. The reaction was cooled to room temperature and the solvent was distilled off under reduced pressure to give a crude product of 6.0 g. ESI-MS (m/z): 251.1 [M+H]+.

Stage 2: preparation of ethyl 4-(4-fluorophenyl)-4,6-dimethyl-2-(triazol-2-yl)-1,4-dihydropyrimidine-5-carboxylate.

Triazole-2-carboximidamide hydrochloride (3.03 g, 18.5 mmol), sodium bicarbonate (3.15 g, 37.5 mmol) was added to N-methylpyrrolidone (40 ml), the reaction was heated to 120°C, added in portions compound (21-1) (3.12 g, 12.5 mmol) and kept for 1 h, LC-MS showed completion of the reaction. The reaction was cooled to room temperature, ethyl acetate (60 ml) was added, washed with water and brine, and dried over anhydrous sodium sulfate. Anhydrous sodium sulfate was removed by filtration, and the solvent was distilled off under reduced pressure to give a crude product. The crude product was purified by flash silica gel column chromatography (petroleum ether:ethyl acetate=10:1) to give the title compound as a yellow solid (2.11 g). The product above 350 mg was separated by chiral chromatography using the following separation conditions: separation column: CHIRALPAK 1C 0.46 cm ID x 15 cm length, mobile phase: hexane/IPA/DEA=90/10/0.1 (vol. /vol.) / flow rate: 1.0 ml/min, wavelength: UV 254 nm, temperature: 35°C.

The following products were obtained by separation: (S)-ethyl 4-(4-fluorophenyl)-4,6-dimethyl-2-(triazol-2yl)-1,4-dihydropyrimidine-5-carboxylate (21-2) 172 mg, e .i.%=99.3%, Rt=3.555 min. ESI-MS (m/z): 360.1 [M+H]+; and (R)-ethyl 4-(4-fluorophenyl)-4,6-dimethyl-2-(triazol-2-yl)-1,4-dihydropyrimidine-5-carboxylate (21-2') 171 mg, e.g. i.%=98.1%, Rt=4.873 min. ESI-MS (m/z): 360.1 [M+H]+.

Step 3-Step 4: 2-((1-(((S)-5-(ethoxycarbonyl)-6-(4-fluorophenyl)-6-methyl-2-(triazol-2-yl)-3,6dihydropyrimidine- 4-yl)methyl)-3,3-difluoropiperidin-4-yl)oxy)acetic acid (10-224).

Using methods similar to those described in Step 2 and Step 3 of Example 1, the title compound 15 mg was obtained by reacting compound (21-2), after the bromination reaction, with 2-((3,3-difluoropiperidin-4-yl) hydroxy)acetic acid (compound 5-5 in example 5).

The structure was characterized in the following way:

1H NMR (400 MHz, DMSO-d6) δ 9.35 (s, 1H), 8.01 (d, J=3.2 Hz, 1H), 7.92 (d, J=3.2 Hz, 1H ), 7.41 (dd, J=8.8, 5.5 Hz, 2H), 7.11 (t, J=8.8 Hz, 2H), 4.18-4.06 (m, 2H) , 3.89-3.53 (m, 6H), 3.04-2.60 (m, 4H), 2.44 (s, 1H), 1.97 (s, 1H), 1.80 (s , 3H), 0.92 (td, J=7.1, 1.4 Hz, 3H). ESI-MS (m/z): 553.2 [M+H]+.

Example 22 Preparation of 2-((1-(((R)-5-(ethoxycarbonyl)-6-(4-fluorophenyl)-6-methyl-2-(triazol-2-yl)3,6-dihydropyrimidin-4 -yl)methyl)-3,3-difluoropiperidin-4-yl)oxy)acetic acid (10-225)

- 26 039977

The title compound 15 mg was prepared from compound (21-2') in Example 21 using methods similar to those described in Step 3-4 of Example 21.

The structure was characterized in the following way:

1H NMR (400 MHz, DMSO-d ₆ ) δ 9.35 (s, 1H), 8.01 (d, J=3.2 Hz, 1H), 7.92 (d, J=3.2 Hz, 1H), 7.41 (dd, J=8.8, 5.5 Hz, 2H), 7.11 (t, J=8.8 Hz, 2H), 4.21-4.11 (m, 2H ), 3.94-3.54 (m, 6H), 3.14-2.70 (m, 4H), 2.45 (s, 1H), 1.97 (s, 1H), 1.80 ( s, 4H), 0.92 (td, J=7.1, 1.4 Hz, 3H). ESI-MS (m/z): 553.2 [M+H]+.

Example 23 Preparation of 2-((1-(((S)-5-(ethoxycarbonyl)-6-(4-fluorophenyl)-6-methyl-2-(triazol-2-yl)3,6-dihydropyrimidin-4 -yl)methyl)-4,4-difluoropyrrolidin-3-yl)oxy)acetic acid (10-211)

Using methods similar to those described in Step 2 and Step 3 of Example 1, the title compound 2 mg was obtained by reacting compound (21-2) in Example 21, after the bromination reaction, with 2-((4,4-difluoropyrrolidine-3 -yl)oxy)acetic acid (compound (12-4) in Example 12).

The structure was characterized in the following way:

¹ H NMR (400 MHz, CDCl3) δ 7.85 (s, 1H), 7.50-7.35 (m, 3H), 6.99 (t, J=8.6 Hz, 2H), 4, 22-4.12 (m, 1H), 3.94-3.84 (m, 3H), 3.30-3.17 (m, 3H), 3.08-2.93 (m, 2H), 2.27-1.99 (m, 1H), 1.91 (s, 3H), 1.68-1.48 (m, 1H), 0.97 (t, J=7.0 Hz, 3H) . ESI-MS (m/z): 539.2 [M+H] ⁺ .

Example 24 Preparation of 2-((1-(((R)-5-(ethoxycarbonyl)-6-(4-fluorophenyl)-6-methyl-2-(triazol-2-yl)3,6-dihydropyrimidin-4 -yl)methyl)-4,4-difluoropyrrolidin-3 -yl)oxy)acetic acid (10-212)

Using methods similar to those described in Step 2 and Step 3 of Example 1, the title compound 4 mg was obtained by reacting compound (21-2') in Example 21, after the bromination reaction, with 2-((4,4-difluoropyrrolidine- 3-yl)oxy)acetic acid (compound (12-4) in Example 12).

The structure was characterized in the following way:

1H NMR (400 MHz, CDCl3) δ 7.85 (s, 1H), 7.51-7.35 (m, 3H), 6.99 (t, J=8.6 Hz, 2H), 4.23 -4.09 (m, 1H), 3.94-3.82 (m, 3H), 3.33-3.17 (m, 3H), 3.13-2.90 (m, 2H), 2 .27-1.98 (m, 1H), 1.91 (s, 3H), 1.68-1.45 (m, 1H), 0.97 (t, J=7.0 Hz, 3H). ESI-MS (m/z): 539.2 [M+H] ⁺ .

