Immunoproteasome-selective inhibitor

Abstract

An immunoproteasome-selective inhibitor, a pharmaceutical composition comprising a compound represented by formula (I-1) or a pharmaceutically acceptable salt thereof, and a method for treating a disease by using the compound.

Description

Immunoproteasome selective inhibitors

[0001] This disclosure claims priority to Chinese patent application number CN202311256595.2, filed with the Patent Office of China on September 26, 2023, with the invention name “immunoproteasome selective inhibitors”, Chinese patent application number CN202311690035.8, filed with the Patent Office of China on December 8, 2023, with the invention name “immunoproteasome selective inhibitors”, Chinese patent application number CN202410330302.9, filed with the Patent Office of China on March 21, 2024, with the invention name “immunoproteasome selective inhibitors”, and Chinese patent application number CN202411297537.9, filed with the Patent Office of China on September 14, 2024, with the invention name “immunoproteasome selective inhibitors”, the entire contents of which are incorporated by reference into this disclosure. Technical Field

[0002] The present disclosure belongs to the field of medicinal chemistry, and specifically relates to an immunoproteasome selective inhibitor compound, a pharmaceutical composition containing the compound, and a method for treating diseases using the compound of the present disclosure. Background Art

[0003] In eukaryotic cells, 80%-90% of proteins are degraded by the ubiquitin-proteasome system. This degradation pathway comprises ubiquitin, ubiquitin-activating enzymes (E1), ubiquitin-conjugating enzymes (E2), ubiquitin ligases (E3), deubiquitinating enzymes (DUBs), and the proteasome, of which the proteasome is crucial for regulating cellular physiological functions and maintaining cellular homeostasis. Proteasome dysfunction has been shown to be associated with tumors, immune diseases, infectious diseases, central nervous system diseases, and other diseases. Therefore, inhibition of the ubiquitin-proteasome system, particularly the proteasome, has become an important area of ​​drug discovery.

[0004] The 26S constitutive proteasome (CP), the most common proteasome in eukaryotic cells, is a barrel-shaped structure consisting of a 20S core and two 19S regulatory subunits at either end. The 20S proteasome is composed of 28 subunits arranged in four homologous heptameric rings, forming an α7-β7-β7-α7 cylindrical structure with two-fold axial symmetry. The proteasome's proteolytic activity is closely associated with the β1c, β2c, and β5c subunits, which possess caspase-like (CL), trypsin-like (TL), and chymotrypsin-like (CT-L) activities, respectively. Upon stimulation by immune cells and / or certain cytokines (such as interferon γ and tumor necrosis factor α), the β1c, β2c, and β5c subunits are replaced by novel subunits β1i (LMP2), β2i (MECL1), and β5i (LMP7), forming the immunoproteasome (IP).

[0005] The β2i and β5i subunits have similar activities to β2 and β5. In contrast, β1i primarily performs chymotrypsin-like (CT-L) activity, primarily promoting the hydrolysis of carboxyl-terminal peptides composed of hydrophobic amino acids (particularly tyrosine, tryptophan, phenylalanine, and leucine). Constitutive and immunoproteasomes exhibit distinct proteolytic properties, with immunoproteasomes preferring to hydrolyze proteins rich in nonpolar amino acids. Some of the immunoproteasome's hydrolysis products serve as anchors for MHC class I molecules and play an important role in antigen presentation. Therefore, the immunoproteasome is responsible for cellular immunity and the regulation of major histocompatibility complex (MHC) class I antigen presentation. Studies have shown that mouse dendritic cells lacking the immunoproteasomes β1i, β2i, and β5i exhibit defects in MHC class I antigen presentation.

[0006] Nowadays, bortezomib ( Bortezomib, PS341), carfilzomib ( Carfilzomib, PR171) and ixazomib ( Ixazomib citrate (MLN-9708) has been approved by the FDA for the treatment of multiple myeloma and mantle cell lymphoma, and several other inhibitors are in clinical trials. However, most of these inhibitors are non-selective proteasome inhibitors, inhibiting both the constitutive proteasome and the immunoproteasome. Their potential is limited to the treatment of hematologic malignancies and carries significant toxic side effects. Because non-selective proteasome inhibitors indiscriminately inhibit multiple active sites of both proteasomes, the development of less toxic, selective immunoproteasome inhibitors is particularly urgent and important for the treatment of hematologic malignancies, neurological diseases, and immune system disorders.

[0007] Recent studies have shown that inhibiting cytokine expression requires the simultaneous inhibition of at least two of the three active subunits of the immunoproteasome: β5i / β2i or β5i / β1i. KZR-616, a tripeptide epoxyketone developed by Kezar Life Sciences, is currently in the clinical stage, and more immunoproteasome selective inhibitors still need to be developed.

[0008] Summary of the Invention

[0009] The present disclosure provides a compound represented by formula (I-1), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof:

[0010] wherein Ring A is selected from a 4-10 membered heterocyclic group, and the 4-10 membered heterocyclic group contains 1-3 heteroatoms selected from N, O or S, and Ring A may be optionally further substituted with one or more selected from =O, OH, halogen, C 1-4 Alkyl or C 1-4 substituted by an alkoxy substituent;

[0011] Y 1 、Y 2 and Y 3 Each independently selected from N or CR y ;

[0012] R y Selected from H, OH, halogen, C 1-4 Alkyl or -OR 2a ;

[0013] R 1 Selected from C 1-4 Alkyl, 3-6 membered heterocyclic group, the C 1-4 The alkyl group may be further optionally substituted with one or more alkyl radicals selected from OH, CN, halogen or -OC 3-6 Cycloalkyl, C 1-4 Alkoxy, -O-halogenated C 1-4 substituted by an alkyl substituent;

[0014] R 2 Selected from-SR 2a 、C 2-4 Alkynyl, C 2-4 Alkenyl, C 1-4 Alkyl, -OR 2a 、C 3-6 Cycloalkyl, -C(=O)-R 2d or a 4-7 membered heterocyclic group, wherein the C 1-4 Alkyl, C 3-6 The cycloalkyl or 4-7 membered heterocyclic group may be further optionally substituted by one or more alkyl radicals selected from OH, CN, halogen, C 1-4 Alkyl, C 1-4 Alkoxy or NR 2b R 2c substituted by a substituent;

[0015] R 2a Selected from C 3-6 Cycloalkyl, C 1-4 Alkyl or halogenated C 1-4 alkyl;

[0016] Or, R 2 With Y 2 and its adjacent carbon atoms together form a 5-8 membered heterocyclic ring;

[0017] Or, R 2 With Y 3 and its adjacent carbon atoms together form a 5-8 membered heterocyclic ring;

[0018] R 2b and R 2c Each independently selected from H or C 1-4 alkyl;

[0019] R 2d Selected from H, OH, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Alkoxy or halogenated C 1-4 Alkyl, and the C 1-4 The alkyl group may be further substituted with one or more radicals selected from deuterium, halogen or C 1-4 substituted by an alkoxy substituent;

[0020] R 3 Selected from H or OH;

[0021] R 4 Selected from H or C 1-4 alkyl;

[0022] R 5 Selected from C 2-4 Alkenyl, C4-6 Cycloalkenyl or C 3-6 Cycloalkyl, and the C 2-4 Alkenyl, C 4-6 Cycloalkenyl or C 3-6 The cycloalkyl group may be further optionally substituted with C 2-4 Alkenyl or C 1-4 Alkyl substituted;

[0023] n is selected from 0 or 1.

[0024] In some embodiments, the ring A is selected from

[0025] In some embodiments, the R 1 Selected from methyl, -CH2OH,

[0026] In some embodiments, the R 2 Selected from -OCH3, -CH2OH, -CHF2, -CH2NHCH3,

[0027] In some embodiments, the Y 1 is selected from N, CH or -CF; Y 2 Selected from CH, -CF, -CO-CHF2; Y 3 is selected from N or CH.

[0028] In some embodiments, the Y 3 With R 2 Together

[0029] In some embodiments, the structural unit Selected from

[0030] In some embodiments, the R 3 is selected from H or OH; preferably, R 3 For OH.

[0031] In some embodiments, the R 4 is selected from H or methyl; preferably, R 4 For H.

[0032] In some embodiments, the R 5 Selected from Preferably, R5 Selected from

[0033] The present disclosure also provides a compound represented by formula (I), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof:

[0034] wherein Ring A is selected from a 6-7 membered heterocyclic group, and the 6-7 membered heterocyclic group contains 1-3 heteroatoms selected from N, O or S, and Ring A may be optionally further substituted with one or more heteroatoms selected from OH, halogen, C 1-4 Alkyl or C 1-4 substituted by an alkoxy substituent;

[0035] R 1 Selected from C 1-4 Alkyl, 3-6 membered heterocyclic group, the C 1-4 The alkyl group may be further optionally substituted with one or more alkyl radicals selected from OH, CN, halogen or -OC 3-6 substituted by a cycloalkyl substituent;

[0036] R 2 Selected from C 1-4 Alkyl, -OR 2a 、C 3-6 Cycloalkyl, -C(=O)-R 2d or a 4-7 membered heterocyclic group, wherein the C 1-4 Alkyl, C 3-6 The cycloalkyl or 4-7 membered heterocyclic group may be further optionally substituted by one or more alkyl radicals selected from OH, CN, halogen, C 1-4 Alkyl, C 1-4 Alkoxy or NR 2b R 2c substituted by a substituent;

[0037] R 2a Selected from C 3-6 Cycloalkyl, C 1-4 Alkyl or halogenated C 1-4 alkyl;

[0038] R 2b and R 2c Each independently selected from H or C 1-4 alkyl;

[0039] R 2d Selected from H, OH, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Alkyl or halogenated C 1-4 alkyl;

[0040] R 3 Selected from H or OH;

[0041] n is selected from 0 or 1.

[0042] The present disclosure also provides a compound represented by formula (I), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof:

[0043] wherein Ring A is selected from a 6-7 membered heterocyclic group, and the 6-7 membered heterocyclic group contains 1-3 heteroatoms selected from N, O or S, and Ring A may be optionally further substituted with one or more heteroatoms selected from OH, halogen, C 1-4 Alkyl, C 1-4 substituted by an alkoxy substituent;

[0044] R 1 Selected from C 1-4 Alkyl, 3-6 membered heterocyclic group, the C 1-4 The alkyl group may be further optionally substituted with one or more alkyl radicals selected from OH, CN, halogen, -OC 3-6 substituted by a cycloalkyl substituent;

[0045] R 2 Selected from C 1-4 Alkyl, -OR 2a 、C 3-6 Cycloalkyl, 4-7 membered heterocyclic group, the C 1-4 Alkyl, C 3-6 The cycloalkyl or 4-7 membered heterocyclic group may be further optionally substituted by one or more alkyl radicals selected from OH, CN, halogen, C 1-4 Alkyl, C 1-4 Alkoxy, NR 2b R 2c substituted by a substituent;

[0046] R 2a Selected from C 3-6 Cycloalkyl, C 1-4 Alkyl, halogenated C 1-4 alkyl;

[0047] R 2b and R 2c Each independently selected from H or C 1-4 alkyl;

[0048] R 3 Selected from H or OH;

[0049] n is selected from 0 or 1.

[0050] In some embodiments, Ring A is selected from

[0051] In some embodiments, R 1Selected from methyl or

[0052] In some embodiments, R 2 Selected from methoxy,

[0053] In some embodiments, R 2 Selected from methoxy,

[0054] In some embodiments, the compound of formula (I), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof are selected from the compounds of formula (II) or pharmaceutically acceptable salts thereof:

[0055] Wherein, X is selected from N or CR x ; R x Selected from H, OH, halogen or C 1-4 alkoxy;

[0056] Y is selected from O, NH or NCH3;

[0057] m is selected from 1 or 2; n is selected from 0 or 1;

[0058] R 1 、R 2 、R 3 Definitions are as for the compounds of formula (I).

[0059] In some embodiments, the compound of formula (I), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof, wherein the compound is selected from the compound represented by general formula (IIIA):

[0060] R 2 Selected from-SR 2a 、C 2-4 Alkynyl, C 2-4 Alkenyl, C 1-4 Alkyl, -OR 2a 、C 3-6 Cycloalkyl, -C(=O)-R 2d or a 4-7 membered heterocyclic group, wherein the C 1-4 Alkyl, C 3-6 The cycloalkyl or 4-7 membered heterocyclic group may be further optionally substituted by one or more alkyl radicals selected from OH, CN, halogen, C 1-4 Alkyl, C 1-4 Alkoxy or NR 2b R 2c substituted by a substituent;

[0061] R 2a Selected from C 3-6 Cycloalkyl, C 1-4 Alkyl or halogenated C 1-4 alkyl;

[0062] R 2b and R 2c Each independently selected from H or C 1-4 alkyl;

[0063] R 2d Selected from H, OH, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Alkoxy or halogenated C 1-4 Alkyl, and the C 1-4 The alkyl group may be further substituted with one or more radicals selected from deuterium, halogen or C 1-4 substituted by an alkoxy substituent;

[0064] R 3 Selected from H or OH.

[0065] In some embodiments, the R 2 Selected from C 1-4 Alkyl, -OR 2a 、C 3-6 Cycloalkyl, 4-7 membered heterocyclic group; the C 1-4 The alkyl group is further replaced by OH, C 1-4 Alkoxy, NR 2b R 2c Substitute; said C 3-6 The cycloalkyl or 4-7 membered heterocyclic group may be further optionally substituted with one or more selected from OH, C 1-4 substituted by an alkyl substituent;

[0066] R 2a Selected from C 3-6 Cycloalkyl or halogenated C 1-4 alkyl;

[0067] R 2b and R 2c Each independently selected from H or C 1-4 alkyl;

[0068] R 3 Selected from H or OH.

[0069] In some embodiments, the R 2 Selected from

[0070] In some embodiments, the R 2Selected from -OCH3, CHF2.

[0071] In some embodiments, the compound of formula (I), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof, wherein the compound is selected from the compound represented by general formula (IIIB):

[0072] Wherein, X is selected from N or CR x ; R x Selected from OH, halogen or C 1-4 alkoxy;

[0073] Y is O;

[0074] m is selected from 1 or 2; n is 1;

[0075] R 1 Selected from C 1-4 Alkyl, and the C 1-4 The alkyl group may be optionally further substituted with one or more selected from -OC 3-6 substituted by a cycloalkyl substituent;

[0076] R 3 Selected from H or OH.

[0077] In some embodiments, X is selected from N or CR x ; R x Selected from OH, F, methoxy.

[0078] In some embodiments, the R 1 Selected from methyl or

[0079] In some embodiments, the compound of formula (I), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof, wherein the compound is selected from the compound represented by general formula (IIIC):

[0080] X is selected from N or CR x ; R x Selected from H, OH, halogen or C 1-4 alkoxy;

[0081] Y is selected from O, NH or NCH3;

[0082] m is selected from 1 or 2;

[0083] R 1 Selected from C 1-4 Alkyl, 3-6 membered heterocyclic group, the C1-4 The alkyl group may be further optionally substituted with one or more alkyl radicals selected from OH, CN, halogen, -OC 3-6 substituted by a cycloalkyl substituent;

[0084] R 3 Selected from H or OH.

[0085] In some embodiments, X is selected from N or CH.

[0086] In some embodiments, the R 1 Selected from methyl, -CH2OH, -CH2CN or

[0087] In some embodiments, the R 3 For H.

[0088] In some embodiments, the compound of formula (I), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof, wherein the compound is selected from the compound represented by formula (IIID):

[0089] Wherein, X is selected from N or CR x ; R x Selected from H, OH, halogen or C 1-4 alkoxy;

[0090] Y is selected from O, NH or NCH3;

[0091] m is selected from 1 or 2; n is selected from 0 or 1;

[0092] R 1 Selected from C 1-4 Alkyl, 3-6 membered heterocyclic group, the C 1-4 The alkyl group may be further optionally substituted with one or more alkyl radicals selected from OH, CN, halogen or -OC 3-6 substituted by a cycloalkyl substituent;

[0093] R 2d Selected from H, OH, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Alkyl or halogenated C 1-4 alkyl;

[0094] R 3 Selected from H or OH.

[0095] In some embodiments, X is selected from N or CH.

[0096] In some embodiments, the R 1Selected from methyl, -CH2OH, -CH2CN or

[0097] In some embodiments, the R 3 For H.

[0098] In some embodiments, the R 2d Selected from C 1-4 Alkyl or halogenated C 1-4 Alkyl; preferably, R 2d Selected from methyl, ethyl or fluoroethyl.

[0099] In some embodiments, the compound of formula (I), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof, wherein the compound is selected from the compound represented by general formula (IIID-1):

[0100] R 2d Selected from methyl, ethyl or fluoroethyl.

[0101] In some embodiments, the compound of formula (I), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof, wherein the compound is selected from the compound represented by general formula (IIIE):

[0102] Wherein, X is N, Y is O, m is selected from 0 or 1, R 2 for R 3 Selected from H or OH, R 4 For H.

[0103] The present disclosure also provides a compound, a stereoisomer, an isotopic derivative, a solvate, a nitrogen oxide, a prodrug, or a pharmaceutically acceptable salt thereof, wherein the compound is selected from:

[0104] The present disclosure also provides a pharmaceutical composition comprising a compound of formula (I-1), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof, and a pharmaceutically acceptable carrier.

[0105] In certain embodiments of the present disclosure, in the pharmaceutical composition, the content of the compound of formula (I-1), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof is selected from 0.1 mg to 1000 mg.

[0106] In certain embodiments of the present disclosure, the content of the compound of formula (I-1), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof in the pharmaceutical composition is 1%-95%.

[0107] In certain embodiments of the present disclosure, in the pharmaceutical composition, the pharmaceutically acceptable carrier includes one or more of a filler, a disintegrant, a binder, a glidant, and a lubricant.

[0108] The present disclosure also provides the use of the compound of formula (I-1), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts, or pharmaceutical compositions thereof for preparing drugs for treating diseases mediated by selective proteasome inhibitors.

[0109] In some embodiments, the disease is a tumor, an infectious disease, an autoimmune disease, or a central nervous system disease, preferably an autoimmune disease.

[0110] In some embodiments, the autoimmune disease is selected from systemic lupus erythematosus, arthritis, myasthenia gravis, polymyositis, dermatomyositis, autoimmune gastritis, autoimmune hepatitis, lupus nephritis, Goodpasture's syndrome, IgA nephropathy, IgG4-related nephropathy, Crohn's disease, ulcerative colitis, psoriasis, or multiple sclerosis.

[0111] The present disclosure also provides a method for preventing or treating a disease mediated by a selective proteasome inhibitor, comprising administering to a subject in need thereof an effective preventive or therapeutic amount of a compound of formula (I-1), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof, or a pharmaceutical composition thereof.

[0112] In some embodiments, the disease is a tumor, an infectious disease, an autoimmune disease, or a central nervous system disease, preferably an autoimmune disease.

[0113] In some embodiments, the autoimmune disease is selected from systemic lupus erythematosus, arthritis, myasthenia gravis, polymyositis, dermatomyositis, autoimmune gastritis, autoimmune hepatitis, lupus nephritis, Goodpasture's syndrome, IgA nephropathy, IgG4-related nephropathy, Crohn's disease, ulcerative colitis, psoriasis, or multiple sclerosis.

[0114] Explanation of terms

[0115] Unless otherwise indicated, the following terms and phrases used in this disclosure are intended to have the following meanings: A particular term or phrase should not be considered ambiguous or unclear without a specific definition, but should be understood according to its ordinary meaning.

[0116] The term "pharmaceutically acceptable" refers to those compounds, materials, compositions and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problems or complications, commensurate with a reasonable benefit / risk ratio.

[0117] The term "pharmaceutically acceptable salt" refers to derivatives of the disclosed compounds prepared with relatively nontoxic acids or bases. These salts can be prepared during compound synthesis, isolation, or purification, or by reacting the purified free form of the compound with a suitable acid or base. When the compound contains relatively acidic functional groups, base addition salts, including those based on alkali and alkaline earth metal cations, are obtained by reaction with alkali metal or alkaline earth metal hydroxides or organic amines. When the compound contains relatively basic functional groups, acid addition salts are obtained by reaction with organic or inorganic acids.

[0118] The term "isotopic derivative" refers to a compound in which at least one atom is replaced by an atom having the same atomic number but a different atomic mass. Examples of isotopes that can be introduced into the compounds of the present disclosure include stable and radioactive isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, chlorine, bromine, and iodine, for example, 2 H (deuterium, D), 3 H (tritium, T), 11 C. 13 C. 14 C. 15 N. 17 O. 18 O. 32 p、 33 p、 33 S. 34 S. 35 S. 36 S. 18 F. 36 Cl, 82 Br, 123 I. 124 I. 125 I. 129 I and 131I, etc., preferably deuterium. Compared to non-deuterated drugs, deuterated drugs have advantages such as reduced toxic side effects, increased drug stability, enhanced therapeutic efficacy, and extended drug biological half-life. All isotopic composition changes of the compounds disclosed herein, whether radioactive or not, are included in the scope of this disclosure. Each available hydrogen atom attached to a carbon atom can be independently replaced by a deuterium atom, wherein the deuterium replacement can be partial or complete, and partial deuterium replacement means that at least one hydrogen is replaced by at least one deuterium.

[0119] The term "nitrogen oxide" or "N-oxide" refers to a derivative formed by further oxidation of the nitrogen atom in a nitrogen-containing group. Common N-oxides include N-oxides of tertiary amines or N-oxides of nitrogen atoms in nitrogen-containing heterocycles.

[0120] The term "prodrug" refers to certain derivatives of the disclosed compounds that have little or no pharmacological activity themselves, which have cleavable groups and decompose into the disclosed compounds through solvent decomposition or under physiological conditions. The types of prodrugs include, but are not limited to, amides, esters, anhydrides, salts, and the like. The "ester" refers to a derivative formed with a suitable alcohol when the disclosed compound contains an acidic group (such as a carboxylic acid); or a derivative formed with a suitable acid (including an organic acid or an inorganic acid) when the disclosed compound contains a hydroxyl group. Methods for preparing prodrugs are well known to those skilled in the art.

[0121] The compounds of the present disclosure can exist in unsolvated as well as solvated forms, including hydrated forms. In general, the solvated forms are equivalent to unsolvated forms and are also encompassed within the scope of the present disclosure.

[0122] The compounds disclosed herein exist as isomers, such as cis-trans isomers, enantiomers, diastereomers, and racemic mixtures and other mixtures thereof, all of which fall within the scope of the present disclosure.

[0123] The term "enantiomer" refers to stereoisomers that are mirror images of one another.

[0124] The term "diastereomer" refers to stereoisomers that have two or more chiral centers and are not mirror images of each other.

[0125] The term "cis-trans isomers" refers to configurations in which a molecule cannot rotate freely about a double bond or a single bond of a ring-forming carbon atom.

[0126] Unless otherwise specified, use a solid wedge key. and dotted wedge key Indicates the absolute configuration of a stereocenter.

[0127] Stereoisomers of the compounds disclosed herein can be prepared by chiral synthesis, chiral reagents, or other conventional techniques. For example, one enantiomer of a compound disclosed herein can be prepared by asymmetric catalysis or chiral auxiliary derivatization techniques. Alternatively, a single stereoisomer of the compound can be obtained from a mixture using chiral resolution techniques. Alternatively, the enantiomer can be prepared directly using chiral starting materials. Separation of optically pure compounds disclosed herein is typically accomplished using preparative chromatography, employing chiral chromatographic columns to achieve the purpose of separating chiral compounds.

[0128] The term "pharmaceutically acceptable carrier" refers to a medium generally accepted in the art for delivering biologically active agents to animals, particularly mammals, and includes adjuvants, excipients, or vehicles, such as diluents, preservatives, fillers, flow regulators, disintegrants, wetting agents, emulsifiers, suspending agents, sweeteners, flavoring agents, fragrances, antibacterial agents, antifungal agents, lubricants, and dispersants, depending on the mode of administration and the nature of the dosage form. Pharmaceutically acceptable carriers are formulated within the purview of those skilled in the art based on a wide range of factors. These include, but are not limited to, the type and nature of the active agent being formulated, the subject to whom the composition containing the agent is to be administered, the intended route of administration of the composition, and the intended therapeutic indication. Pharmaceutically acceptable carriers include both aqueous and non-aqueous media, as well as a variety of solid and semisolid dosage forms. In addition to the active agent, such carriers include a variety of different ingredients and additives, and the inclusion of additional ingredients in a formulation for various reasons (e.g., to stabilize the active agent, binders, etc.) is well known to those skilled in the art. The term "excipient" generally refers to a carrier, diluent and / or vehicle required to formulate an effective pharmaceutical composition.

[0129] The term "effective preventive or therapeutic amount" means that the compound of the present invention or its pharmaceutically acceptable salt refers to a sufficient amount of the compound to treat the disorder at a reasonable effect / risk ratio applicable to any medical treatment and / or prevention. However, it should be recognized that the total daily dosage of the compound represented by formula (I) or its pharmaceutically acceptable salt and composition of the present invention must be determined by the attending physician within the scope of sound medical judgment. For any specific patient, the specific therapeutically effective dosage level must be determined based on a variety of factors, including the disorder being treated and the severity of the disorder; the activity of the specific compound used; the specific composition used; the patient's age, weight, general health, sex and diet; the administration time, route of administration and excretion rate of the specific compound used; the duration of treatment; drugs used in combination with or concurrently with the specific compound used; and similar factors well known in the medical field. For example, it is practiced in the art to start the dose of the compound at a level lower than that required to obtain the desired therapeutic effect and gradually increase the dose until the desired effect is achieved.

[0130] The term "optionally substituted" as used herein refers to two situations in which one or more hydrogen atoms of the substituted group may be "substituted" or "unsubstituted" by one or more substituents.

[0131] In this disclosure, “R 2 With Y 3 and its adjacent carbon atoms together form a 5-8 membered heterocyclic ring" refers to a structural unit R on the left side of the center 2 With Y 3 and its adjacent carbon atoms together form a 5-8 membered heterocyclic ring, the 5-8 membered heterocyclic ring does not contain a phenyl structure on the right side; for example, when Y 3 With R 2 Together When the corresponding R 2 With Y 3 and its adjacent carbon atoms together form a 7-membered heterocyclic ring; the 5-8-membered heterocyclic ring contains 1-3 heteroatoms selected from N, O or S. Unless otherwise specified, "ring" refers to saturated, partially saturated or unsaturated monocyclic and polycyclic rings.

[0132] Unless otherwise specified, the term "heterocyclyl" refers to a substituted or unsubstituted saturated or unsaturated non-aromatic ring containing 1-3 heteroatoms selected from N, O, or S. The "heterocyclyl" described in the present disclosure refers to a non-aromatic cyclic group containing at least one heteroatom as a ring atom and derived by removing one hydrogen atom; including saturated or partially saturated monocyclic heterocyclyls and polycyclic heterocyclyls; the heterocyclyl group is not related to the connection position (i.e., it can be connected through a carbon atom or a heteroatom). The polycyclic heterocyclyl includes fused heterocyclyl, spiro heterocyclyl, and bridged heterocyclyl.

[0133] The "fused heterocyclic group" refers to a cyclic structure formed by two or more rings sharing two adjacent ring atoms, and at least one of the rings is a heterocyclic ring; the fused heterocyclic group includes a cyclic structure formed by the fusion of a monoheterocyclic group with a monoheterocyclic group, a cycloalkyl group, an aryl group, or a heteroaryl group, and also includes a cyclic structure formed by the fusion of a heteroaryl group with a cycloalkyl group or a monoheterocyclic group.

[0134] The "spiro heterocyclic group" refers to a cyclic structure formed by two or more rings sharing a ring atom, and at least one ring is a heterocyclic ring.

[0135] The "bridged heterocyclic group" refers to a cyclic structure formed by two or more rings sharing non-adjacent ring atoms, and at least one ring is a heterocyclic ring.

[0136] The heterocyclic group disclosed herein is preferably a 4-10 membered monoheterocyclic group, a 5-10 membered fused heterocyclic group, a 5-10 membered spiro heterocyclic group or a 5-10 membered bridged heterocyclic group; further, the heterocyclic group is preferably a 4-7 membered monoheterocyclic group, a 6-10 membered spiro heterocyclic group or a 7-9 membered bridged heterocyclic group; the heterocyclic group is optionally further substituted by one or more selected from =O, halogen, C 1-4 Alkyl, C 1-4 The alkoxy group is substituted by a substituent, and examples of heterocyclic groups include but are not limited to: wait.

[0137] Unless otherwise specified, the term "aryl" refers to an unsaturated, typically aromatic hydrocarbon group which may be a single ring or multiple rings fused together. Examples of 6-10 membered aryl groups include, but are not limited to, phenyl and naphthyl.

[0138] Unless otherwise specified, the term "heteroaryl" means a stable monocyclic or polycyclic aromatic hydrocarbon, preferably containing carbon atoms and 1, 2, 3 or 4 ring heteroatoms independently selected from N, O and S. Preferably it is a 5-12 membered heteroaryl, more preferably a 5-10 membered heteroaryl;

[0139] Unless otherwise specified, "cycloalkyl" refers to a saturated monocyclic or polycyclic hydrocarbon group. 3-8 Cycloalkyl, more preferably C 3-6 Cycloalkyl. Examples of cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and the like.

[0140] Unless otherwise specified, the term "alkyl" is used to denote a straight-chain or branched saturated hydrocarbon group. 1-6 Alkyl, more preferably C 1-4 Examples of the alkyl group include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, pentyl, isopentyl, neopentyl, n-hexyl, and the like.

[0141] Unless otherwise specified, the term "halogen" means a fluorine, chlorine, bromine or iodine atom.

[0142] Unless otherwise specified, the term "haloalkyl" refers to an alkyl group substituted with a halogen, preferably a haloC 1-4 Alkyl groups, examples include but are not limited to -CF3, -CH2CF3, -CH2CHF2, -CH2CH2F, and the like.

[0143] The term "alkoxy" refers to an alkyl group defined in the present disclosure connected to another group through an oxygen atom, i.e., "-O-alkyl". 1-6 Alkoxy" (structure is -OC 1-6 alkyl), "C 1-4"alkoxy", specific examples include but are not limited to methoxy, ethoxy, propoxy, 1-methylethoxy, butoxy, 2-methylbutoxy, 3-methylbutoxy, 1,1-dimethylpropoxy, 1,2-dimethylpropoxy, etc.; preferably, the "alkoxy" described in the present disclosure is preferably C 1-4 Alkoxy, more preferably C 1-2 Alkoxy.

[0144] Unless otherwise specified, the term "alkenyl" refers to a radical derived from a straight-chain or branched alkene (containing at least one carbon-carbon double bond) by removing a hydrogen atom, including "C 2-6 Alkenyl", "C 2-5 Alkenyl", "C 2-4 Alkenyl", "C 2-3 "Alkenyl", specific examples include but are not limited to: -CH=CH2, -CH=CHCH3, -C(CH2)=CH2, -CH=CHCH2CH3, -CH2CH=CHCH3, etc.

[0145] Unless otherwise specified, the term "alkynyl" refers to a radical derived from a straight-chain or branched alkyne (containing at least one carbon-carbon triple bond) by removing a hydrogen atom, including "C 2-5 Alkynyl", "C 2-4 Alkynyl", "C 2-3 "Alkynyl", specific examples include but are not limited to: -C≡CH, -C≡CHCH3, HC≡CHCH2-, HC≡CC≡C-, etc.

[0146] Specifically, all combinations of substituents and / or variations thereof disclosed herein are permissible only if such combinations result in stable compounds. BRIEF DESCRIPTION OF THE DRAWINGS

[0147] The drawings described herein are used to provide further understanding of the present disclosure and constitute a part of the present disclosure. The exemplary embodiments of the present disclosure and their descriptions are used to explain the present disclosure and do not constitute improper limitations on the present disclosure.

[0148] Figure 1 is a graph showing the arthritis score of the CAIA mouse model;

[0149] Figure 2 shows the cumulative urine protein values ​​in the systemic lupus erythematosus mouse model;

[0150] Figure 3 shows the lymphadenopathy and skin lesion scores in the systemic lupus erythematosus mouse model. DETAILED DESCRIPTION

[0151] To make the objectives, technical solutions, and advantages of the present disclosure more clearly understood, the present disclosure is further described in detail below with reference to the accompanying drawings and examples. It is apparent that the described examples are only some of the examples of the present disclosure, and not all of them. All other examples derived by those skilled in the art based on the present disclosure are intended to fall within the scope of protection of the present disclosure.

[0152] The naming of the title compounds in this disclosure is converted from the compound structure with the help of Chemdraw. If there is an inconsistency between the compound name and the compound structure, it can be determined by integrating relevant information and reaction routes; if it cannot be confirmed by other means, the given compound structure shall prevail. The preparation method of some compounds in this disclosure quotes the preparation method of the aforementioned similar compounds. Those skilled in the art should be aware that when using or referring to the preparation method cited, the feed ratio of the reactants, the reaction solvent, the reaction temperature, etc. can be appropriately adjusted according to the different reactants.

[0153] Summary of experimental instruments:

[0154] The structures of the compounds were confirmed by nuclear magnetic resonance (NMR) and / or liquid chromatography-mass spectrometry (LC-MS) or ultra-performance liquid chromatography-mass spectrometry (UPLC-MS). NMR chemical shifts (δ) are given in parts per million (ppm). NMR measurements were performed using a Bruker Neo 400M or Bruker Ascend 400 NMR instrument. The solvents used were deuterated dimethyl sulfoxide (DMSO-d6), deuterated methanol (CD3OD), deuterated chloroform (CDCl3), and deuterated water (D2O). The internal standard was tetramethylsilane (TMS).

[0155] Liquid chromatography-mass spectrometry (LC-MS) was performed using an Agilent 1260-6125B single quadrupole mass spectrometer (electrospray ionization as the ion source).

[0156] Ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS) was performed using a Waters UPLC H-class SQD mass spectrometer (electrospray ionization as the ion source).

[0157] HPLC analysis was performed using Waters e2695-2998 or Waters ARC and Agilent 1260 or Agilent Poroshell HPH high performance liquid chromatography.

[0158] Preparative HPLC was performed using a Waters 2555-2489 (10 μm, ODS 250 cm×5 cm) or a GILSON Trilution LC.

[0159] Chiral HPLC was performed using Waters Acquity UPC2 and a Daicel Chiralpak AD-H column (5 μm, 4.6×250 mm).

[0160] Supercritical fluid chromatography (SFC) was performed using a Waters SFC 80Q.

[0161] The starting materials and intermediates used in the examples can be synthesized according to methods known in the art or purchased commercially. Experimental procedures in the examples where specific conditions are not specified were generally performed under conventional conditions or as recommended by the raw material or product manufacturers. Reagents where the specific sources are not specified were purchased commercially.

[0162] The reaction progress in the examples can be monitored by conventional methods such as thin layer chromatography (TLC) and LC-MS. The eluent system for column chromatography and the developing solvent system for thin layer chromatography used for purification can be composed of one or more of the following solvents: dichloromethane, methanol, n-hexane, ethyl acetate, petroleum ether, ethyl acetate, acetone, dichloromethane, etc. The volume ratio of the solvent is adjusted according to the polarity of the compound, and a small amount of alkaline or acidic reagents such as triethylamine, acetic acid, trifluoroformic acid, etc. can also be added for adjustment.

[0163] Unless otherwise specified, all reactions in the examples were carried out under continuous magnetic stirring in a dry nitrogen or argon atmosphere, with dry solvents and reaction temperatures expressed in degrees Celsius or °C.