Example 25 Preparation of (E)-3-((R)-4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazole-2

- 27 039977 yl)-3,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-226)

Step 1: Preparation of (S)-tert-butyl 2-formylmorpholine-4-carboxylate (25-2).

While cooling in an ice bath, (S)-tert-butyl 2-(hydroxymethyl)morpholine-4-carboxylate (25-1) (1.0 g, 4.6 mmol) was dissolved in dichloromethane (10 ml), the reagent was added in portions Dess-Martin (2.9 g, 6.9 mmol) and the reaction was stirred at 15°C for 4 h. stirred for 30 min and the layers were allowed to separate and separated from each other. The organic layer was washed with saturated sodium bicarbonate and brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give a crude product of 980 mg. The crude product was used in the next reaction without purification.

Step 2: Preparation of (R,E)-tert-butyl 2-(3-(tert-butoxy)-3-oxoprop-1-en-1-yl)morpholine-4carboxylate (25-3).

At room temperature, NaH (60%, 182 mg, 4.55 mmol) was dispersed in dry tetrahydrofuran (7 ml), stirred for 5 min, then a solution of tert-butyl diethylphosphonoacetate (1.21 g, 4.78 mmol) was slowly added in portions ) in dry tetrahydrofuran (3 ml) and the reaction was stirred at room temperature for 2 hours. Then, the reaction solution was added to a solution of compound (252) (980 mg, 4.55 mmol) in dry tetrahydrofuran (5 ml) and the reaction was stirred in overnight at room temperature after addition in portions. The solvent was distilled off under reduced pressure and the residue was purified by column chromatography to give a colorless oil (520 mg) which precipitated at room temperature to give a solid residue, HPLC detection showed Z/E<l/35, e.i. (enantiomeric excess) was 94%. ESI-MS (m/z): 214.1 [M+1-100]+.

Step 3: preparation of (R,E)-3-(mofolin-2-yl)acrylic acid trifluoroacetate salt (25-4).

At room temperature, compound (25-3) (520 mg, 1.66 mmol) was dissolved in dichloromethane (10 ml), trifluoroacetic acid (5 ml) was added and the reaction was carried out at room temperature for 3 hours. Insoluble matters were filtered off and the filtrate was concentrated , giving the crude product of the title compound 423 mg, which was used directly in the next reaction without purification. ESI-MS (m/z): 158.1 [M+H] ⁺ .

Step 4: Preparation of (E)-3-((R)-4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2yl)-3 ,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-226).

At room temperature, (R)-ethyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triazol-2-yl)-1,4dihydropyrimidine-5-carboxylate (400 mg, 0.87 mmol), compound (25-4) (423 mg, 1.66 mmol) and N,Ndiisopropylethylamine (451 mg, 3.49 mmol) were added to dichloromethane (10 ml) and the reaction was carried out at room temperature overnight. The reaction solution was concentrated to give a crude product, which was purified by preparative liquid chromatography to give the title compound 200 mg. Z/E<1/35, e.i. is 95.5%.

The structure was characterized in the following way:

¹ H NMR (400 MHz, DMSO-d6) δ 12.50 (s, 1H), 9.63 (s, 1H), 8.02 (d, J=3.12 Hz, 1H), 7.95 ( d, J=3.16 Hz, 1H), 7.44-7.40 (m, 2H), 7.18 (td, J=8.48, 2.64 Hz, 1H), 6.73 (dd , J=15.80, 4.08 Hz, 1H), 6.05 (s, 1H), 5.93 (dd, J=15.80, 1.76 Hz, 1H), 4.24-4, 21 (m, 1H), 3.98-3.92(m, 5H), 3.68 (td, J=10.92, 1.72 Hz, 1H), 2.95 (d, J=11, 12 Hz, 1H), 2.83 (d, J=10.92 Hz, 1H), 2.40(td, J=11.16, 2.68 Hz, 1H), 2.07 (t, J= 10.64 Hz, 1H), 1.04 (t, J=7.08 Hz, 3H). ESI-MS (m/z): 535.2 [M+H] ⁺ .

Example 26 Preparation of (E)-3-((R)-1-(((R)-6-(2-chloro-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(triazol-2yl)-3 ,6-dihydropyrimidin-4-yl)methyl)-5,5-difluoropiperidin-3-yl)acrylic acid (10-230)

- 28 039977

Example 17 (608 mg) was separated by chiral chromatography using the following separation conditions: separation column: CHIRALPAK IG 0.46 cm ID x 15 cm length, mobile phase: hexane/EtOH/HOAc=75/25/0.1 (v/v/v), flow rate: 1.0 ml/min, wavelength: UV 254 nm, temperature: 35°C. The title compound 277 mg was obtained by separation, ee%=99.5%, R _t =15.656 min and the structural characterization data were as follows:

¹ H NMR (400 MHz, DMSO-d6) δ 12.32 (brs, 1H), 9.55 (s, 1H), 8.00 (d, J=3.12 Hz, 1H), 7.94 ( d, J=3.12 Hz, 1H), 7.45-7.41 (m, 2H), 7.19 (td, J=8.48 Hz, 2.68 Hz, 1H), 6.76 ( dd, J=15, 85 Hz, 6.64 Hz, 1H), 6.06 (s, 1H), 5.82 (dd, J=15, 85 Hz, 1.16 Hz, 1H), 4.08 (d, J=16.53 Hz, 1H), 4.01 (d, J=16.53 Hz, 1H), 3.98-3.92 (m, 2H), 3.26-3.13 ( m, 1H), 2.90 (br d, J=11.08 Hz, 1H), 2.79-2.69 (m, 2H), 2.29 (t, J=10.80 Hz, 2H) , 1.94-1.77 (m, 1H), 1.04 (t, J=7.12 Hz, 3H). ESI-MS (m/z): 569.2 [M+H]+.

Example 27 Preparation of (E)-3-((R)-4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(methoxycarbonyl)-2-(triazol2-yl)-3 ,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-227)

At room temperature, (R)-methyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triazol-2-yl)-1,4dihydropyrimidine-5-carboxylate (400 mg, 0.90 mmol) and compound (25-4) (488 mg, 1.80 mmol) in Example 25 were dissolved in dichloromethane (10 ml), K,K-diisopropylethylamine (696 mg, 5.40 mmol) was added and the reaction was carried out at room temperature during the night. The reaction solution was concentrated to give a crude product, which was purified by preparative liquid chromatography to give the title compound 205 mg.

The structure was characterized in the following way:

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 12.44 (s, 1H), 9.68 (s, 1H), 7.98 (dd, J=27.6, 3.1 Hz, 2H) , 7.48-7.36 (m, 2H), 7.18 (td, J=8.5, 2.6 Hz, 1H), 6.73 (dd, J=15.8, 4.1 Hz , 1H), 6.04 (s, 1H), 5.93 (dd, J=15.8, 1.6 Hz, 1H), 4.23 (d, J=9.3 Hz, 1H), 4 .01-3.90 (m, 3H), 3.68 (t, J=10.2 Hz, 1H), 3.52 (s, 3H), 2.94 (d, J=11.0 Hz, 1H), 2.82 (d, J=11.1 Hz, 1H), 2.41 (dd, J=11.0, 8.6 Hz, 1H), 2.08 (t, J=10.7 Hz, 1H). ESI-MS (m/z): 521.1 [M+H] ⁺ .