[0164] Unless otherwise specified, percentages (%) used in this disclosure refer to mass percentages for solid-liquid mixtures and solid-solid mixtures, and volume percentages for liquid-liquid mixtures. Ratios of components in mixed solvents and eluents used in this disclosure refer to volume ratios.

[0165] The compounds disclosed herein can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combining them with other chemical synthesis methods, and equivalent substitutions well known to those skilled in the art. Preferred embodiments include, but are not limited to, the examples disclosed herein.

[0166] Intermediate INT-1:

[0167] (S)-2-Amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate

[0168] Reaction route:

[0169] Steps:

[0170] Step A: Dissolve (S)-2-((tert-Butoxycarbonyl)amino)-3-(cyclopent-1-en-1-yl)propanoic acid (96 g, 0.37 mol) in 500 mL of a 1 / 1 tetrahydrofuran / dichloromethane mixture, cool to 0°C, and sequentially add isobutyl chloroformate (61.6 g, 0.45 mol) and N-methylmorpholine (45.6 g, 0.45 mol). After addition, react at 0°C under nitrogen for 1 hour. Dissolve N,O-dimethylhydroxylamine hydrochloride (44 g, 0.45 mol) in dichloromethane, add triethylamine (49.4 g, 0.491 mol) at 0°C, and stir for 0.5 hour. Slowly add this mixture to the above reaction mixture at 0-5°C, warm to room temperature, and continue reacting for 16 hours.

[0171] The reaction of the raw materials was monitored by thin layer chromatography. Water and dichloromethane (100 mL) were added to the reaction solution, stirred and separated, and the organic phase was separated, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated. The residue was purified by column chromatography (eluent: petroleum ether / ethyl acetate = 30 / 1 to 3 / 1) to give 90 g of tert-butyl (S)-(3-(cyclopent-1-en-1-yl)-1-(methoxy(methyl)amino)-1-carbonylpropyl-2-yl)carbamate.

[0172] 1 H NMR (400MHz, CDCl3): δ5.46(s,1H),5.08(m,1H),4.81(m,1H),3.93(m,1H),3.78 (s,3H),3.21(s,3H),2.49(m,1H),2.15-2.35(m,4H),1.83(m,2H),1.43(s,9H).

[0173] Step B: To a solution of tert-butyl (S)-(3-(cyclopent-1-en-1-yl)-1-(methoxy(methyl)amino)-1-carbonylprop-2-yl)carbamate (30 g, 0.1 mol) in tetrahydrofuran (300 mL) was added a solution of isopropenylmagnesium bromide in tetrahydrofuran (500 mL, 0.5 mol) dropwise at 0°C. After the addition was complete, the reaction mixture was stirred at 0°C for 2 hours, then quenched with saturated aqueous ammonium chloride (500 mL), and the resulting mixture was extracted with ethyl acetate (400 mL × 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography (eluent: petroleum ether / ethyl acetate = 100 / 1) to give 15 g of tert-butyl (S)-(1-(cyclopent-1-en-1-yl)-4-methyl-3-carbonylpent-4-en-2-yl)carbamate.

[0174] 1H NMR (400MHz, CDCl3): δ6.06(s,1H),5.89(s,1H),5.41(m,1H),5.13-5.18(m,2H),2.56 (m,1H),2.25-2.36(m,4H),2.15(m,1H),1.89(s,3H),1.75-1.83(m,2H),1.41(s,9H).

[0175] Step C: At room temperature and nitrogen, tert-butyl (S)-(1-(cyclopent-1-en-1-yl)-4-methyl-3-carbonylpent-4-en-2-yl)carbamate (5 g, 21.81 mmol) was dissolved in N,N-dimethylformamide (200 mL), cooled to -20°C, and 10% sodium hypochlorite solution (29.4 mL, 39.4 mmol) was added dropwise over a period of about 30-40 minutes, followed by reaction at 0°C for 1 hour.

[0176] After thin-layer chromatography monitoring showed the disappearance of the starting material, water was added to dilute the mixture, and the mixture was extracted twice with a mixed solvent of ethyl acetate / petroleum ether = 10 / 1 (200 mL). The organic phases were combined, washed with saturated brine (20 mL), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure. The resulting residue was purified by column chromatography (eluent: petroleum ether / ethyl acetate = 250 / 1) to give 1.3 g of (S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-carbonylpropyl-2-yl-carbamic acid tert-butyl ester.

[0177] MS (ESI) M / Z: 318.3 [M+Na] + .

[0178] Step D: Dissolve (S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-carbonylprop-2-yl-carbamic acid tert-butyl ester (1.3 g, 4.41 mmol) in dichloromethane (15 mL), cool to 0°C, add 5 mL of trifluoroacetic acid, and stir at 0°C for 0.5 hours.

[0179] The reaction of the raw materials was monitored by thin layer chromatography, and the mixture was concentrated to dryness below 25°C. Dichloromethane was then added to continue concentration, which was repeated three times. Finally, the mixture was pulled dry with an oil pump to obtain 1.3 g of (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate.

[0180] 1H NMR (400MHz, DMSO-d6): δ8.13(s,3H),5.53(s,1H),4.16(m,1H),3.10(m,1H),3.1 2(m,1H),2.62(m,1H),2.49(m,1H),2.15-2.25(m,4H),1.83(m,2H),1.43(s,3H).

[0181] Intermediate INT-2:

[0182] (2-Morpholinoacetyl)-L-Alanine

[0183] Reaction route:

[0184] Steps:

[0185] Step A: Dissolve tert-Butyloxycarbonyl-L-alanine (20 g, 0.11 mol) in N,N-dimethylformamide (50 mL) and add sodium carbonate (17 g, 0.16 mol). Cool the reaction system to 10°C and add benzyl bromide (16 g, 0.11 mol) dropwise. Stir at room temperature for 2 hours.

[0186] The reaction solution was poured into water (150 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to obtain 30 g of tert-butyloxycarbonyl-L-alanine benzyl ester, which was used directly in the next step.

[0187] Step B: Dissolve tert-butyloxycarbonyl-L-alanine benzyl ester (28 g, 0.10 mol) in dichloromethane (100 mL), slowly add 8 M hydrogen chloride in dioxane solution (10 mL), and stir at room temperature for 1 hour.

[0188] After the reaction of the raw materials was completed by monitoring by thin layer chromatography, the reaction solution was spin-dried, slurried with ethyl acetate and filtered to obtain 18.5 g of L-alanine benzyl ester hydrochloride solid, which was directly used in the next reaction.

[0189] Step C: Dissolve 4-morpholineacetic acid (10 g, 68.9 mmol) and L-alanine benzyl ester hydrochloride (16 g, 75.8 mmol) in N,N-dimethylformamide (30 mL). Add N,N-diisopropylethylamine (26.7 g, 0.21 mol) and O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (31.4 g, 82.7 mmol) in sequence and stir at room temperature for 2 hours.

[0190] The reaction mixture was poured into 100 mL of water and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified on a silica gel column (eluent: petroleum ether / ethyl acetate = 2 / 1) to obtain 15 g of (2-morpholinoacetyl)-L-alanine benzyl ester.

[0191] MS (ESI) M / Z: 306.82 [M+H] + .

[0192] Step D: Dissolve (2-morpholinoacetyl)-L-alanine benzyl ester (15 g, 48.9 mmol) in methanol (50 mL), add 10% palladium carbon (1.5 g), replace the hydrogen atmosphere three times, and hydrogenate at room temperature and normal pressure for 1 hour.

[0193] After the reaction of the raw materials was monitored by thin layer chromatography, the mixture was filtered and the filter cake was washed with a small amount of methanol. The filtrates were combined and dried to obtain 12 g of (2-morpholinoacetyl)-L-alanine, which was used directly in the next reaction.

[0194] Intermediate INT-3:

[0195] (S)-2-Amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate

[0196] Reaction route:

[0197] Steps:

[0198] Step A: Crude (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (0.6 g, 1.9 mmol) was dissolved in methyl tert-butyl ether (6 mL), and p-toluenesulfonic acid (0.33 g, 1.9 mmol) was added at 0°C. The suspension was stirred at 0°C for 1 hour, filtered, and the filter cake was dried under vacuum to give 0.56 g of (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate.

[0199] MS (ESI) M / Z: 196.1 [M+H] + .

[0200] 1H NMR (400MHz, DMSO-d6): δ8.04(s,2H),7.48(d,J=8.0Hz,2H),7.12(d,J=7.9Hz,2H),5.53(s,1H),4.16(dd,J=8. 3,4.9Hz,1H),3.20(d,J=4.9Hz,1H),3.08(d,J=4.8Hz,1H),2.58(dd,J=14.6,4.0Hz,1H),2.42(dd,J=14.6,8.1 Hz,1H),2.29(s,3H),2.28-2.12(m,4H),1.83(p,J=7.0,6.5Hz,2H),1.44(s,3H).

[0201] Intermediate INT-4:

[0202] Methyl (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate

[0203] Reaction route:

[0204] Steps:

[0205] Step A: Dissolve (R)-2-(N-(N'-Benzylprolyl)amino)diphenylmethanone-nickel(II)-glycine complex (5 g, 10.04 mmol) and 4-(difluoromethoxy)benzaldehyde (5.18 g, 30.11 mmol, 3.98 mL) in methanol (50 mL). Add sodium methoxide in methanol (5.5 M, 9.12 mL) and stir at 20°C for 12 hours. Then, add dilute hydrochloric acid (2 M, 100.36 mL) and heat the reaction to 50°C and stir for 2 hours.

[0206] After LC-MS monitoring indicated complete reaction of the starting material, the reaction mixture was concentrated under reduced pressure to remove the methanol and filtered. The reaction mixture was then extracted with dichloromethane (100 mL x 5). The organic phase was discarded, and the aqueous phase was concentrated under reduced pressure. The residue was purified without further purification to yield 8 g of crude (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropionic acid hydrochloride.

[0207] MS (ESI) M / Z: 248.0 [M+H] + .

[0208] Step B: Dissolve (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid hydrochloride (8 g, 32.36 mmol) in methanol (120 mL) at 0°C, add thionyl chloride (11.55 g, 97.09 mmol, 7.05 mL) dropwise, and stir at 50°C for 12 hours.

[0209] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was diluted with water (30 mL), the pH adjusted to 9 with aqueous ammonia, and then extracted with ethyl acetate (40 mL x 2). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified without further purification to yield 2.3 g of methyl (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate.

[0210] MS (ESI) M / Z: 262.2 [M+H] + .

[0211] 1 H NMR (400MHz, CD3OD): δ7.43(d,J=8.6Hz,2H),7.15(d,J=8.6Hz,2H),7.03-6.62(m,1H),4.98(d,J=4.6Hz,1H),3.69(s,3H),3.63(d,J=4.5Hz,1H).

[0212] Intermediate INT-5:

[0213] Benzyl (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropionate

[0214] Reaction route:

[0215] Steps:

[0216] Step A: (R)-2-(N-(N'-Benzylprolyl)amino)diphenylmethanone-nickel(II)-glycine complex (3 g, 6.02 mmol) and 4-(difluoromethoxy)benzaldehyde (5.18 g, 30.10 mmol) were dissolved in methanol (30 mL). A methanolic solution of sodium methoxide (5.5 M, 5.57 mL) was added and stirred at 20°C for 12 hours. Dilute hydrochloric acid (2 M, 60.2 mL) was then added and the reaction was heated to 50°C and stirred for 2 hours.

[0217] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was concentrated under reduced pressure to remove the methanol and filtered. The reaction solution was then extracted with dichloromethane (100 mL x 5). The organic phase was discarded, and the aqueous phase was purified via cation exchange resin (Dowex 50WX8) to yield 1.3 g of (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropionic acid.

[0218] MS (ESI) M / Z: 248.0 [M+H] + .

[0219] Step B: Dissolve (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid (200 mg, 0.81 mmol) in benzyl alcohol (2 mL) at 0°C, add thionyl chloride (481.83 mg, 4.05 mmol, 0.92 mL) dropwise, and stir at 90°C for 12 hours.

[0220] LC-MS / MS monitoring indicated a small amount of residual starting material. The reaction mixture was quenched with water (10 mL). The aqueous phase was adjusted to pH 9 with aqueous ammonia and extracted with ethyl acetate (20 mL x 2). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 1 / 1) to yield 150 mg of benzyl (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate.

[0221] MS (ESI) M / Z: 338.1 [M+H] + .

[0222] 1 H NMR (400MHz, CDCl3): δ7.38-7.29(m,5H),7.19(dd,J=6.6,2.9Hz,2H),7.08-7.02(m,2 H), 6.46 (t, J = 73.9Hz, 1H), 5.08 (s, 2H), 4.80 (d, J = 5.6Hz, 1H), 3.60 (d, J = 5.6Hz, 1H).

[0223] Intermediate INT-6:

[0224] 3-tert-Butyl-4-methyl-(4S,5R)-5-(4-bromophenyl)-2,2-dimethyloxazolidine-3,4-dicarboxylate

[0225] Reaction route:

[0226] Steps:

[0227] Step A: Dissolve (S)-3-(4-bromophenyl)-2-(tert-butoxycarbonylamino)propanoic acid (20 g, 58.11 mmol) in N,N-dimethylformamide (400 mL), add iodomethane (24.74 g, 174.32 mmol, 10.85 mL) and potassium carbonate (16.06 g, 116.21 mmol), and stir at 25°C for 12 hours.

[0228] After LC-MS monitoring showed that the raw material reaction was complete, water (800 mL) was added to dilute the reaction mixture, and then extracted with ethyl acetate (400 mL×2). The organic phases were combined, washed with saturated brine (300 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain 23.3 g of (S)-3-(4-bromophenyl)-2-(tert-butoxycarbonylamino)propionic acid methyl ester.

[0229] MS (ESI) M / Z: 358.0 [M+H] + .

[0230] Step B: Dissolve (S)-methyl 3-(4-bromophenyl)-2-(tert-butoxycarbonylamino)propanoate (28 g, 78.16 mmol) in acetonitrile (500 mL), add di-tert-butyl dicarbonate (20.47 g, 93.80 mmol, 21.55 mL) and 4-dimethylaminopyridine (954.89 mg, 7.82 mmol), and stir at 70°C for 2 hours.

[0231] After LC-MS monitoring indicated that the reaction of the starting materials was complete, the reaction solution was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (eluent: petroleum ether / ethyl acetate, 5 / 1 gradient) to obtain 27.6 g of methyl (S)-2-(bis(tert-butyloxycarbonyl)amino)-3-(4-bromophenyl)propanoate.

[0232] MS (ESI) M / Z: 458.1 [M+H] + .

[0233] 1 H NMR (400MHz, CDCl3): δ1.41(s,18H),3.18(dd,J=14.01,10.19Hz,1H),3.39(dd,J=14.01,5.19H z,1H),3.75(s,3H),5.12(dd,J=10.19,5.19Hz,1H),7.07(d,J=8.34Hz,2H),7.37-7.42(m,2H).

[0234] Step C: Methyl (S)-2-(bis(tert-butoxycarbonyl)amino)-3-(4-bromophenyl)propanoate (15 g, 32.73 mmol) was dissolved in carbon tetrachloride (300 mL). N-bromosuccinimide (6.87 g, 38.62 mmol) was added. The reaction solution was then irradiated with an incandescent lamp and stirred at reflux at 80°C for 1 hour. The reaction solution was concentrated under reduced pressure, and the residue was dissolved in acetone (750 mL). Silver nitrate (10.04 g, 59.10 mmol) was added. The mixture was then stirred at 20°C for 12 hours in the dark.

[0235] After thin-layer chromatography monitoring indicated complete reaction of the starting material, the reaction solution was filtered and concentrated under reduced pressure. The residue (100 mL) was diluted with saturated aqueous ammonium chloride (100 mL) and extracted with ethyl acetate (100 mL x 2). The combined organic phases were washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: petroleum ether / ethyl acetate, 3 / 1 gradient) to yield 7.14 g of 3-tert-butyl-4-methyl-(4S,5R)-5-(4-bromophenyl)-2-oxooxazolidine-3,4-dicarboxylate.

[0236] MS (ESI) M / Z: 400.1 [M+H] + .

[0237] 1 H NMR (400MHz, CDCl3): δ1.51 (s, 9H), 3.91 (s, 3H), 4.60 (d, J = 4.41Hz, 1H), 5.36 (d, J = 4.41Hz, 1H), 7.25-7.31 (m, 2H), 7.58-7.62 (m, 2H).

[0238] Step D: Dissolve 3-tert-butyl-4-methyl-(4S,5R)-5-(4-bromophenyl)-2-oxooxazolidine-3,4-dicarboxylate (7.1 g, 17.74 mmol) in methanol (140 mL), add cesium carbonate (1.16 g, 3.55 mmol) at 0°C, and stir at 0°C for 1 hour.

[0239] After LC-MS monitoring indicated complete reaction of the starting materials, potassium bisulfate aqueous solution was added to adjust the pH to 2-3. The mixture was extracted with ethyl acetate (20 mL x 2). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: petroleum ether / ethyl acetate, 3 / 1 gradient) to yield 3.7 g of (2S,3R)-3-(4-bromophenyl)-2-(tert-butoxycarbonylamino)-3-hydroxy-propionic acid methyl ester.

[0240] MS (ESI) M / Z: 374.0 [M+H] + .

[0241] 1 H NMR (400MHz, CDCl3): δ7.50 (d, J = 8.28Hz, 2H), 7.26-7.31 (m, 2H), 5.23 (m, 2H), 4.48-4.59 (m, 1H), 3.80 (s, 3H), 1.36 (br s, 6H), 1.26-1.31 (m, 3H).

[0242] Step E: Methyl (2S,3R)-3-(4-bromophenyl)-2-(tert-butoxycarbonylamino)-3-hydroxy-propionate (4.5 g, 12.02 mmol) and 2,2-dimethoxypropane (5.40 g, 51.88 mmol, 6.36 mL) were dissolved in acetone (65 mL), and then boron trifluoride etherate (170.67 mg, 1.20 mmol, 147.89 μL) was added dropwise and stirred at 20 °C for 2 hours.

[0243] After LC-MS monitoring indicated complete reaction of the starting materials, aqueous sodium bicarbonate (1 mL) and water (40 mL) were added to the reaction solution, followed by extraction with ethyl acetate (50 mL x 2). The combined organic phases were washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: petroleum ether / ethyl acetate, 10 / 1 gradient) to yield 4.29 g of 3-tert-butyl-4-methyl-(4S,5R)-5-(4-bromophenyl)-2,2-dimethyloxazolidine-3,4-dicarboxylate.

[0244] MS (ESI) M / Z: 414.0 [M+H] + .

[0245] 1 H NMR (400MHz, DMSO-d6): δ7.60(d,J=8.5Hz,2H),7.40(d,J=8.5Hz,2H),5.06(d,J=8.1 Hz,1H),4.14-3.99(m,1H),3.71-3.61(m,3H),1.67-1.56(m,6H),1.44-1.28(m,9H).

[0246] Example 1:

[0247] (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-carbonylpropan-2-yl)-3-(4-cyclopropyloxyphenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0248] Reaction route:

[0249] Steps:

[0250] Step A: Dissolve p-iodophenol (8.5 g, 38.63 mmol) in N-methylpyrrolidone (150 mL). Add cesium carbonate (25.2 g, 77.26 mmol) and cyclopropyl bromide (11.7 g, 96.63 mmol) under nitrogen protection, and stir at 130°C overnight.

[0251] Thin-layer chromatography revealed 40%-50% residual starting material. The reaction system was cooled to room temperature, and ethyl acetate and water were added, stirred, and the layers separated. Extraction was then performed with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated and purified by column chromatography to yield 2.5 g of 1-cyclopropyloxy-4-iodobenzene.

[0252] 1 H NMR (400MHz, CDCl3): δ7.53-7.57(m,2H),6.81-6.83(m,2H),3.68(m,1H),0.75-0.78(m,4H).

[0253] Step B: Zinc powder (890 mg, 13.67 mmol) was added to N,N-dimethylacetamide (25 mL). Trimethylsilyl chloride (100 mg, 0.91 mmol) was added under nitrogen and stirred for 30 minutes. Iodine (117 mg, 0.46 mmol) was then added and stirring continued for 10 minutes. After cooling to 10-20°C, a solution of (R)-methyl 2-(tert-butyloxycarbonyl)amino)-3-iodopropionate (4.5 g, 13.67 mmol) in N,N-dimethylacetamide (10 mL) was slowly added and stirred for half an hour. Finally, 1-cyclopropyloxy-4-iodobenzene (1.2 g, 4.56 mmol), 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (112.5 mg, 0.274 mmol), and tris(dibenzylideneacetone)palladium (125 mg, 0.137 mmol) were added and the reaction was stirred at 50°C overnight.

[0254] Completion of the reaction was monitored by thin-layer chromatography. The reaction system was cooled to room temperature, water and ethyl acetate were added, and the filtrate was extracted three times with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was spin-dried to dryness. The crude product was purified by column chromatography to yield 700 mg of methyl (S)-2-(tert-butyloxycarbonyl)amino)-3-(4-cyclopropyloxyphenyl)propionate.

[0255] MS (ESI) M / Z: 335.9 [M+H] +.

[0256] Step C: A solution of methyl (S)-2-(tert-butoxycarbonyl)amino)-3-(4-cyclopropyloxyphenyl)propanoate (700 mg, 2.08 mmol) in dichloromethane (10 mL) was cooled to 0°C, trifluoroacetic acid (10 mL) was added, and the mixture was stirred at 0°C for 30 minutes.

[0257] The reaction of the raw materials was monitored by thin layer chromatography, concentrated to dryness below 25°C and replaced with dichloromethane three times to obtain 700 mg of (S)-2-amino-3-(4-cyclopropyloxyphenyl)propionic acid methyl ester trifluoroacetate.

[0258] Step D: Dissolve (2-morpholinoacetyl)-L-alanine (441 mg, 2.0 mmol) and (S)-2-amino-3-(4-cyclopropyloxyphenyl)propionic acid methyl ester trifluoroacetate (441 mg, 2.0 mmol) in N,N-dimethylformamide (12 mL), add 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (841 mg, 2.2 mmol), cool to 0-5°C, add N,N-diisopropylethylamine (439 mg, 3.4 mmol) to adjust the pH to 7, and warm to room temperature for 1 hour.

[0259] The reaction was monitored for completion by thin-layer chromatography. Ethyl acetate and water were added to the reaction mixture, stirred, and the layers separated. Extraction was then performed with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by column chromatography to yield 200 mg of methyl (S)-3-(4-cyclopropyloxyphenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[0260] MS (ESI) M / Z: 581.0 [M+H] + .

[0261] Step E: Dissolve (S)-methyl 3-(4-cyclopropyloxyphenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate (250 mg, 0.58 mmol) in methanol (6 mL), add lithium hydroxide monohydrate (24.2 mg, 0.58 mmol) under ice-cooling, and warm to room temperature for 1 hour.

[0262] The reaction was monitored for completion by thin-layer chromatography. The reaction solution was extracted with ethyl acetate / petroleum ether (1 / 1), and the aqueous phase was adjusted to pH 7 with 1N dilute hydrochloric acid and lyophilized to obtain 200 mg of (S)-3-(4-cyclopropyloxyphenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionic acid lithium salt.

[0263] Step F: (S)-3-(4-cyclopropyloxyphenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionic acid lithium salt (200 mg, 0.4 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (135 mg, 0.4 mmol) were dissolved in N,N-dimethylformamide (20 mL), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (209 mg, 0.54 mmol) was added, and the mixture was cooled to 0-5°C. N,N-diisopropylethylamine (65 mg, 0.50 mmol) was added to adjust the pH to 7, and the mixture was heated to room temperature for 1 hour.

[0264] The reaction was monitored for completion by thin-layer chromatography. Ethyl acetate and water were added to the reaction solution, stirred, and the layers separated, followed by extraction with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by column chromatography to yield 28 mg of (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-carbonylprop-2-yl)-3-(4-cyclopropyloxyphenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide.

[0265] MS (ESI) M / Z: 597.5 [M+H] + .

[0266] 1 HNMR (400MHz, DMSO-d6): δ8.26(d,J=7.2Hz,1H),8.03(d,J=8.0Hz,1H),7.73(d,J=7.6H z,1H),7.10(d,J=8.4Hz,2H),6.88(d,J=8.4Hz,2H),5.39(s,1H),4.45-4.55(m,2H),4.2 5(m,1H),3.83(m,1H),3.55-3.58(m,4H),3.15(m,1H),2.75-2.91(m,3H),2.63(m,1H),2 .35(m,5H),2.15-2.24(m,5H),1.85(m,2H),1.35(s,3H),1.15(d,J=7.6Hz,3H),0.75(m, 2H),0.61(m,2H).

[0267] Example 2:

[0268] (S)-3-(4-(azetidin-1-yl)phenyl)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0269] Reaction route:

[0270] Steps:

[0271] Step A: Add azetidine hydrochloride (2 g, 21.3 mmol) to dioxane (60 mL), followed by 1,4-dibromobenzene (4.93 g, 19.4 mmol) and 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (2.17 g, 3.88 mmol). Replace the atmosphere with nitrogen twice, then add sodium tert-butoxide (5.6 g, 58.2 mmol) and tris(dibenzylideneacetone)palladium (1.78 g, 1.94 mmol). Heat the temperature to 60°C and react for 3 hours.

[0272] The reaction mixture was cooled to room temperature and filtered. The filtrate was poured into water and extracted three times with ethyl acetate. The combined ethyl acetate phases were washed three times with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and spin-dried. The crude product was purified by silica gel column chromatography to yield 1.7 g of 1-(4-bromophenyl)azetidine.

[0273] 1 H NMR (400MHz, CDCl3): δ7.28 (d, J = 8.8 Hz, 2H), 6.32 (d, J = 8.8 Hz, 2H), 3.85 (t, J = 7.2 Hz, 4H), 2.36 (m, 2H).

[0274] Step B: Add zinc powder (1.4 g, 21.2 mmol) to N,N-dimethylformamide (30 mL). Under nitrogen, add trimethylsilyl chloride (77 mg, 0.71 mmol). Stir for 30 minutes, then add iodine (180 mg, 0.71 mmol). Continue stirring for 10 minutes. Then, add (R)-methyl 2-tert-butoxycarbonylamino-3-iodopropionate (9.3 g, 28.3 mmol). Continue stirring for 30 minutes, then add 1-(4-bromophenyl)azetidine (1.5 g, 7.07 mmol), 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (174 mg, 0.42 mmol), and tris(dibenzylideneacetone)palladium (194 mg, 0.21 mmol). Heat the temperature to 50°C and stir overnight.

[0275] The reaction mixture was cooled to room temperature and filtered. The filtrate was poured into water and extracted three times with ethyl acetate. The combined ethyl acetate phases were washed three times with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and spin-dried. The crude product was purified by silica gel column chromatography to obtain 1.8 g of (S)-3-(4-(azetidin-1-yl)phenyl)-2-tert-butoxycarbonyl-aminopropionic acid methyl ester.

[0276] MS (ESI) M / Z: 335.3 [M+H] + .

[0277] Step C: Dissolve (S)-3-(4-(azetidin-1-yl)phenyl)-2-tert-butoxycarbonyl-aminopropionic acid methyl ester (250 mg, 0.75 mmol) in dichloromethane (5 mL), add trifluoroacetic acid (0.4 mL, 3.74 mmol), and stir at room temperature for 1 hour.

[0278] After thin layer chromatography monitoring showed the disappearance of the starting material, the reaction solution was concentrated to dryness to obtain 250 mg of (S)-3-(4-(azetidin-1-yl)phenyl)-2-aminopropionic acid methyl ester trifluoroacetate.

[0279] MS (ESI) M / Z: 235.0 [M+H] + .

[0280] Step D: (2-Morpholinoacetyl)-L-alanine (160 mg, 0.72 mmol) and (S)-3-(4-(azetidin-1-yl)phenyl)-2-aminopropionic acid methyl ester trifluoroacetate (250 mg, 0.72 mmol) were dissolved in N,N-dimethylformamide (5 mL), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (330 mg, 0.86 mmol) was added, and then the pH was adjusted to about 7 with N,N-diisopropylethylamine and stirred at room temperature for 30 minutes.

[0281] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to yield 150 mg of methyl (S)-3-(4-(azetidin-1-yl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate.

[0282] MS (ESI) M / Z: 433.5 [M+H] + .

[0283] Step E: Dissolve (S)-methyl 3-(4-(azetidin-1-yl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate (130 mg, 0.30 mmol) in tetrahydrofuran / water (2 mL / 2 mL), add lithium hydroxide monohydrate (25 mg, 0.60 mmol), and stir at room temperature for 1 hour.

[0284] After the reaction of the raw materials was monitored by thin layer chromatography, the pH was adjusted to 7-8 with 1 M hydrochloric acid and lyophilized to obtain 120 mg of crude (S)-3-(4-(azetidin-1-yl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionic acid lithium salt, which was directly used in the next reaction.

[0285] Step F: (S)-3-(4-(azetidin-1-yl)phenyl)-2-((S)-2-(2-morpholinoacetamido)propionamido)propanoic acid lithium salt (120 mg, 0.29 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (90 mg, 0.29 mmol) were dissolved in N,N-dimethylformamide (5 mL), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (132 mg, 0.35 mmol) was added, and then the pH was adjusted to about 7 with N,N-diisopropylethylamine and stirred at room temperature for 30 minutes.

[0286] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to yield 21.66 mg of (S)-3-(4-(azetidin-1-yl)phenyl)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide.

[0287] MS (ESI) M / Z: 596.5 [M+H] + .

[0288] 1H NMR (400MHz, CDCl3): δ7.54(br,1H),7.06(d,J=8.0Hz,2H),6.63(m,1H),6.3 5(d,J=8.4Hz,2H),6.05(d,J=6.4Hz,1H),5.31(m,1H),4.58-4.35(m,3H),3.8 3(m,4H),3.73(m,4H),3.27(m,1H),3.13-2.82(m,5H),2.63-2.40(m,5H),2.3 5(m,2H),2.27-2.15(m,5H),1.83(m,2H),1.50(s,3H),1.36(d,J=6.8Hz,3H).

[0289] Example 3:

[0290] (S)-N-((S)-3-(cyclopent-1-enyl-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-3-(4-difluoromethoxy)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0291] Reaction route:

[0292] Steps:

[0293] Step A: Add zinc powder (2.4 g, 37.05 mmol) to N,N-dimethylformamide (30 mL). Add trimethylsilyl chloride (161 mg, 1.48 mmol) under nitrogen and stir for 30 minutes. Then add iodine (188 mg, 0.74 mmol) and continue stirring for 10 minutes. Cool the reaction system to 10-20°C and slowly add a solution of methyl (R)-2-(tert-butoxycarbonyl)amino)-3-iodopropionate (12.2 g, 37.05 mmol) in N,N-dimethylformamide (10 mL) and stir for half an hour. Finally, 1-(difluoromethoxy)-4-iodobenzene (2 g, 7.41 mmol), 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (180.5 mg, 0.44 mmol) and tris(dibenzylideneacetone)palladium (203.4 mg, 0.22 mmol) were added, and the temperature was raised to 50°C and stirred for reaction overnight.

[0294] The reaction was monitored for completion by thin-layer chromatography. The reaction mixture was filtered after adding ethyl acetate and water, and the filtrate was extracted three times with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by column chromatography to yield 1.3 g of methyl (S)-2-((tert-butoxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)propanoate.

[0295] MS (ESI) M / Z: 368.3 [M+Na] + .

[0296] Step B: Dissolve (S)-methyl 2-((tert-butoxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)propanoate (1.3 g, 3.75 mmol) in methanol (6 mL). Add lithium hydroxide monohydrate (320 mg, 0.58 mmol) in water (3 mL) under ice-cooling, and warm to room temperature for 1 hour.

[0297] The reaction was monitored for completion by thin-layer chromatography. The reaction solution was back-extracted with a 1 / 1 mixture of ethyl acetate and petroleum ether. The aqueous phase was adjusted to pH 2 with 1N HCl and extracted three times with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to dryness to yield 900 mg of (S)-2-((tert-butoxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)propanoic acid.

[0298] MS (ESI) M / Z: 331.3 [M+H] + .

[0299] Step C: Dissolve (S)-2-((tert-Butoxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)propanoic acid (900 mg, 2.72 mmol) in N,N-dimethylformamide (15 mL), add potassium bicarbonate (350 mg, 3.53 mmol) and benzyl bromide (560 mg, 3.26 mmol) in sequence, and stir at room temperature for 1 hour.

[0300] Completion of the reaction was monitored by thin-layer chromatography. The reaction mixture was quenched with water and extracted three times with ethyl acetate / petroleum ether (1 / 1). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. Column chromatography of the crude product yielded 800 mg of benzyl (S)-2-((tert-butoxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)propanoate.

[0301] MS (ESI) M / Z: 422.1 [M+H] + .

[0302] Step D: Dissolve benzyl (S)-2-((tert-butoxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)propanoate (800 mg, 2.41 mmol) in dichloromethane (10 mL), cool to 0°C, add 8M hydrogen chloride solution in 1,4-dioxane (3 mL), and stir at room temperature for 1 hour.

[0303] The reaction of the raw materials was monitored by thin layer chromatography. The reaction solution was dried and replaced with dichloromethane three times to obtain 700 mg of (S)-2-amino-3-(4-(difluoromethoxy)phenyl)propionic acid benzyl ester hydrochloride.

[0304] Step E: Dissolve (S)-benzyl 2-amino-3-(4-(difluoromethoxy)phenyl)propanoate hydrochloride (500 mg, 1.39 mmol) in N,N-dimethylformamide (10 mL), add (2-morpholinoacetyl)-L-alanine (332 mg, 1.53 mmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (691 mg, 1.80 mmol), cool to 0-5°C, add N,N-diisopropylethylamine (361 mg, 2.8 mmol) to adjust the pH to 7, and warm to room temperature for 1 hour.

[0305] Completion of the reaction of the starting materials was monitored by thin-layer chromatography. The reaction system was quenched with water and extracted three times with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by column chromatography to yield 600 mg of benzyl (S)-3-(4-(difluoromethoxy)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate.

[0306] MS (ESI) M / Z: 520.1 [M+H] + .

[0307] Step F: Dissolve benzyl (S)-3-(4-(difluoromethoxy)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate (600 mg, 1.16 mmol) in methanol (10 mL), add 10% palladium on carbon (60 mg), and hydrogenate under normal pressure for 1 hour.

[0308] The reaction was monitored by thin layer chromatography until completion, palladium-carbon was removed by filtration, and the filtrate was concentrated to obtain 500 mg of (S)-3-(4-(difluoromethoxy)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionic acid.

[0309] MS (ESI) M / Z: 430.3 [M+H] + .

[0310] Step G: (S)-3-(4-(difluoromethoxy)phenyl)-2-((S)-2-(2-morpholinoacetamido)propionamido)propanoic acid (200 mg, 0.16 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (162 mg, 0.5 mmol) were dissolved in N,N-dimethylformamide (15 mL), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (231 mg, 0.6 mmol) was added, and the mixture was cooled to 0-5°C. Diisopropylethylamine (120 mg, 0.93 mmol) was added to adjust the pH to 7, and the mixture was warmed to room temperature for 1 hour.

[0311] The reaction of the raw materials was monitored by thin layer chromatography. Ethyl acetate and water were added to the reaction solution, stirred and separated, and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated. 17.6 mg of (S)-N-((S)-3-(cyclopent-1-enyl-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-3-(4-difluoromethoxy)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide was obtained by thin layer chromatography.