- 29 039977

Example 28 Preparation of (E)-3-((R)-4-(((R)-6-(2-bromo-4-fluorophenyl)-5-(methoxycarbonyl)-2-(triazol2-yl)-3 ,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-229)

At room temperature, (R)-methyl 6-(bromomethyl)-4-(2-bromo-4-fluorophenyl)-2-(triazol-2-yl)-1,4dihydropyrimidine-5-carboxylate (400 mg, 0.82 mmol) and compound (25-4) (443 mg, 1.63 mmol) in Example 25 were dissolved in dichloromethane (10 ml), then K,K-diisopropylethylamine (635 mg, 4.92 mmol) was added and the reaction was carried out at room temperature temperature during the night. The reaction solution was concentrated to give a crude product, which was purified by preparative liquid chromatography to give the title compound 200 mg.

The structure was characterized in the following way:

1H NMR (400 MHz, DMSO-d ₆ ) δ 12.49 (s, 1H), 9.68 (s, 1H), 8.01 (d, J=3.1 Hz, 1H), 7.95 ( d, J=3.1 Hz, 1H), 7.57 (dd, J=8.6, 2.6 Hz, 1H), 7.38 (dd, J=8.7, 6.2 Hz, 1H ), 7.22 (td, J=8.5, 2.6 Hz, 1H), 6.73 (dd, J=15.8, 4.1 Hz, 1H), 6.02 (s, 1H) , 5.93 (dd, J=15.8, 1.7 Hz, 1H), 4.27-4.19 (m, 1H), 4.01-3.89 (m, 3H), 3.68 (t, J=10.2 Hz, 1H), 3.52 (s, 3H), 2.94 (d, J=11.0 Hz, 1H), 2.83 (d, J=11.3 Hz , 1H), 2.41 (dd, J=11.1, 8.4 Hz, 1H), 2.08 (t, J=10.6 Hz, 1H). ESI-MS (m/z): 567.1 [M+H]+.

Example 29 Preparation of (E)-3-((R)-4-(((R)-6-(2-bromo-3-fluorophenyl)-5-(methoxycarbonyl)-2-(triazol2-yl)-3 ,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-236)

The title compound mg was prepared using methods similar to those described in Example 27 and replacing (R)-methyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triazol-2-yl )1,4-dihydropyrimidine-5-carboxylate on (R)-methyl 4-(2-bromo-3-fluorophenyl)-6-(bromomethyl)-2-(triazol-2yl)-1,4-dihydropyrimidine-5- carboxylate.

The structure was characterized in the following way:

¹ H NMR (400 MHz, DMSO-d6) δ 9.70 (s, 1H), 8.02 (d, J=3.2 Hz, 1H), 7.95 (d, J=3.2 Hz, 1H), 7.39 (m, J=7.9, 5.5 Hz, 1H), 7.31-7.17 (m, 2H), 6.74 (dd, J=15.8, 4, 1 Hz, 1H), 6.09 (s, 1H), 5.93 (d, J=15.9 Hz, 1H), 4.274.19 (m, 1H), 4.02 (m, J=16, 8, 9.4 Hz, 1H), 3.95 (d, J=7.7 Hz, 2H), 3.68 (td, J=11.4, 2.5 Hz, 1H), 3.51 ( s, 3H), 2.99-2.92 (m, 1H), 2.83 (d, J=11.4 Hz, 1H), 2.42 (td, J=11.4, 3.3 Hz , 1H), 2.09 (t, J=10.6 Hz, 1H), 1.41 (s, 1H). ESI-MS (m/z): 567.0 [M+H] ⁺ .

Example 30 Preparation of (E)-3-((R)-4-(((R)-6-(2-chloro-3,4-difluorophenyl)-5-(methoxycarbonyl)-2(triazol-2-yl )-3,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-237)

- 30 039977

The title compound 25 mg was prepared using methods similar to those described in Example 27 and replacing (R)-methyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triazole-2- yl) 1,4-dihydropyrimidine-5-carboxylate on (R)-methyl 6-(bromomethyl)-4-(2-chloro-3,4-difluorophenyl)-2(triazol-2-yl)-1,4- dihydropyrimidine-5-carboxylate.

The structure was characterized in the following way:

1H NMR (400 MHz, CDCl ₃ ) δ 7.79 (d, J=3.1 Hz, 1H), 7.45 (d, J=3.1 Hz, 1H), 7.12-6.94 ( m, 2H), 6.78 (dd, J=15.7, 4.0 Hz, 1H), 6.11 (s, 1H), 6.06 (d, J=15.6 Hz, 1H), 4.53 (s, 1H), 4.31 (d, J=15.8 Hz, 1H), 4.13 (d, J=15.3 Hz, 1H), 4.01 (s, 2H), 3.54 (s, 3H), 3.27 (d, J=70.2 Hz, 2H), 2.76 (s, 1H). ESI-MS (m/z): 539.2 [M+H]+.

Example 31 Preparation of (E)-3-((R)-4-(((R)-6-(2-bromo-3,4-difluorophenyl)-5-(methoxycarbonyl)-2(triazol-2-yl )-3,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-238)

The title compound 70 mg was prepared using methods similar to those described in Example 27 and replacing (R)-methyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triazole-2- yl) 1,4-dihydropyrimidine-5-carboxylate on (R)-methyl 4-(2-bromo-3,4-difluorophenyl)-6-(bromomethyl)-2(triazol-2-yl)-1,4- dihydropyrimidine-5-carboxylate.

The structure was characterized in the following way:

% NMR (400 MHz, CDCl ₃ ) δ 9.66 (s, 1H), 7.85 (d, J=3.1 Hz, 1H), 7.47 (d, J=3.1 Hz, 1H) , 7.12-7.01 (m, 2H), 6.92-6.87 (m, 1H), 6.19 (s, 1H), 6.13-6.09 (m, 1H), 4 .50-4.35 (m, 1H), 4.20-4.05 (m, 3H), 4.00-3.80 (m, 2H), 3.62 (s, 3H), 3.05 -2.75 (m, 2H), 2.71-2.55 (m, 1H), 2.30-2.15 (m, 1H). ESI-MS (m/z): 584.0 [M+H] ⁺ .

Example 32 Preparation of (E)-3-((R)-4-(((S)-6-(3,4-difluoro-2-methylphenyl)-5-(methoxycarbonyl)-2(triazol-2-yl )-3,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-239)

- 31 039977

The title compound 22 mg was prepared using methods similar to those described in Example 27 and replacing (R)-methyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triazole-2- yl) 1,4-dihydropyrimidine-5-carboxylate on (S)-methyl 6-(bromomethyl)-4-(3,4-difluoro-2-methylphenyl)-2(triazol-2-yl)-1,4- dihydropyrimidine-5-carboxylate.

The structure was characterized in the following way:

¹ H NMR (400 MHz, CDCl,) δ 7.93 (dd, J=3.2, 1.5 Hz, 1H), 7.72 (t, J=2.7 Hz, 1H), 7.02 (m, J=7.2, 3.3 Hz, 2H), 6.82 (m, J=15.8, 4.2 Hz, 1H), 6.16-5.96 (m, 1H), 5.91 (s, 1H), 4.37 (d, J=9.8 Hz, 1H), 4.20-3.75 (m, 4H), 3.60 (s, 3H), 3.14 -2.64 (m, 2H), 2.55 (d, J=2.4 Hz, 3H), 2.60-2.41 (m, J=11.7, 5.8 Hz, 1H), 2.29-2.20 (m, J=10.6 Hz, 1H). ESI-MS (m/z): 519.2 [M+H] ⁺ .

Example 33 Preparation of (E)-3-((R)-4-(((R)-6-(2-chloro-3-fluorophenyl)-5-(methoxycarbonyl)-2-(triazol2-yl)-3 ,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-240)

mg was obtained using methods similar to those

The title compound as described in Example 27 and replacing (R)-methyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triazol-2-yl)1,4-dihydropyrimidin-5- carboxylate on (R)-methyl 6-(bromomethyl)-4-(2-chloro-3-fluorophenyl)-2-(triazol-2yl)-1,4-dihydropyrimidine-5-carboxylate.

The structure was characterized in the following way:

1H NMR (400 MHz, CDCI3) δ 7.84 (d, J=3.1 Hz, 1H), 7.48 (s, 1H), 7.22-7.16 (m, J=16.5 Hz , 2H), 7.06 (s, 1H), 6.89 (d, J=15.7 Hz, 1H), 6.25 (s, 1H), 6.11 (d, J=15.7 Hz , 1H), 4.50 (s, 1H), 4.22 (d, J=15.6 Hz, 1H), 4.03 (dd, J=31.8, 9.3 Hz, 3H), 3 .60 (s, 3H), 3.04 (s, 2H), 2.70 (s, 1H), 2.35 (s, 1H). ESI-MS (m/z): 521.2 [M+H]+.

Example 34 Preparation of (E)-3-((R)-4-(((R)-6-(2,4-dichlorophenyl)-5-(methoxycarbonyl)-2-(triazol-2yl)-3.6 -dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-241)

The title compound 400 mg was prepared using methods similar to those described in Example 27 and replacing (R)-methyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triazol-2-yl )1,4-dihydropyrimidine-5-carboxylate on (R)-methyl 6-(bromomethyl)-4-(2,4-dichlorophenyl)-2-(triazol-2-yl)1,4-dihydropyrimidine-5-carboxylate .

The structure was characterized in the following way:

1H NMR (400 MHz, CDC13) δ 9.64 (s, 1H), 7.86 (d, J=3.1 Hz, 1H), 7.47 (d, J=3.1 Hz, 1H), 7.42 (d, J=2.1 Hz, 1H), 7.28 (d, J=6.5 Hz, 1H), 7.19 (dd, J=8.4, 2.1 Hz, 1H ), 6.93 (dd, J=15.7, 4.1 Hz, 1H), 6.22 (s, 1H), 6.12 (dd, J=15.7, 1.8 Hz, 1H) , 4.40 (d, J=9.9 Hz, 1H), 4.12-4.01 (m, 2H), 3.96-3.85 (m, 2H), 3.62 (s, 3H ), 2.89-2.75 (m, 2H), 2.60 (td, J=10.9, 2.8 Hz, 1H), 2.23 (t, J=10.7 Hz, 1H) . ESI-MS (m/z): 537.2 [M+H] ⁺ .

Example 35 Preparation of (E)-3-((R)-4-(((S)-6-(4-fluoro-2-methylphenyl)-5-(methoxycarbonyl)-2-(triazol2-yl)-3 ,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-242)

- 32 039977

The title compound 80 mg was prepared using methods similar to those described in Example 27 and replacing (R)-methyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triazole-2- yl)1,4-dihydropyrimidine-5-carboxylate on (S)-methyl 6-(bromomethyl)-4-(4-fluoro-2-methylphenyl)-2-(triazol2-yl)-1,4-dihydropyrimidine-5 -carboxylate.

The structure was characterized in the following way:

1H NMR (400 MHz, CDCl ₃ ) δ 9.59 (s, 1H), 7.82 (d, J=3.1 Hz, 1H), 7.44 (s, 1H), 7.13 (t, J=7.1 Hz, 1H), 6.90 (t, J=4.1 Hz, 2H), 6.80 (t, J=3.6 Hz, 1H), 6.12 (dd, J= 15.7, 1.2 Hz, 1H), 5.96 (s, 1H), 4.40 (s, 1H), 4.05 (d, J=11.3 Hz, 2H), 3.93 ( d, J=16.0 Hz, 2H), 3.61 (s, 3H), 2.81 (s, 2H), 2.63 (s, 3H), 2.57 (s, 1H), 2, 20 (s, 1H). ESIMS (m/z): 501.2 [M+H]+.

Example 36 Preparation of (E)-3-((R)-4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(methoxycarbonyl)-2-(4methyltriazol-2-yl) -3,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-243)

The title compound 400 mg was prepared using methods similar to those described in Example 27 and replacing (R)-methyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triazol-2-yl )1,4-dihydropyrimidine-5-carboxylate on (R)-methyl 6-(bromomethyl)-4-(2-methyl-4-fluorophenyl)-2-(4methyltriazol-2-yl)-1,4-dihydropyrimidine- 5-carboxylate.

The structure was characterized in the following way:

¹ H NMR (400 MHz, CDCl ₃ ) δ 9.65 (s, 1H), 7.29-7.25 (m, 1H), 7.13 (dd, J=8.6, 2.6 Hz, 1H), 7.01 (s, 1H), 6.93-6.88 (m, 2H), 6.19 (s, 1H), 6.10 (d, J=15.7 Hz, 1H), 4.38 (s, 1H), 4.06 (d, J=12.5 Hz, 2H), 3.90-3.86 (m, 2H), 3.60 (s, 3H), 2.81 (s, 2H), 2.60 (s, 1H), 2.45 (s, 3H), 2.20 (s, 1H). ESI-MS (m/z): 535.1 [M+H] ⁺ .

Example 37 Preparation of (E)-3-((R)-4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(methoxycarbonyl)-2-(4-(trifluoromethyl)triazole -2-yl)-3,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-244)

- 33 039977

The title compound 80 mg was prepared using methods similar to those described in Example 27 and replacing (R)-methyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triazol-2-yl )-1,4dihydropyrimidine-5-carboxylate on (R)-methyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(4-(trifluoromethyl)triazol-2-yl)-1, 4-dihydropyrimidine-5-carboxylate.