[0312] MS (ESI) M / Z: 607.5 [M+H] + .

[0313] 1 HNMR (400MHz, DMSO-d6): δ8.34(d,J=7.6Hz,1H),8.11(d,J=8.4Hz,1H),7.73(d,J=7.6 Hz,1H),7.21(d,J=8.4Hz,2H),7.13(t,1H),7.03(d,J=8.4Hz,2H),5.41(s,1H),4.45- 4.55(m,2H),4.25(m,1H),3.55-3.58(m,4H),3.15(m,1H),2.75-2.91(m,4H),2.63(m, 1H),2.35(m,5H),2.15-2.24(m,5H),1.85(m,2H),1.35(s,3H),1.15(d,J=7.6Hz,3H).

[0314] Example 4:

[0315] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-3-(4-(difluoromethoxy)phenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0316] Reaction route:

[0317] Steps:

[0318] Step A: Potassium hydroxide (14.1 g, 252 mmol) was dissolved in ethanol (200 mL), and glycine (7.9 g, 105 mmol) and 4-(difluoromethoxy)benzaldehyde (21 g, 0.125 mmol) were added respectively. The mixture was stirred at room temperature for 16 hours. The reaction was monitored by thin layer chromatography. The mixture was concentrated to dryness and slurried with isopropyl ether to obtain crude 2-amino-3-(4-(difluoromethoxy)phenyl)-3-hydroxypropionic acid, which was directly used in the next step.

[0319] MS (ESI) M / Z: 247.99 [M+H] + .

[0320] Step B: Dissolve the crude 2-amino-3-(4-(difluoromethoxy)phenyl)-3-hydroxypropanoic acid from the previous step in water (40 mL). Add sodium bicarbonate (13.3 g, 0.157 mmol) under ice-cooling. Add dropwise a solution of phenylacetyl chloride (12.9 g, 84 mmol) in acetone (40 mL). After addition, warm to room temperature and react for 16 hours.

[0321] After the reaction of the raw materials was monitored by thin layer chromatography, the mixture was washed twice with a 1 / 1 mixed solvent of petroleum ether / ethyl acetate. The aqueous phase was adjusted to pH 1-2 with 6N hydrochloric acid, ethyl acetate was added, stirred and separated, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and purified by column chromatography to obtain 4.3 g of 3-(4-(difluoromethoxy)phenyl)-3-hydroxy-2-(2-phenylacetylamino)propionic acid.

[0322] MS (ESI) M / Z: 366.2 [M+1] + .

[0323] Step C: Dissolve 3-(4-(difluoromethoxy)phenyl)-3-hydroxy-2-(2-phenylacetylamino)propionic acid (4.3 g, 3.41 mmol) in pure water (100 mL), add 30% sodium hydroxide to adjust the pH to 7-7.5, raise the temperature to 38°C, add penicillin hydrolase (430 mg), and react at pH 7-7.5 for 48 hours.

[0324] The reaction solution was adjusted to pH 5-6 with 6N HCl, 430 mg of activated carbon was added, the temperature was raised to 60°C and stirred for 1 hour, the activated carbon was removed by filtration, and the solution was extracted three times with ethyl acetate. The aqueous phase was directly used for the next reaction.

[0325] Step D: Add the aqueous phase from step C into a reaction flask, adjust the pH to 10-11 with 30% sodium hydroxide under ice bath, add di-tert-butyl dicarbonate (5.1 g, 23.48 mmol), and react at room temperature for 6 hours.

[0326] The reaction of the raw materials was monitored by thin-layer chromatography. The reaction solution was washed twice with petroleum ether. The aqueous phase was adjusted to pH 1-2 with 3N hydrochloric acid in an ice bath. Ethyl acetate was added, stirred, and separated and extracted. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to dryness to obtain 1.8 g of (2S)-2-(tert-butoxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)-3-hydroxypropionic acid.

[0327] MS (ESI) M / Z: 315.35 [M+H] + .

[0328] Step E: 1.8 g of (2S)-2-(tert-butoxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)-3-hydroxypropionic acid was separated by chiral column to give 86 mg of (2S,3R)-2-(tert-butoxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)-3-hydroxypropionic acid and 56 mg of (2S,3S)-2-(tert-butoxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)-3-hydroxypropionic acid.

[0329] Separation conditions: Superchiral S-AD preparative column, 2.1 cm ID (inner diameter) × 25 cm length, 5 μm; mobile phase: n-hexane / ethanol / formic acid = 80 / 20 / 0.05; flow rate: 20 mL / min; temperature: 35°C.

[0330] Step F: (2S,3R)-2-(tert-Butyloxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)-3-hydroxypropanoic acid (86 mg, 0.23 mmol) was dissolved in N,N-dimethylformamide (6 mL), potassium carbonate (95 mg, 0.69 mmol) and iodomethane (42.5 mg, 0.30 mmol) were added respectively, and the reaction was stirred at room temperature for 16 hours.

[0331] The reaction of the raw materials was monitored by thin layer chromatography. Ethyl acetate and water were added to the reaction solution, stirred and separated, and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to obtain 80 mg of methyl (2S,3R)-2-(tert-butoxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)-3-hydroxypropanoate.

[0332] Step G: Methyl (2S,3R)-2-(tert-butoxycarbonyl)amino)-3-(4-(difluoromethoxy)phenyl)-3-hydroxypropanoate (80 mg, 0.81 mmol) was dissolved in dichloromethane (3 mL). Trifluoroacetic acid (3 mL) was added under ice-cooling and the mixture was warmed to room temperature for 1 hour.

[0333] The reaction of the raw materials was monitored by thin layer chromatography, the reaction solution was concentrated to dryness, and the crude product was replaced with tetrahydrofuran 2-3 times to obtain 80 mg of (2S,3R)-2-amino-3-(4-(difluoromethoxy)phenyl)-3-hydroxypropionic acid methyl ester trifluoroacetate.

[0334] MS (ESI) M / Z: 262.2 [M+H] + .

[0335] Step H: Dissolve (2S,3R)-2-amino-3-(4-(difluoromethoxy)phenyl)-3-hydroxypropionic acid methyl ester trifluoroacetate (80 mg, 0.3 mmol) in N,N-dimethylformamide (5 mL), add (2-morpholinoacetyl)-L-alanine (74 mg, 0.33 mmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (151 mg, 0.39 mmol), cool to 0-5°C, add N,N-diisopropylethylamine (79 mg, 0.61 mmol) to adjust the pH to 7, and warm to room temperature for 1 hour.

[0336] The reaction of the raw materials was monitored by thin layer chromatography. Ethyl acetate and water were added to the reaction solution, stirred and separated, and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness to obtain 70 mg of (2S,3R)-3-(4-(difluoromethoxy)phenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionic acid methyl ester.

[0337] MS (ESI) M / Z: 460.47 [M+H] + .

[0338] Step I: Methyl (2S,3R)-3-(4-(difluoromethoxy)phenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate (70 mg, 0.15 mmol) was dissolved in tetrahydrofuran (2 mL), and lithium hydroxide (9.6 mg, 0.22 mmol) in 2 mL of water was added under ice-cooling. The mixture was reacted at room temperature for 1 hour. The reaction was completed by thin layer chromatography. The mixture was washed twice with petroleum ether / ethyl acetate = 1 / 1 under alkaline conditions. The aqueous phase was adjusted to pH = 7 with 1N hydrochloric acid and lyophilized to obtain 70 mg of lithium (2S,3R)-3-(4-(difluoromethoxy)phenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionate.

[0339] MS (ESI) M / Z: 446.5 [M+H] + .

[0340] Step J: Lithium (2S,3R)-3-(4-(difluoromethoxy)phenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionate (70 mg, 0.16 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (45.6 mg, 0.143 mmol) were dissolved in N,N-dimethylformamide (4 mL), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (71 mg, 0.185 mmol) was added, the mixture was cooled to 0-5°C, N,N-diisopropylethylamine (37 mg, 0.29 mmol) was added to adjust the pH to 7, and the mixture was warmed to room temperature for 1 hour.

[0341] The reaction of the raw materials was monitored by thin layer chromatography. Ethyl acetate and water were added to the reaction solution, stirred and separated, and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated. The mixture was purified by thin layer chromatography to obtain 13.42 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-3-(4-(difluoromethoxy)phenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide.

[0342] MS (ESI) M / Z: 623.6 [M+H] +

[0343] 1H NMR (400MHz, DMSO-d6): δ8.01(d,J=7.6Hz,1H),7.90(d,J=8.8Hz,1H),7.79(d,J=7.6Hz,1H),7 .36(d,J=8.8Hz,2H),7.19(t,1H),7.01(d,J=8.4Hz,2H),5.68(d,J=4.4Hz,1H),5.43(m,1H),4 .98(m,1H),4.55(m,1H),4.41(m,1H),4.31(m,1H),3.41-3.52(m,5H),3.15(m,1H),2.81-2.95 (m,2H),2.25-2.35(m,5H),2.15-2.23(m,5H),1.75(m,2H),1.31(s,3H),1.14(d,J=6.8Hz,3H).

[0344] Example 5:

[0345] (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionamido)-3-(4-methoxyphenyl)propionamide

[0346] Reaction route:

[0347] Steps:

[0348] Step A: Dissolve N-Fmoc-3-cyclopropyloxy-L-alanine (3 g, 6.81 mol) in N,N-dimethylformamide (10 mL) and add sodium bicarbonate (1.14 g, 13.61 mmol). Cool the reaction system to 10°C and add benzyl bromide (1.4 g, 8.17 mmol) dropwise. Stir at room temperature for 2 hours.

[0349] The reaction solution was poured into water (30 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to obtain 3.2 g of crude N-Fmoc-3-cyclopropyloxy-L-alanine benzyl ester, which was used directly in the next step.

[0350] Step B: Dissolve N-Fmoc-3-cyclopropyloxy-L-alanine benzyl ester (3 g, 6.56 mmol) in N,N-dimethylformamide (6 mL), add piperidine (1.5 mL), and stir at room temperature for 1 hour.

[0351] After completion of the reaction, the reaction mixture was poured into 30 mL of water and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by silica gel column chromatography to yield 1.6 g of 3-cyclopropyloxy-L-alanine benzyl ester.

[0352] MS (ESI) M / Z: 236.0 [M+H] + .

[0353] Step C: Dissolve morpholineacetic acid (500 mg, 3.40 mmol) and 3-cyclopropyloxy-L-alanine benzyl ester (0.8 g, 3.40 mmol) in N,N-dimethylformamide (3 mL). Add N,N-diisopropylethylamine (1.3 g, 10.2 mmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (1.55 g, 4.08 mmol) in sequence and stir at room temperature for 1 hour.

[0354] The reaction mixture was poured into water (10 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 1.2 g of benzyl (S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propanoate.

[0355] MS (ESI) M / Z: 362.9 [M+H] + .

[0356] Step D: Dissolve benzyl (S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propanoate (1.2 g, 3.31 mmol) in methanol (10 mL), add 10% palladium on carbon (0.2 g), and hydrogenate at room temperature and normal pressure for 30 minutes.

[0357] After the reaction of the raw materials was monitored by thin layer chromatography, the mixture was filtered and the filter cake was washed with a small amount of methanol. The filtrates were combined and dried to obtain 880 mg of (S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionic acid, which was used directly in the next reaction.

[0358] Step E: (S)-3-Cyclopropyloxy-2-(2-morpholinoacetylamino)propionic acid (500 mg, 1.84 mmol) and S-2-amino-3-(4-methoxyphenyl)propionic acid benzyl ester hydrochloride (591 mg, 1.84 mmol) were dissolved in N,N-dimethylformamide (5 mL), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (837 mg, 2.20 mmol) was added, and the pH was adjusted to about 7 with N,N-diisopropylethylamine and stirred at room temperature for 30 minutes.

[0359] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 550 mg of benzyl (S)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionylamino)-3-(4-methoxyphenyl)propanoate.

[0360] MS (ESI) M / Z: 540.4 [M+H] + .

[0361] Step F: Dissolve benzyl (S)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionylamino)-3-(4-methoxyphenyl)propanoate (550 mg, 1.02 mmol) in methanol (10 mL), add 10% palladium on carbon (0.1 g), and hydrogenate at room temperature and atmospheric pressure for 30 minutes.

[0362] The reaction of the starting material was monitored by thin-layer chromatography, and the mixture was filtered and the filter cake was washed with a small amount of methanol. The filtrates were combined and dried to obtain 370 mg of (S)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionylamino)-3-(4-methoxyphenyl)propionic acid, which was used directly in the next reaction.

[0363] Step G: (S)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionamido)-3-(4-methoxyphenyl)propanoic acid (100 mg, 0.22 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (70 mg, 0.22 mmol) were dissolved in N,N-dimethylformamide (5 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (101 mg, 0.27 mmol) was added, and the pH was adjusted to about 7 with N,N-diisopropylethylamine. The mixture was stirred at room temperature for 30 minutes.

[0364] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 19.38 mg of (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionamido)-3-(4-methoxyphenyl)propionamide.

[0365] MS (ESI) M / Z: 627.6 [M+H] + .

[0366] 1 H NMR (400MHz, CDCl3): δ7.80(br,1H),7.14(d,J=8.0Hz,2H),6.82(m,3H),6.17(br,1H),5.33(m,1H),4.57(m,2H),4.46(m,1H),3.9 3-3.56(m,9H),3.36-3.25(m,2H),3.12-2.85(m,4H),2.63-2.36(m,3H),2.20-2.10(m,5H),1.84(m,2H),1.49(s,3H),0.47(m,4H).

[0367] Example 6:

[0368] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionamido)-3-hydroxy-3-(4-methoxyphenyl)propionamide

[0369] Reaction route:

[0370] Steps:

[0371] Step A: (S)-3-Cyclopropyloxy-2-(2-morpholinoacetylamino)propionic acid (120 mg, 0.44 mmol) and (2S,3R)-2-amino-3-hydroxy-3-(4-methoxyphenyl)propionic acid benzyl ester hydrochloride (150 mg, 0.44 mmol) were dissolved in N,N-dimethylformamide (5 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (202 mg, 0.53 mmol) was added, and then the pH was adjusted to about 7 with N,N-diisopropylethylamine and stirred at room temperature for 30 minutes.

[0372] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 100 mg of benzyl (2S,3R)-2-((S)-3-cyclopropyl-2-(2-morpholinoacetylamino)propionylamino)-3-hydroxy-3-(4-methoxyphenyl)propanoate.

[0373] MS (ESI) M / Z: 556.5 [M+H] + .

[0374] Step B: Dissolve benzyl (2S,3R)-2-((S)-3-cyclopropyl-2-(2-morpholinoacetylamino)propionylamino)-3-hydroxy-3-(4-methoxyphenyl)propanoate (100 mg, 0.18 mmol) in methanol (5 mL), add 10% palladium on carbon (0.1 g), and hydrogenate at 20°C under normal pressure for 20 minutes.

[0375] The reaction of the starting material was monitored by thin-layer chromatography, and the mixture was filtered and the filter cake was washed with a small amount of methanol. The combined filtrates were spin-dried to obtain 84 mg of (2S,3R)-2-((S)-3-cyclopropyl-2-(2-morpholinoacetylamino)propionylamino)-3-hydroxy-3-(4-methoxyphenyl)propionic acid, which was used directly in the next reaction.

[0376] Step C: (2S,3R)-2-((S)-3-cyclopropyl-2-(2-morpholinoacetylamino)propionamido)-3-hydroxy-3-(4-methoxyphenyl)propanoic acid (84 mg, 0.18 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (56 mg, 0.18 mmol) were dissolved in N,N-dimethylformamide (5 mL), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (82 mg, 0.22 mmol) was added, and then the pH was adjusted to about 7 with N,N-diisopropylethylamine and stirred at room temperature for 30 minutes.

[0377] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 6.2 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionamido)-3-hydroxy-3-(4-methoxyphenyl)propionamide.

[0378] MS (ESI) M / Z: 643.5 [M+H] + .

[0379] 1H NMR (400MHz, CDCl3): δ7.84(br,1H),7.23(d,J=8.4Hz,2H),7.20(m,1H),6.99(d,J=6.8Hz,1H) ,6.84(d,J=8.4Hz,2H),5.47(m,1H),5.21(m,1H),4.65(m,1H),4.56(m,1H),4.45(m,1H),3.85( m,1H),3.78(s,3H),3.72(m,4H),3.56(m,1H),3.32(m,1H),3.26(m,1H),3.14(m,2H),2.90(m, 1H),2.70-2.51(m,4H),2.36-2.18(m,5H),1.87(m,2H),1.51(s,3H),1.26(m,2H),0.50(m,4H).

[0380] Example 7:

[0381] (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-2-((S)-2-(2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetamido)propionamido)-3-(4-methoxyphenyl)propionamide

[0382] Reaction route:

[0383] Steps:

[0384] Step A: Dissolve ethyl bromoacetate (9.2 g, 55.0 mmol) in tetrahydrofuran (50 mL) and add zinc powder (4 g, 60.0 mmol). Stir at room temperature for 30 minutes. Add a solution of tetrahydropyran-4-one (5 g, 50.0 mmol) in tetrahydrofuran dropwise to the reaction system. Reflux the reaction for 4 hours.

[0385] The reaction system was cooled to room temperature, quenched with water, and the acid was adjusted to neutral. Most of the tetrahydrofuran was removed by swirl. The aqueous phase was extracted three times with ethyl acetate. The combined organic phases were washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated. The crude product was purified by silica gel column chromatography to yield 5.5 g of ethyl 2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetate.

[0386] 1H NMR (400MHz, CDCl3): δ4.18 (q, J = 7.2Hz, 2H), 3.84 (m, 2H), 3.73 (m, 2H), 2.49 (s, 2H), 1.65 (m, 4H), 1.29 (d, J = 7.2Hz, 3H).

[0387] Step B: Dissolve ethyl 2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetate (2 g, 10.63 mmol) in ethanol (10 mL), add lithium hydroxide monohydrate (871 mg, 21.25 mmol), and stir at 75°C for overnight.

[0388] The reaction system was cooled to room temperature and filtered. The filtrate was adjusted to neutral with hydrochloric acid and dried to obtain 3 g of crude 2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetic acid, which was used directly in the next reaction.

[0389] MS (ESI) M / Z: 159.02 [MH] - .

[0390] Step C: 2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetic acid (1.5 g, 9.3 mmol) and L-alanine benzyl ester hydrochloride (2.2 g, 10.03 mmol) were dissolved in N,N-dimethylformamide (5 mL). N,N-diisopropylethylamine (3.6 g, 28.10 mmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (4.3 g, 11.24 mmol) were added in sequence and stirred at room temperature for 1 hour.

[0391] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 1.2 g of benzyl (S-2-(2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetamido)propanoate.

[0392] MS (ESI) M / Z: 322.26 [M+H] + .

[0393] Step D: Dissolve benzyl (S)-2-(2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetamido)propanoate (1.2 g, 3.73 mmol) in methanol (10 mL), add 10% palladium on carbon (0.1 g), and hydrogenate at room temperature and normal pressure for 1 hour.

[0394] The reaction of the raw materials was monitored by thin-layer chromatography, and the mixture was filtered and the filter cake was washed with a small amount of methanol. The filtrates were combined and dried to obtain 1 g of (S)-2-(2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetylamino)propionic acid, which was used directly in the next reaction.

[0395] Step E: (S)-2-(2-(4-Hydroxytetrahydro-2H-pyran-4-yl)acetamido)propanoic acid (860 mg, 3.72 mmol) and (S)-2-amino-3-(4-methoxyphenyl)propionic acid benzyl ester hydrochloride (1.2 g, 3.72 mmol) were dissolved in N,N-dimethylformamide (5 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (1.7 g, 4.46 mmol) was added, and the pH was adjusted to about 7 with N,N-diisopropylethylamine. The mixture was stirred at room temperature for 30 minutes.

[0396] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 1 g of benzyl (S)-2-((S)-2-(2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetylamino)propionamido)-3-(4-methoxyphenyl)propanoate.

[0397] MS (ESI) M / Z: 499.44 [M+H] + .

[0398] Step F: Dissolve benzyl (S)-2-((S)-2-(2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetylamino)propionamido)-3-(4-methoxyphenyl)propanoate (1 g, 2.01 mmol) in methanol (5 mL), add 10% palladium on carbon (0.1 g), and hydrogenate at room temperature and atmospheric pressure for 1 hour.

[0399] The reaction was monitored by thin-layer chromatography, followed by filtration, and the filter cake was washed with a small amount of methanol. The combined filtrates were spin-dried to obtain 0.8 g of (S)-2-((S)-2-(2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetylamino)propionamido)-3-(4-methoxyphenyl)propionic acid, which was used directly in the next reaction.

[0400] Step G: (S)-2-((S)-2-(2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetamido)propionamido)-3-(4-methoxyphenyl)propanoic acid (100 mg, 0.25 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (76 mg, 0.25 mmol) were dissolved in N,N-dimethylformamide (5 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (112 mg, 0.29 mmol) was added, and the pH was adjusted to about 7 with N,N-diisopropylethylamine. The mixture was stirred at room temperature for 30 minutes.

[0401] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 26.18 mg of (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-2-((S)-2-(2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetylamino)propionamido)-3-(4-methoxyphenyl)propionamide.

[0402] MS (ESI) M / Z: 586.5 [M+H] + .

[0403] 1 H NMR (400MHz, CDCl3): δ7.17(d,J=8.4Hz,2H), 6.83(d,J=8.0Hz,2H), 6.71(d,J=7.2Hz, 1H),6.21(d,J=7.6Hz,1H),5.84(d,J=6.4Hz,1H),5.27(m,1H),4.52(m,2H),4.42(m,1 H),3.88-3.70(m,7H),3.22(d,J=4.8Hz,1H),3.02(m,1H),2.90(m,2H),2.48(m,1H),2 .33(s,2H),2.27-2.13(m,6H),1.73-1.25(m,6H),1.50(s,3H),1.36(d,J=6.8Hz,3H).

[0404] Example 8:

[0405] (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-2-((S)-2-(2-(4-fluorotetrahydro-2H-pyran-4-yl)acetamido)propionamido)-3-(4-methoxyphenyl)propionamide

[0406] Reaction route:

[0407] Steps:

[0408] Step A: Dissolve benzyl bromoacetate (5 g, 22.0 mmol) in tetrahydrofuran (10 mL) and add zinc powder (1.6 g, 24.0 mmol). Stir at room temperature for 30 minutes. Add a solution of tetrahydropyran-4-one (2 g, 20.0 mmol) in tetrahydrofuran dropwise to the reaction system. Reflux the reaction for 4 hours.

[0409] The reaction system was cooled to room temperature, quenched with water, and the acid was adjusted to neutral. Most of the tetrahydrofuran was removed by swirl. The aqueous phase was extracted three times with ethyl acetate. The combined organic phases were washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated. The crude product was purified by silica gel column chromatography to yield 3.4 g of 2-(4-hydroxytetrahydro-2H-pyran-4-yl)benzyl acetate.

[0410] Step B: Dissolve benzyl 2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetate (1.5 g, 5.99 mmol) in dichloromethane (10 mL), cool to 0°C, add diethylaminosulfur trifluoride (1.93 g, 11.99 mmol), and stir at room temperature overnight.

[0411] The reaction system was quenched with water and extracted three times with dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated. The crude product was purified by silica gel column chromatography to obtain 600 mg of 2-(4-fluorotetrahydro-2H-pyran-4-yl)benzyl acetate.

[0412] 1 H NMR (400MHz, CDCl3): δ7.34 (m, 5H), 5.15 (s, 2H), 3.78 (m, 4H), 2.70 (d, J = 18.4Hz, 2H), 1.86 (m, 4H).

[0413] Step C: Dissolve benzyl 2-(4-fluorotetrahydro-2H-pyran-4-yl)acetate (600 mg, 2.38 mmol) in methanol (5 mL), add 10% palladium on carbon (0.1 g), and hydrogenate at room temperature and normal pressure for 1 hour.

[0414] The reaction of the raw materials was monitored by thin layer chromatography, and the mixture was filtered and the filter cake was washed with a small amount of methanol. The filtrates were combined and dried to obtain 390 mg of 2-(4-fluorotetrahydro-2H-pyran-4-yl)acetic acid, which was used directly in the next reaction.

[0415] Step D: 2-(4-Fluorotetrahydro-2H-pyran-4-yl)acetic acid (385 mg, 2.37 mmol) and L-alanine benzyl ester hydrochloride (512 mg, 2.37 mmol) were dissolved in N,N-dimethylformamide (5 mL). N,N-diisopropylethylamine (920 mg, 7.12 mmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (1.1 g, 2.85 mmol) were added in sequence and stirred at room temperature for 1 hour.

[0416] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 700 mg of benzyl (S)-2-(2-(4-fluorotetrahydro-2H-pyran-4-yl)acetamido)propanoate.

[0417] Step E: Dissolve benzyl (S)-2-(2-(4-fluorotetrahydro-2H-pyran-4-yl)acetamido)propanoate (700 mg, 2.16 mmol) in methanol (10 mL), add 10% palladium on carbon (0.1 g), and hydrogenate at room temperature and normal pressure for 1 hour.

[0418] The reaction of the raw materials was monitored by thin layer chromatography, and the mixture was filtered and the filter cake was washed with a small amount of methanol. The filtrates were combined and dried to obtain 500 mg of (S)-2-(2-(4-hydroxytetrahydro-2H-pyran-4-yl)acetylamino)propionic acid, which was used directly in the next reaction.

[0419] Step F: (S)-2-(2-(4-Fluorotetrahydro-2H-pyran-4-yl)acetamido)propanoic acid (250 mg, 1.07 mmol) and (S)-2-amino-3-(4-methoxyphenyl)propionic acid benzyl ester hydrochloride (345 mg, 1.07 mmol) were dissolved in N,N-dimethylformamide (5 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (490 mg, 1.29 mmol) was added, and the pH was adjusted to about 7 with N,N-diisopropylethylamine. The mixture was stirred at room temperature for 30 minutes.

[0420] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 1 g of benzyl (S)-2-((S)-2-(2-(4-fluorotetrahydro-2H-pyran-4-yl)acetylamino)propionamido)-3-(4-methoxyphenyl)propanoate.

[0421] MS (ESI) M / Z: 501.2 [M+H] + .

[0422] Step G: Dissolve benzyl (S)-2-((S)-2-(2-(4-fluorotetrahydro-2H-pyran-4-yl)acetylamino)propionamido)-3-(4-methoxyphenyl)propanoate (370 mg, 0.74 mmol) in methanol (5 mL), add 10% palladium on carbon (0.1 g), and hydrogenate at room temperature and atmospheric pressure for 1 hour.

[0423] The reaction was monitored by thin-layer chromatography, followed by filtration, and the filter cake was washed with a small amount of methanol. The combined filtrates were spin-dried to obtain 300 mg of (S)-2-((S)-2-(2-(4-fluorotetrahydro-2H-pyran-4-yl)acetylamino)propionamido)-3-(4-methoxyphenyl)propionic acid, which was used directly in the next reaction.

[0424] Step H: (S)-2-((S)-2-(2-(4-fluorotetrahydro-2H-pyran-4-yl)acetamido)propionamido)-3-(4-methoxyphenyl)propanoic acid (100 mg, 0.24 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (75 mg, 0.24 mmol) were dissolved in N,N-dimethylformamide (5 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (111 mg, 0.29 mmol) was added, and the pH was adjusted to about 7 with N,N-diisopropylethylamine. The mixture was stirred at room temperature for 30 minutes.

[0425] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 40.44 mg of (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-2-((S)-2-(2-(4-fluorotetrahydro-2H-pyran-4-yl)acetylamino)propionamido)-3-(4-methoxyphenyl)propionamide.

[0426] MS (ESI) M / Z: 588.5 [M+H] + .

[0427] 1 H NMR (400MHz, CDCl3): δ7.16(d,J=8.8Hz,2H),6.82(d,J=8.8Hz,2H),6.60(d ,J=7.2Hz,1H),6.24(m,1H),5.95(m,1H),5.29(m,1H),4.52(m,2H),4.42(m ,1H),3.85-3.64(m,6H),3.26(m,1H),3.04-2.85(m,3H),2.57-2.42(m,3H) ,2.26-2.11(m,5H),1.92-1.63(m,6H),1.49(m,4H),1.34(d,J=6.8Hz,3H).

[0428] Example 9:

[0429] (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-2-((S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetamido)propionamido)-3-(4-methoxyphenyl)propionamide

[0430] Reaction route:

[0431] Steps:

[0432] Step A: Dissolve triethyl phosphonoacetate (67.2 g, 0.3 mol) in tetrahydrofuran (200 mL), cool to 0°C, and add sodium hydroxide (12 g, 0.3 mol) portionwise. Stir at 0°C for 30 minutes, then add a solution of tetrahydropyran-4-one (10 g, 0.1 mol) in tetrahydrofuran dropwise. Stir at room temperature for 1 minute.

[0433] The reaction system was quenched with saturated ammonium chloride, and most of the tetrahydrofuran was removed by swirl. The aqueous phase was extracted three times with ethyl acetate. The combined organic phases were washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by silica gel column chromatography to obtain 6.1 g of ethyl (tetrahydropyran-4-ylidene)acetate.

[0434] Step B: Dissolve ethyl (tetrahydropyran-4-ylidene)acetate (6 g, 35.3 mmol) in methanol (60 mL), cool to zero degree, add sodium methoxide (5.72 g, 0.11 mol), and stir at room temperature for 3 hours.

[0435] The reaction mixture was slowly poured into ice water containing 12 g of potassium bisulfate and extracted three times with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by silica gel column chromatography to obtain 1.6 g of methyl 2-(4-methoxytetrahydro-2H-pyran-4-yl)acetate.

[0436] 1 H NMR (400MHz, CDCl3): δ3.85-3.66(m,7H),3.26(s,3H),2.54(s,2H),1.85-1.70(m,4H).

[0437] Step C: Dissolve methyl 2-(4-methoxytetrahydro-2H-pyran-4-yl)acetate (800 mg, 4.26 mmol) in tetrahydrofuran / water (2 mL / 2 mL), add lithium hydroxide monohydrate (255 mg, 6.38 mmol), and stir at room temperature for 1 hour.

[0438] Completion of the reaction was monitored by thin-layer chromatography. The pH was adjusted to 3-4 with potassium bisulfate and extracted three times with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated to yield 720 mg of crude 2-(4-methoxytetrahydro-2H-pyran-4-yl)acetic acid, which was used directly in the next reaction.

[0439] Step D: 2-(4-Methoxytetrahydro-2H-pyran-4-yl)acetic acid (700 mg, 4.02 mmol) and L-alanine benzyl ester hydrochloride (868 mg, 4.02 mmol) were dissolved in N,N-dimethylformamide (5 mL). N,N-diisopropylethylamine (1.6 g, 12.07 mmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (1.8 g, 4.83 mmol) were added in sequence and stirred at room temperature for 1 hour.

[0440] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to yield 1.2 g of benzyl (S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetamido)propanoate.

[0441] MS (ESI) M / Z: 336.4 [M+H] + .

[0442] Step E: Dissolve benzyl (S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetamido)propanoate (700 mg, 2.09 mmol) in tetrahydrofuran / water (2 mL / 2 mL), add lithium hydroxide monohydrate (132 mg, 3.14 mmol), and stir at room temperature for 1 hour.

[0443] The reaction was monitored by thin-layer chromatography, and the pH was adjusted to 3-4 with potassium bisulfate. The mixture was extracted six times with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to yield 390 mg of crude (S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetamido)propanoic acid, which was used directly in the next reaction.

[0444] Step F: (S)-2-(2-(4-Methoxytetrahydro-2H-pyran-4-yl)acetamido)propanoic acid (390 mg, 1.59 mmol) and (S)-2-amino-3-(4-methoxyphenyl)propionic acid benzyl ester hydrochloride (512 mg, 1.59 mmol) were dissolved in N,N-dimethylformamide (5 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (725 mg, 1.91 mmol) was added, and the pH was adjusted to about 7 with N,N-diisopropylethylamine. The mixture was stirred at room temperature for 30 minutes.

[0445] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 490 mg of benzyl (S)-2-((S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetylamino)propionamido)-3-(4-methoxyphenyl)propanoate.

[0446] MS (ESI) M / Z: 513.6 [M+H] + .

[0447] Step G: Dissolve benzyl (S)-2-((S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetylamino)propionamido)-3-(4-methoxyphenyl)propanoate (490 mg, 0.96 mmol) in methanol (5 mL), add 10% palladium on carbon (0.1 g), and hydrogenate at room temperature and atmospheric pressure for 1 hour.

[0448] The reaction was monitored by thin-layer chromatography, followed by filtration, and the filter cake was washed with a small amount of methanol. The combined filtrates were spin-dried to obtain 360 mg of (S)-2-((S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetylamino)propionamido)-3-(4-methoxyphenyl)propionic acid, which was used directly in the next reaction.

[0449] Step H: (S)-2-((S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetamido)propionamido)-3-(4-methoxyphenyl)propanoic acid (180 mg, 0.43 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (132 mg, 0.43 mmol) were dissolved in N,N-dimethylformamide (5 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (194 mg, 0.51 mmol) was added, and the pH was adjusted to about 7 with N,N-diisopropylethylamine. The mixture was stirred at room temperature for 30 minutes.

[0450] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 94.3 mg of (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-2-((S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetylamino)propionamido)-3-(4-methoxyphenyl)propionamide.

[0451] MS (ESI) M / Z: 600.5 [M+H] + .

[0452] 1 H NMR (400MHz, CDCl3): δ7.15 (d, J = 8.4Hz, 2H), 7.00 (m, 1H), 6.82 (d, J = 8.4Hz, 2H), 6.67 (d, J = 7. 2Hz,1H),6.05(d,J=6.4Hz,1H),5.30(m,1H),4.52(m,2H),4.42(m,1H),3.78(s,3H),3.70(m,4 H),3.27(d,J=4.8Hz,1H),3.21(s,3H),2.96(m,2H),2.89(d,J=4.8Hz,1H),2.48(m,1H),2.40( m,1H),2.27-2.12(m,5H),1.81(m,4H),1.66-1.52(m,2H),1.49(s,3H),1.33(d,J=7.2Hz,3H).

[0453] Example 10:

[0454] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-3-hydroxy-3-(4-methoxyphenyl)-2-((S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetamido)propionamido)propionamide

[0455] Reaction route:

[0456] Steps:

[0457] Step A: Dissolve (S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetamido)propanoic acid (200 mg, 0.82 mmol) and benzyl (2S,3R)-2-amino-3-hydroxy-3-(4-methoxyphenyl)propanoate hydrochloride (308 mg, 0.82 mmol) in N,N-dimethylformamide (5 mL). Add 2-(7-azobenzotriazole)-N,N,N,N-tetramethyluronium hexafluorophosphate (370 mg, 0.98 mmol) and adjust the pH to about 7 with N,N-diisopropylethylamine. Stir at room temperature for 30 minutes.

[0458] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The combined organic phases were washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 320 mg of benzyl (2S,3R)-3-hydroxy-3-(4-methoxyphenyl)-2-((S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetylamino)propionamido)propanoate.

[0459] MS (ESI) M / Z: 529.5 [M+H] + .

[0460] Step B: Dissolve benzyl (2S,3R)-3-hydroxy-3-(4-methoxyphenyl)-2-((S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetamido)propionamido)propanoate (320 mg, 0.61 mmol) in methanol (5 mL), add 10% palladium on carbon (0.1 g), and hydrogenate at room temperature and atmospheric pressure for 1 hour.