The structure was characterized in the following way:

¹ H NMR (400 MHz, CDCla) δ 9.70 (s, 1H), 7.86 (s, 1H), 7.27 (s, 1H), 7.17 (dd, J=8.6, 2 .6 Hz, 1H), 6.96 (ddd, J=11.3, 10.6, 3.4 Hz, 2H), 6.24 (s, 1H), 6.18 (d, J=15, 4 Hz, 1H), 4.38 (d, J=10.4 Hz, 1H), 4.19-4.10 (m, 1H), 4.02 (d, J=11.1 Hz, 1H) , 3.83 (dd, J=13.3, 9.8 Hz, 2H), 3.63 (s, 3H), 2.95 (d, J=10.8 Hz, 1H), 2.66 ( e, J=11.6 Hz, 1H), 2.51-2.33 (m, 2H). ESI-MS (m/z): 589.1 [M+H] ⁺ .

Example 38 Preparation of (E)-3-((R)-4-(((S)-6-(2-(difluoromethyl)-4-fluorophenyl)-5-(methoxycarbonyl)-2(triazol-2-yl )-3,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)acrylic acid (10-245)

The title compound 70 mg was prepared using methods similar to those described in Example 27 and replacing (R)-methyl 6-(bromomethyl)-4-(2-chloro-4-fluorophenyl)-2-(triazole-2- yl) 1,4-dihydropyrimidine-5-carboxylate on (S)-methyl 6-(bromomethyl)-4-(2-(difluoromethyl)-4-fluorophenyl)-2(triazol-2-yl)-1,4- dihydropyrimidine-5-carboxylate.

The structure was characterized in the following way:

¹ H NMR (400 MHz, CDC^y 9.68 (s, 1H), 7.83 (d, J=3.1 Hz, 1H), 7.69-7.55 (m, 1H), 7, 46 (d, J=2.9 Hz, 1H), 7.40 (d, J=7.5 Hz, 1H), 7.20 (s, 1H), 7.08 (t, J=7.5 Hz, 1H), 6.96 (dd, J=15.7, 3.9 Hz, 1H), 6.15 (dd, J=15.7, 1.5 Hz, 1H), 6.08 (s , 1H), 4.40 (d, J=5.0 Hz, 1H), 4.16 (d, J=17.6 Hz, 1H), 4.00 (d, J=11.4 Hz, 1H ), 3.93-3.80 (m, 2H), 3.62 (s, 3H), 2.93 (d, J=9.2 Hz, 1H), 2.65 (d, J=9, 7 Hz, 1H), 2.46 (t, J=11.1 Hz, 1H), 2.36 (d, J=9.6 Hz, 1H). ESI-MS (m/z): 537.2 [M+H]+.

Example 39 Preparation of (E)-3-((R)-4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(methoxycarbonyl)-2-(triazol2-yl)-3 ,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)but-2-enoic acid (10-246)

Step 1: Preparation of (S)-4-(tert-butoxycarbonyl)morpholine-2-carboxylic acid (39-2).

At room temperature, (S)-tert-butyl 2-(hydroxymethyl)morpholine-4-carboxylate (39-1) (5 g, 23.01 mmol) was dissolved in acetone (250 ml) and saturated sodium bicarbonate solution (75 ml) was added ). The reaction was cooled to 0°C in an ice bath, sodium bromide (474 mg, 4.6 mmol) and tetramethylpiperidinyloxy (65 mg, 0.46 mmol) were added, followed by the slow addition of trichloroisocyanuric acid (10.7 g, 46.03 mmol) and the reaction was carried out at room temperature overnight. Isopropanol (15 ml) was added to the reaction, stirred for 30 min, filtered with suction, and the filter cake was discarded. The filtrate was concentrated, saturated sodium carbonate solution (75 ml) was added, extracted with ethyl acetate (50 mlx2) and the organic phase was discarded. The aqueous phase was neutralized with 6N hydrochloric acid and extracted with ethyl acetate (50 mlx3). Organic

- 34 039977 the sk phase was combined, dried over anhydrous sodium sulfate, the drying agent was filtered off and the filtrate was concentrated to give the title compound 3 g, which was used directly in the next reaction without purification. ESI-MS (m/z): 176.1 [M+1-56]+.

Step 2: Preparation of (S)-tert-butyl 2-(methoxy(methyl)carbamoyl)morpholine-4-carboxylate (39-3).

At room temperature, compound (39-2) (2 g, 8.65 mmol) was dissolved in dichloromethane (20 ml), hexafluorophosphate 2-(7-azabenzotriazol-1-yl)-N,N,N',N'- was added tetramethyluronium (3.95 g, 10.38 mmol) and the reaction was carried out at room temperature for 30 minutes. N,Ndiisopropylethylamine (2.57 g, 19.89 mmol) and N,O-dimethylhydroxylamine hydrochloride (1.01 g, 10.38 mmol) were added and the reaction was carried out overnight. Water (20 ml) was added to the reaction and extracted with dichloromethane (20 mlx3). The organic phase was combined, which was washed successively with 0.05N hydrochloric acid (20 ml), saturated sodium bicarbonate solution, water and saturated sodium chloride solution and dried over anhydrous sodium sulfate, the drying agent was filtered off and the filtrate was concentrated to give the title compound (2 g ), which was used directly in the next reaction without purification. ESI-MS (m/z): 219.1 [M+1-56]+.

Step 3: Preparation of (S)-tert-butyl 2-acetylmorpholine-4-carboxylate (39-4).

At room temperature, compound (39-3) (2 g, 7.29 mmol) was dissolved in anhydrous tetrahydrofuran (40 ml), cooled to -20° C. under nitrogen protection, methylmagnesium bromide (3M, 7.29 ml, 21. 87 mmol) and the reaction was carried out at -20°C for 4 hours. Saturated ammonium chloride (20 ml) was added to the reaction and extracted with ethyl acetate (20 mlx3). The organic phase was combined, washed successively with 0.05N hydrochloric acid (20 ml), saturated sodium bicarbonate solution, water and saturated sodium chloride solution and dried over anhydrous sodium sulfate, the drying agent was filtered off and the filtrate was concentrated to give the title compound (1.5 d), which was used directly in the next reaction without purification. ESIMS (m/z): 174.1 [M+1-56]+.

Step 4: Preparation of (R,E)-tert-butyl 2-(4-(tert-butoxy)-4-oxobut-2-en-2-yl)morpholine-4carboxylate (39-5).

The title compound (1.1 g) was obtained using methods similar to those described in Step 2 of Example 25 and replacing (S)-tert-butyl 2-formylmorpholine-4-carboxylate with compound (39-4). ESI-MS (m/z): 172.1 [M+1-100-56] ⁺ .

Step 5: preparation of (R,E)-3-(mofolin-2-yl)but-2-enoic acid trifluoroacetate salt (39-6).

The title compound (491 mg) was obtained using methods similar to those described in step 3 of Example 25 and replacing (R,E)-tert-butyl 2-(3-(tert-butoxy)-3-oxoprop-1- en-1yl)morpholine-4-carboxylate for compound (39-5). ESI-MS (m/z): 172.1 [M+H] ⁺ .