[0461] The reaction of the starting material was monitored by thin-layer chromatography, and the mixture was filtered and the filter cake was washed with a small amount of methanol. The combined filtrates were spin-dried to obtain 260 mg of (2S,3R)-3-hydroxy-3-(4-methoxyphenyl)-2-((S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetylamino)propionamido)propionic acid, which was used directly in the next reaction.

[0462] Step C: (2S,3R)-3-hydroxy-3-(4-methoxyphenyl)-2-((S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetamido)propionamido)propanoic acid (130 mg, 0.30 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (92 mg, 0.30 mmol) were dissolved in N,N-dimethylformamide (5 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (135 mg, 0.36 mmol) was added, and the pH was adjusted to about 7 with N,N-diisopropylethylamine. The mixture was stirred at room temperature for 30 minutes.

[0463] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 35.48 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-3-hydroxy-3-(4-methoxyphenyl)-2-((S)-2-(2-(4-methoxytetrahydro-2H-pyran-4-yl)acetamido)propionamido)propionamide.

[0464] MS (ESI) M / Z: 616.5 [M+H] + .

[0465] 1 H NMR (400MHz, CDCl3): δ7.24(m,2H),6.95-6.82(m,4H),5.45(m,1H),5.21(m,1H),4.61(m,2H),4.40(m,1H), 3.78(s,3H),3.68(m,4H),3.26(d,J=4.8Hz,1H),3.19(s,3H),2.90(d,J=4.8Hz,1H),2.56(m,1H),2.4 0(s,2H),2.38-2.02(m,6H),1.88-1.74(m,5H),1.67-1.42(m,2H),1.51(s,3H),1.28(d,J=7.2Hz,3H).

[0466] Example 11:

[0467] (S)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetylamino)propionamide-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-3-(4-methoxyphenyl)propionamide

[0468] Reaction route:

[0469] Steps:

[0470] Step A: Dissolve homomorpholine (500 mg, 4.93 mmol) in acetonitrile (10 mL), add potassium carbonate (3.41 g, 24.7 mmol) and tert-butyl bromoacetate (964 mg, 4.93 mmol), and stir at room temperature for 16 hours.

[0471] Completion of the reaction was monitored by thin-layer chromatography. The reaction mixture was diluted with ethyl acetate and filtered. The filtrate was stirred with water to separate the layers, and then extracted twice with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated to yield 1 g of tert-butyl 2-(1,4-oxazepan-4-yl)acetate.

[0472] MS (ESI) M / Z: 216.3 [M+H] + .

[0473] Step B: Dissolve tert-butyl 2-(1,4-oxazepan-4-yl)acetate (1 g, 1.56 mmol) in dichloromethane (15 mL), add trifluoroacetic acid (15 mL) under ice-cooling, and react at room temperature for 2 hours.

[0474] The reaction of the raw materials was monitored by thin layer chromatography. The reaction solution was concentrated and replaced with dichloromethane three times to obtain 1 g of 2-(1,4-oxazepan-4-yl)acetic acid.

[0475] MS (ESI) M / Z: 159.7 [M+H] + .

[0476] Step C: Dissolve 2-(1,4-oxazepan-4-yl)acetic acid (1 g, 3.71 mmol) and (S)-alanine benzyl ester hydrochloride (730 mg, 3.41 mmol) in N,N-dimethylformamide (20 mL), add 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (1.75 g, 4.61 mmol). Cool to 0-5°C, add N,N-diisopropylethylamine (540 mg, 4.20 mmol) to adjust the pH to 7, and warm to room temperature for 1 hour.

[0477] Completion of the reaction was monitored by thin-layer chromatography. The reaction mixture was quenched with water and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by column chromatography to yield 1.2 g of (2-(1,4-oxazepan-4-yl)acetyl)-L-alanine benzyl ester as a pale yellow oil.

[0478] MS (ESI) M / Z: 320.1 [M+H] + .

[0479] Step D: Dissolve (2-(1,4-oxazepan-4-yl)acetyl)-L-alanine benzyl ester (1.2 g, 0.30 mmol) in methanol (20 mL), add 10% palladium on carbon (120 mg), and hydrogenate under normal pressure for 1.5 hours.

[0480] The reaction of the raw materials was monitored by thin layer chromatography, palladium carbon was removed by filtration, and the filtrate was concentrated to obtain 700 mg of (2-(1,4-oxazepan-4-yl)acetyl)-L-alanine.

[0481] MS (ESI) M / Z: 231.1 [M+H] + .

[0482] Step E: Dissolve (2-(1,4-oxazepan-4-yl)acetyl)-L-alanine (550 mg, 2.39 mmol) and benzyl (S)-2-amino-3-(4-methoxyphenyl)propionate (620 mg, 2.17 mmol) in N,N-dimethylformamide (10 mL). Add 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (1.07 g, 2.82 mmol). Cool to 0-5°C, add N,N-diisopropylethylamine (561 mg, 4.35 mmol) to adjust the pH to 7, and warm to room temperature for 1 hour.

[0483] Completion of the reaction was monitored by thin-layer chromatography. Ethyl acetate and water were added to the reaction mixture, stirred, and the layers separated. Extraction was then performed with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by column chromatography to yield 800 mg of benzyl (S)-2-((S)-2-[2-(1,4-oxazepan-4-yl)acetylamino)propionamido]-3-(4-methoxyphenyl)propanoate.

[0484] MS (ESI) M / Z: 498.3 [M+H] + .

[0485] Step F: Dissolve benzyl (S)-2-((S)-2-[2-(1,4-oxazepan-4-yl)acetylamino)propionamido]-3-(4-methoxyphenyl)propanoate (800 mg, 1.61 mmol) in methanol (20 mL), add 10% palladium on carbon (80 mg), and hydrogenate at normal pressure for 1 hour.

[0486] The reaction of the starting materials was monitored by thin layer chromatography. The palladium on carbon was removed by filtration and the filtrate was concentrated to obtain 500 mg of (S)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetamido)-3-(4-methoxyphenyl)propanoic acid.

[0487] MS (ESI) M / Z: 572.6 [M+H] + .

[0488] Step G: (S)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetamido)-3-(4-methoxyphenyl)propanoic acid (140 mg, 0.34 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (100 mg, 0.31 mmol) were dissolved in N,N-dimethylformamide (10 mL), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (155 mg, 0.41 mmol) was added, and the mixture was cooled to 0-5°C. N,N-diisopropylethylamine (81 mg, 0.63 mmol) was added to adjust the pH to 7, and the mixture was warmed to room temperature for 1 hour.

[0489] The reaction of the raw materials was monitored by thin-layer chromatography. Ethyl acetate and water were added to the reaction solution, stirred, and the layers were separated, followed by extraction with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated. The crude product was purified by column chromatography to obtain 25.93 mg of (S)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetylamino)propionamide-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-3-(4-methoxyphenyl)propionamide.

[0490] MS (ESI) M / Z: 585.5 [M+H] + .

[0491] 1 HNMR (400MHz, DMSO-d6): δ8.29(d,J=7.2Hz,1H), 8.08(d,J=8.4Hz,1H), 7.75(d,J=7.6Hz,1H ),7.12(d,J=8.4Hz,2H),6.79(d,J=8.4Hz,2H),5.39(s,1H),4.41-4.52(m,2H),4.25(m,1H), 3.72(s,3H),3.65(m,2H),3.55(m,2H),3.21(m,1H),2.95-3.03(m,3H),2.88(m,1H),2.55-2 .65(m,5H),2.41(m,1H),2.15-2.25(m,5H),1.78(m,4H),1.35(s,3H),1.15(d,J=6.8Hz,3H).

[0492] Example 12:

[0493] (2S,3R)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetamido)propionamide)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-3-hydroxy-3-(4-methoxyphenyl)propionamide

[0494] Reaction route:

[0495] Steps:

[0496] Step A: Dissolve (2-(oxazepan-4-yl)acetyl)-L-alanine (150 mg, 0.65 mmol) in N,N-dimethylformamide (10 mL). Add benzyl (2S,3R)-2-amino-3-hydroxy-3-(4-methoxyphenyl)propanoate (178 mg, 0.59 mmol), 1H-benzotriazol-1-yloxytripyrrolidinyl hexafluorophosphate (462 mg, 0.88 mmol), and 1-hydroxybenzotriazole (120 mg, 0.88 mmol) in this order. Cool the reaction system to 0-5°C, add N,N-diisopropylethylamine (153 mg, 1.18 mmol) to adjust the pH to 7, and warm to room temperature for 0.5 hours.

[0497] Completion of the reaction of the raw materials was monitored by thin-layer chromatography. Ethyl acetate and water were added to the reaction mixture, stirred, and the layers separated. Extraction was then performed with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated to yield 300 mg of benzyl (2S,3R)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetamido)-3-hydroxy-3-(4-methoxyphenyl)propanoate.

[0498] MS (ESI) M / Z: 514.5 [M+H] + .

[0499] Step B: Dissolve benzyl (2S,3R)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetamido)-3-hydroxy-3-(4-methoxyphenyl)propanoate (300 mg, 0.58 mmol) in methanol (10 mL), add 10% palladium on carbon (30 mg), and hydrogenate at normal pressure for 1 hour.

[0500] The reaction of the starting materials was monitored by thin layer chromatography, palladium carbon was removed by filtration, and the mixture was concentrated to obtain 150 mg of (2S,3R)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetamido)-3-hydroxy-3-(4-methoxyphenyl)propanoic acid.

[0501] MS (ESI) M / Z: 424.49 [M+H] + .

[0502] Step C: (2S,3R)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetamido)-3-hydroxy-3-(4-methoxyphenyl)propanoic acid (100 mg, 0.47 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (58.5 mg, 0.21 mmol) was dissolved in N,N-dimethylformamide (4 mL), and 1H-benzotriazol-1-yloxytripyrrolidino hexafluorophosphate (167.7 mg, 0.32 mmol) and 1-hydroxybenzotriazole (43.4 mg, 0.322 mmol) were added sequentially. The mixture was cooled to 0-5°C, and N,N-diisopropylethylamine (55.4 mg, 0.43 mmol) was added to adjust the pH to 7. The mixture was then allowed to warm to room temperature and reacted for 1 hour.

[0503] The reaction of the starting materials was monitored by thin-layer chromatography. Ethyl acetate and water were added to the reaction mixture, stirred, and the layers separated. The mixture was then extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by column chromatography to yield 13.4 mg of (2S,3R)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetamido)propionamide)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-3-hydroxy-3-(4-methoxyphenyl)propionamide.

[0504] MS (ESI) M / Z: 601.5 [M+H] + .

[0505] 1 HNMR (400MHz, DMSO-d6): δ7.91(m,2H),7.80(m,1H),7.24(d,J=8.8Hz,2H),6.79 (d,J=8.4Hz,2H),5.52(d,J=8.8Hz,2H),5.41(s,1H),4.91(m,1H),4.55(m,1H), 4.25(m,2H),3.81(s,3H),3.75(m,2H),3.58(m,2H),2.95-3.11(m,4H),2.61(m, 2H),2.31(m,2H),2.241(m,5H),1.78(m,4H),1.35(s,3H),1.15(d,J=6.8Hz,3H).

[0506] Example 13:

[0507] (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-3-(4-cyclopropylphenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0508] Reaction route:

[0509] Steps:

[0510] Step A: Add (S)-3-(4-bromophenyl)-2-(tert-butoxycarbonyl)amino)propanoic acid (2.1 g, 5.86 mmol) to toluene (30 mL). Under nitrogen, add cyclopropylboronic acid (1.5 g, 17.85 mmol), sodium carbonate (1.86 g, 17.58 mmol), and tetrakistriphenylphosphine palladium (670 mg, 0.586 mmol) in sequence. Replace the mixture with nitrogen three times and heat to 100°C for 16 hours.

[0511] When LC / MS showed a 30% conversion, the reaction system was cooled to room temperature, ethyl acetate was added, stirred, filtered, and the filtrate was concentrated. The crude product was purified by HPLC to obtain 280 mg of (S)-2-(tert-butyloxycarbonyl)amino)-3-(4-cyclopropylphenyl)propanoic acid.

[0512] MS (ESI) M / Z: 306.1 [M+1] + .

[0513] Step B: Dissolve (S)-2-(tert-Butyloxycarbonyl)amino)-3-(4-cyclopropylphenyl)propanoic acid (280 mg, 0.68 mmol) in N,N-dimethylformamide (6 mL), add potassium carbonate (187 mg, 1.36 mmol) and iodomethane (145 mg, 0.76 mmol) in sequence, and stir at room temperature for 16 hours.

[0514] Completion of the reaction was monitored by thin-layer chromatography. Ethyl acetate and water were added to the reaction mixture, stirred, and separated into layers. The mixture was then extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated to yield 260 mg of methyl (S)-2-(tert-butyloxycarbonyl)amino)-3-(4-cyclopropylphenyl)propionate.

[0515] Step C: Dissolve (S)-methyl 2-(tert-butoxycarbonyl)amino)-3-(4-cyclopropylphenyl)propanoate (260 mg, 0.81 mmol) in dichloromethane (3 mL), add trifluoroacetic acid (3 mL) under ice-cooling, and warm to room temperature for reaction for 1 hour.

[0516] The reaction of the raw materials was monitored by thin layer chromatography. The reaction solution was concentrated to obtain 180 mg of (S)-3-(4-cyclopropylphenyl)-2-(2,2,2-trifluoroacetyl)-14-azepinyl)propionic acid methyl ester trifluoroacetate.

[0517] Step D: Dissolve (S)-3-(4-cyclopropylphenyl)-2-(2,2,2-trifluoroacetyl)-14-azepine)propionic acid methyl ester trifluoroacetate (180 mg, 0.63 mmol) in N,N-dimethylformamide (6 mL), add (2-morpholinoacetyl)-L-alanine (165 mg, 0.76 mmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (316 mg, 0.82 mmol), cool to 0-5°C, add N,N-diisopropylethylamine (165 mg, 1.27 mmol) to adjust the pH to 7, and warm to room temperature for 1 hour.

[0518] Completion of the reaction was monitored by thin-layer chromatography. Ethyl acetate and water were added to the reaction mixture, stirred, and the layers separated. Extraction was then performed with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by column chromatography to yield 180 mg of methyl (S)-3-(4-cyclopropylphenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[0519] MS (ESI) M / Z: 418.57 [M+H] + .

[0520] Step E: Dissolve (S)-methyl 3-(4-cyclopropylphenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate (210 mg, 0.50 mmol) in tetrahydrofuran (2 mL). Add a solution of lithium hydroxide monohydrate (31.7 mg, 0.75 mmol) in water (2 mL) under ice-cooling and react at room temperature for 1 hour.

[0521] The reaction of the raw materials was monitored by thin layer chromatography. The mixture was back-extracted twice with a 1 / 1 mixture of petroleum ether and ethyl acetate. The aqueous phase was adjusted to pH 7 with 1N hydrochloric acid and lyophilized to obtain 180 mg of 3-(4-cyclopropylphenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionic acid lithium salt.

[0522] MS (ESI) M / Z: 404.6 [M+H] + .

[0523] Step F: 3-(4-cyclopropylphenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionic acid lithium salt (50 mg, 0.37 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (106 mg, 0.33 mmol) were dissolved in N,N-dimethylformamide (4 mL), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (164 mg, 0.43 mmol) was added, and the mixture was cooled to 0-5°C. N,N-diisopropylethylamine (85.9 mg, 0.66 mmol) was added to adjust the pH to 7, and the mixture was heated to room temperature for 1 hour.

[0524] The reaction of the raw materials was monitored by thin-layer chromatography. Ethyl acetate and water were added to the reaction solution, stirred, and the layers were separated, and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated. The crude product was purified by column chromatography and preparative column chromatography to obtain 27.21 mg of (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-3-(4-cyclopropylphenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide.

[0525] MS (ESI) M / Z: 581.7 [M+H] + .

[0526] 1 HNMR (400MHz, DMSO-d6): δ8.27(d,J=7.6Hz,1H),8.05(d,J=8.4Hz,1H),7.74(d,J=7.6H z,1H),7.05(d,J=8.0Hz,2H),6.92(d,J=8.0Hz,2H),5.39(s,1H),4.49(m,2H),4.25(m,1 H),3.55(m,4H),3.15(m,1H),2.81-2.95(m,4H),2.65(m,1H),2.21-2.35(m,5H),2.11- 2.15(m,5H),1.78(m,3H),1.45(s,3H),1.14(d,J=6.8Hz,3H),0.91(m,2H),0.58(m,2H).

[0527] Example 14:

[0528] (S)-N-((S)-3-(cyclopent-1-enyl-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)-3-(4-(oxetan-3-yl)phenyl)propionamide

[0529] Reaction route:

[0530] Steps:

[0531] Step A: Add zinc powder (1.2 g, 18.50 mmol) to N,N-dimethylformamide (20 mL). Under nitrogen, add trimethylsilyl chloride (100 mg, 0.85 mmol) and stir for 30 minutes. Then, add iodine (100 mg, 0.42 mmol) and continue stirring for 10 minutes. Cool to 10-20°C and slowly add a solution of methyl (R)-2-(tert-butyloxycarbonyl)amino)-3-iodopropionate (6.1 g, 37.05 mmol) in N,N-dimethylformamide (10 mL). Stir for half an hour. Finally, add 1-(difluoromethoxy)-4-iodobenzene (1 g, 3.65 mmol), 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (91 mg, 0.22 mmol), and tris(dibenzylideneacetone)palladium (101.4 mg, 0.11 mmol). Heat the temperature to 50°C and react overnight.

[0532] Completion of the reaction was monitored by thin-layer chromatography. The reaction mixture was filtered after adding ethyl acetate and water, and extracted three times with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by column chromatography to yield 200 mg of methyl (S)-2-(tert-butyloxycarbonyl)amino)-3-(4-(oxetan-3-yl)phenyl)propanoate.

[0533] MS (ESI) M / Z: 336.4 [M+H] + .

[0534] Step B: Dissolve (S)-methyl 2-(tert-butoxycarbonyl)amino)-3-(4-(oxetan-3-yl)phenyl)propanoate (200 mg, 0.59 mmol) in dichloromethane (3 mL), add trifluoroacetic acid (3 mL) under ice-cooling, and warm to room temperature for 1 hour.

[0535] The reaction of the raw materials was monitored by thin layer chromatography. The reaction solution was concentrated to obtain 180 mg of (S)-2-amino-3-(4-(oxetan-3-yl)phenyl)propionic acid methyl ester trifluoroacetate.

[0536] MS (ESI) M / Z: 236.1 [M+H] +.

[0537] Step C: Dissolve (S)-2-amino-3-(4-(oxetan-3-yl)phenyl)propanoic acid methyl ester trifluoroacetate (150 mg, 0.63 mmol) in N,N-dimethylformamide (6 mL), add (2-morpholinoacetyl)-L-alanine (165 mg, 0.76 mmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (316 mg, 0.82 mmol), cool to 0-5°C, add N,N-diisopropylethylamine (165 mg, 1.27 mmol) to adjust the pH to 7, and warm to room temperature for 1 hour.

[0538] Completion of the reaction of the starting materials was monitored by thin-layer chromatography. The reaction solution was diluted with water and extracted three times with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated. The crude product was purified by column chromatography to yield 50 mg of methyl (S)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)-3-(4-(oxetan-3-yl)phenyl)propanoate.

[0539] MS (ESI) M / Z: 434.5 [M+H] + .

[0540] Step D: Methyl (S)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)-3-(4-(oxetan-3-yl)phenyl)propanoate (50 mg, 0.09 mmol) was dissolved in tetrahydrofuran (2 mL). An aqueous solution of lithium hydroxide monohydrate (4.3 mg, 0.16 mmol) (2 mL) was added under ice-cooling and the mixture was reacted at room temperature for 1 hour.

[0541] The reaction of the raw materials was monitored by thin-layer chromatography. The mixture was back-extracted twice with a 1 / 1 mixture of petroleum ether and ethyl acetate. The aqueous phase was adjusted to pH 7 with 1N hydrochloric acid and lyophilized to obtain 50 mg of (S)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)-3-(4-(oxetan-3-yl)phenyl)propionic acid lithium salt.

[0542] MS (ESI) M / Z: 420.4 [M+H] + .

[0543] Step E: (S)-2-((S)-2-(2-morpholinoacetylamino)propionamido)-3-(4-(oxetan-3-yl)phenyl)propionic acid lithium salt (50 mg, 0.11 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (35 mg, 0.1 mmol) were dissolved in N,N-dimethylformamide (4 mL), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (53 mg, 0.14 mmol) was added, the mixture was cooled to 0-5°C, N,N-diisopropylethylamine (28 mg, 0.21 mmol) was added to adjust the pH to 7, and the mixture was warmed to room temperature for 1 hour.

[0544] The reaction of the raw materials was monitored by thin-layer chromatography. Ethyl acetate and water were added to the reaction solution, stirred, and separated, and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated. The crude product was purified by column chromatography and preparative column chromatography to obtain 3.02 mg of (S)-N-((S)-3-(cyclopent-1-enyl-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)-3-(4-(oxetane-3-yl)phenyl)propionamide.

[0545] MS (ESI) M / Z: 597.7 [M+H] + .

[0546] 1 HNMR (400MHz, DMSO-d6): δ8.31(d,J=7.6Hz,1H),8.09(d,J=8.4Hz,1H),7.78 (b,1H),7.20-7.30(m,4H),5.30-5.41(m,1H),4.45-4.55(m,5H),4.21(m,2H) ,3.41-3.51(m,4H),3.15(m,1H),2.91-3.10(m,4H),2.65-2.75(m,1H),2.21- 2.35(m,5H),2.11-2.15(m,5H),1.75-1.83(m,3H),1.36(s,3H),1.17(d,3H).

[0547] Example 15:

[0548] (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-3-(4-(hydroxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0549] Reaction route:

[0550] Steps:

[0551] Step A: Add p-iodobenzyl alcohol (3 g, 12.82 mmol) to tetrahydrofuran (20 mL), then add sodium hydroxide (770 mg, 19.23 mmol) at 0°C. Stir and react at 0°C for half an hour, then add 4-methoxybenzyl chloride (2.8 g, 17.95 mmol). Warm to room temperature and react overnight.

[0552] The reaction solution was poured into saturated ammonium chloride and extracted three times with ethyl acetate. The ethyl acetate phases were combined, washed three times with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and spin-dried to obtain 4.9 g of crude 1-iodo-4-((4-methoxybenzyloxy)methyl)benzene, which was used directly in the next reaction.

[0553] Step B: Zinc powder (1.1 g, 16.94 mmol) was added to N,N-dimethylformamide (10 mL). Trimethylsilyl chloride (61 mg, 0.57 mmol) was added under nitrogen. After stirring for 30 minutes, iodine (143 mg, 0.57 mmol) was added. The reaction was stirred for 10 minutes. Then, (R)-2-tert-butoxycarbonyl-amino-3-iodopropionic acid methyl ester (5.6 g, 16.94 mmol) was added. After stirring for 30 minutes, 1-iodo-4-((4-methoxybenzyloxy)methyl)benzene (2 g, 5.65 mmol), 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (140 mg, 0.34 mmol), and tris(dibenzylideneacetone)palladium (155 mg, 0.17 mmol) were added in sequence. The temperature was raised to 50°C and the reaction was stirred overnight.

[0554] The reaction mixture was cooled to room temperature and filtered. The filtrate was poured into water and extracted three times with ethyl acetate. The combined ethyl acetate phases were washed three times with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and spin-dried. The crude product was purified by silica gel column chromatography to obtain 2 g of methyl (S)-2-(tert-butoxycarbonylamino)-3-(4-((4-methoxybenzyloxy)methyl)phenyl)propanoate.

[0555] MS (ESI) M / Z: 430.50 [M+H] + .

[0556] Step C: Dissolve (S)-methyl 2-(tert-butoxycarbonylamino)-3-(4-((4-methoxybenzyloxy)methyl)phenyl)propanoate (300 mg, 0.70 mmol) in dichloromethane (1.5 mL), add 8M hydrogen chloride solution in dioxane (0.1 mL), and stir at room temperature for 20 minutes.

[0557] After thin layer chromatography monitoring showed that the starting material disappeared, 140 mg of (S)-2-amino-3-(4-((4-methoxybenzyloxy)methyl)phenyl)propionic acid methyl ester hydrochloride was obtained by filtration.

[0558] Step D: (S)-2-(2-Morpholinoacetylamino)propionic acid (83 mg, 0.38 mmol) and (S)-2-amino-3-(4-((4-methoxybenzyloxy)methyl)phenyl)propionic acid methyl ester hydrochloride (140 mg, 0.38 mmol) were dissolved in N,N-dimethylformamide (5 mL), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (174 mg, 0.46 mmol) was added, and then the pH was adjusted to about 7 with N,N-diisopropylethylamine and stirred at room temperature for 30 minutes.

[0559] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified on a silica gel column to obtain 75 mg of methyl 3-(4-((4-methoxybenzyloxy)methyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate.

[0560] MS (ESI) M / Z: 550.3 [M+Na] + .

[0561] Step E: Dissolve methyl 3-(4-((4-methoxybenzyloxy)methyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate (65 mg, 0.12 mmol) in dichloromethane (1.5 mL), add trifluoroacetic acid (0.5 mL), and stir at room temperature for 20 minutes.

[0562] After thin layer chromatography monitoring showed the disappearance of the starting material, the reaction solution was spin-dried to obtain 70 mg of methyl (S)-3-(4-(hydroxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionate trifluoroacetate.

[0563] MS (ESI) M / Z: 408.4 [M+H] + .

[0564] Step F: Dissolve (S)-methyl 3-(4-(hydroxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate trifluoroacetate (70 mg, 0.12 mmol) in tetrahydrofuran / water (2 mL / 2 mL), add lithium hydroxide monohydrate (15 mg, 0.38 mmol), and stir at room temperature for 1 hour.

[0565] After the reaction of the raw materials was completed by monitoring by thin layer chromatography, the pH was adjusted to 7-8 with 1N hydrochloric acid and lyophilized to obtain 70 mg of crude (S)-3-(4-(hydroxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionic acid lithium salt, which was directly used in the next reaction.

[0566] Step G: (S)-3-(4-(Hydroxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoic acid lithium salt (70 mg, 0.12 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one trifluoroacetate (38 mg, 0.12 mmol) were dissolved in N,N-dimethylformamide (5 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (56 mg, 0.15 mmol) was added, and the pH was adjusted to about 7 with N,N-diisopropylethylamine. The mixture was stirred at room temperature for 30 minutes.

[0567] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 15.08 mg of (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-3-(4-(hydroxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide.

[0568] MS (ESI) M / Z: 571.8 [M+H] + .

[0569] 1 H NMR (400MHz, DMSO-d6): δ8.31(d,J=7.2Hz,1H),8.06(d,J=8.4Hz,1H),7.74(d ,J=8.0Hz,1H),7.16(m,4H),5.40(br,1H),5.12(m,1H),4.53-4.42(m,4H),4.2 7(m,1H),3.56(m,4H),3.17(m,1H),3.01-2.84(m,4H),2.67(m,1H),2.37(m,4 H),2.26(m,4H),1.80(m,2H),1.38(s,3H),1.23(m,2H),1.15(d,J=6.8Hz,3H).

[0570] Example 16:

[0571] (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-2-((S)-2-(2-morpholinylacetamido)propionamido)-3-(4-propionylphenyl)propionamide

[0572] Reaction route:

[0573] Steps:

[0574] Step A: Methyl 4-iodobenzoate (5 g, 19.08 mmol) and tetraisopropyl titanate (7.59 g, 26.71 mmol) were dissolved in tetrahydrofuran (100 mL). The reaction system was cooled to -10°C in an ice-salt bath. Ethylmagnesium bromide (3.4 M, 19.64 mL, 66.78 mmol) was added dropwise. The reaction was stirred at -10°C for 0.5 hour, then heated to 25°C and stirred for 16 hours.

[0575] After thin-layer chromatography monitoring indicated complete reaction of the starting materials, the reaction system was quenched with saturated aqueous ammonium chloride (20 mL) at 0°C. The mixture was filtered through a pad of celite, and the filtrate was extracted with ethyl acetate (200 mL x 2). The organic phases were combined, washed with saturated brine (100 mL x 1), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to yield 3.5 g of 1-(4-iodophenyl)cyclopropanol.

[0576] MS (ESI) M / Z: 261.0 [M+H] + .

[0577] Step B: Zinc powder (477.49 mg, 7.3 mmol) was suspended in N,N-dimethylformamide (20 mL). Elemental iodine (46.32 mg, 0.18 mmol) was added and stirred at 25°C for 0.5 hour until the reddish-brown color disappeared. Methyl (R)-2-(tert-butyloxycarbonyl)amino)-3-iodopropionate (1.2 g, 3.65 mmol) was added to the reaction system. Stirring was continued at 25°C for 1 hour. 1-(4-iodophenyl)cyclopropanol (1.04 g, 4.02 mmol), tris(dibenzylideneacetone)dipalladium (167.12 mg, 0.18 mmol), and 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (149.84 mg, 0.36 mmol) were then added to the reaction system. The temperature was raised to 50°C and stirred for 16 hours.

[0578] After thin-layer chromatography monitoring indicated complete reaction of the starting materials, the reaction solution was filtered through a pad of celite. Water (20 mL) and ethyl acetate (50 mL) were added to the filtrate. The organic phase was washed with water (20 mL x 2) and saturated brine (20 mL x 1), then dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography to yield 550 mg of methyl (S)-2-((tert-butoxycarbonyl)amino)-3-(4-propionylphenyl)propionate.

[0579] MS (ESI) M / Z: 336.2 [M+H] + .

[0580] 1H NMR(400MHz,Chloroform-d)δ7.90(d,J=8.2Hz,2H),7.22(d,J=7.9Hz,2H),4.99(d,J=8.4Hz,1H),4.62(d,J=7.2Hz,1H),3.72 (s,3H),3.19(dd,J=13.7,5.8Hz,1H),3.09(dd,J=13.8,6.4Hz,1H),2.98(q,J=7.2Hz,2H),1.41(s,9H),1.22(t,J=7.2Hz,3H).

[0581] Step C: Dissolve (S)-methyl 2-((tert-butoxycarbonyl)amino)-3-(4-propionylphenyl)propionate (550 mg, 1.64 mmol) in dichloromethane (5 mL). Cool the reaction system to 0°C in an ice-water bath. Add trifluoroacetic acid (1 mL, 13.42 mmol) dropwise. Warm the reaction system to 25°C and stir for 0.5 h.

[0582] After thin layer chromatography monitoring showed that the raw materials had reacted completely, the reaction solution was concentrated under reduced pressure to obtain 385 mg of crude (S)-2-amino-3-(4-propionylphenyl)propionic acid methyl ester, which was directly used in the next reaction.

[0583] MS (ESI) M / Z: 236.1 [M+H] + .

[0584] Step D: Methyl (S)-2-amino-3-(4-propionylphenyl)propionate (385 mg, 1.64 mmol), (2-morpholinoacetyl)-L-alanine (354.62 mg, 1.64 mmol), benzotriazole-N,N,N',N'-tetramethyluronium hexafluorophosphate (932.93 mg, 2.46 mmol) and 1-hydroxybenzotriazole (332.40 mg, 2.46 mmol) were dissolved in N,N-dimethylformamide (5 mL). The reaction system was cooled to 0°C in an ice-water bath, and diisopropylethylamine (1.36 mL, 8.2 mmol) was added dropwise. The reaction system was warmed to 25°C and stirred for 4 hours.

[0585] After LCMS monitoring showed that the starting material had reacted completely, the reaction system was quenched with saturated aqueous sodium bicarbonate (10 mL). The mixture was extracted with ethyl acetate (20 mL × 2 times). The organic phases were combined, washed with saturated brine (20 mL × 1 time), then dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography to obtain 440 mg of methyl (S)-2-((S)-2-(2-morpholinoacetamido)propionamido)-3-(4-propionylphenyl)propionate.

[0586] MS (ESI) M / Z: 434.2 [M+H] + .

[0587] Step E: Methyl (S)-2-((S)-2-(2-morpholinoacetamido)propionamido)-3-(4-propionylphenyl)propanoate (440 mg, 1.01 mmol) was dissolved in methanol (6 mL) and water (2 mL). The reaction system was cooled to 0°C in an ice-water bath. Lithium hydroxide monohydrate (42 mg, 1.01 mmol) was added, and the reaction system was heated to 25°C and stirred for 2 hours.

[0588] After LCMS monitoring showed that the raw material reaction was complete, the reaction solution was concentrated under reduced pressure, and after adding water (10 mL), the pH of the aqueous phase was adjusted to 7 with 1N hydrochloric acid, and freeze-dried to obtain 420 mg of (S)-2-((S)-2-(2-morpholinoacetamido)propionamido)-3-(4-propionylphenyl)propionic acid crude product, which was directly used in the next reaction.

[0589] MS (ESI) M / Z: 420.2 [M+H] + .

[0590] Step F: (S)-2-((S)-2-(2-morpholinoacetamido)propionamido)-3-(4-propionylphenyl)propanoic acid (200 mg, 0.48 mmol), (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (181.18 mg, 0.48 mmol), benzotriazole-N,N,N',N'-tetramethyluronium hexafluorophosphate (273.05 mg, 0.72 mmol) and 1-hydroxybenzotriazole (97.29 mg, 0.72 mmol) were dissolved in N,N-dimethylformamide (5 mL). The reaction system was cooled to 0°C in an ice-water bath. Diisopropylethylamine (0.4 mL, 2.4 mmol) was added dropwise. The reaction system was warmed to 25°C and stirred for 2 hours.

[0591] After TLC monitoring indicated complete reaction of the starting materials, the reaction system was quenched with saturated aqueous sodium bicarbonate (10 mL). The mixture was extracted with ethyl acetate (20 mL x 2). The organic phases were combined, washed with saturated brine (10 mL x 1), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The crude product was purified by preparative HPLC and preparative TLC to yield 115 mg of (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-2-((S)-2-(2-morpholinoacetamido)propionamido)-3-(4-propionylphenyl)propionamide.

[0592] MS (ESI) M / Z: 597.3 [M+H] + .

[0593] 1H NMR (400MHz, DMSO-d6) δ8.35(d,J=7.3Hz,1H),8.13(d,J=8.5Hz,1H),7.83(d,J=8.2Hz,2H),7.70( d,J=7.8Hz,1H),7.34(d,J=8.1Hz,2H),5.40(s,1H),4.55(ddt,J=21.1,13.2,5.9Hz,2H),4.26(p,J =6.9Hz,1H),3.53(t,J=4.6Hz,4H),3.18(d,J=5.3Hz,1H),3.06-2.95(m,4H),2.94-2.72(m,3H),2. 39-2.16(m,10H),1.79(p,J=7.5Hz,2H),1.37(s,3H),1.14(d,J=7.0Hz,3H),1.07(t,J=7.2Hz,3H).

[0594] Example 17:

[0595] (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanon-2-yl)-3-(4-(methoxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0596] Reaction route:

[0597] Steps:

[0598] Step A: Add p-iodobenzyl alcohol (3 g, 12.82 mmol) to tetrahydrofuran (20 mL), add sodium hydroxide (770 mg, 19.23 mmol) at 0°C, stir at 0°C for half an hour, add iodomethane (2.4 g, 16.67 mmol), and warm to room temperature for 2 hours.