Step 6: Preparation of (E)-3-((R)-4-(((R)-6-(2-chloro-4-fluorophenyl)-5-(methoxycarbonyl)-2-(triazol-2yl)-3 ,6-dihydropyrimidin-4-yl)methyl)mofolin-2-yl)but-2-enoic acid (10-246).

The title compound (180 mg) was obtained using methods similar to those described in Example 27 and replacing (R,E)-3-(mofolin-2-yl)acrylic acid trifluoroacetate salt with compound (39-6).

The structure was characterized in the following way:

1H NMR (400 MHz, CDCl ₃ ) δ 7.85 (d, J=3.1 Hz, 1H), 7.51 (d, J=3.0 Hz, 1H), 7.33 (dd, J= 8.6, 2.0 Hz, 1H), 7.14 (dd, J=8.5, 2.6 Hz, 1H), 6.95 (td, J=8.3, 2.6 Hz, 1H ), 6.18 (s, 1H), 6.04 (s, 1H), 4.37-4.22 (m, 2H), 4.104.07 (m, 2H), 3.62 (s, 3H) , 3.27 (s, 1H), 2.83 (s, 1H), 2.43 (s, 1H), 2.08 (s, 3H). ESI-MS (m/z): 535.1 [M+H]+ .

Additional compounds can be obtained by methods similar to those in the examples above.

In the following pharmacological tests, the compound of the present invention was compared with the compound of the present invention and the compound GLS4 (control compound 1), the compound of example 9 in WO 2015144093 (control compound 2) and the compound of example 5 in WO 2014037480 (control compound 3) described in the paragraph of the state of the art of the present invention of the present application in order to adequately illustrate the advantages of the examples of the present invention.

The structure of control compound 1 is

the structure of control compound 2 is

- 35 039977

F

and the structure of control compound 3 is

F

Experimental example 1: test for biological activity.

The inhibitory effect of the compounds of the present invention on hepatitis B virus (HBV) was tested. The cytotoxicity and effects on the viral nucleic acid (DNA) replication level (HBV) of the compounds of the present invention were tested at the virus-cell level.

Test method.

HepG2.2.15 cells in the logarithmic growth phase were seeded in a 96-well plate at a cell concentration of 40 cells per μl. Cells were kept in a 5% CO ₂ incubator at 37°C for 3 days; and culture medium was replaced with new culture medium (200 μl/well) before adding compounds. The stock solution concentration of each exemplary compound is 200 μM. With 200 µM as the highest concentration, the solution was diluted to various concentrations of DMSO and 1 µl of the test compound was added to the appropriate culture medium well and the final test compound concentrations were 0.06, 0.24, 0.98, 3.9, 15, 6, 62.5, 250, 1000 nM (used to calculate the semi-effective concentration (EC ₅₀ )). The test results are shown in table. 1-1 and tab. 1-2.

- 36 039977

As shown in Table. 1-1, the test compounds have an effective inhibitory activity on hepatitis B virus (HBV).

As shown in Table. 1-2, the anti-HBV activity of the compounds having one configuration of the present invention is approximately 10 times that of the control compound 2, indicating that the compounds of the present invention have a more potent hepatitis B virus (HBV) inhibitory activity.

The remaining compounds of the present invention have the same inhibitory activity as above.

Experimental example 2: determination of cytotoxicity.

Test compounds were diluted to 30 mM DMSO, with 30 mM as the highest concentration, compounds were subjected to three-fold serial dilutions to various concentrations. 0.2 μl of compounds at various concentrations were added to a 384-well plate, HepG2.2.15 cells having a concentration of 2000 cells per 50 μl were added to each well, and the highest concentration of test compounds was 150 μM. 1 µl DMSO was added to the respective wells for control. The tablet was kept in a 5% CO ₂ incubator at 37°C for 4 days; and 50 μl CellTiter-Glo was added to each well after 4 days. The plates were read for detection and the semi-cytotoxic concentration (CC ₅₀ ) was calculated. The test results are as shown in table. 2.

- 37 039977

table 2

Connection no. SS ₅₀ (µM) 10-7 example 6 129.5 10-34 example 9 79.6 10-36 example 10 >150.0 10-40 example 11 73.5 10-42 examples 12 >150.0 10-88 example 1 61.1 Isomer B of example 4 51.3 Isomer A of example 4 43.9 10-95 example 5 131.3 10-136 example 16 122.3 10-162 examples 15 >150.0 10-180 example 14 >150.0 10-182 examples 13 >150.0 10-226 example 25 >150.0 10-230 example 26 137.0 10-227 example 27 >150.0 10-229 example 28 >150.0 10-237 example 30 >150.0 10-238 example 31 >150.0 10-239 example 32 >150.0 10-240 example 33 >150.0

The test compounds of the present invention have relatively low cytotoxicity and relatively high safety. The remaining compounds of the present invention have similar safety characteristics.

Experimental Example 3: test for hERG inhibitory effect.

In cardiac muscle cells hERG (human heart specific potassium channel gene), the encoded potassium channel mediates delayed potassium current rectification (IKr). IKr inhibition is the most important mechanism for drug-induced QT prolongation. In the hERG test, the evaluation criterion is as follows: if the IC ₅₀ of a compound is greater than 10 μM, then the compound is determined not to have any inhibitory effect on hERG.

Using the Predictor™ hERG fluorescent polarization assay, the effect of the compounds on the hERG potassium ion channel was determined. The test results are as shown in table. 3 below.

-38039977

Table 3

Connection no. 1C ₅₀ value (µM) Control compound 1 2.85 Control Compound 2 1-10 10-36 example 10 >10 10-40 example 11 >10 10-42 examples 12 >10 10-88 example 1 >10 Isomer B of example 4 >10 Isomer A of example 4 >10 10-93 example 2 >10 10-95 example 5 >10 10-226 example 25 >10 10-227 example 27 >10 10-230 example 26 >10

As described above, control compound 1 and control compound 2 have varying degrees of cardiotoxicity (having a significant inhibitory effect on the hERG potassium ion channel in cardiac muscle cells) and thus have a potential risk of causing arrhythmia; while the test compounds of the present invention do not have an inhibitory effect on the hERG potassium ion channel and thus do not have significant cardiotoxicity, thereby providing greater safety. The remaining compounds of the present invention have similar safety.

Experimental example 4: in vivo study of pharmacokinetic parameters (PK) in rats.

The test compounds were administered to male SD rats intravenously (iv) and by gavage (po), respectively, the doses of iv and po administration were 1 and 2 mg/kg, respectively, the solvent system for iv administration was 5% DMSO: 5% solyutol: 90% saline and po solvent system was 0.5% MS. Blood was collected at various time points after iv administration and po administration for PK studies. Plasma samples and liver tissue samples were subjected to protein precipitation followed by LC-MS/MS analysis. The mass spectrometer was an API 5500 and the liquid chromatograph was a Waters ACQUITY I CLASS system; the chromatographic column was an Agela ASB Ci ₈ column (2.1 mm x 50 mm, 1.9 µm); mobile phase A was water+0.1% formic acid and phase B was acetonitrile; the flow rate was 0.4 ml/min and the column temperature was 40°C. The ion source was an ESI source in positive ion mode and the scanning method was selective recording of selected multiple ion decay reactions (MRM). The test results are shown in the following table.