[0599] The reaction solution was poured into saturated ammonium chloride and extracted three times with ethyl acetate. The ethyl acetate phases were combined, washed three times with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered and spin-dried to obtain 3 g of crude 1-iodo-4-(methoxymethyl)benzene, which was used directly in the next reaction.

[0600] Step B: Add zinc powder (1.6 g, 24.19 mmol) to N,N-dimethylformamide (10 mL). Under nitrogen, add trimethylsilyl chloride (87 mg, 0.81 mmol). Stir for 30 minutes, then add iodine (205 mg, 0.81 mmol). Continue stirring for 10 minutes. Then, add (R)-methyl 2-tert-butoxycarbonylamino-3-iodopropionate (8 g, 24.19 mmol). Continue stirring for 30 minutes, then add 1-iodo-4-(methoxymethyl)benzene (2 g, 8.06 mmol), 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (200 mg, 0.48 mmol), and tris(dibenzylideneacetone)palladium (220 mg, 0.24 mmol). Heat the temperature to 50°C and stir overnight.

[0601] The reaction mixture was cooled to room temperature and filtered. The filtrate was poured into water and extracted three times with ethyl acetate. The combined ethyl acetate phases were washed three times with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and spin-dried. The crude product was purified by silica gel column chromatography to yield 1.7 g of (S)-3-(4-(methoxymethyl)phenyl)-2-tert-butoxycarbonyl-aminopropionic acid methyl ester.

[0602] MS (ESI) M / Z: 324.51 [M+H] + .

[0603] Step C: Dissolve (S)-3-(4-(methoxymethyl)phenyl)-2-tert-butoxycarbonyl-aminopropionic acid methyl ester (700 mg, 2.16 mmol) in dichloromethane (5 mL), add a solution of hydrogen chloride in dioxane (2 mL, 8 mol / L), and stir at room temperature for 30 minutes.

[0604] After thin layer chromatography monitoring showed the disappearance of the starting material, the reaction solution was concentrated to dryness to obtain 510 mg of (S)-3-(4-(methoxymethyl)phenyl)-2-aminopropionic acid methyl ester hydrochloride.

[0605] Step D: (S)-2-(2-Morpholinoacetylamino)propionic acid (250 mg, 1.15 mmol) and (S)-3-(4-(methoxymethyl)phenyl)-2-aminopropionic acid methyl ester hydrochloride (300 mg, 1.15 mmol) were dissolved in N,N-dimethylformamide (5 mL), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (526 mg, 1.39 mmol) was added, and then the pH was adjusted to about 7 with N,N-diisopropylethylamine and stirred at room temperature for 30 minutes.

[0606] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 200 mg of methyl 3-(4-(methoxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate.

[0607] MS (ESI) M / Z: 422.4 [M+H] + .

[0608] Step E: Dissolve methyl 3-(4-(methoxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate (200 mg, 0.47 mmol) in tetrahydrofuran / water (2 mL / 2 mL), add lithium hydroxide monohydrate (40 mg, 0.95 mmol), and stir at room temperature for 1 hour.

[0609] After the reaction of the raw materials was monitored by thin layer chromatography, the pH was adjusted to 7-8 with 1N hydrochloric acid and lyophilized to obtain 190 mg of crude lithium salt of 3-(4-(methoxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionic acid, which was used directly in the next reaction.

[0610] Step F: 3-(4-(Methoxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionic acid lithium salt (190 mg, 0.46 mmol) and (S)-2-amino-3-cyclopentenyl-1-((R)-2-methyloxiran-2-yl)propyl-1-one trifluoroacetate (142 mg, 0.46 mmol) were dissolved in N,N-dimethylformamide (5 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (210 mg, 0.55 mmol) was added, and then the pH was adjusted to about 7 with N,N-diisopropylethylamine and stirred at room temperature for 30 minutes.

[0611] The reaction mixture was poured into water (20 mL) and extracted three times with ethyl acetate. The organic phases were combined, washed twice with a saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The resulting residue was purified by silica gel column chromatography to obtain 13.42 mg of (S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-propanone-2-yl)-3-(4-(methoxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide.

[0612] MS (ESI) M / Z: 585.6 [M+H] + .

[0613] 1 H NMR (400MHz, CDCl3): δ7.24(m,4H),7.15(br,1H),6.83(br,1H),6.19(m,1H),5.3 4(m,1H),4.56(m,2H),4.40(s,3H),4.33(m,1H),3.78(m,4H),3.38(s,3H),3.25( d,J=4.8Hz,1H),3.03(d,J=7.2Hz,3H),2.89(d,J=4.8Hz,1H),2.74-2.43(m,3H), 2.26-2.13(m,5H),1.84(m,2H),1.64(m,2H),1.49(s,3H),1.35(d,J=7.2Hz,3H).

[0614] Example 18:

[0615] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-cyclopropyloxyphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0616] Reaction route:

[0617] Steps:

[0618] Step A: Dissolve 4-bromophenol (5 g, 28.90 mmol) and vinyl acetate (7.46 g, 86.70 mmol) in toluene (50 mL). Under nitrogen protection, add 1,5-cyclooctadiene iridium chloride dimer (0.19 g, 0.29 mmol) and sodium carbonate (1.84 g, 17.34 mmol). Stir the reaction system at 110 ° C for 16 hours.

[0619] After thin-layer chromatography monitoring indicated complete reaction of the starting materials, water (50 mL) and ethyl acetate (50 mL) were added to the reaction system. The aqueous phase was extracted with ethyl acetate (30 mL x 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography to yield 4.8 g of 1-bromo-4-vinyloxybenzene.

[0620] MS (ESI) M / Z: 199.0 [M+H] + .

[0621] Step B: Dissolve 1-bromo-4-vinyloxybenzene (4.8 g, 24.12 mmol) and chloroiodomethane (17.02 g, 96.48 mmol) in dichloromethane (100 mL). Cool the reaction system to 0°C in an ice-water bath. Slowly add a n-hexane solution of diethylzinc (1 M, 5.96 g, 48.24 mmol, 48.24 mL) dropwise. Keep the reaction temperature below 5°C. After the addition is complete, continue stirring at 0°C for 4 hours.

[0622] After thin-layer chromatography monitoring indicated complete reaction of the starting materials, saturated aqueous ammonium chloride (10 mL) was slowly added dropwise to the reaction system at 0°C to quench the reaction. Water (50 mL) was added, and the aqueous phase was extracted with dichloromethane (50 mL x 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography to yield 4.9 g of 1-bromo-4-cyclopropyloxybenzene.

[0623] MS (ESI) M / Z: 213.0 [M+H] + .

[0624] Step C: 1-bromo-4-cyclopropyloxybenzene (4.9 g, 23.00 mmol) was dissolved in tetrahydrofuran (50 mL), and the reaction system was placed in a dry ice ethanol bath and cooled to -78°C. A cyclohexane solution of n-butyllithium (2.5 M, 10.12 mL, 25.3 mmol) was slowly added dropwise. After the addition was complete, stirring was continued at -78°C for 0.5 hour. N,N-dimethylformamide (3.36 g, 46 mmol, 3.55 mL) was added, and stirring was continued at -78°C for 0.5 hour.

[0625] After TLC monitoring showed that the raw materials had reacted completely, methanol (5 mL) was slowly added dropwise to the reaction system at -78°C to quench the reaction, and the reaction solution was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to obtain 3.4 g of 4-cyclopropyloxybenzaldehyde.

[0626] MS (ESI) M / Z: 163.0 [M+H] + .

[0627] Step D: Under nitrogen protection, (R)-2-(N-(N'-benzylprolyl)amino)diphenylmethanone-nickel(II)-glycine complex (2 g, 4.01 mmol) and 4-cyclopropyloxybenzaldehyde (1.30 g, 8.02 mmol) were dissolved in methanol (20 mL), and a methanol solution of sodium methoxide (5.5 M, 2.23 mL, 12.03 mmol) was added dropwise at 25°C. After stirring at 25°C for 12 hours, dilute hydrochloric acid (2 M, 20.00 mL) was added, and the mixture was stirred at 50°C for 2 hours.

[0628] After LC-MS monitoring showed that the raw materials had reacted completely, the reaction solution was concentrated under reduced pressure to remove methanol, the aqueous phase was washed with dichloromethane (15 mL×5), and the aqueous phase was freeze-dried to obtain 0.95 g of crude (2S,3R)-2-amino-3-(4-cyclopropyloxyphenyl)-3-hydroxypropionic acid hydrochloride.

[0629] MS (ESI) M / Z: 238.1 [M+H] + .

[0630] Step E: At 0 ° C, (2S, 3R) -2-amino-3- (4-cyclopropyloxyphenyl) -3-hydroxypropanoic acid hydrochloride (0.95 g, 4.01 mmol) was dissolved in benzyl alcohol (10 mL), thionyl chloride (0.95 g, 8.02 mmol) was added dropwise, and then stirred at 90 ° C for 5 hours.

[0631] LC-MS / MS monitoring indicated a small amount of starting material remaining. Water (10 mL) was added to the reaction mixture to quench it. The aqueous phase was adjusted to pH 9 with aqueous ammonia and extracted with ethyl acetate (20 mL x 2). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to yield 150 mg of benzyl (2S,3R)-2-amino-3-(4-cyclopropyloxyphenyl)-3-hydroxypropanoate.

[0632] MS (ESI) M / Z: 328.1 [M+H] + .

[0633] Step F: Benzyl (2S,3R)-2-amino-3-(4-cyclopropyloxyphenyl)-3-hydroxypropanoate (105 mg, 0.32 mmol) and (2-morpholinoacetyl)-L-alanine (69.19 mg, 0.32 mmol) were dissolved in tetrahydrofuran (5 mL). The reaction system was cooled to 0°C in an ice-water bath. 3-(diethoxyphosphoryloxy)-1,2,3-benzotriazine-4-one (143.63 mg, 0.48 mmol) and sodium carbonate (135.66 mg, 1.28 mmol) were added and stirring was continued at 25°C for 12 hours.

[0634] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was quenched with water (10 mL). The aqueous phase was extracted with dichloromethane (20 mL x 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography to yield 150 mg of benzyl (2S,3R)-3-(4-cyclopropyloxyphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[0635] MS (ESI) M / Z: 526.2 [M+H] + .

[0636] Step G: Dissolve benzyl (2S,3R)-3-(4-cyclopropyloxyphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate (150 mg, 0.29 mmol) in methanol (5 mL), add 10% wet palladium on carbon (30 mg), and stir under a hydrogen atmosphere (15 psi) at 25°C for 1 hour.

[0637] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was filtered through a pad of celite, and the filtrate was concentrated under reduced pressure to yield 120 mg of crude (2S,3R)-3-(4-cyclopropyloxyphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionic acid, which was used directly in the next step.

[0638] MS (ESI) M / Z: 436.2 [M+H] + .

[0639] Step H: (2S,3R)-3-(4-cyclopropyloxyphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoic acid (120 mg, 0.28 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (102.89 mg, 0.28 mmol) were dissolved in tetrahydrofuran (5 mL). The reaction system was cooled to 0°C in an ice-water bath. 3-(diethoxyphosphoryloxy)-1,2,3-benzotriazine-4-one (125.67 mg, 0.42 mmol) and sodium carbonate (0.12 g, 1.12 mmol) were added and stirring was continued at 25°C for 12 hours.

[0640] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was quenched with water (10 mL). The aqueous phase was extracted with dichloromethane (20 mL x 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography to yield 150 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-cyclopropyloxyphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide.

[0641] MS (ESI) M / Z: 613.3 [M+H] + .

[0642] 1H NMR (400MHz, CD3OD): δ7.27(d,J=8.7Hz,2H),6.95(d,J=8.7Hz,2H),5.47-5.42(m,1H),4.99(d,J=4.8Hz,1H),4.66 (dd,J=8.4,5.3Hz,1H),4.52(d,J=4.8Hz,1H),4.45(q,J=7.1Hz,1H),3.74(tt,J=6.0,3.0Hz,1H),3.71-.64(m,4H) ,3.12(d,J=5.0Hz,1H),3.06(d,J=15.8Hz,1H),2.98(d,J=15.8Hz,1H),2.88(d,J=5.0Hz,1H),2.53-2.46(m,5H),2 .36-2.20(m,5H),1.85(p,J=7.5Hz,2H),1.41(s,3H),1.31(d,J=7.1Hz,3H),0.82-0.72(m,2H),0.72-0.61(m,2H).

[0643] Example 19:

[0644] (2S,3R)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-cyclopropyloxyphenyl)-3-hydroxypropionamide

[0645] Reaction route:

[0646] Steps:

[0647] Step A: Dissolve 1,4-oxazepane (3 g, 29.66 mmol) in acetonitrile (40 mL), add benzyl 2-bromoacetate (6.79 g, 29.66 mmol) and potassium carbonate (12.30 g, 88.98 mmol), and stir at 20°C for 4 hours.

[0648] After LC-MS monitoring showed that the reaction of the starting material was complete, the mixture was diluted with ethyl acetate (20 mL), filtered, and the filtrate was concentrated under reduced pressure to obtain 7.39 g of crude 2-(1,4-oxazepan-4-yl)benzyl acetate, which was used directly in the next step.

[0649] MS (ESI) M / Z: 250.1 [M+H] + .

[0650] Step B: Dissolve benzyl 2-(1,4-oxazepan-4-yl)acetate (7.39 g, 29.64 mmol) in methanol (70 mL), add 10% wet palladium on carbon (320 mg), and stir under a hydrogen atmosphere (15 psi) at 25°C for 1 hour.

[0651] After LC-MS monitoring showed that the reaction of the starting materials was complete, the reaction solution was filtered through a pad of celite, and the filtrate was concentrated under reduced pressure to obtain 0.47 g of crude 2-(1,4-oxazacycloheptane-4-yl)acetic acid, which was used directly in the next step.

[0652] MS (ESI) M / Z: 160.1 [M+H] + .

[0653] Step C: At 0°C, 2-(1,4-oxazacycloheptane-4-yl)acetic acid (2 g, 12.56 mmol) was dissolved in N,N-dimethylformamide (20 mL), and L-alanine benzyl ester hydrochloride (2.71 g, 12.56 mmol), benzotriazole-N,N,N',N'-tetramethyluronium hexafluorophosphate (7.14 g, 18.84 mmol), 1-hydroxybenzotriazole (2.55 g, 18.84 mmol) and N,N-diisopropylethylamine (1.97 g, 15.25 mmol) were added and stirred at 25°C for 4 hours.

[0654] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was added to a saturated aqueous sodium bicarbonate solution (10 mL). The aqueous phase was extracted with ethyl acetate (20 mL x 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography to yield 2.8 g of N-(2-(1,4-oxazepan-4-yl)acetyl)-L-alanine benzyl ester.

[0655] MS (ESI) M / Z: 321.1 [M+H] + .

[0656] Step D: Dissolve N-(2-(1,4-oxazepan-4-yl)acetyl)-L-alanine benzyl ester (0.61 g, 1.92 mmol) in methanol (30 mL), add 10% wet palladium on carbon (93 mg), and stir under a hydrogen atmosphere (15 psi) at 25°C for 1 hour.

[0657] After LC-MS monitoring indicated that the reaction of the starting materials was complete, the reaction solution was filtered through a pad of celite, and the filtrate was concentrated under reduced pressure to obtain 1.95 g of crude N-(2-(1,4-oxazepan-4-yl)acetyl)-L-alanine, which was used directly in the next step.

[0658] MS (ESI) M / Z: 231.1 [M+H] + .

[0659] Step E: Benzyl (2S,3R)-2-amino-3-(4-cyclopropyloxyphenyl)-3-hydroxypropanoate (100 mg, 0.27 mmol) and N-(2-(1,4-oxazepan-4-yl)acetyl)-L-alanine (62.17 mg, 0.27 mmol) were dissolved in tetrahydrofuran (5 mL). The reaction system was cooled to 0°C in an ice-water bath. 3-(diethoxyphosphoryloxy)-1,2,3-benzotriazine-4-one (121.18 mg, 0.41 mmol) and sodium carbonate (114.47 mg, 1.08 mmol) were added and stirring was continued at 25°C for 12 hours.

[0660] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was quenched with water (10 mL). The aqueous phase was extracted with dichloromethane (20 mL x 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography to yield 114 mg of benzyl (2S,3R)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetylamino)propionylamino)-3-(4-cyclopropyloxyphenyl)-3-hydroxypropanoate.

[0661] MS (ESI) M / Z: 540.2 [M+H] + .

[0662] Step F: Dissolve benzyl (2S,3R)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetylamino)propionylamino)-3-(4-cyclopropyloxyphenyl)-3-hydroxypropanoate (114 mg, 0.21 mmol) in methanol (10 mL), add 10% wet palladium on carbon (22 mg), and stir under a hydrogen atmosphere (15 psi) at 25°C for 1 hour.

[0663] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was filtered through a pad of celite, and the filtrate was concentrated under reduced pressure to yield 94 mg of crude (2S,3R)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetylamino)propionylamino)-3-(4-cyclopropyloxyphenyl)-3-hydroxypropanoic acid, which was used directly in the next step.

[0664] MS (ESI) M / Z: 450.2 [M+H] + .

[0665] Step G: (2S,3R)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetylamino)propionylamino)-3-(4-cyclopropyloxyphenyl)-3-hydroxypropanoic acid (94 mg, 0.21 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (77.17 mg, 0.21 mmol) were dissolved in tetrahydrofuran (5 mL). The reaction system was cooled to 0°C in an ice-water bath. 3-(diethoxyphosphoryloxy)-1,2,3-benzotriazine-4-one (94.25 mg, 0.32 mmol) and sodium carbonate (89.03 mg, 0.84 mmol) were added and stirring was continued at 25°C for 12 hours.

[0666] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was quenched with water (10 mL). The aqueous phase was extracted with dichloromethane (20 mL x 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography to yield 60 mg of (2S,3R)-2-((S)-2-(2-(1,4-oxazepan-4-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-cyclopropyloxyphenyl)-3-hydroxypropionamide.

[0667] MS (ESI) M / Z: 627.3 [M+H] + .

[0668] 1H NMR (400MHz, CD3OD): δ7.30-7.23(m,2H),6.98-6.93(m,2H),5.44(s,1H),4.99(d,J=4.7Hz,1H),4.66(dd,J=8. 5,5.2Hz,1H),4.52(d,J=4.7Hz,1H),4.45(q,J=7.0Hz,1H),3.80(t,J=6.0Hz,2H),3.72(dt,J=8.7,3.8Hz,3H),3 .25-3.15(m,2H),3.12(d,J=5.4Hz,1H),2.87(d,J=5.0Hz,1H),2.78-2.70(m,4H),2.49(dd,J=14.1,5.2Hz,1H), 2.32-2.20(m,5H),1.95-1.79(m,4H),1.41(s,3H),1.32(d,J=7.1Hz,3H),0.82-0.70(m,2H),0.70-0.61(m,2H).

[0669] Example 20:

[0670] (2S,3R)-3-(4-Acetylphenyl)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0671] Reaction route:

[0672] Steps:

[0673] Step A: Under nitrogen protection, methyl (2S,3R)-3-(4-bromophenyl)-2-(tert-butoxycarbonylamino)-3-hydroxypropanoate (300 mg, 801.65 μmol), tributyl(1-ethoxyvinyl)stannane (710 mg, 1.97 mmol) and bistriphenylphosphine palladium dichloride (56.27 mg, 80.17 μmol) were dissolved in dioxane (5 mL), and the reaction solution was heated to 90°C and stirred for 12 hours.

[0674] After LC-MS monitoring showed that the raw material reaction was complete, the reaction solution was quenched with a saturated aqueous solution of potassium fluoride (10 mL), extracted with ethyl acetate (10 mL), and the organic phase was decompressed to obtain a crude product. The product was purified by flash silica gel column chromatography (12 g silica gel column, ethyl acetate / petroleum ether eluent, 0-15% gradient, 40 mL / min) to obtain 170 mg of (2S, 3R)-3-(4-acetylphenyl)-2-(tert-butoxycarbonylamino)-3-hydroxypropionic acid methyl ester.

[0675] MS (ESI) M / Z: 338.1 [M+H] + .

[0676] 1 H NMR (400MHz, CDCl3): δ7.95(d,J=8.3Hz,2H),7.49(d,J=8.3Hz,2H),5.33(br s,2H),4.54-4.64(m,1H),3.79(s,3H),2.60(s,3H),1.32(br s,9H).

[0677] Step B: Methyl (2S,3R)-3-(4-acetylphenyl)-2-(tert-butoxycarbonylamino)-3-hydroxypropanoate (100 mg, 296.41 μmol) was dissolved in dichloromethane (1.5 mL) and trifluoroacetic acid (1.5 mL) and stirred at 0°C for 1 hour.

[0678] After thin layer chromatography monitoring showed that the raw materials had reacted completely, the reaction solution was concentrated under reduced pressure to obtain 70 mg of crude trifluoroacetic acid methyl (2S,3R)-3-(4-acetylphenyl)-2-amino-3-hydroxypropanoate.

[0679] MS (ESI) M / Z: 238.1 [M+H] + .

[0680] Step C: At 0°C, trifluoroacetate of methyl (2S,3R)-3-(4-acetylphenyl)-2-amino-3-hydroxypropanoate (70 mg, 295.05 μmol) and (2-morpholinoacetyl)-L-alanine (76.56 mg, 354.06 μmol) were dissolved in N,N-dimethylformamide (1 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (224.37 mg, 590.09 μmol) and N,N-diisopropylethylamine (190.66 mg, 1.48 mmol) were added sequentially and stirred at 0°C for 1 hour.

[0681] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (12 g silica gel column, methanol / dichloromethane eluent, 0-3% gradient, 30 mL / min) to afford 190 mg of methyl (2S,3R)-3-(4-acetylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[0682] MS (ESI) M / Z: 436.2 [M+H] + .

[0683] Step D: Methyl (2S,3R)-3-(4-acetylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate (190 mg, 436.31 μmol) was dissolved in acetonitrile (5 mL) and water (0.5 mL) at 0°C, and lithium bromide (189.46 mg, 2.18 mmol) and N,N-diisopropylethylamine (169.17 mg, 1.31 mmol) were added sequentially. The mixture was stirred at 25°C for 12 hours.

[0684] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was separated by preparative HPLC (column: C18 150×30 mm; mobile phase: [water (ammonia + ammonium bicarbonate)-acetonitrile]; gradient: 6% to 36% phase B over 7 minutes) to yield 53.5 mg of (2S,3R)-3-(4-acetylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid.

[0685] MS (ESI) M / Z: 422.1 [M+H] + .

[0686] 1 H NMR (400MHz, CD3OD): δ7.92(d,J=8.3Hz,2H),7.54(d,J=8.3Hz,2H),5.36(d,J=2.8Hz,1H),4.65(d,J=2.8Hz,1H),4. 44(q,J=7.0Hz,1H),3.67(t,J=4.6Hz,4H),2.95-3.10(m,2H),2.58(s,3H),2.43-2.53(m,4H),1.28(d,J=7.0Hz,3H).

[0687] Step E: (2S,3R)-3-(4-Acetylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoic acid (42 mg, 99.66 μmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (43.94 mg, 119.59 μmol) were dissolved in N,N-dimethylformamide (1 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (75.79 mg, 199.32 μmol) and N,N-diisopropylethylamine (64.40 mg, 498.29 μmol) were added sequentially at 0°C and the mixture was stirred at 0°C for 1 hour.

[0688] After LC-MS monitoring showed that the raw materials were completely reacted, the reaction solution was separated and purified by preparative HPLC to obtain 9.93 mg of (2S,3R)-3-(4-acetylphenyl)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide.

[0689] MS (ESI) M / Z: 599.3 [M+H] + .

[0690] 1 H NMR (400MHz, CD3OD): δ7.92(d,J=8.3Hz,2H),7.52(d,J=8.3Hz,2H),5.46(s,1H),5.16(d,J =4.0Hz,1H),4.66-4.71(m,1H),4.61(d,J=4.3Hz,1H),4.39-4.47(m,1H),3.68(t,J=4.6Hz, 4H),3.17(d,J=5.0Hz,1H),2.94-3.15(m,2H),2.91(d,J=5.0Hz,1H),2.59(s,3H),2.44-2. 55(m,5H),2.25-2.36(m,5H),1.86(quin,J=7.6Hz,2H),1.41(s,3H),1.29(d,J=7.0Hz,3H).

[0691] Example 21:

[0692] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)-3-(4-propionylphenyl)propionamide

[0693] Reaction route:

[0694] Steps:

[0695] Step A: Methyl 4-iodobenzoate (10 g, 38.16 mmol) and tetraisopropoxytitanium (15.18 g, 53.43 mmol, 15.77 mL) were dissolved in tetrahydrofuran (200 mL), and ethylmagnesium bromide (3 M, 44.52 mL) was added at -10°C, and the mixture was stirred at -10°C for 0.5 hours and then at 25°C for 12 hours.

[0696] After thin-layer chromatography monitoring indicated complete reaction of the starting materials, the reaction mixture was quenched with saturated aqueous ammonium chloride (60 mL) and extracted with ethyl acetate (60 mL x 2). The organic phases were combined, washed with saturated brine (60 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: petroleum ether / ethyl acetate, 10 / 1 gradient) to yield 6.66 g of crude 1-(4-iodophenyl)cyclopropanol.

[0697] MS (ESI) M / Z: 261.0 [M+H] + .

[0698] Step B: Zinc powder (2.93 g, 44.81 mmol) and iodine (322.05 mg, 1.27 mmol) were added to N,N-dimethylformamide (120 mL). The mixture was stirred at 25°C under nitrogen for 0.5 hour until the reddish-brown color disappeared. Methyl (2R)-2-(tert-butoxycarbonylamino)-3-iodopropionate (8.35 g, 25.38 mmol) was then added, and the mixture was stirred at 25°C for 1 hour. 1-(4-iodophenyl)cyclopropanol (6.6 g, 25.38 mmol), tris(dibenzylideneacetone)dipalladium (1.16 g, 1.27 mmol), and 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (1.04 g, 2.54 mmol) were then added, and the mixture was stirred at 50°C for 12 hours.

[0699] After thin-layer chromatography monitoring showed that the raw materials had reacted completely, the reaction solution was filtered through diatomaceous earth, the filtrate was added with water (60 mL), extracted with ethyl acetate (90 mL×2), the organic phases were combined, washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: petroleum ether / ethyl acetate eluent, 5 / 1 gradient) to give 2 g of (S)-2-(tert-butoxycarbonylamino)-3-(4-propionylphenyl)propionic acid methyl ester.

[0700] MS (ESI) M / Z: 336.2 [M+H] + .

[0701] 1 H NMR (400MHz, DMSO-d6): δ7.93-7.83(m,2H),7.46-7.30(m,3H),4.30-4.18(m,1H),3.72-3.58(m,3H),3.13-2.86(m,4H),1.26(br s,9H),1.14-1.02(m,J=7.2Hz,3H).

[0702] Step C: Dissolve (S)-methyl 2-(tert-butoxycarbonylamino)-3-(4-propionylphenyl)propanoate (820 mg, 2.44 mmol) in acetonitrile (15 mL), add di-tert-butyl dicarbonate (800.38 mg, 3.67 mmol, 842.51 μL) and 4-dimethylaminopyridine (29.87 mg, 244.49 μmol), and stir at 70 ° C for 12 hours.

[0703] After LC-MS monitoring indicated that the reaction of the starting materials was complete, the mixture was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (eluent: petroleum ether / ethyl acetate, 5 / 1 gradient) to obtain 1.03 g of methyl (S)-2-(bis(tert-butyloxycarbonyl)amino)-3-(4-propionylphenyl)propanoate.

[0704] MS (ESI) M / Z: 436.2 [M+H] + .

[0705] 1 H NMR (400MHz, DMSO-d6): δ7.89(d,J=8.0Hz,2H),7.31(d,J=8.3Hz,2H),5.24(dd,J=10.9,4.9Hz,1H),3.71(s,3H),3 .39-3.46(m,1H),3.17-3.20(m,1H),3.00(q,J=7.3Hz,2H),1.99-2.03(m,3H),1.33(s,18H),1.10(t,J=7.3Hz,3H).

[0706] Step D: Dissolve (S)-methyl 2-(bis(tert-butoxycarbonyl)amino)-3-(4-propionylphenyl)propanoate (1.03 g, 2.37 mmol) and N-bromosuccinimide (496.70 mg, 2.79 mmol) in carbon tetrachloride (20 mL) and stir at 80°C under UV light for 1 hour. Filter the mixture, and concentrate the filtrate under reduced pressure. Add acetone (20 mL) and silver nitrate (760 mg, 4.47 mmol) to the residue and stir at 20°C for 12 hours.

[0707] After thin-layer chromatography monitoring indicated complete reaction of the starting materials, the mixture was concentrated under reduced pressure, diluted with ethyl acetate (10 mL), and filtered. The filtrate was washed with saturated aqueous ammonium chloride (30 mL) and saturated brine (30 mL), and the organic phase was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: petroleum ether / ethyl acetate, 3 / 1 gradient) to yield 312 mg of 3-tert-butyl-4-methyl-(4S,5R)-2-oxo-5-(4-propionylphenyl)oxazolidine-3,4-dicarboxylate.

[0708] MS (ESI) M / Z: 378.2 [M+H] + .

[0709] 1 H NMR (400MHz, DMSO-d6): δ8.04(d,J=8.3Hz,2H),7.62(d,J=8.5Hz,2H),5.79-5.82(m,1H),4. 75(d,J=5.8Hz,1H),3.80(s,3H),3.07(q,J=7.0Hz,2H),1.41(s,9H),1.09(t,J=7.0Hz,3H).

[0710] Step E: A mixture of 3-tert-butyl-4-methyl-(4S,5R)-2-oxo-5-(4-propionylphenyl)oxazolidine-3,4-dicarboxylate (300 mg, 794.94 μmol) and cesium carbonate (51.80 mg, 158.99 μmol) was added to methanol (10 mL) and stirred at 20°C for 1 hour.

[0711] After thin-layer chromatography monitoring indicated complete reaction of the starting material, the mixture was diluted with water (10 mL). The pH was adjusted to 2-3 with aqueous potassium bisulfate (3 mL), and the mixture was extracted with ethyl acetate (15 mL x 2). The organic phases were combined and concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: petroleum ether / ethyl acetate, 2 / 1 gradient) to yield 150 mg of methyl (2S,3R)-2-(tert-butoxycarbonylamino)-3-hydroxy-3-(4-propionylphenyl)propanoate.

[0712] MS (ESI) M / Z: 352.2 [M+H] + .

[0713] 1 H NMR (400MHz, CDCl3): δ7.96(d,J=8.28Hz,2H),7.48(d,J=8.28Hz,2H),5.32(br s,2H),4.53-4.63(m,1H),3.79(s,3H),3.00(d,J=7.28Hz,2H),1.19-1.35(m,12H).

[0714] Step F: Dissolve methyl (2S,3R)-2-(tert-butoxycarbonylamino)-3-hydroxy-3-(4-propionylphenyl)propanoate (150 mg, 426.87 μmol) in dichloromethane (1 mL). Add trifluoroacetic acid (1.54 g, 13.46 mmol, 1 mL) dropwise at 0°C and stir at 0°C for 1 hour.

[0715] After monitoring by thin layer chromatography showed that the starting material had reacted completely, the mixture was concentrated under reduced pressure to obtain 170 mg of crude trifluoroacetic acid salt of (2S,3R)-2-amino-3-hydroxy-3-(4-propionylphenyl)propionic acid methyl ester without purification.

[0716] MS (ESI) M / Z: 252.1 [M+H] + .

[0717] Step G: Methyl (2S,3R)-2-amino-3-hydroxy-3-(4-propionylphenyl)propanoate (160 mg, 437.99 μmol) was dissolved in N,N-dimethylformamide (3 mL). (2-morpholinoacetyl)-L-alanine (113.65 mg, 525.59 μmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (249.81 mg, 656.99 μmol) and N,N-diisopropylethylamine (452.85 mg, 3.50 mmol, 610.31 μL) were added at 0°C and stirred at 0°C for 1 hour.

[0718] After LC-MS monitoring indicated complete reaction of the starting materials, the mixture was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (eluent: petroleum ether / ethyl acetate, 3 / 1 gradient) to yield 160 mg of crude product, methyl (2S,3R)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)-3-(4-propionylphenyl)propanoate.

[0719] MS (ESI) M / Z: 450.3 [M+H] + .

[0720] Step H: Methyl (2S,3R)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)-3-(4-propionylphenyl)propanoate (160 mg, 355.95 μmol) was dissolved in acetonitrile (2 mL) and water (0.2 mL). Lithium bromide (247.30 mg, 2.85 mmol) and triethylamine (108.06 mg, 1.07 mmol, 148.63 μL) were added at 0°C and stirred at 20°C for 12 hours.

[0721] After LC-MS monitoring indicated complete reaction of the starting material, the mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC (column: Welch Xtimate C18 150 × 30 mm × 5 μm; mobile phase: [water (ammonium bicarbonate)-acetonitrile]; gradient: 8% to 38% phase B over 7 minutes) to yield 65 mg of (2S,3R)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)-3-(4-propionylphenyl)propanoic acid.

[0722] MS (ESI) M / Z: 436.3 [M+H] + .

[0723] 1 H NMR (400MHz, CD3OD): δ7.93(d,J=8.28Hz,2H),7.55(d,J=8.28Hz,2H),5.39(d,J=2.51Hz,1H),4.73(d,J=2.76Hz,1H),4.44(d,J=7.03Hz,1H),3.71(br t,J=4.64Hz,4H),3.12-3.23(m,2H),3.04(d,J=7.28Hz,2H),2.54-2.69(m,4H),1.27(d,J=7.03Hz,3H),1.18(t,J=7.28Hz,3H).

[0724] Step I: (2S,3R)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)-3-(4-propionylphenyl)propanoic acid (60 mg, 137.78 μmol) was dissolved in N,N-dimethylformamide (1.5 mL), and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxirane- To the mixture was added 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (78.58 mg, 206.67 μmol) and N,N-diisopropylethylamine (53.42 mg, 413.35 μmol, 72.00 μL), and stirred at 0°C for 0.5 hours.

[0725] After LC-MS monitoring indicated complete reaction of the starting materials, the mixture was concentrated under reduced pressure. The residue was separated and purified by preparative HPLC to yield 20 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)-3-(4-propionylphenyl)propionamide.

[0726] MS (ESI) M / Z: 613.4 [M+H] + .

[0727] 1H NMR (400MHz, CD3OD): δ1.18(t,J=7.28Hz,3H),1.29(d,J=7.03Hz,3H),1.41(s,3H),1.83-1.91(m,2H),2.24-2 .37(m,5H),2.42-2.55(m,5H),2.89-2.94(m,1H),2.97(s,1H),2.95-2.99(m,1H),3.00-3.08(m,2H),3.02-3.0 4(m,1H),3.17(d,J=5.02Hz,1H),3.67(t,J=4.52Hz,4H),4.43(d,J=7.03Hz,1H),4.60(d,J=4.27Hz,1H),4.68 (dd,J=8.53,5.02Hz,1H),5.15(d,J=4.02Hz,1H),5.46(s,1H),7.51(d,J=8.28Hz,2H),7.92(d,J=8.28Hz,2H).

[0728] Example 22:

[0729] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-hydroxy-3-(4-(2-methoxyacetyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0730] Reaction route:

[0731] Steps:

[0732] Step A: Dissolve 1-(4-bromophenyl)-2-methoxy-ethanone (5 g, 21.83 mmol) in toluene (75 mL), add ethylene glycol (2.72 g, 43.77 mmol, 2.44 mL) and toluene-4-sulfonic acid (415.00 mg, 2.41 mmol), and stir at 140 ° C for 4 hours through a water separator.