Table 4

Connection no. Route of administration dosing AUCjnf Stakh mg/kg h*ng/ml ng/ml Control compound 2 iv 1.00 2150 2470 10-226 example 25 iv 1.00 2400 2890

According to the data in Table. 4 Compared to control compound 2, the compound of the present invention (e.g., 10-226 of Example 25) administered intravenously at 1.00 mg/kg has better drug exposure (AUC _INF ) and higher drug-39039977 blood concentration. (Cmax in vivo and thus has the best pharmacokinetic parameters. __________ Table 5. Pharmacokinetic parameters in plasma _______

Connection no. Route of administration dose auc _inf With _tach mg/kg h*ng/ml ng/ml Control Compound 2 ro 2.00 897 1250 10-226 example 25 ro 2.00 2290 2180

According to the data in Table. 5, compared with control compound 2, the compound of the present invention (e.g., 10-226 of example 25) administered by gavage at 2.00 mg/kg has better drug exposure (AUC _INF ) and higher blood drug concentration (C _max ) in vivo and thus has the best absorption parameters.

Table 6. Pharmacokinetic parameters in the liver

Connection no. Route of administration dose auc _inf Stakh mg/kg h*ng/ml ng/ml Control Compound 2 ro 2.00 1490 547 10-226 example 25 ro 2.00 3030 1690

According to the data in Table. 6, compared with control compound 2, the compound of the present invention (e.g., 10-226 of Example 25) administered by gavage at 2.00 mg/kg has better drug exposure (AUC _INF ) and higher drug concentration in blood (C _max ) in vivo, which further indicates that the compound of the present invention (for example, 10-226 of example 25) has better absorption parameters. In addition, the bioavailability (F) of the compound of the present invention (e.g., 10-226 of Example 25) administered by gavage at 2.00 mg/kg is 47.6%, which is significantly greater than the bioavailability (20.9% ) control compound 2.

Table 7

Connection no. Route of administration sample dose auc _inf Stakh mg/q h*ng/m l ng/m l Control compound 2 ro plasma 2.00 897 1250 liver 2.00 1490 547 10-230 example 26 ro plasma 2.00 236 179 liver 2.00 2340 1520

As shown in Table. 7, the degree of exposure of the compound of the present invention (for example, 10-230 of example 26) in the liver in vivo is 10 times the degree of exposure in plasma and the drug concentration in the liver in vivo is 10 times the concentration in plasma; while the in vivo liver exposure of control compound 2 is 1.5 times the plasma exposure and the in vivo liver drug concentration is 0.5 of the plasma concentration. The above indicates that the compound of the present invention has excellent drug exposure (AUCINF) and drug concentration (C _max ) in the liver in vivo and thus has liver-targeting properties.

Experimental Example 5 In vivo study of pharmacokinetic parameters (PK) in beagle dogs.

The test compounds were administered to male beagle dogs intravenously (iv) and by gavage (po), respectively, the doses of iv and po administration were 0.5 mg/kg and 2.5 mg/kg, respectively, the system of solvents for iv administration was 5% DMSO: 5% Solutol: 90% saline and the po solvent system was 0.5% MS. Blood was collected at various time points after iv administration and po administration for PK studies. Plasma samples and liver tissue samples were subjected to protein precipitation followed by LC-MS/MS analysis. Mass spectrometer was API 5500 and liquid chromatograph was Waters

- 40 039977

ACQUITY I CLASS system; the chromatographic column was an Agela ASB C _i8 column (2.1 mm x 50 mm, 1.9 µm); mobile phase A was water+0.1% formic acid and phase B was acetonitrile; the flow rate was 0.4 ml/min and the column temperature was 40°C. The ion source was an ESI source in positive ion mode and the scanning method was selective recording of selected multiple ion decay reactions (MRM). The test results are shown in the following table.

Table 8

Connection no. Route of administration Dose auc _inf AUC _last With _tach mg/q h*ng/m l h*ng/m l ng/m L Control iv 0.50 ISO 1120 2130 connection 3 10-227 example 27 iv 0.50 8750 8690 2310

According to the data in Table. 8, compared with control compound 3, the compound of the present invention (for example, 10-227 of example 27) administered intravenously at 0.50 mg/kg has better drug exposure (AUC _INF ) and higher blood drug concentration (C _max ) in vivo and thus has the best pharmacokinetic parameters.

Table 9

Connection no. Route of administration Dose auc _inf AUCi _as t Stakh mg/q h*ng/m l h*ng/m l ng/m L Control compound 3 ro 2.50 2360 2330 1210 10-227 example 27 ro 2.50 33200 32900 7200

As shown in Table. 9, the plasma exposure of a compound of the present invention (e.g., 10-227 of Example 27) administered by gavage at 2.50 mg/kg is approximately 14 times that of control compound 3, and the blood drug concentration is approximately 6 times the concentration of control compound 3, which further indicates that the compounds of the present invention (for example, 10-227 of example 27) have better absorption properties. In addition, the bioavailability (F) of a compound of the present invention (e.g., 10-227 of Example 27) administered by gavage at 2.50 mg/kg was 75.9%, which is significantly greater than the bioavailability (41.7% ) control compound 3.

Experimental Example 6 In vivo study of pharmacokinetic parameters (PK) in cynomolgus monkeys.

The test compounds were administered to male cynomolgus monkeys intravenously (iv) and by gavage (po), respectively, the doses of iv and po administration were 0.5 and 2.5 mg/kg, respectively, the system of solvents for iv administration was 5 % DMSO: 5% Solutol: 90% saline and the po solvent system was 0.5% MS. Blood was collected at various time points after iv administration and po administration for PK studies. Plasma samples and liver tissue samples were subjected to protein precipitation followed by LC-MS/MS analysis. The mass spectrometer was an API 5500 and the liquid chromatograph was a Waters ACQUITY I CLASS system; the chromatographic column was an Agela ASB C _i8 column (2.1 mm x 50 mm, 1.9 µm); mobile phase A was water+0.1% formic acid and phase B was acetonitrile; the flow rate was 0.4 ml/min and the column temperature was 40°C. The ion source was an ESI source in positive ion mode and the scanning method was selective recording of selected multiple ion decay reactions (MRM). The test results are shown in the following table.

-41 039977

Table 10

Connection no. Route of administration Dose mg/kg auc _inf h*ng/ml AUC _last h*ng/ml Stakh ng/ml Control compound 3 iv 0.50 623 623 1607 10-227 examples 27 iv 0.50 2463 2441 1673

According to the data in Table. 10, in a PK study in cynomolgus monkeys, compared to control compound 3, the compound of the present invention (e.g., 10-227 of Example 27) administered intravenously at 0.50 mg/kg had better drug exposure (AUCINF) and greater concentration of the drug in the blood (C _max ) in vivo and thus has the best pharmacokinetic parameters.

Table 11

Connection no. Route of administration Dose auc _inf AUC _last Stakh mg/kg g h*ng/m L h*ng/m l ng/m L Control connection 3 ro 2.50 211 208 32 10-227 example 27 ro 2.50 4566 4417 1069

As shown in Table. 11, in a PK study in cynomolgus monkeys, the plasma exposure of a compound of the present invention (e.g., 10-227 of Example 27) administered by gavage at 2.50 mg/kg is approximately 21 times the exposure of control compound 3 and the blood drug concentration is approximately 33 times that of the control compound 3, further indicating that the compound of the present invention (eg, 10-227 of Example 27) has superior absorption properties. In addition, the bioavailability (F) of the compound of the present invention (e.g., 10-227 of Example 27) administered by gavage at 2.50 mg/kg is 37.2%, which is significantly greater than the bioavailability (6.77% ) control compound 3.

Experimental Example 7: CYP450 Enzyme Induction Study.

Cell extraction.

Tube HepG2 C3A/pCYP3A4-Luc, C8 cells were removed from the liquid nitrogen reservoir, cells were removed in a sterile water bath at 37°C; and the tube was gently shaken until the ice was completely melted. The recovered cells were transferred to a 15 ml sterile centrifuge tube and 5-10 ml of pre-grated basal cell culture medium was added at 37°C; the cells spontaneously settled within 2 min and were centrifuged (1000 rpm) for 8 min. The supernatant was discarded and the cells were suspended in 10 ml of pre-grated cell culture medium. The cell suspension was transferred to a 10 cm cell culture dish and kept in a 5% CO2 incubator at 37°C. The original cell culture medium was replaced with selective cell culture medium after 24 h.

cell proliferation.

Cells were recovered after growing to 80-90% of the culture dish and then transferred to a 15 ml sterile centrifuge tube. Centrifugation was performed at 1000 rpm for 8 min and the cells were harvested. The supernatant was discarded and the cells were suspended with 3 ml of pre-rubbed complete medium. The cell suspension was subcultured at a ratio of 1:3 or 1:5.

CYP3A4 induction test.

The cell plate was received on the first day. 5 µl of 1xMatrigel was added to a 384-well white cell plate and centrifugation was performed at 600 rpm for 1 minute. The culture dish was unloaded, the culture medium was discarded, the cells were washed with 1 ml of PBS, which was then aspirated, 2 ml of 0.25% pancreatin was added, and the cells were kept in the incubator for 2-3 minutes. 5 ml of cell culture medium was added to terminate after complete trypsinolysis of the cells and then transferred to a centrifuge tube. Centrifugation was carried out at 1000 rpm for 8 min. The supernatant was discarded, the cells were suspended and counted, and the suspension was diluted to 4 x 10 ⁵ cells per ml. The cell suspension was placed in a 384-well white plate for

- 42 039977 cells at 25 µl per well. The cell plate was centrifuged at 300 rpm for 1 min and kept in a 5% CO ₂ incubator at 37° C. for 24 hours. On the second day, 300 nl of 100X compounds were transferred to the cell plate from the compound plate. The cell plate was centrifuged at 300 rpm for 1 min and kept in a 5% CO ₂ incubator at 37° C. for 72 hours. On the fifth day, the cell plate and Bright-Glo luciferase reagent were removed and equilibrated to room temperature. Bright-Glo luciferase reagent was added to the cell plate (30 μl per well). The cell plate was centrifuged at 1000 rpm for 1 minute and kept at room temperature for 2 minutes. The fluorescent signal was measured on a plate reader.

Data processing.

The concentration curve of the test compounds was built using the Prism 5 software and the EC ₅₀ values were calculated.

Table 12

Compound no. 10-227 examples 27 Control connection 3 rifampicin EC ₅₀ (µM) 122.8 5,641 1.828

As shown in Table. 12, compared to control compound 3 and rifampicin, the compound of the present invention (eg 10-227 of example 27) has less inductive effect on CYP450 isoform 3A4 and thus has better safety.

As shown in the figure, at a concentration of 10 μM, compared to the control compound 3 and rifampicin, the compound of the present invention (for example, 10-227 of example 27) has a lower inductive effect on the CYP450 isoform 3A4, which is approximately 29% of the inductive effect of rifampicin; while the inductive effect of control compound 3 on CYP450 isoform 3A4 at the same concentration is comparable to that of rifampicin. The data above indicates that the compound of the present invention (eg 10-227 of example 27) has better safety.

In addition to what is described in the invention, as described above, various modifications of the invention will be apparent to a person skilled in the art. It is intended that these modifications fall within the scope of the appended claims. Each reference cited in the present invention (including all patents, patent applications, journal articles, books and any other descriptions) is incorporated into the present invention by reference in its entirety.

Claims (9)

CLAIM 1. A compound or its stereoisomer or tautomer, where the compound has the following structure: where R ¹ represents C _1-6 alkyl; R ² is H; Q is -(CR ^a R ^a ) _g - or -O-; each R ^a and R ^a are H; R ⁴ and R ⁵ are H or halogen; R ⁶ are H; each R ^b at each occurrence is independently selected from the group consisting of H, C _1-6 haloalkyl, and C _1-6 alkyl; each R ^c at each occurrence is independently selected from the group consisting of Cl, Br, C _1-6 haloalkyl, and C _1-6 alkyl; R ⁶ is attached to the ring carbon atom(s) designated * or ** in the general formula; g is 1; i is equal to 1; m is 0; and t is 1. - 43 039977 2. The compound according to claim 1 or its stereoisomer or tautomer, where each R ^b at each occurrence is independently selected from the group consisting of H and C _1-6 alkyl; each R ^c at each occurrence is independently selected from the group consisting of Cl, Br and C _1-6 alkyl. 3. The compound according to claim 1 or 2 or its stereoisomer or tautomer, where the compound has the following structure: 4. The compound or its stereoisomer or tautomer, where the compound is selected from the group consisting of - 44 039977 5. A pharmaceutical composition having antiviral activity, comprising a prophylactically or therapeutically effective amount of a compound according to any one of claims 1 to 4, or a stereoisomer or tautomer thereof, and one or more pharmaceutically acceptable carriers, and the pharmaceutical composition is a solid, liquid or transdermal formulation. 6. A process for the preparation of a pharmaceutical composition according to claim 5, comprising mixing a compound according to any one of claims 1 to 4, or a stereoisomer or tautomer thereof, and one or more pharmaceutically acceptable carriers. 7. The use of a compound according to any one of claims 1 to 4, or a stereoisomer or tautomer thereof, or a pharmaceutical composition according to claim 5, for the preparation of a medicament for the prevention or treatment of a viral disease. 8. Use according to claim 7, wherein the drug is an oral, intravenous, intra-arterial, subcutaneous, intraperitoneal, intramuscular or transdermal drug. 9. Use according to claim 7, wherein the viral disease is selected from the group consisting of viral hepatitis type A, viral hepatitis type B, viral hepatitis type C, influenza, herpes, and acquired immunodeficiency syndrome (AIDS).