[0733] After TLC monitoring indicated that the reaction of the starting materials was complete, the reaction solution was concentrated under reduced pressure. The residue was purified by basic flash silica gel column chromatography (eluent: ethyl acetate) to obtain 4.2 g of 2-(4-bromophenyl)-2-(methoxymethyl)-1,3-dioxolane.

[0734] MS (ESI) M / Z: 273.0 [M+H] + .

[0735] 1H NMR (400MHz, CDCl3): δ7.51-7.46(m,2H),7.42-7.37(m,2H),4.16-4.07(m,2H),3.90-3.80(m,2H),3.58(s,2H),3.40(s,3H).

[0736] Step B: Dissolve 2-(4-bromophenyl)-2-(methoxymethyl)-1,3-dioxolane (3.67 g, 13.44 mmol) in tetrahydrofuran (40 mL). Under a nitrogen atmosphere, slowly add a cyclohexane solution of n-butyllithium (2.5 M, 6.98 mL) dropwise at -78°C and stir for 30 minutes. Then, slowly add N,N-dimethylformamide (2.95 g, 40.31 mmol, 3.10 mL) dropwise at -78°C and stir for 10 minutes.

[0737] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was quenched with saturated aqueous ammonium chloride (30 mL) at 0°C and extracted with ethyl acetate (35 mL x 2). The combined organic phases were washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: petroleum ether / ethyl acetate, 2 / 1 gradient) to yield 2.4 g of 4-(2-(methoxymethyl)-1,3-dioxolan-2-yl)benzaldehyde.

[0738] MS (ESI) M / Z: 223.2 [M+H] + .

[0739] 1 H NMR (400MHz, CDCl3): δ10.03(s,1H),7.88(d,J=7.5Hz,2H),7.70(d,J=7.6Hz,2H),4.19-4.12(m,2H),3.93-3.84(m,2H),3.62(s,2H),3.40(s,3H).

[0740] Step C: (R)-2-(N-(N'-Benzylprolyl)amino)diphenylmethanone-nickel(II)-glycine complex (500 mg, 1.00 mmol) and 4-(2-(methoxymethyl)-1,3-dioxolan-2-yl)benzaldehyde (669.12 mg, 3.01 mmol) were dissolved in methanol (6 mL), and a methanol solution of sodium methoxide (5.5 M, 912.38 μL) was added. After stirring at 20°C for 12 hours, dilute hydrochloric acid (2 M, 10.00 mL) was added, and the mixture was stirred at 50°C for 2 hours.

[0741] After LC-MS monitoring indicated complete reaction of the starting material, the reaction mixture was concentrated under reduced pressure to remove the methanol and filtered. The filtrate was extracted with dichloromethane (20 mL x 5). The organic phase was discarded, and the aqueous phase was concentrated under reduced pressure. The residue was purified without further purification to yield 870 mg of crude (2S,3R)-2-amino-3-hydroxy-3-(4-(2-methoxyacetyl)phenyl)propanoic acid hydrochloride.

[0742] MS (ESI) M / Z: 298.1 [M+H] + .

[0743] Step D: Dissolve (2S,3R)-2-amino-3-hydroxy-3-(4-(2-methoxyacetyl)phenyl)propanoic acid hydrochloride (870 mg, 2.93 mmol) in methanol (16 mL) at 0°C, and add thionyl chloride (1.04 g, 8.78 mmol, 637.62 μL) dropwise. Stir at 50°C for 12 hours.

[0744] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was diluted with water (10 mL), the pH adjusted to 9 with aqueous ammonia, and then extracted with ethyl acetate (10 mL x 2). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified without further purification to yield 130 mg of methyl (2S,3R)-2-amino-3-hydroxy-3-(4-(2-methoxyacetyl)phenyl)propanoate.

[0745] MS (ESI) M / Z: 268.1 [M+H] + .

[0746] Step E: At 0°C, (2-morpholinoacetyl)-L-alanine (130.82 mg, 604.99 μmol) and (2S,3R)-2-amino-3-hydroxy-3-(4-(2-methoxyacetyl)phenyl)propionic acid methyl ester (147 mg, 549.99 μmol) were dissolved in N,N-dimethylformamide (2 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (418.25 mg, 1.10 mmol) and N,N-diisopropylethylamine (213.25 mg, 1.65 mmol, 287.39 μL) were added and stirred at 0°C for 30 min.

[0747] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was diluted with water (10 mL) and extracted with ethyl acetate (10 mL x 2). The combined organic phases were washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: petroleum ether / ethyl acetate, 0 / 1 gradient) to yield 96 mg of methyl (2S,3R)-3-hydroxy-3-(4-(2-methoxyacetyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[0748] MS (ESI) M / Z: 466.2 [M+H] + .

[0749] Step F: Methyl (2S,3R)-3-hydroxy-3-(4-(2-methoxyacetyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate (95 mg, 204.08 μmol) was dissolved in acetonitrile (2 mL) and water (0.2 mL), and lithium bromide (88.62 mg, 1.02 mmol) and N,N-diisopropylethylamine (79.13 mg, 612.25 μmol, 106.64 μL) were added and stirred at 20 °C for 12 hours.

[0750] After thin-layer chromatography monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was purified by preparative high-performance liquid chromatography (column: C18, 150 × 30 mm; mobile phase: [water (ammonia water + ammonium bicarbonate)-acetonitrile]; gradient: 5% to 35% phase B over 7 minutes) to yield 30 mg of (2S,3R)-3-hydroxy-3-(4-(2-methoxyacetyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid.

[0751] MS (ESI) M / Z: 629.4 [M+H] + .

[0752] 1H NMR (400MHz, CD3OD): δ7.88(d,J=8.3Hz,2H),7.53(d,J=8.2Hz,2H),5.45(s,1H),5.15(d,J=4.2Hz ,1H),4.80-4.76(m,2H),4.68(dd,J=5.0,8.7Hz,1H),4.60(d,J=4.2Hz,1H),4.42(d,J=7.0Hz,1H) ,3.67(t,J=4.6Hz,4H),3.49-3.46(m,3H),3.17(d,J=5.1Hz,1H),3.07-2.95(m,2H),2.93-2.90(m ,1H),2.54-2.43(m,5H),2.35-2.25(m,5H),1.91-1.81(m,2H),1.41(s,3H),1.28(d,J=7.0Hz,3H).

[0753] Step G: (2S,3R)-3-hydroxy-3-(4-(2-methoxyacetyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoic acid (54 mg, 119.61 μmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (52.74 mg, 143.53 μmol) were dissolved in N,N-dimethylformamide (1 mL) at 0°C and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (90.96 mg, 239.22 μmol) was added. Then, N,N-diisopropylethylamine (77.29 mg, 598.05 μmol, 104.17 μL) was added and the reaction mixture was stirred at 0° C. for 30 minutes.

[0754] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was purified by preparative HPLC to yield 16.09 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-hydroxy-3-(4-(2-methoxyacetyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide.

[0755] MS (ESI) M / Z: 629.3 [M+H] + .

[0756] 1H NMR (400MHz, CD3OD): δ7.89(d,J=8.3Hz,2H),7.53(d,J=8.3Hz,2H),5.46(s,1H),5.15(d,J=4.0Hz ,1H),4.79(s,2H),4.68(dd,J=8.8,5.0Hz,1H),4.60(d,J=4.3Hz,1H),4.43(q,J=7.0Hz,1H),3.68 (t,J=4.6Hz,4H),3.48(s,3H),3.18(d,J=5.0Hz,1H),2.93-3.07(m,2H),2.91(d,J=5.0Hz,1H),2. 42-2.55(m,5H),2.24-2.36(m,5H),1.86(quin,J=7.3Hz,2H),1.42(s,3H),1.29(d,J=7.0Hz,3H).

[0757] Example 23:

[0758] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-hydroxy-3-(4-(methoxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0759] Reaction route:

[0760] Steps:

[0761] Step A: Under nitrogen at -78°C, dissolve 1-bromo-4-(methoxymethyl)benzene (5 g, 24.87 mmol) in tetrahydrofuran (50 mL) and add a 2.5 M solution of n-butyllithium in cyclohexane (9.95 mL). Stir at -78°C for 30 minutes, then add N,N-dimethylformamide (4.75 g, 64.98 mmol, 5.00 mL). Slowly warm the mixture to 25°C and stir for 30 minutes.

[0762] Liquid chromatography-mass spectrometry (LC-MS) monitoring indicated the formation of the product. Water (50 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (50 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and then filtered. The filtrate was concentrated under reduced pressure. The resulting mixture was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 5 / 1) to yield 3.3 g of 4-(methoxymethyl)benzaldehyde.

[0763] MS (ESI) M / Z: 150.9 [M+H] + .

[0764] 1 H NMR (400MHz, CDCl3): δ10.01 (s, 1H), 7.85-7.89 (m, 2H), 7.50 (d, J = 7.99Hz, 2H), 4.54 (s, 2H), 3.43 (s, 3H).

[0765] Step B: 4-(Methoxymethyl)benzaldehyde (778.59 mg, 5.18 mmol) and (R)-2-(N-(N'-benzylprolyl)amino)diphenylmethanone-nickel(II)-glycine complex (500 mg, 1.04 mmol) were dissolved in methanol (7.5 mL), and a methanol solution of sodium methoxide (280.07 mg, 5.18 mmol) was added at 25° C. After stirring at 25° C. for 12 hours, dilute hydrochloric acid (2 M, 10.00 mL) was added, and the mixture was stirred at 50° C. for 2 hours.

[0766] After LC-MS monitoring showed that the raw materials were completely reacted, the reaction solution was concentrated under reduced pressure to remove methanol, and a white solid was generated. The solid was filtered, and the filtrate was washed with dichloromethane (15 mL×5). The aqueous phase was concentrated under reduced pressure to obtain 280 mg of crude (2S,3R)-2-amino-3-hydroxy-3-(4-(methoxymethyl)phenyl)propionic acid hydrochloride.

[0767] MS (ESI) M / Z: 226.1 [M+H] + .

[0768] Step C: (2S,3R)-2-amino-3-hydroxy-3-(4-(methoxymethyl)phenyl)propanoic acid hydrochloride (280 mg, 1.07 mmol) was dissolved in methanol (5 mL), thionyl chloride (254.58 mg, 2.14 mmol, 155.42 μL) was added, and the mixture was stirred at 50° C. for 12 hours.

[0769] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure, the residue was dissolved in water (10 mL), the pH was adjusted to 9 with aqueous ammonia, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to yield 195 mg of methyl (2S,3R)-2-amino-3-hydroxy-3-(4-(methoxymethyl)phenyl)propanoate.

[0770] MS (ESI) M / Z: 240.1 [M+H] + .

[0771] Step D: At 0°C, methyl (2S,3R)-2-amino-3-hydroxy-3-(4-(methoxymethyl)phenyl)propanoate (195 mg, 814.99 μmol) and (2-morpholinoacetyl)-L-alanine (176.23 mg, 814.99 μmol) were dissolved in tetrahydrofuran (3 mL), and 3-(diethoxyphosphoryloxy)-1,2,3-benzotriazine-4-one (268.25 mg, 896.49 μmol) and sodium carbonate (172.76 mg, 1.63 mmol) were added sequentially, and the mixture was stirred at 25°C for 12 hours.

[0772] After LC-MS monitoring indicated complete reaction of the starting materials, water (15 mL) was added to the reaction solution, which was then extracted with dichloromethane, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The resulting mixture was purified by silica gel column chromatography (eluent: ethyl acetate) to yield 279.2 mg of methyl (2S,3R)-3-hydroxy-3-(4-(methoxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[0773] MS (ESI) M / Z: 438.3 [M+H] + .

[0774] Step E: Methyl (2S,3R)-3-hydroxy-3-(4-(methoxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate (279 mg, 637.74 μmol) was dissolved in acetonitrile (3 mL) and water (0.1 mL) at 0°C, and triethylamine (193.60 mg, 1.91 mmol, 266.30 μL) and lithium bromide (276.94 mg, 3.19 mmol) were added and stirred at 25°C for 12 hours.

[0775] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The resulting mixture was separated by preparative HPLC (separation column: C18 150×30 mm; mobile phase: [water (formic acid)-acetonitrile]; gradient: phase B from 0% to 26% over 7 minutes) to yield 158.9 mg of (2S,3R)-3-hydroxy-3-(4-(methoxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid.

[0776] MS (ESI) M / Z: 424.2 [M+H] + .

[0777] Step F: (2S,3R)-3-hydroxy-3-(4-(methoxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoic acid (158.9 mg, 338.46 μmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (136 .81 mg, 372.30 μmol) was dissolved in N,N-dimethylformamide (2 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (141.56 mg, 372.30 μmol) and N,N-diisopropylethylamine (218.71 mg, 1.69 mmol, 294.76 μL) were added in sequence, and the mixture was stirred at 0°C for 0.5 h.

[0778] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The resulting mixture was separated and purified by preparative HPLC to yield 70 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-hydroxy-3-(4-(methoxymethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide.

[0779] MS (ESI) M / Z: 601.2 [M+H] + .

[0780] 1 H NMR (400MHz, CD3OD): δ7.36(d,J=8.28Hz,2H),7.26(d,J=8.03Hz,2H),5.45(s,1H),5.08(d,J=4 .27Hz,1H),4.67(dd,J=8.41,5.14Hz,1H),4.56(d,J=4.27Hz,1H),4.39-4.47(m,3H),3.67-3.7 2(m,4H),3.36(s,3H),3.13(d,J=4.77Hz,1H),2.96-3.08(m,2H),2.89(d,J=4.77Hz,1H),2.44- 2.54(m,5H),2.23-2.36(m,5H),1.86(quin,J=7.34Hz,2H),1.42(s,3H),1.29(d,J=7.03Hz,3H).

[0781] Example 24:

[0782] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-hydroxy-3-(4-(methoxyethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0783] Reaction route:

[0784] Steps:

[0785] Step A: Under nitrogen protection, (R)-2-(N-(N'-benzylprolyl)amino)diphenylmethanone-nickel(II)-glycine complex (600 mg, 1.20 mmol) and 4-(2-methoxyethyl)benzaldehyde (593.26 mg, 3.61 mmol) were dissolved in methanol (10 mL), and a methanol solution of sodium methoxide (5.5 M, 1.09 mL) was added dropwise at 25°C. After stirring at 25°C for 12 hours, dilute hydrochloric acid (2 M, 10.00 mL) was added, and the mixture was stirred at 50°C for 2 hours.

[0786] After LC-MS monitoring showed that the raw materials had reacted completely, the reaction solution was concentrated under reduced pressure to remove methanol, and a white solid was generated. The solid was filtered, and the filtrate was washed with dichloromethane (15 mL×5). The aqueous phase was concentrated under reduced pressure to obtain 900 mg of crude (2S,3R)-2-amino-3-hydroxy-3-(4-(2-methoxyethyl)phenyl)propionic acid hydrochloride.

[0787] MS (ESI) M / Z: 240.1 [M+H] + .

[0788] 1 H NMR (400MHz, D2O): δ7.30-7.48(m,4H),5.39(br s,1H),4.10-4.27(m,1H),3.72(br t,J=6.0Hz,2H),3.19-3.36(m,3H),2.89(br t,J=6.0Hz).

[0789] Step B: At 0°C, (2S,3R)-2-amino-3-hydroxy-3-(4-(2-methoxyethyl)phenyl)propanoic acid hydrochloride (800 mg, 3.34 mmol) was dissolved in methanol (15 mL), and thionyl chloride (10.03 mmol, 728.54 μL) was added dropwise, followed by stirring at 50°C for 12 hours.

[0790] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure, the residue was dissolved in water (10 mL), the pH was adjusted to 9 with aqueous ammonia, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to yield 280 mg of methyl (2S,3R)-2-amino-3-hydroxy-3-(4-(2-methoxyethyl)phenyl)propanoate.

[0791] MS (ESI) M / Z: 254.1 [M+H] + .

[0792] 1 H NMR (400MHz, CD3OD): δ7.19-7.40 (m, 4H), 5.03 (d, J = 4.4Hz, 1H), 3.78 (d, J = 4.4Hz, 1H),3.67-3.73(m,3H),3.60(t,J=6.9Hz,2H),3.32(s,3H),2.86(t,J=6.8Hz,2H).

[0793] Step C: At 0°C, methyl (2S,3R)-2-amino-3-hydroxy-3-(4-(2-methoxyethyl)phenyl)propanoate (280 mg, 1.11 mmol) and (2-morpholinoacetyl)-L-alanine (286.84 mg, 1.33 mmol) were dissolved in N,N-dimethylformamide (3 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (630.48 mg, 1.66 mmol) and N,N-diisopropylethylamine (428.61 mg, 3.32 mmol, 577.64 μL) were added in sequence, and the mixture was stirred at 0°C for 0.5 hour.

[0794] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (20 g silica gel column, methanol / dichloromethane eluent, 0-6% gradient, 35 mL / min) to afford 480 mg of methyl (2S,3R)-3-hydroxy-3-(4-(methoxyethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[0795] MS (ESI) M / Z: 452.3 [M+H] + .

[0796] Step D: Methyl (2S,3R)-3-hydroxy-3-(4-(methoxyethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate (480 mg, 1.06 mmol) was dissolved in acetonitrile (10 mL) and water (0.1 mL) at 0°C, and lithium bromide (923.30 mg, 10.63 mmol) and triethylamine (322.72 mg, 3.19 mmol, 443.91 μL) were added sequentially. The mixture was stirred at 25°C for 12 hours.

[0797] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was separated by preparative HPLC (column: C18 150×30 mm; mobile phase: [water (ammonia + ammonium bicarbonate)-acetonitrile]; gradient: 7% to 37% phase B over 7 minutes) to yield 110 mg of (2S,3R)-3-hydroxy-3-(4-(methoxyethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid.

[0798] MS (ESI) M / Z: 438.3 [M+H] + .

[0799] Step E: (2S,3R)-3-hydroxy-3-(4-(methoxyethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoic acid (110 mg, 251.44 μmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (110. 87 mg, 301.72 μmol) was dissolved in N,N-dimethylformamide (2 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (143.41 mg, 377.16 μmol) and N,N-diisopropylethylamine (97.49 mg, 754.31 μmol) were added sequentially at 0°C, and the mixture was stirred at 0°C for 0.5 h.

[0800] After LC-MS / MS monitoring showed that the raw materials had reacted completely, the reaction solution was separated and purified by preparative HPLC to obtain 23.7 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-hydroxy-3-(4-(methoxyethyl)phenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide.

[0801] MS (ESI) M / Z: 615.3 [M+H] + .

[0802] 1 H NMR (400MHz, CD3OD): δ7.29(d,J=8.1Hz,2H),7.16(d,J=8.0Hz,2H),5.45(s,1H),5.04(d,J=4.4Hz,1H),4.67(dd, J=8.4,5.2Hz,1H),4.53(d,J=4.4Hz,1H),4.42(q,J=7.2Hz,1H),3.65-3.77(m,4H),3.58(t,J=6.9Hz,2H),3.32(br s,3H),3.13(d,J=5.0Hz,1H),3.02(d,J=14.1Hz,2H),2.89(d,J=5.0Hz,1H),2.79-2.85(m,2H),2.44-2.56(m,5H),2.28(br t,J=7.8Hz,5H),1.86(s,2H),1.42(s,3H),1.29(d,J=7.0Hz,3H).

[0803] Example 25:

[0804] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-hydroxy-3-(4-methylthiophenyl)-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0805] Reaction route:

[0806] Steps:

[0807] Step A: Dissolve (R)-2-(N-(N'-Benzylprolyl)amino)diphenylmethanone-nickel(II)-glycine complex (500 mg, 1.00 mmol) and 4-(difluoromethoxy)-2-fluoro-benzaldehyde (458.29 mg, 3.01 mmol, 400.60 μL) in methanol (10 mL). Add sodium methoxide in methanol (5 M, 802.89 μL) and stir at 20°C for 0.5 h. Add dilute hydrochloric acid (2 M, 10.00 mL) to the reaction mixture and stir at 50°C for 2 h.

[0808] After LC-MS monitoring indicated complete reaction of the starting material, the reaction mixture was concentrated under reduced pressure to remove the methanol and filtered. The aqueous phase was extracted with dichloromethane (20 mL x 5). The organic phase was discarded, and the aqueous phase was filtered and concentrated under reduced pressure. No purification was required to obtain 650 mg of crude (2S,3R)-2-amino-3-hydroxy-3-(4-methylthiophenyl)propionic acid hydrochloride.

[0809] MS (ESI) M / Z: 228.1 [M+H] + .

[0810] Step B: Dissolve (2S,3R)-2-amino-3-hydroxy-3-(4-methylthiophenyl)propanoic acid hydrochloride (650 mg, 2.86 mmol) in methanol (12 mL), add thionyl chloride (1.36 g, 11.44 mmol, 830.88 μL), and stir at 50°C for 12 hours.

[0811] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was concentrated under reduced pressure, the residue was dissolved in water (10 mL), the pH was adjusted to 9 with aqueous ammonia, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to yield 170 mg of crude (2S,3R)-2-amino-3-hydroxy-3-(4-methylthiophenyl)propionic acid methyl ester.

[0812] MS (ESI) M / Z: 242.1 [M+H] + .

[0813] Step C: Methyl (2S,3R)-2-amino-3-hydroxy-3-(4-methylthiophenyl)propanoate (170 mg, 704.50 μmol) was dissolved in N,N-dimethylformamide (3 mL). At 0°C, (2-morpholinoacetyl)-L-alanine (152.34 mg, 704.50 μmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (401.81 mg, 1.06 mmol) and N,N-diisopropylethylamine (273.15 mg, 2.11 mmol, 368.13 μL) were added in sequence and the mixture was stirred at 0°C for 0.5 h.

[0814] After LC-MS monitoring indicated complete reaction of the starting materials, the mixture was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (eluent: petroleum ether / ethyl acetate, 0 / 1 gradient) to yield 240 mg of crude product, methyl (2S,3R)-3-hydroxy-3-(4-methylthiophenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[0815] MS (ESI) M / Z: 440.1 [M+H] + .

[0816] Step D: Methyl (2S,3R)-3-hydroxy-3-(4-methylthiophenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate (240 mg, 546.04 μmol) was dissolved in acetonitrile (2 mL) and water (0.2 mL), and lithium bromide (237.11 mg, 2.73 mmol) and triethylamine (165.76 mg, 1.64 mmol, 228.01 μL) were added at 0°C, and then stirred at 20°C for 12 hours.

[0817] After LC-MS / MS monitoring showed that the starting material had reacted completely, the mixture was concentrated under reduced pressure, and the residue was separated by preparative HPLC (separation column: C18 150×30 mm; mobile phase: [water (ammonia water + ammonium bicarbonate)-acetonitrile]; gradient: phase B increased from 10% to 40% in 7 minutes) to give 80 mg of (2S,3R)-3-hydroxy-3-(4-methylthiophenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionic acid.

[0818] MS (ESI) M / Z: 426.2 [M+H] + .

[0819] 1 H NMR (400MHz, CD3OD): δ7.32(d,J=8.28Hz,2H),7.17(d,J=8.28Hz,2H),5.24(d,J=2.76Hz,1H),4.61(d,J=3.01Hz,1H), 4.41-4.50(m,1H),3.69(t,J=4.64Hz,4H),2.93-3.12(m,2H),2.46-2.55(m,4H),2.44(s,3H),1.30(d,J=7.03Hz,3H).

[0820] Step E: (2S,3R)-3-hydroxy-3-(4-methylthiophenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid (70 mg, 164.51 μmol) was dissolved in N,N-dimethylformamide (1.5 mL), and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxirane- In the mixture was added 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (93.83 mg, 246.77 μmol) and N,N-diisopropylethylamine (63.79 mg, 493.54 μmol, 85.97 μL), and stirred at 0°C for 0.5 hours.

[0821] After LC-MS monitoring indicated complete reaction of the starting material, the mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC to yield 43 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-hydroxy-3-(4-methylthiophenyl)-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide.

[0822] MS (ESI) M / Z: 603.3 [M+H] + .

[0823] 1 H NMR (400MHz, CD3OD): δ7.30(d,J=8.28Hz,2H),7.17(d,J=8.53Hz,2H),5.45(s,1H),4. 99-5.05(m,1H),4.67(dd,J=8.53,5.27Hz,1H),4.55(d,J=4.52Hz,1H),4.46(q,J=7.03 Hz,1H),3.69(t,J=4.52Hz,4H),3.03-3.14(m,2H),2.87-3.00(m,2H),2.41-2.54(m,8 H),2.23-2.35(m,5H),1.86(quin,J=7.40Hz,2H),1.43(s,3H),1.31(d,J=7.03Hz,3H).

[0824] Example 26:

[0825] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-ethynylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0826] Reaction route:

[0827] Steps:

[0828] Step A: Under nitrogen protection, 3-tert-butyl-4-methyl-(4S,5R)-5-(4-bromophenyl)-2,2-dimethyloxazolidine-3,4-dicarboxylate (500 mg, 1.21 mmol), tributyl(ethynyl)stannane (760.53 mg, 2.41 mmol) and bistriphenylphosphine palladium dichloride (84.71 mg, 120.69 μmol) were dissolved in dioxane (8 mL), and the reaction solution was heated to 90 ° C and stirred for 12 hours.

[0829] After LC-MS monitoring showed that the raw material reaction was complete, the reaction solution was quenched with a saturated aqueous solution of potassium fluoride (20 mL), extracted with ethyl acetate (20 mL), and the organic phase was concentrated under reduced pressure to give a crude product, which was subjected to flash silica gel chromatography (12 g silica gel column, ethyl acetate / petroleum ether eluent, 0-3% gradient, 40 mL / min) to give 300 mg of 3-tert-butyl-4-methyl-(4S,5R)-5-(4-ethynylphenyl)-2,2-dimethyloxazolidine-3,4-dicarboxylate.

[0830] MS (ESI) M / Z: 359.9 [M+H] + .

[0831] 1 H NMR (400MHz, CDCl3): δ7.51(d,J=8.0Hz,2H),7.35(d,J=8.3Hz,2H),5.01-5.10(m,1H),4.11-4 .31(m,1H),3.77(s,3H),3.08-3.13(m,1H),1.69-1.78(m,6H),1.55-1.59(m,2H),1.40(s,7H).

[0832] Step B: Dissolve 3-tert-butyl-4-methyl-(4S,5R)-5-(4-ethynylphenyl)-2,2-dimethyloxazolidine-3,4-dicarboxylate (300 mg, 834.69 μmol) in dichloromethane (3 mL) and trifluoroacetic acid (3 mL) and stir at 0°C for 1 hour.

[0833] After LC-MS monitoring showed that the raw materials had reacted completely, the reaction solution was concentrated under reduced pressure to obtain 180 mg of crude trifluoroacetic acid salt of (2S,3R)-2-amino-3-hydroxy-3-(4-ethynylphenyl)propionic acid methyl ester.

[0834] MS (ESI) M / Z: 219.7 [M+H] + .

[0835] Step C: At 0°C, (2S,3R)-2-amino-3-hydroxy-3-(4-ethynylphenyl)propionic acid methyl ester trifluoroacetate (180 mg, 821.03 μmol) and (2-morpholinoacetyl)-L-alanine (213.04 mg, 985.24 μmol) were dissolved in N,N-dimethylformamide (4 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (624.36 mg, 1.64 mmol) and N,N-diisopropylethylamine (530.56 mg, 4.11 mmol) were added sequentially, and the mixture was stirred at 0°C for 1 hour.

[0836] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (20 g silica gel column, methanol / dichloromethane eluent, 0-5% gradient, 40 mL / min) to yield 330 mg of crude product, methyl (2S,3R)-3-(4-ethynylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[0837] MS (ESI) M / Z: 417.9 [M+H] + .

[0838] Step D: Methyl (2S,3R)-3-(4-ethynylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoate (330 mg, 790.51 μmol) was dissolved in acetonitrile (5 mL) and water (0.5 mL) at 0°C, and lithium bromide (686.51 mg, 7.91 mmol) and N,N-diisopropylethylamine (306.50 mg, 2.37 mmol) were added sequentially. The mixture was stirred at 25°C for 12 hours.

[0839] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was separated by preparative HPLC (column: C18 150×30 mm; mobile phase: [water (ammonia + ammonium bicarbonate)-acetonitrile]; gradient: phase B from 7% to 37% over 7 minutes) to yield 87 mg of (2S,3R)-3-(4-ethynylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid.

[0840] MS (ESI) M / Z: 404.2 [M+H] + .

[0841] Step E: (2S,3R)-3-(4-ethynylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoic acid (85 mg, 210.69 μmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (92.91 mg, 252.83 μmol) were dissolved in N,N-dimethylformamide (2 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (160.22 mg, 421.39 μmol) and N,N-diisopropylethylamine (136.15 mg, 1.05 mmol) were added sequentially at 0°C and the mixture was stirred at 0°C for 1 hour.

[0842] After LC-MS / MS monitoring showed that the raw materials had reacted completely, the reaction solution was separated and purified by preparative HPLC to obtain 27.62 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxoprop-2-yl)-3-(4-ethynylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide.

[0843] MS (ESI) M / Z: 581.4 [M+H] + .

[0844] 1 H NMR (400MHz, CD3OD): δ7.37(s,4H),5.46(s,1H),5.10(d,J=3.9Hz,1H),4.68(dd,J= 8.5,5.1Hz,1H),4.58(d,J=3.9Hz,1H),4.44(q,J=7.0Hz,1H),3.69(t,J=4.6Hz,4H) ,3.48(s,1H),3.16(d,J=5.0Hz,1H),2.93-3.08(m,2H),2.87-2.92(m,1H),2.42-2. 54(m,5H),2.25-2.34(m,5H),1.82-1.91(m,2H),1.44(s,3H),1.29(d,J=7.0Hz,3H).

[0845] Example 27:

[0846] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-2-((S)-3-difluoromethoxy-2-(2-morpholinoacetylamino)propionamido)-3-hydroxy-3-(4-methoxyphenyl)propionamide

[0847] Reaction route:

[0848] Steps:

[0849] Step A: Dissolve N-tert-Butyloxycarbonyl-L-serine benzyl ester (9.5 g, 32.17 mmol), cuprous iodide (612.63 mg, 3.22 mmol), and sodium sulfate (456.91 mg, 3.22 mmol) in acetonitrile (100 mL), and then add 2,2-difluoro-2-fluorosulfonylacetic acid (8.59 g, 48.25 mmol, 4.99 mL). The mixture is stirred at 60°C for 2 hours.

[0850] After the reaction of the starting materials was complete as monitored by thin-layer chromatography, the reaction solution was extracted with ethyl acetate (200 mL). The organic phase was washed with saturated brine (200 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting mixture was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 3 / 1) to provide 1.49 g of N-tert-butoxycarbonyl-O-difluoromethyl-L-serine benzyl ester.

[0851] MS (ESI) M / Z: 368.1 [M+H] + .

[0852] 1 H NMR (400MHz, CDCl3): δ7.32-7.42(m,5H),5.97-6.36(m,1H),5.36(br d,J=8.0Hz,1H),5.17-5.26(m,2H),4.54-4.62(m,1H),4.31(dd,J=10.2,2.4Hz,1H),4.14(dd,J=10.3,3.0Hz,1H),1.46(s,9H).

[0853] Step B: Dissolve N-tert-Butyloxycarbonyl-O-difluoromethyl-L-serine benzyl ester (1.49 g, 4.31 mmol) in dichloromethane (15 mL) and trifluoroacetic acid (3 mL) and stir the mixture at 25°C for 2 hours.

[0854] Liquid chromatography-mass spectrometry monitoring showed the formation of the product, and the reaction solution was concentrated under reduced pressure to obtain 1.55 g of trifluoroacetic acid salt of O-difluoromethyl-L-serine benzyl ester.

[0855] MS (ESI) M / Z: 246.1 [M+H] + .

[0856] Step C: Dissolve O-difluoromethyl-L-serine benzyl ester trifluoroacetate (1.55 g, 4.31 mmol) and 2-morpholinoacetic acid (751.55 mg, 5.18 mmol) in dichloromethane (15 mL) and N,N-dimethylformamide (8 mL), add 4-(4,6-dimethoxytriazin-2-yl)-4-methylmorpholine hydrochloride (2.39 g, 8.63 mmol) and N-methylmorpholine (1.31 g, 12.94 mmol, 1.42 mL), and stir the mixture at 25°C for 2 hours.

[0857] LC-MS / MS monitoring showed the formation of the product, and the reaction solution was concentrated under reduced pressure. The resulting mixture was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 1 / 1) to obtain 1.51 g of O-difluoromethyl-N-(2-morpholinoacetyl)-L-serine benzyl ester.

[0858] MS (ESI) M / Z: 373.2 [M+H] + .

[0859] Step D: Dissolve O-difluoromethyl-N-(2-morpholinoacetyl)-L-serine benzyl ester (700 mg, 1.88 mmol) in methanol (10 mL) and add 10% wet palladium on carbon (200.06 mg). The reaction mixture was purged with hydrogen three times and the mixture was stirred under a hydrogen atmosphere (15 psi) at 25°C for 4 hours.

[0860] After LC-MS monitoring showed that the reaction of the raw materials was complete, the reaction solution was filtered and the filtrate was concentrated under reduced pressure to obtain 540 mg of O-difluoromethyl-N-(2-morpholinoacetyl)-L-serine.

[0861] MS (ESI) M / Z: 283.1 [M+H] + .

[0862] Step E: O-Difluoromethyl-N-(2-morpholinoacetyl)-L-serine (260 mg, 703.61 μmol) and (2S,3R)-2-amino-3-hydroxy-3-(4-methoxyphenyl)propionic acid methyl ester trifluoroacetate (238.71 mg, 703.61 μmol) were dissolved in N,N-dimethylformamide (3 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (321.04 mg, 844.33 μmol) and N,N-diisopropylethylamine (454.68 mg, 3.52 mmol, 612.78 μL) were added. The mixture was stirred at 0°C for 0.5 h.

[0863] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The resulting mixture was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 0 / 1) to yield 220 mg of methyl (2S,3R)-2-((S)-3-difluoromethoxy-2-(2-morpholinoacetylamino)propionylamino)-3-hydroxy-3-(4-methoxyphenyl)propanoate.

[0864] MS (ESI) M / Z: 489.9 [M+H] + .

[0865] Step F: Methyl (2S,3R)-2-((S)-3-difluoromethoxy-2-(2-morpholinoacetylamino)propionylamino)-3-hydroxy-3-(4-methoxyphenyl)propanoate (100 mg, 204.30 μmol) was dissolved in acetonitrile (1 mL) and water (0.1 mL). Triethylamine (62.02 mg, 612.91 μmol, 85.31 μL) and lithium bromide (88.71 mg, 1.02 mmol) were then added. The mixture was stirred at 0°C for 2 hours.

[0866] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The resulting mixture was separated by preparative HPLC (separation column: Xtimate C18 150×40mm×10μm; mobile phase: [water(formic acid)-acetonitrile]; gradient: 8% to 38% phase B over 10 minutes) to yield 28.5 mg of (2S,3R)-2-((S)-3-difluoromethoxy-2-(2-morpholinoacetylamino)propionylamino)-3-hydroxy-3-(4-methoxyphenyl)propanoic acid.

[0867] MS (ESI) M / Z: 476.3 [M+H] + .

[0868] Step G: (2S,3R)-2-((S)-3-Difluoromethoxy-2-(2-morpholinoacetylamino)propionamido)-3-hydroxy-3-(4-methoxyphenyl)propanoic acid (28 mg, 58.89 μmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (23.80 mg, 64.78 μmol) were dissolved in N,N-dimethylformamide (0.5 mL) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (24.63 mg, 64.78 μmol) and N,N-diisopropylethylamine (38.06 mg, 294.46 μmol, 51.29 μL) were added. The mixture was stirred at 0°C for 0.5 hours.

[0869] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The resulting mixture was separated and purified by preparative HPLC to yield 15 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-2-((S)-3-difluoromethoxy-2-(2-morpholinoacetylamino)propionylamino)-3-hydroxy-3-(4-methoxyphenyl)propionamide.

[0870] MS (ESI) M / Z: 675.3 [M+H] +.

[0871] 1 H NMR (400MHz, CD3OD): δ7.28(d,J=8.5Hz,2H),6.84(d,J=8.8Hz,2H),6.21-6.60(m,1H),5.44(s,1H),5.01(d,J=4.8Hz, 1H),4.72(t,J=5.1Hz,1H),4.66(dd,J=8.3,5.5Hz,1H),4.57(d,J=4.5Hz,1H),4.05-4.15(m,2H),3.77(s,3H),3.75(br t,J=4.5Hz,4H),3.11(d,J=4.8Hz,1H),2.89(d,J=5.0Hz,1H),2.66-2.75(m ,4H),2.45-2.53(m,1H),2.22-2.33(m,6H),1.81-1.89(m,2H),1.41(s,3H).

[0872] Example 28:

[0873] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide

[0874] Reaction route:

[0875] Steps:

[0876] Step A: Under nitrogen protection, (R)-2-(N-(N'-benzylprolyl)amino)diphenylmethanone-nickel(II)-glycine complex (500 mg, 1.00 mmol) and 4-(difluoromethyl)benzaldehyde (470.08 mg, 3.01 mmol) were dissolved in methanol (10 mL). A methanol solution of sodium methoxide (5.5 M, 912.38 μL) was added dropwise at 25°C. After stirring at 25°C for 12 hours, dilute hydrochloric acid (2 M, 10 mL) was added and stirred at 50°C for 2 hours.

[0877] After LC-MS monitoring showed that the raw materials had reacted completely, the reaction solution was concentrated under reduced pressure to remove methanol, and a white solid was generated. The solid was filtered, and the filtrate was washed with dichloromethane (15 mL×5). The aqueous phase was concentrated under reduced pressure to obtain 800 mg of crude (2S,3R)-2-amino-3-(4-difluoromethylphenyl)-3-hydroxypropionic acid hydrochloride.

[0878] MS (ESI) M / Z: 232.0 [M+H] + .

[0879] 1 H NMR (400MHz, D2O): δ7.46-7.90(m,4H),6.62-7.09(m,1H),5.46(br s,1H),4.25(br s,1H).

[0880] Step B: At 0°C, (2S,3R)-2-amino-3-(4-difluoromethylphenyl)-3-hydroxypropanoic acid hydrochloride (750 mg, 2.60 mmol) was dissolved in methanol (15 mL), and thionyl chloride (926.25 mg, 7.79 mmol, 565.48 μL) was added dropwise, followed by stirring at 50°C for 12 hours.

[0881] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure, the residue was dissolved in water (10 mL), the pH was adjusted to 9 with aqueous ammonia, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to yield 320 mg of methyl (2S,3R)-2-amino-3-(4-difluoromethylphenyl)-3-hydroxypropanoate.

[0882] MS (ESI) M / Z: 246.1 [M+H] + .

[0883] 1 H NMR (400MHz, CD3OD): δ7.54(br s,4H),6.55-7.00(m,1H),5.09(br s,1H),3.61-3.87(m,4H).

[0884] Step C: At 0°C, methyl (2S,3R)-2-amino-3-(4-difluoromethylphenyl)-3-hydroxypropanoate (310 mg, 1.26 mmol) and (2-morpholinoacetyl)-L-alanine (328.02 mg, 1.52 mmol) were dissolved in N,N-dimethylformamide (3 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (721.01 mg, 1.90 mmol) and N,N-diisopropylethylamine (490.14 mg, 3.79 mmol, 660.57 μL) were added sequentially, and the mixture was stirred at 0°C for 0.5 hour.

[0885] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (20 g silica gel column, methanol / dichloromethane eluent, 0-6% gradient, 35 mL / min) to yield 650 mg of crude product, methyl (2S,3R)-3-(4-difluoromethylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[0886] MS (ESI) M / Z: 444.3 [M+H] + .

[0887] Step D: Methyl (2S,3R)-3-(4-difluoromethylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate (650 mg, 1.47 mmol) was dissolved in acetonitrile (15 mL) and water (0.1 mL) at 0°C, and lithium bromide (1.02 g, 11.73 mmol) and triethylamine (444.98 mg, 4.40 mmol, 612.07 μL) were added sequentially and stirred at 25°C for 12 hours.

[0888] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was separated by preparative HPLC (column: C18 150×30 mm; mobile phase: [water (ammonia + ammonium bicarbonate)-acetonitrile]; gradient: 5% to 35% phase B over 7 minutes) to yield 230 mg of (2S,3R)-3-(4-difluoromethylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid.

[0889] MS (ESI) M / Z: 430.2 [M+H] + .

[0890] Step E: (2S,3R)-3-(4-difluoromethylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid (110 mg, 256.16 μmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (112.96 mg, 3 07.40 μmol) was dissolved in N,N-dimethylformamide (2 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (146.10 mg, 384.24 μmol) and N,N-diisopropylethylamine (99.32 mg, 768.49 μmol, 133.85 μL) were added sequentially at 0°C, and the mixture was stirred at 0°C for 0.5 h.

[0891] After LC-MS / MS monitoring showed that the raw materials had reacted completely, the reaction solution was separated and purified by preparative HPLC to obtain 81.0 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethylphenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide.

[0892] MS (ESI) M / Z: 607.2 [M+H] + .

[0893] 1 H NMR (400MHz, CD3OD): δ7.44-7.54(m,4H),6.58-6.88(m,1H),5.42-5.51(m,1H),5.14(d, J=3.9Hz,1H),4.68(dd,J=8.6,5.1Hz,1H),4.60(d,J=4.1Hz,1H),4.44(q,J=7.1Hz,1H),3 .69(t,J=4.6Hz,4H),3.14(d,J=5.0Hz,1H),2.93-3.08(m,2H),2.89(d,J=5.0Hz,1H),2.4 4-2.57(m,5H),2.24-2.37(m,5H),1.82-1.91(m,2H),1.42(s,3H),1.29(d,J=7.2Hz,3H).

[0894] Example 29:

[0895] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide

[0896] Reaction route:

[0897] Steps:

[0898] Step A: Dissolve (S)-N-benzyloxycarbonylaziridine-2-carboxylic acid methyl ester (5 g, 21.26 mmol) and cyclopropanol (2.05 g, 35.28 mmol) in dichloromethane (30 mL), add boron trifluoride etherate (603.34 mg, 4.25 mmol, 522.83 μL) and stir at 0°C for 2 hours.

[0899] After complete reaction of the starting materials as monitored by thin-layer chromatography and liquid chromatography-mass spectrometry, the reaction mixture was quenched with saturated aqueous sodium bicarbonate (4 mL) at 0°C and extracted with dichloromethane (40 mL x 2). The combined organic phases were washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: petroleum ether / ethyl acetate, 3 / 1 gradient) to yield 4.5 g of N-benzyloxycarbonyl-O-cyclopropyl-L-serine methyl ester.

[0900] MS (ESI) M / Z: 293.7 [M+H] + .

[0901] 1 H NMR (400MHz, DMSO-d6): δ7.78-7.71(m,1H)7.41-7.27(m,5H)5.04(s,2H)4.35-4.27(m,1H)3.72-3.57(m,5H)3.32-3.25(m,1H)0.50-0.36(m,4H).

[0902] Step B: Dissolve N-benzyloxycarbonyl-O-cyclopropyl-L-serine methyl ester (4.5 g, 15.34 mmol) in methanol (50 mL), add 10% wet palladium carbon (1.64 g), and stir under hydrogen atmosphere (15 psi) at 25°C for 4 hours.

[0903] After thin layer chromatography monitoring indicated that the reaction of the starting material was complete, the reaction solution was filtered and concentrated under reduced pressure to obtain 2.4 g of O-cyclopropyl-L-serine methyl ester without purification.

[0904] MS (ESI) M / Z: 160.1 [M+H] + .

[0905] 1 H NMR (400MHz, DMSO-d6): δ3.62(s,3H)3.60-3.47(m,3H)3.29-3.24(m,1H)1.75(br s,2H)0.44-0.38(m,4H).

[0906] Step C: 2-Morpholineacetic acid (2.1 g, 14.47 mmol) was dissolved in N,N-dimethylformamide (23 mL), and 1-hydroxybenzotriazole (2.15 g, 15.91 mmol) and benzotriazole-N,N,N,N-tetramethyluronium hexafluorophosphate (6.04 g, 15.91 mmol) were added. O-cyclopropyl-L-serine methyl ester (2.30 g, 14.47 mmol) and N,N-diisopropylethylamine (2.80 g, 21.70 mmol, 3.78 mL) were added at 0° C. The reaction mixture was stirred at 20° C. for 30 minutes.

[0907] After thin-layer chromatography monitoring indicated complete reaction of the starting material, the mixture was diluted with dichloromethane (10 mL) and concentrated under reduced pressure. It was then diluted with water (40 mL) and extracted with ethyl acetate (50 mL x 2). The organic phases were combined, washed with saturated brine (80 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: dichloromethane / methanol, 20 / 1 gradient) to yield 3.1 g of O-cyclopropyl-N-(2-morpholinoacetyl)-L-serine methyl ester.

[0908] MS (ESI) M / Z: 287.1 [M+H] + .

[0909] 1 H NMR (400MHz, DMSO-d6): δ7.95(s,1H)4.56(ddd,J=4.0,4.9,8.5Hz,1H)3.81(dd,J=5.2,10.0Hz,1H)3.68(dd,J=3. 9,10.0Hz,1H)3.64(s,3H)3.59(t,J=4.6Hz,4H)3.32-3.27(m,1H)3.04-2.92(m,2H)2.48-2.40(m,4H)0.43(s,4H).

[0910] Step D: Dissolve O-cyclopropyl-N-(2-morpholinoacetyl)-L-serine methyl ester (3.1 g, 10.83 mmol) in tetrahydrofuran (30 mL) and water (10 mL), add lithium hydroxide monohydrate (908.67 mg, 21.65 mmol), and stir at 20°C for 1 hour.

[0911] After LC-MS monitoring showed that the reaction of the starting material was complete, the reaction solution was filtered and concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (eluent: dichloromethane / methanol, 20 / 1 gradient) to obtain 2.57 g of O-cyclopropyl-N-(2-morpholinoacetyl)-L-serine.

[0912] MS (ESI) M / Z: 273.1 [M+H] + .

[0913] 1 H NMR (400MHz, DMSO-d6): δ7.68 (d, J=7.7Hz, 1H) 4.08 (td, J=3.9, 7.7Hz, 1H) 3.76-3.69 (m, 1H) 3.65 (dd, J= 3.5, 9.5Hz, 1H) 3.63-3.53 (m, 4H) 3.27-3.20 (m, 1H) 2.90 (d, J = 2.0Hz, 2H) 2.47-2.36 (m, 4H) 0.44-0.30 (m, 4H).

[0914] Step E: O-cyclopropyl-N-(2-morpholinoacetyl)-L-serine (74 mg, 271.76 μmol) and methyl (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate (70.99 mg, 271.76 μmol) were dissolved in N,N-dimethylformamide (1 mL) at 0°C. 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (206.66 mg, 543.52 μmol) was added, followed by the slow dropwise addition of N,N-diisopropylethylamine (175.62 mg, 1.36 mmol, 236.68 μL). The reaction mixture was stirred at 0°C for 30 minutes.

[0915] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (eluent: dichloromethane / methanol, 20 / 1 gradient) to yield 115 mg of methyl (2S,3R)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate.

[0916] MS (ESI) M / Z: 516.2 [M+H] + .

[0917] Step F: Methyl (2S,3R)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionamido)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate (115 mg, 223.08 μmol) was dissolved in acetonitrile (2 mL) and water (0.2 mL), lithium bromide (96.87 mg, 1.12 mmol) and N,N-diisopropylethylamine (86.50 mg, 669.25 μmol, 116.57 μL) were added, and the mixture was stirred at 20 °C for 12 hours.

[0918] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was purified by preparative HPLC (separation column: Boston Green ODS 150 × 30 mm × 5 μm; mobile phase: [water (formic acid)-acetonitrile]; gradient: 15% to 45% phase B over 6 minutes) to yield 22 mg of (2S,3R)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid.

[0919] MS (ESI) M / Z: 502.1 [M+H] + .

[0920] Step G: (2S,3R)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid (22 mg, 43.87 μmol) was dissolved in N,N-dimethylformamide (1 mL) at 0°C, and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2- To the mixture were added 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (33.36 mg, 87.74 μmol), 2-(2-((2-methyl-oxiran-2-yl)propan-1-one)-p-toluenesulfonate (19.34 mg, 52.64 μmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (33.36 mg, 87.74 μmol), followed by the slow dropwise addition of N,N-diisopropylethylamine (28.35 mg, 219.35 μmol, 38.21 μL). The reaction mixture was stirred at 0°C for 30 minutes.

[0921] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The residue was separated and purified by preparative HPLC to yield 12.79 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-2-((S)-3-cyclopropyloxy-2-(2-morpholinoacetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide.

[0922] MS (ESI) M / Z: 679.4 [M+H] + .

[0923] 1H NMR (400MHz, DMSO-d6): δ8.04(br d,J=7.3Hz,1H)7.91-7.72(m,2H)7.38-7.34(m,2H)7.20-6.99(m,3H)5.71(d,J=4.5Hz,1H)5.39(s,1H)4.97(t,J=4.2Hz,1H)4.59(br d,J=6.1Hz,1H)4.45(br dd,J=3.5,9.0Hz,2H)3.61-3.49(m,6H)3.29-3.24(m,1H)3.14(d,J=5.0Hz,1H)2.97(br d,J=5.1Hz,3H)2.45-2.32(m,5H)2.30-2.19(m,5H)1.79(t,J=7.4Hz,2H)1.36(s,3H)0.48-0.36(m,4H).

[0924] Example 30:

[0925] (2S,3R)-2-((S)-2-(2-(2-oxa-7-azaspiro[3.5]non-7-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide

[0926] Reaction route:

[0927] Steps:

[0928] Step A: Dissolve 2-oxa-7-azaspiro[3.5]nonane (300 mg, 2.36 mmol) and benzyl 2-bromoacetate (540.33 mg, 2.36 mmol, 370.09 μL) in acetonitrile (5 mL), add potassium carbonate (977.99 mg, 7.08 mmol), and stir at 20°C for 3.5 hours.

[0929] After LC / MS monitoring indicated complete reaction of the starting material, the reaction mixture was diluted with water (10 mL) and extracted with ethyl acetate (10 mL x 2). The combined organic phases were washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified without further purification to yield 600 mg of benzyl 2-(2-oxa-7-azaspiro[3.5]nonan-7-yl)acetate.

[0930] MS (ESI) M / Z: 276.1 [M+H] + .

[0931] 1 H NMR (400MHz, CDCl3): δ7.40-7.31(m,5H)5.16(s,2H)4.40(s,4H)3.24(s,2H)2.48(br t,J=4.9Hz,4H)1.91(t,J=5.6Hz,4H).

[0932] Step B: Dissolve benzyl 2-(2-oxa-7-azaspiro[3.5]nonan-7-yl)acetate (600.00 mg, 2.18 mmol) in methanol (12 mL), add 10% wet palladium on carbon (46.3 mg), and stir under a hydrogen atmosphere (15 psi) at 25°C for 12 hours.

[0933] After TLC monitoring showed that the reaction of the starting material was complete, the reaction solution was filtered and concentrated under reduced pressure. The residue was purified without further purification to obtain 400 mg of 2-(2-oxa-7-azaspiro[3.5]non-7-yl)acetic acid.

[0934] MS (ESI) M / Z: 186.1 [M+H] + .

[0935] 1 H NMR (400MHz, DMSO-d6): δ4.28(s,4H)3.14(s,2H)2.68(br s,4H)1.86(br t,J=5.6Hz,4H).

[0936] Step C: 2-(2-oxa-7-azaspiro[3.5]nonan-7-yl)acetic acid (400 mg, 2.16 mmol) was dissolved in N,N-dimethylformamide (2 mL) and dichloromethane (4 mL), and L-alanine benzyl ester hydrochloride (558.93 mg, 2.59 mmol), 4-(4,6-dimethoxytriazin-2-yl)-4-methylmorpholine hydrochloride (1.27 g, 4.32 mmol), and N-methylmorpholine (458.72 mg, 4.54 mmol, 498.61 μL) were added at 0° C. The reaction mixture was then stirred at 20° C. for 4 hours.

[0937] After monitoring by LC-MS / MS and TLC indicated complete reaction of the starting material, the mixture was diluted with dichloromethane (10 mL) and concentrated under reduced pressure. The mixture was then diluted with water (10 mL) and extracted with ethyl acetate (10 mL x 2). The organic phases were combined, washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: petroleum ether / ethyl acetate, 0 / 1 gradient) to yield 470 mg of N-(2-(2-oxa-7-azaspiro[3.5]nonan-7-yl)acetyl)-L-alanine benzyl ester.

[0938] MS (ESI) M / Z: 347.1 [M+H] + .

[0939] 1 H NMR(400MHz,DMSO-d6):δ8.02(br d,J=7.5Hz,1H)7.40-7.33(m,5H)5.12(d,J=2.1Hz,2H)4.39(quin,J=7.3Hz,1H)4.25(s,4H)2.96-2.88(m,1H)2.85-2.72(m,1H)2.30(br s,4H)1.74(br t,J=5.5Hz,4H)1.32(d,J=7.3Hz,3H).

[0940] Step D: Dissolve N-(2-(2-oxa-7-azaspiro[3.5]nonan-7-yl)acetyl)-L-alanine benzyl ester (470 mg, 1.36 mmol) in methanol (8 mL), add 10% wet palladium on carbon (28.8 mg), and stir under a hydrogen atmosphere (15 psi) at 25°C for 12 hours.

[0941] After TLC monitoring indicated that the reaction of the starting material was complete, the reaction solution was filtered and concentrated under reduced pressure. The residue was purified without further purification to obtain 330 mg of N-(2-(2-oxa-7-azaspiro[3.5]nonan-7-yl)acetyl)-L-alanine.

[0942] MS (ESI) M / Z: 257.1 [M+H] + .

[0943] 1 H NMR (400MHz, DMSO-d6): δ7.87(d,J=7.5Hz,1H)4.26(s,4H)4.24-4.16(m,1H)2.98-2.88(m,1H)2.87-2.71(m,1H)2.43-2.23(m,4H)1.79(br t,J=5.4Hz,4H)1.29-1.25(m,3H).

[0944] Step E: N-(2-(2-oxa-7-azaspiro[3.5]nonan-7-yl)acetyl)-L-alanine (75 mg, 292.63 μmol) and methyl (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate (76.44 mg, 292.63 μmol) were dissolved in N,N-dimethylformamide (2 mL) at 0°C. 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (333.80 mg, 877.89 μmol) was added, followed by the slow dropwise addition of N,N-diisopropylethylamine (189.10 mg, 1.46 mmol, 254.85 μL). The reaction mixture was stirred at 0°C for 30 minutes.

[0945] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (eluent: dichloromethane / methanol, 20 / 1 gradient) to yield 140 mg of methyl (2S,3R)-2-((S)-2-(2-(2-oxa-7-azaspiro[3.5]nonan-7-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate.

[0946] MS (ESI) M / Z: 500.2 [M+H] + .

[0947] Step F: Methyl (2S,3R)-2-((S)-2-(2-(2-oxa-7-azaspiro[3.5]nonan-7-yl)acetamido)propionamido)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate (140 mg, 280.28 μmol) was dissolved in acetonitrile (2 mL) and water (0.2 mL), lithium bromide (121.70 mg, 1.40 mmol) and N,N-diisopropylethylamine (108.67 mg, 840.83 μmol, 146.46 μL) were added, and the mixture was stirred at 20 °C for 12 hours.

[0948] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was purified by preparative HPLC (separation column: 2-Phenomenex Gemini C18 75×40mm×3μm; mobile phase: [water (ammonia + ammonium bicarbonate)-acetonitrile]; gradient: 5% to 35% phase B over 8 minutes) to yield 60 mg of (2S,3R)-2-((S)-2-(2-(2-oxa-7-azaspiro[3.5]nonan-7-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid.

[0949] MS (ESI) M / Z: 486.2 [M+H] +.

[0950] Step G: (2S,3R)-2-((S)-2-(2-(2-oxa-7-azaspiro[3.5]nonan-7-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid (57 mg, 117.41 μmol) was dissolved in N,N-dimethylformamide (1.5 mL) at 0°C, and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-( (R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (51.77 mg, 140.89 μmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (89.29 mg, 234.82 μmol) were added, followed by the slow dropwise addition of N,N-diisopropylethylamine (75.87 mg, 587.05 μmol, 102.25 μL). The reaction mixture was stirred at 0°C for 30 minutes.

[0951] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The residue was purified by preparative HPLC to yield 27.90 mg of (2S,3R)-2-((S)-2-(2-(2-oxa-7-azaspiro[3.5]non-7-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide.

[0952] MS (ESI) M / Z: 663.2 [M+H] + .

[0953] 1H NMR (400MHz, DMSO-d6): δ7.99(d,J=7.7Hz,1H)7.92(d,J=9.1Hz,1H)7.72(d,J=7.6Hz,1H)7.38-7.32(m,2H)7.06-7.00(m,2H)7.18(s,1H)5.68(br s,1H)5.38(s,1H)4.96(d,J=3.6Hz,1H)4.61-4.53(m,1H)4.42(dd,J=3.7 ,8.9Hz,1H)4.33(t,J=7.3Hz,1H)4.24(s,4H)3.18-3.07(m,1H)2.95(d,J =5.1Hz,1H)2.86(d,J=15.6Hz,1H)2.76(d,J=15.5Hz,1H)2.44-2.35(m,1 H)2.30-2.13(m,9H)1.82-1.67(m,6H)1.35(s,3H)1.13(d,J=6.9Hz,3H).

[0954] Example 31:

[0955] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxy-3-fluorophenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propionamide

[0956] Reaction route:

[0957] Steps:

[0958] Step A: Dissolve (R)-2-(N-(N'-Benzylprolyl)amino)diphenylmethanone-nickel(II)-glycine complex (500 mg, 1.00 mmol) and 4-difluoromethoxy-2-fluorobenzaldehyde (572.42 mg, 3.01 mmol) in methanol (10 mL). Add sodium methoxide in methanol (5 M, 1.124 mL) and stir at 20°C for 12 hours. Add dilute hydrochloric acid (2 M, 10.00 mL) to the reaction mixture and stir at 50°C for 2 hours.

[0959] After LC-MS monitoring indicated complete reaction of the starting material, the reaction mixture was concentrated under reduced pressure to remove the methanol and filtered. The reaction solution was then extracted with dichloromethane (20 mL x 5). The organic phase was discarded, and the aqueous phase was filtered and concentrated under reduced pressure. Without further purification, 800 mg of crude (2S,3R)-2-amino-3-(4-difluoromethoxy-3-fluorophenyl)-3-hydroxypropionic acid hydrochloride was obtained.

[0960] MS (ESI) M / Z: 266.0 [M+H] + .

[0961] Step B: Dissolve (2S,3R)-2-amino-3-(4-difluoromethoxy-3-fluorophenyl)-3-hydroxypropanoic acid hydrochloride (800 mg, 3.02 mmol) in methanol (16 mL), add thionyl chloride (1.44 g, 12.07 mmol, 876.44 μL), and stir at 50 °C for 12 hours.

[0962] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure, the residue was dissolved in water (10 mL), the pH was adjusted to 9 with aqueous ammonia, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to yield 240 mg of crude (2S,3R)-2-amino-3-(4-difluoromethoxy-3-fluorophenyl)-3-hydroxypropionic acid methyl ester.

[0963] MS (ESI) M / Z: 280.1 [M+H] + .

[0964] Step C: Methyl (2S,3R)-2-amino-3-(4-difluoromethoxy-3-fluorophenyl)-3-hydroxypropanoate (240 mg, 859.56 μmol) and (2-morpholinoacetyl)-L-alanine (185.87 mg, 859.56 μmol) were dissolved in tetrahydrofuran (5 mL), and 3-(diethoxyphosphoryloxy)-1,2,3-benzotriazine-4-one (282.92 mg, 945.52 μmol) and sodium carbonate (182.21 mg, 1.72 mmol) were added. The mixture was stirred at 20 °C for 12 hours.

[0965] After LC-MS monitoring indicated complete reaction of the starting material, the mixture was diluted with water (10 mL) and extracted with dichloromethane (20 mL x 2). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (eluent: dichloromethane / methanol, 10 / 1 gradient) to yield 500 mg of methyl (2S,3R)-3-(4-difluoromethoxy-3-fluorophenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[0966] MS (ESI) M / Z: 478.2 [M+H] + .

[0967] Step D: Methyl (2S,3R)-3-(4-difluoromethoxy-3-fluorophenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate (500 mg, 1.05 mmol) was dissolved in acetonitrile (5 mL) and water (0.5 mL), and lithium bromide (454.75 mg, 5.24 mmol) and triethylamine (317.92 mg, 3.14 mmol, 437.30 μL) were added at 0°C, and then stirred at 25°C for 12 hours.

[0968] After LC-MS / MS monitoring showed that the starting material had reacted completely, the mixture was concentrated under reduced pressure, the excess acetonitrile was spun off, and the residue was separated by preparative high performance liquid chromatography (separation column: C18 150×30 mm; mobile phase: [water (ammonia water + ammonium bicarbonate)-acetonitrile]; gradient: phase B from 10% to 40% in 7 minutes) to give 60 mg of (2S,3R)-3-(4-difluoromethoxy-3-fluorophenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid.

[0969] MS (ESI) M / Z: 464.1 [M+H] + .

[0970] 1 H NMR (400MHz, CD3OD): δ7.25-7.33(m,1H),7.14-7.23(m,2H),6.79(s,1H),5.23(d,J=3.01Hz,1H),4.39- 4.51(m,2H),3.69(t,J=4.52Hz,4H),3.01(d,J=5.27Hz,2H),2.39-2.56(m,4H),1.30(d,J=7.03Hz,3H).

[0971] Step E: (2S,3R)-3-(4-difluoromethoxy-3-fluorophenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionic acid (60 mg, 129.48 μmol) was dissolved in N,N-dimethylformamide (1.5 mL), and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methylepoxy)-1-ol was added at 0°C. In the mixture, 1,2-dimethylthiazolylpyrazol-2-yl)propan-1-one p-toluenesulfonate (57.09 mg, 155.37 μmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (73.85 mg, 194.21 μmol) and N,N-diisopropylethylamine (33.47 mg, 258.95 μmol, 45.10 μL) were stirred at 0°C for 0.5 hours.

[0972] After LC-MS monitoring indicated complete reaction of the starting materials, the mixture was concentrated under reduced pressure. The residue was separated and purified by preparative HPLC to yield 40 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxy-3-fluorophenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide.

[0973] MS (ESI) M / Z: 641.3 [M+H] + .

[0974] 1 H NMR (400MHz, CD3OD): δ7.28(d,J=11.80Hz,1H),7.17-7.22(m,2H),6.62-7.02(m,1H),5.45(s,1H),5.05(d,J=4.27Hz,1H),4.65-4.71(m,1H ),4.56(d,J=4.27Hz,1H),4.43(d,J=7.03Hz,1H),3.65-3.74(m,4H),3.18(d,J=5.02Hz,1H),2.89-3.07(m,3H),2.45-2.54(m,5H),2.28(br t,J=7.65Hz,5H),1.86(t,J=7.40Hz,2H),1.43(s,3H),1.30(d,J=7.03Hz,3H).

[0975] Example 32:

[0976] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxy-2-fluorophenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide

[0977] Reaction route:

[0978] Steps:

[0979] Step A: Dissolve (R)-2-(N-(N'-Benzylprolyl)amino)diphenylmethanone-nickel(II)-glycine complex (700 mg, 1.00 mmol) and 4-difluoromethoxy-2-fluorobenzaldehyde (801.39 mg, 3.01 mmol) in methanol (15 mL). Add sodium methoxide in methanol (5 M, 1.124 mL) and stir at 20°C for 12 hours. Add dilute hydrochloric acid (2 M, 15.00 mL) to the reaction mixture and stir at 50°C for 2 hours.

[0980] After LC-MS monitoring indicated complete reaction of the starting material, the reaction mixture was concentrated under reduced pressure to remove the methanol and filtered. The reaction solution was then extracted with dichloromethane (20 mL x 5). The organic phase was discarded, and the aqueous phase was filtered and concentrated under reduced pressure. No purification was required to obtain 1.06 g of crude (2S,3R)-2-amino-3-(4-difluoromethoxy-2-fluorophenyl)-3-hydroxypropanoic acid hydrochloride.

[0981] MS (ESI) M / Z: 266.1 [M+H] + .

[0982] Step B: Dissolve (2S,3R)-2-amino-3-(4-difluoromethoxy-2-fluorophenyl)-3-hydroxypropanoic acid hydrochloride (1.06 g, 1.00 mmol) in methanol (20 mL), add thionyl chloride (1.90 g, 15.99 mmol, 1.16 mL) and stir at 50 °C for 12 hours.

[0983] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was concentrated under reduced pressure, the residue was dissolved in water (10 mL), the pH was adjusted to 9 with aqueous ammonia, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to yield 640 mg of crude (2S,3R)-2-amino-3-(4-difluoromethoxy-2-fluorophenyl)-3-hydroxypropionic acid methyl ester.

[0984] MS (ESI) M / Z: 280.2 [M+H] + .

[0985] Step C: Methyl (2S,3R)-2-amino-3-(4-difluoromethoxy-2-fluorophenyl)-3-hydroxypropanoate (250 mg, 895.38 μmol) was dissolved in N,N-dimethylformamide (5 mL), and (2-morpholinoacetyl)-L-alanine (193.61 mg, 895.38 μmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (510.67 mg, 1.34 mmol) and N,N-diisopropylethylamine (347.16 mg, 2.69 mmol, 467.87 μL) were added at 0°C and stirred at 0°C for 0.5 h.

[0986] After LC-MS monitoring indicated complete reaction of the starting materials, the mixture was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (eluent: petroleum ether / ethyl acetate, 0 / 1 gradient) to yield 170 mg of crude product, methyl (2S,3R)-3-(4-difluoromethoxy-2-fluorophenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[0987] MS (ESI) M / Z: 477.9 [M+H] + .

[0988] Step D: Methyl (2S,3R)-3-(4-difluoromethoxy-2-fluorophenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate (170 mg, 356.07 μmol) was dissolved in acetonitrile (2 mL) and water (0.2 mL), and lithium bromide (154.62 mg, 1.78 mmol) and triethylamine (108.09 mg, 1.07 mmol, 148.68 μL) were added at 0°C, and then stirred at 20°C for 12 hours.

[0989] After LC-MS / MS monitoring showed that the starting material had reacted completely, the mixture was concentrated under reduced pressure, the excess acetonitrile was spun off, the mixture was diluted with water, and the residue was separated by preparative high performance liquid chromatography (separation column: C18 150×30 mm; mobile phase: [water (ammonia water + ammonium bicarbonate)-acetonitrile]; gradient: phase B from 10% to 40% in 7 minutes) to give 80 mg of (2S,3R)-3-(4-difluoromethoxy-2-fluorophenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid.

[0990] MS (ESI) M / Z: 464.2 [M+H] + .

[0991] Step E: (2S,3R)-3-(4-difluoromethoxy-2-fluorophenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionic acid (70 mg, 151.06 μmol) was dissolved in N,N-dimethylformamide (1.5 mL), and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methylepoxy)-1-ol was added at 0°C. In the mixture, 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (86.15 mg, 226.58 μmol) and N,N-diisopropylethylamine (39.05 mg, 302.11 μmol, 52.62 μL) were stirred at 0°C for 0.5 hours.

[0992] After LC-MS monitoring indicated complete reaction of the starting materials, the mixture was concentrated under reduced pressure. The residue was separated and purified by preparative HPLC to yield 20 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxy-2-fluorophenyl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide.

[0993] MS (ESI) M / Z: 641.4 [M+H] + .

[0994] 1 H NMR (400MHz, CD3OD): δ7.56 (t, J = 8.41Hz, 1H), 6.65-7.07 (m, 3H), 5.46 (s, 1H), 5.39 (d, J = 4. 27Hz,1H),4.69(dd,J=8.41,5.14Hz,1H),4.64(d,J=4.27Hz,1H),4.42(d,J=7.03Hz,1H),3.7 0(t,J=4.52Hz,4H),3.18(d,J=5.27Hz,1H),2.93-3.08(m,2H),2.91(d,J=5.02Hz,1H),2.45- 2.57(m,5H),2.24-2.37(m,5H),1.86(t,J=7.40Hz,2H),1.43(s,3H),1.30(d,J=7.03Hz,3H).

[0995] Example 33:

[0996] (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(6-difluoromethoxypyridin-3-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide

[0997] Reaction route:

[0998] Steps:

[0999] Step A: Under nitrogen protection, (R)-2-(N-(N'-benzylprolyl)amino)diphenylmethanone-nickel(II)-glycine complex (500 mg, 1.00 mmol) and 6-difluoromethoxynicotinaldehyde (521.23 mg, 3.01 mmol) were dissolved in methanol (7 mL). A methanol solution of sodium methoxide (5 M, 1.00 mL) was added dropwise at 25°C. After stirring at 25°C for 12 hours, dilute hydrochloric acid (2 M, 10.00 mL) was added, and the mixture was stirred at 50°C for 1 hour.

[1000] After LC-MS monitoring showed that the raw materials were completely reacted, the reaction solution was concentrated under reduced pressure to remove methanol, and a white solid was generated. The solid was filtered, and the filtrate was extracted with dichloromethane (20 mL×5). The aqueous phase was concentrated under reduced pressure to obtain 690 mg of crude (2S,3R)-2-amino-3-(6-difluoromethoxypyridin-3-yl)-3-hydroxypropionic acid hydrochloride.

[1001] MS (ESI) M / Z: 249.0 [M+H] + .

[1002] 1 H NMR (400MHz, D2O): δ8.24(d,J=1.5Hz,1H),7.92-8.02(m,1H),7.13-7.52(m,1H)7.10(d,J=8.8Hz,1H),5.32-5.39(m,1H),4.10-4.20(m,1H).

[1003] Step B: At 0 ° C, (2S, 3R) -2-amino-3- (6-difluoromethoxypyridin-3-yl) -3-hydroxypropanoic acid hydrochloride (690 mg, 2.78 mmol) was dissolved in methanol (10 mL), thionyl chloride (992.28 mg, 8.34 mmol, 605.79 μ L) was added dropwise, and then stirred at 50 ° C for 12 hours.

[1004] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure, the residue was dissolved in water (10 mL), the pH was adjusted to 9 with aqueous ammonia, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to yield 180 mg of methyl (2S,3R)-2-amino-3-(6-difluoromethoxypyridin-3-yl)-3-hydroxypropanoate.

[1005] MS (ESI) M / Z: 263.0 [M+H] + .

[1006] 1 H NMR (400MHz, CD3OD): δ8.12-8.27(m,1H),7.81-7.95(m,1H),7.32-7.75(m, 1H),6.89-7.05(m,1H),4.96-5.11(m,1H),3.60-3.76(m,1H),3.35(s,3H).

[1007] Step C: Methyl (2S,3R)-2-amino-3-(6-difluoromethoxypyridin-3-yl)-3-hydroxypropanoate (180 mg, 686.47 μmol) and (2-morpholinoacetyl)-L-alanine (163.28 mg, 755.12 μmol) were dissolved in tetrahydrofuran (2 mL), and 1,2,3-benzotriazine-4-one (225.95 mg, 755.12 μmol) and sodium carbonate (145.52 mg, 1.37 mmol) were added in sequence. The mixture was stirred at 25 °C for 12 hours.

[1008] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was diluted with water (4 mL) and extracted with dichloromethane (4 mL x 3). The organic phases were combined and concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (20 g silica gel column, methanol / dichloromethane eluent, 0-3% gradient, 35 mL / min) to afford 150 mg of methyl (2S,3R)-3-(6-difluoromethoxypyridin-3-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[1009] MS (ESI) M / Z: 461.1 [M+H] + .

[1010] 1H NMR (400MHz, CD3OD): δ8.22(d,J=2.3Hz,1H),7.89(dd,J=8.5,2.3Hz,1H),7.30-7.71(m,1H),6.89(d,J=8.5Hz,1H),5.35(d,J=2.8Hz,1H),4 .77(d,J=2.8Hz,1H),4.43-4.52(m,1H),3.75-3.82(m,3H),3.64-3.74(m,4H),2.90-3.10(m,2H),2.39-2.56(m,4H),1.30(d,J=7.0Hz,3H).

[1011] Step D: Methyl (2S,3R)-3-(6-difluoromethoxypyridin-3-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate (150 mg, 325.78 μmol) was dissolved in acetonitrile (5 mL) and water (0.5 mL) at 0°C, and lithium bromide (282.93 mg, 3.26 mmol) and N,N-diisopropylethylamine (126.32 mg, 977.35 μmol) were added sequentially, and the mixture was stirred at 25°C for 12 hours.

[1012] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was separated by preparative HPLC (column: Welch Xtimate C18 150×30mm×5μm; mobile phase: [water(ammonium bicarbonate)-acetonitrile]; gradient: 5% to 35% phase B over 7 minutes) to yield 110 mg of (2S,3R)-3-(6-difluoromethoxypyridin-3-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid.

[1013] MS (ESI) M / Z: 447.2 [M+H] + .

[1014] 1 H NMR (400MHz, CD3OD): δ8.18(d,J=2.0Hz,1H),7.87(dd,J=8.4,2.4Hz,1H),7.27-7.71(m,1H),6.86(d,J=8.3Hz,1H),5. 26(d,J=3.3Hz,1H),4.39-4.52(m,2H),3.67-3.72(m,4H),2.97-3.07(m,2H),2.44-2.52(m,4H),1.31(d,J=7.0Hz,3H).

[1015] Step E: (2S,3R)-3-(6-difluoromethoxypyridin-3-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid (110 mg, 246.41 μmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (10 8.66 mg, 295.70 μmol) was dissolved in N,N-dimethylformamide (2 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (187.39 mg, 492.83 μmol) and N,N-diisopropylethylamine (159.24 mg, 1.23 mmol) were added sequentially at 0°C, and the mixture was stirred at 0°C for 1 hour.

[1016] After LC-MS / MS monitoring showed that the raw materials were completely reacted, the reaction solution was separated and purified by preparative HPLC to obtain 90.15 mg of (2S,3R)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(6-difluoromethoxypyridin-3-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide.

[1017] MS (ESI) M / Z: 624.3 [M+H] + .

[1018] 1 H NMR (400MHz, CD3OD): δ8.16(d,J=2.4Hz,1H),7.88(dd,J=8.5,2.4Hz,1H),7.30-7.70(m,1H),6.90(d ,J=8.6Hz,1H),5.45(s,1H),5.08(d,J=4.6Hz,1H),4.67(dd,J=8.7,5.0Hz,1H),4.59(d,J=4.6Hz,1H ),4.45(q,J=7.0Hz,1H),3.67-3.73(m,4H),3.16(d,J=5.0Hz,1H),2.96-3.09(m,2H),2.91(d,J=5.0 Hz,1H),2.46-2.54(m,5H),2.25-2.32(m,5H),1.82-1.91(m,2H),1.42(s,3H),1.31(d,J=7.0Hz,3H).

[1019] Example 34:

[1020] (2S,3S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(5-difluoromethoxypyridin-2-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide

[1021] Reaction route:

[1022] Steps:

[1023] Step A: Under nitrogen, zinc powder (5.23 g, 79.98 mmol) was added to NN, dimethylformamide (180 mL), followed by iodine (509.88 mg, 2.01 mmol). After stirring at 25°C for 30 minutes, the reddish-brown color disappeared. Methyl (R)-2-(tert-butoxycarbonylamino)-3-iodopropionate (13.22 g, 40.18 mmol) was then added to the reaction mixture, which was stirred at 25°C for 1 hour. 2-Bromo-5-difluoromethoxypyridine (9 g, 40.18 mmol), tris(dibenzylideneacetone)palladium (1.84 g, 2.01 mmol), and 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (1.65 g, 4.02 mmol) were then added, and the mixture was stirred at 50°C for 12 hours.

[1024] After thin-layer chromatography monitoring indicated complete reaction of the starting material, the product was filtered through celite. Water (100 mL) was added to the filtrate, which was then extracted with ethyl acetate (100 mL x 2). The organic phases were combined, washed with saturated brine (200 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (80 g silica gel column, ethyl acetate / petroleum ether eluent, 0-15% gradient, 40 mL / min) to yield 6.82 g of methyl (S)-2-(tert-butoxycarbonylamino)-3-(5-difluoromethoxypyridin-2-yl)propanoate.

[1025] MS (ESI) M / Z: 347.2 [M+H] + .

[1026] 1H NMR (400MHz, CD3OD): δ8.35(d,J=2.26Hz,1H),7.56(dd,J=8.53,2.51Hz,1H),7.33(d,J=8.53Hz,1H),6.70-7.10(m,1H),4.57(br dd,J=8.66,5.14Hz,1H),3.65-3.74(m,3H),3.23-3.29(m,1H),3.12(dd,J=14.05,8.78Hz,1H),1.38(s,9H).

[1027] Step B: Methyl (S)-2-(tert-butoxycarbonylamino)-3-(5-difluoromethoxypyridin-2-yl)propanoate (6.82 g, 19.69 mmol) was dissolved in acetonitrile (120 mL), and di-tert-butyl dicarbonate (5.16 g, 23.63 mmol) and 4-dimethylaminopyridine (240.58 mg, 1.97 mmol) were added. The system was stirred at 70 ° C for 2 hours.

[1028] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (12 g silica gel column, ethyl acetate / petroleum ether eluent, 0-15% gradient, 40 mL / min) to afford 7.2 g of methyl (S)-2-(bis(tert-butoxycarbonyl)amino)-3-(5-difluoromethoxypyridin-2-yl)propanoate.

[1029] MS (ESI) M / Z: 447.2 [M+H] + .

[1030] 1 H NMR (400MHz, CD3OD): δ8.35(d,J=2.74Hz,1H),7.57(dd,J=8.46,2.74Hz,1H),7.29(d,J=8.46Hz,1H),6.70-7.12(m,1H),5 .41(dd,J=10.37,4.77Hz,1H),3.76(s,3H)3.57(dd,J=14.13,4.83Hz,1H),3.39(dd,J=14.07,10.49Hz,1H),1.41(s,18H).

[1031] Step C: Dissolve (S)-methyl 2-(bis(tert-butoxycarbonyl)amino)-3-(5-difluoromethoxypyridin-2-yl)propanoate (7 g, 15.68 mmol) in carbon tetrachloride (140 mL), add N-bromosuccinimide (4.19 g, 23.52 mmol), and then irradiate the reaction solution with an incandescent lamp and reflux at 90°C with stirring for 12 hours.

[1032] After LC-MS monitoring showed that the raw material reaction was complete, the reaction solution was filtered and concentrated under reduced pressure to obtain 7 g of crude product (2R)-2-(bis(tert-butoxycarbonyl)amino)-3-bromo-3-(5-difluoromethoxypyridin-2-yl)propionic acid methyl ester.

[1033] MS (ESI) M / Z: 525.1 [M+H] + .

[1034] Step D: Methyl (2R)-2-(bis(tert-butoxycarbonyl)amino)-3-bromo-3-(5-difluoromethoxypyridin-2-yl)propanoate (7 g, 13.32 mmol) was dissolved in acetone (350 mL), and silver nitrate (4.25 g, 25.02 mmol) was added. The system was stirred at 30 °C in the dark for 12 h.

[1035] After LC-MS monitoring indicated that the starting material had reacted, the reaction solution was filtered and concentrated under reduced pressure. The reaction solution was diluted with ethyl acetate (100 mL) and saturated aqueous ammonium chloride (100 mL), and the layers were separated. The organic phase was washed with saturated brine (200 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (80 g silica gel column, ethyl acetate / petroleum ether eluent, 0-35% gradient, 80 mL / min) to obtain 3.03 g of crude 3-tert-butyl-4-methyl-(4S,5S)-5-(5-difluoromethoxypyridin-2-yl)-2-oxooxazolidine-3,4-dicarboxylate.

[1036] MS (ESI) M / Z: 389.1 [M+H] + .

[1037] Step E: Dissolve 3-tert-butyl-4-methyl-(4S,5S)-5-(5-difluoromethoxypyridin-2-yl)-2-oxooxazolidine-3,4-dicarboxylate (2.65 g, 6.82 mmol) in methanol (50 mL), add cesium carbonate (222.35 mg, 682.43 μmol) and stir at 0°C for 1 hour.

[1038] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was quenched with aqueous potassium bisulfate (20 mL) and extracted with ethyl acetate (20 mL x 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (40 g silica gel column, ethyl acetate / petroleum ether eluent, 0-30% gradient, 60 mL / min) to afford 1.4 g of methyl (2S,3S)-2-(tert-butoxycarbonylamino)-3-(5-difluoromethoxypyridin-2-yl)-3-hydroxypropanoate.

[1039] MS (ESI) M / Z: 363.1 [M+H]+ .

[1040] 1 H NMR (400MHz, CD3OD): δ8.38(d,J=2.03Hz,1H),7.50-7.69(m,2H),6.74-7.12(m,1H),4.97-5.05(m,1H),4.66(br dd,J=8.88,5.42Hz,1H),3.62(s,3H),1.39(s,9H).

[1041] Step F: Methyl (2S,3S)-2-(tert-butoxycarbonylamino)-3-(5-difluoromethoxypyridin-2-yl)-3-hydroxypropanoate (600 mg, 1.66 mmol) was dissolved in dichloromethane (12 mL) and trifluoroacetic acid (6 mL) and stirred at 25°C for 0.5 hours.

[1042] After LC-MS monitoring showed that the raw materials had reacted completely, the reaction solution was concentrated under reduced pressure to obtain 400 mg of crude trifluoroacetic acid salt of (2S,3S)-2-amino-3-(5-difluoromethoxypyridin-2-yl)3-hydroxypropionic acid methyl ester.

[1043] MS (ESI) M / Z: 263.0 [M+H] + .

[1044] Step G: Methyl (2S,3S)-2-amino-3-(5-difluoromethoxypyridin-2-yl) 3-hydroxypropanoate trifluoroacetate (400 mg, 1.53 mmol) and (2-morpholinoacetyl)-L-alanine (329.86 mg, 1.53 mmol) were dissolved in N,N-dimethylformamide (8 mL), cooled to 0°C, and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (1.16 g, 3.05 mmol) and N,N-diisopropylethylamine (985.79 mg, 7.63 mmol) were added in sequence, and the mixture was stirred at 0°C for 30 minutes.

[1045] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (4 g silica gel column, methanol / dichloromethane eluent, 0-4% gradient, 20 mL / min) to yield 600 mg of crude product, methyl (2S,3S)-3-(5-difluoromethoxypyridin-2-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate.

[1046] MS (ESI) M / Z: 461.2 [M+H] + .

[1047] Step H: At 0°C, methyl (2S,3S)-3-(5-difluoromethoxypyridin-2-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoate (600 mg, 1.30 mmol) was dissolved in acetonitrile (10 mL) and water (1 mL). Lithium bromide (1.13 g, 13.03 mmol) and N,N-diisopropylethylamine (505.26 mg, 3.91 mmol) were added sequentially and stirred at 25°C for 12 hours.

[1048] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was separated by preparative HPLC (column: Welch Xtimate C18 150×30mm×5μm; mobile phase: [water(ammonium bicarbonate)-acetonitrile]; gradient: 5% to 35% phase B over 7 minutes) to yield 144 mg of (2S,3S)-3-(5-difluoromethoxypyridin-2-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propanoic acid.

[1049] MS (ESI) M / Z: 447.2 [M+H] + .

[1050] 1 H NMR (400MHz, CD3OD): δ8.33(s,1H),7.52-7.61(m,2H),6.71-7.12(m,1H),5.04(d,J=5.5Hz,1H),4 .67(d,J=5.8Hz,1H),4.41(q,J=7.2Hz,1H),3.72(t,J=4.6Hz,4H),3.05(d,J=2.5Hz,2H),2.54(br d,J=3.5Hz,4H),1.31(d,J=7.0Hz,3H).

[1051] Step I: (2S,3S)-3-(5-Difluoromethoxypyridin-2-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionamido)propanoic acid (144 mg, 322.58 μmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (142 mg, 387 μmol) were dissolved in N,N-dimethylformamide (3 mL). 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (245.31 mg, 645.16 μmol) and N,N-diisopropylethylamine (208.45 mg, 1.61 mmol) were added sequentially at 0°C, and the mixture was stirred at 0°C for 1 hour.

[1052] After LC-MS / MS monitoring showed that the raw materials had reacted completely, the reaction solution was separated and purified by preparative HPLC to obtain 80.68 mg of (2S,3S)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(5-difluoromethoxypyridin-2-yl)-3-hydroxy-2-((S)-2-(2-morpholinoacetylamino)propionylamino)propionamide.

[1053] MS (ESI) M / Z: 624.3 [M+H] + .

[1054] 1 H NMR (400MHz, CD3OD): δ8.36(d,J=2.5Hz,1H),7.56-7.61(m,1H),7.49-7.54(m,1H),6.75-7.15(m,1H),5.40(br s,1H),4.93(d,J=6.5Hz,1H),4.77(d,J=6.5Hz,1H),4.64(dd,J=8.7,4.9Hz,1H),4.38(q,J=7.3Hz,1H),3.68-3.74(m,4H),3.18-3.21(m,1H ),2.95-3.07(m,2H),2.89-2.92(m,1H),2.44-2.55(m,5H),2.19-2.32(m,5H),1.84(quin,J=7.2Hz,2H),1.43(s,3H),1.30(d,J=7.0Hz,3H).

[1055] Example 35:

[1056] (2S,3R)-2-((2S)-2-(2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide

[1057] Reaction route:

[1058] Steps:

[1059] Step A: Dissolve 3-oxa-6-azabicyclo[3.1.1]heptane hydrochloride (800 mg, 5.90 mmol) in acetonitrile (16 mL), add ethyl 2-bromoacetate (985.32 mg, 5.90 mmol, 652.96 μL) and potassium carbonate (2.45 g, 17.70 mmol), and stir at 20°C for 4 hours.

[1060] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was diluted with water (10 mL) and extracted with ethyl acetate (10 mL). The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The residue was purified without further purification to yield 1.01 g of ethyl 2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetate.

[1061] MS (ESI) M / Z: 186.1 [M+H] + .

[1062] 1 H NMR (400MHz, CDCl3): δ1.25(t,J=7.15Hz,3H)1.80(d,J=8.28Hz,1H)2.64-2.77(m ,1H)3.47(s,2H)3.63(d,J=6.27Hz,2H)3.74(d,J=11.04Hz,2H)4.11-4.21(m,4H).

[1063] Step B: Ethyl 2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetate (1 g, 5.40 mmol) was dissolved in tetrahydrofuran (15 mL) and water (5 mL), lithium hydroxide monohydrate (453.12 mg, 10.80 mmol) was added, and stirred at 25°C for 1 hour.

[1064] After monitoring by thin layer chromatography showed that the starting material had reacted completely, the mixture was concentrated under reduced pressure to remove tetrahydrofuran and then directly freeze-dried to obtain 2.32 g of crude 2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetic acid.

[1065] MS (ESI) M / Z: 158.1 [M+H] + .

[1066] 1 H NMR (400MHz, DMSO-d6): δ1.86(br d,J=9.29Hz,1H)2.78-2.92(m,1H)3.46(br s,2H)3.81-3.95(m,4H)4.18(br d,J=11.54Hz,2H).

[1067] Step C: 2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetic acid (2.9 g, 16.01 mmol, LiOH) was dissolved in dichloromethane (30 mL) and N,N-dimethylformamide (15 mL) at 0°C, and L-alanine benzyl ester hydrochloride (3.45 g, 16.01 mmol), 4-(4,6-dimethoxytriazin-2-yl)-4-methylmorpholine hydrochloride (9.44 g, 32.02 mmol) and N-methylmorpholine (3.40 g, 33.63 mmol, 3.70 mL) were added, and the reaction mixture was stirred at 20°C for 4 hours.

[1068] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was separated by flash silica gel column chromatography (eluent: dichloromethane / methanol, 10 / 1 gradient) and high performance liquid chromatography (separation column: Xtimate C18 150×40mm×10μm; mobile phase: [water(formic acid)-acetonitrile]; gradient: 5% to 35% phase B over 2 minutes) to yield 0.79 g of N-(2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetyl)-L-alanine benzyl ester.

[1069] MS (ESI) M / Z: 319.1 [M+H] + .

[1070] Step D: N-(2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetyl)-L-alanine benzyl ester (300 mg, 942.31 μmol) was dissolved in methanol (6 mL), 10% wet palladium on carbon (100.28 mg) was added, and the mixture was stirred under a hydrogen atmosphere (15 psi) at 25°C for 2 hours.

[1071] After thin-layer chromatography monitoring indicated complete reaction of the starting material, the reaction mixture was filtered, the filter cake was washed with methanol (10 mL x 2), and the filtrate was concentrated under reduced pressure. The residue was purified without further purification to yield 120 mg of crude N-(2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetyl)-L-alanine.

[1072] Step E: At 0°C, N-(2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetyl)-L-alanine (117 mg, 512.61 μmol) and methyl (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate (133.90 mg, 512.61 μmol) were dissolved in N,N-dimethylformamide (3 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (389.82 mg, 1.03 mmol) and N,N-diisopropylethylamine (331.25 mg, 2.56 mmol, 446.43 μL) were added sequentially. The mixture was stirred at 0°C for 30 minutes.

[1073] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (eluent: dichloromethane / methanol, 10 / 1 gradient) to yield 290 mg of methyl (2S,3R)-2-((2S)-2-(2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate.

[1074] MS (ESI) M / Z: 471.9 [M+H] + .

[1075] Step F: Methyl (2S,3R)-2-((2S)-2-(2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetamido)propionamido)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate (290 mg, 276.81 μmol) was dissolved in acetonitrile (2 mL) and water (0.2 mL). Lithium bromide (120.20 mg, 1.38 mmol) and N,N-diisopropylethylamine (107.32 mg, 830.42 μmol, 144.64 μL) were added and the reaction mixture was stirred at 20 °C for 12 hours.

[1076] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was separated by HPLC (separation column: Xtimate C18 150×40mm×10μm; mobile phase: [water(formic acid)-acetonitrile]; gradient: 5% to 35% phase B over 10 minutes) to yield 115 mg of (2S,3R)-2-((2S)-2-(2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid.

[1077] MS (ESI) M / Z: 458.2 [M+H] + .

[1078] 1 H NMR (400MHz, CD3OD): δ7.43 (d, J = 8.7Hz, 2H) 7.05 (d, J = 8.6Hz, 2H) 6.98-6.59 (m, 1H) 5.26 (d, J = 3.1Hz, 1H) 4.57 (d, J = 3.2Hz, 1H) 4.35 (br d,J=12.6Hz,2H)4.30(br d,J=7.0Hz,1H)4.28-4.20(m,2H)4.20-3.90(m,4H)3.19-2.98(m,1H)2.18(br d,J=8.5Hz,1H)1.29(d,J=7.3Hz,3H).

[1079] Step G: (2S,3R)-2-((2S)-2-(2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid (115 mg, 251.41 μmol) was dissolved in N,N-dimethylformamide (1.5 mL) at 0°C, and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1 To the mixture were added 2-((R)-2-methyloxiran-2-yl)propan-1-one p-toluenesulfonate (110.86 mg, 301.69 μmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (191.19 mg, 502.82 μmol), followed by the slow dropwise addition of N,N-diisopropylethylamine (162.46 mg, 1.26 mmol, 218.95 μL). The reaction mixture was stirred at 0°C for 30 minutes.

[1080] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The residue was separated and purified by preparative high-performance liquid chromatography to yield 33.14 mg of (2S,3R)-2-((2S)-2-(2-(3-oxa-6-azabicyclo[3.1.1]hept-6-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide.

[1081] MS (ESI) M / Z: 635.3 [M+H] + .

[1082] 1H NMR (400MHz, CD3OD): δ7.43(d,J=8.7Hz,2H)7.08(d,J=8.6Hz,2H)7.02-6.59(m,1H)5.46(s,1 H)5.06(d,J=4.5Hz,1H)4.73-4.65(m,1H)4.59(d,J=4.5Hz,1H)4.40(q,J=7.3Hz,1H)4.32(br d,J=12.0Hz,2H)4.03(br s,6H)3.15(d,J=5.0Hz,1H)3.02(br s,1H)2.92(d,J=5.0Hz,1H)2.54(br d,J=5.2Hz,1H)2.35-2.24(m,5H)2.16-2.05(m,1H)1.94-1.81(m,2H)1.44(s,3H)1.33(d,J=7.2Hz,3H).

[1083] Example 36:

[1084] (2S,3R)-2-((2S)-2-(2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide

[1085] Reaction route:

[1086] Steps:

[1087] Step A: Dissolve 3-oxa-8-azabicyclo[3.2.1]octane hydrochloride (1 g, 6.68 mmol) and potassium carbonate (2.77 g, 20.05 mmol) in acetonitrile (15 mL), cool to 0°C, add 2-benzyl bromoacetate (1.53 g, 6.68 mmol) to the above system, stir at 0°C for 2 hours, then heat to 60°C and stir for 12 hours.

[1088] After thin layer chromatography monitoring showed that the raw material reaction was complete, the reaction solution was diluted with water (30 mL), extracted with ethyl acetate (15 mL), and the organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain 1.67 g of 2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetate.

[1089] MS (ESI) M / Z: 262.1 [M+H] + .

[1090] 1H NMR (400MHz, DMSO-d6): δ7.27-7.42(m,5H),5.11(s,2H),3.55(d,J=10.3Hz,2H),3.39(br d,J=9.0Hz,2H),3.16(s,2H),3.08(br s,2H),1.78-1.88(m,2H),1.66-1.74(m,2H).

[1091] Step B: Benzyl 2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetate (2.15 g, 8.23 ​​mmol) was dissolved in methanol (30 mL), 10% wet palladium on carbon (875.58 mg) was added, and the system was stirred under a hydrogen atmosphere (15 psi) at 25°C for 12 hours.

[1092] After LC-MS monitoring showed that the raw material reaction was complete, the reaction solution was filtered, the filter cake was washed with methanol (20 mL), and the filtrate was concentrated under reduced pressure to obtain 1.5 g of crude 2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetic acid.

[1093] MS (ESI) M / Z: 171.7 [M+H] + .

[1094] 1 H NMR (400MHz, DMSO-d6): δ3.71 (d, J = 11.0Hz, 2H), 3.47 (br d, J = 9.8Hz, 2H), 3.35 (br s, 2H), 3.09 (s, 2H), 1.86-1.97 (m, 2H), 1.74-1.83 (m, 2H).

[1095] Step C: 2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetic acid (1.5 g, 8.76 mmol) and L-alanine benzyl ester hydrochloride (2.27 g, 10.51 mmol) were dissolved in N,N-dimethylformamide (10 mL) and dichloromethane (20 mL). 4-(4,6-dimethoxytriazin-2-yl)-4-methylmorpholine hydrochloride (5.16 g, 17.52 mmol) and N-methylmorpholine (1.92 g, 19.01 mmol) were added sequentially at 0°C and stirred at 25°C for 12 hours.

[1096] After LC-MS monitoring showed that the reaction of the raw materials was complete, the reaction solution was concentrated under reduced pressure. The mixture was diluted with water (20 mL) and extracted with ethyl acetate (10 mL x 2). The organic phases were combined, washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain a crude product, which was purified by flash silica gel chromatography (40 g silica gel column, ethyl acetate / petroleum ether eluent, 0-100% gradient, 80 mL / min) to obtain 1.67 g of N-(2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetyl)-L-alanine benzyl ester.

[1097] MS (ESI) M / Z: 332.8 [M+H] + .

[1098] 1 H NMR (400MHz, DMSO-d6): δ8.16 (br d,J=7.5Hz,1H),7.29-7.41(m,5H),5.07-5.18(m,2H),4.43(quin,J=7.3Hz,1H),3.59(dd,J=9.9,6.4Hz,2H),3.33-3.41(m,2H),2.97(br d,J=17.3Hz,2H),2.80-2.92(m,2H),1.65-1.83(m,4H),1.36(d,J=7.3Hz,3H).

[1099] Step D: N-(2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetyl)-L-alanine benzyl ester (1.67 g, 5.02 mmol) was dissolved in methanol (20 mL), 10% wet palladium on carbon (534.67 mg) was added, and the system was stirred under a hydrogen atmosphere (15 psi) at 25°C for 12 hours.

[1100] After LC-MS monitoring showed that the raw material reaction was complete, the reaction solution was filtered, the filter cake was washed with methanol (20 mL), and the filtrate was concentrated under reduced pressure to obtain 1.26 g of crude N-(2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetyl)-L-alanine.

[1101] MS (ESI) M / Z: 242.7 [M+H] + .

[1102] 1H NMR (400MHz, DMSO-d6): δ8.07(br d,J=7.6Hz,1H),4.21-4.30(m,1H),3.62(dd,J=10.2,6.0Hz,2H),3.43(br d,J=9.8Hz,3H),3.02(br s,2H),2.81-2.92(m,2H),1.68-1.87(m,4H),1.31(d,J=7.3Hz,3H).

[1103] Step E: Methyl (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate (145 mg, 555.08 μmol) and N-(2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetyl)-L-alanine (161.38 mg, 666.10 μmol) were dissolved in N,N-dimethylformamide (3 mL) at 0°C. 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (422.12 mg, 1.11 mmol) and N,N-diisopropylethylamine (358.70 mg, 2.78 mmol) were added sequentially and the mixture was stirred at 0°C for 0.5 h.

[1104] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was concentrated under reduced pressure. The residue was purified by flash silica gel column chromatography (12 g silica gel column, methanol / dichloromethane eluent, 0-3% gradient, 40 mL / min) to afford 300 mg of methyl (2S,3R)-2-((2S)-2-(2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate.

[1105] MS (ESI) M / Z: 485.9 [M+H] + .

[1106] Step F: Methyl (2S,3R)-2-((2S)-2-(2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetamido)propionamido)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate (300 mg, 617.95 μmol) was dissolved in acetonitrile (6 mL) and water (0.6 mL) at 0°C. Lithium bromide (536.69 mg, 6.18 mmol) and N,N-diisopropylethylamine (239.59 mg, 1.85 mmol) were added sequentially and the mixture was stirred at 25°C for 12 hours.

[1107] After LC-MS monitoring indicated complete reaction of the starting material, the reaction solution was concentrated under reduced pressure. The residue was separated by preparative HPLC (column: C18 150×30 mm; mobile phase: [water(formic acid)-acetonitrile]; gradient: 3% to 33% phase B over 7 minutes) to yield 144 mg of (2S,3R)-2-((2S)-2-(2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid.

[1108] MS (ESI) M / Z: 472.3 [M+H] + .

[1109] 1 H NMR (400MHz, CD3OD): δ7.42(d,J=8.5Hz,2H),7.03(d,J=8.8Hz,2H),6.58-6.97(m,1H),5.25(d,J=3.3Hz,1H), 4.55(d,J=3.3Hz,1H),4.44(q,J=6.8Hz,1H),3.66-3.77(m,2H),3.50(ddd,J=18.1,10.6,1.9Hz,2H),3.09(br s,1H),3.01(d,J=2.3Hz,3H),1.87-1.96(m,4H),1.33(d,J=7.3Hz,3H).

[1110] Step G: (2S,3R)-2-((2S)-2-(2-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid (50 mg, 86.97 μmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1- Ketone p-toluenesulfonate (38.35 mg, 104.36 μmol) was dissolved in N,N-dimethylformamide (1 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (66.13 mg, 173.93 μmol) and N,N-diisopropylethylamine (56.20 mg, 434.83 μmol) were added sequentially at 0°C, and the mixture was stirred at 0°C for 1 hour.

[1111] After LC-MS monitoring showed that the raw materials were completely reacted, the reaction solution was separated and purified by preparative HPLC to obtain 30.65 mg of (2S,3R)-2-((2S)-2-(2-(3-oxa-8-azabicyclo[3.2.1]oct-8-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide.

[1112] MS (ESI) M / Z: 649.4 [M+H] + .

[1113] 1 H NMR (400MHz, CD3OD): δ7.41(d,J=8.5Hz,2H),7.05(d,J=8.5Hz,2H),6.60-7.00(m,1H),5.45(s,1H),5.06(d,J=4.5Hz,1H),4.68(dd,J=8.5,5.0Hz,1H ),4.57(d,J=4.5Hz,1H),4.42-4.49(m,1H),3.69-3.76(m,2H),3.49(ddd,J =15.9,10.4,1.5Hz,2H),3.15(d,J=5.0Hz,1H),2.88-3.07(m,5H),2.51(br dd,J=14.2,4.4Hz,1H),2.23-2.34(m,5H),1.83-1.96(m,6H),1.42(s,3H),1.33(d,J=7.0Hz,3H).

[1114] Example 37:

[1115] (2S,3R)-2-((2S)-2-(2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide

[1116] Reaction route:

[1117] Steps:

[1118] Step A: Dissolve 6-oxa-3-azabicyclo[3.1.1]heptane hydrochloride (0.55 g, 4.06 mmol) in acetonitrile (15 mL), add benzyl 2-bromoacetate (0.93 g, 4.06 mmol) and potassium carbonate (1.68 g, 12.18 mmol), and stir at 20°C for 4 hours.

[1119] After LC-MS monitoring showed that the reaction of the starting materials was complete, the mixture was diluted with ethyl acetate (20 mL), filtered, and the filtrate was concentrated under reduced pressure to obtain 1 g of crude 2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetate, which was used directly in the next step.

[1120] MS (ESI) M / Z: 248 [M+H] + .

[1121] Step B: Dissolve benzyl 2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetate (1 g, 4.04 mmol) in methanol (10 mL), add 10% wet palladium on carbon (43 mg), and stir under a hydrogen atmosphere (15 psi) at 25°C for 1 hour.

[1122] After LC-MS monitoring showed that the reaction of the starting materials was complete, the reaction solution was filtered through a pad of celite and the filtrate was concentrated under reduced pressure to obtain 0.48 g of crude 2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetic acid, which was used directly in the next step.

[1123] MS (ESI) M / Z: 158 [M+H] + .

[1124] Step C: At 0°C, 2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetic acid (0.48 g, 3.05 mmol) was dissolved in N,N-dimethylformamide (5 mL), and L-alanine benzyl ester hydrochloride (0.66 g, 3.05 mmol) and O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (2.32 g, 6.1 mmol) and N,N-diisopropylethylamine (1.97 g, 15.25 mmol) were added and stirred at 25°C for 4 hours.

[1125] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was added to a saturated aqueous sodium bicarbonate solution (10 mL). The aqueous phase was extracted with ethyl acetate (20 mL x 2). The organic phases were combined, washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 1 / 1) to yield 0.61 g of N-(2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetyl)-L-alanine benzyl ester.

[1126] MS (ESI) M / Z: 319 [M+H] + .

[1127] Step D: Dissolve N-(2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetyl)-L-alanine benzyl ester (0.61 g, 1.92 mmol) in methanol (10 mL), add 10% wet palladium on carbon (20 mg), and stir under a hydrogen atmosphere (15 psi) at 25°C for 1 hour.

[1128] After LC-MS monitoring indicated that the reaction of the starting materials was complete, the reaction solution was filtered through a pad of celite, and the filtrate was concentrated under reduced pressure to obtain 0.4 g of crude N-(2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetyl)-L-alanine, which was used directly in the next step.

[1129] MS (ESI) M / Z: 229 [M+H] + .

[1130] Step E: Benzyl (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate (100 mg, 0.30 mmol) and N-(2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetyl)-L-alanine (82.17 mg, 0.36 mmol) were dissolved in tetrahydrofuran (5 mL). The reaction system was cooled to 0°C in an ice-water bath. 3-(diethoxyphosphoryloxy)-1,2,3-benzotriazine-4-one (98.74 mg, 0.33 mmol) and N,N-diisopropylethylamine (77.54 mg, 0.60 mmol) were added and stirring was continued at 25°C for 12 hours.

[1131] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was quenched with water (10 mL). The aqueous phase was extracted with dichloromethane (20 mL x 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: dichloromethane / methanol = 20 / 1) to yield 120 mg of benzyl (2S,3R)-2-((2S)-2-(2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate.

[1132] MS (ESI) M / Z: 548 [M+H] + .

[1133] Step F: Dissolve benzyl (2S,3R)-2-((2S)-2-(2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate (120 mg, 0.22 mmol) in methanol (10 mL), add 10% wet palladium on carbon (20 mg), and stir under a hydrogen atmosphere (15 psi) at 25°C for 1 hour.

[1134] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was filtered through a pad of celite, and the filtrate was concentrated under reduced pressure to yield 100 mg of crude (2S,3R)-2-((2S)-2-(2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid, which was used directly in the next step.

[1135] MS (ESI) M / Z: 458 [M+H] + .

[1136] Step G: (2S,3R)-2-((2S)-2-(2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid (100 mg, 0.22 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1 -Ketone p-toluenesulfonate (80.84 mg, 0.22 mmol) was dissolved in tetrahydrofuran (5 mL), and the reaction system was placed in an ice-water bath and cooled to 0 ° C. 3-(diethoxyphosphoryloxy)-1,2,3-benzotriazine-4-one (72.41 mg, 0.24 mmol) and N,N-diisopropylethylamine (56.87 mg, 0.44 mmol) were added, and stirring was continued at 25 ° C for 12 hours.

[1137] After LC-MS / MS monitoring showed that the raw material had reacted completely, water (10 mL) was added to the reaction solution to quench the reaction. The aqueous phase was extracted with dichloromethane (20 mL×2). The organic phases were combined, washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The product was purified by silica gel column chromatography to give 54 mg of (2S,3R)-2-((2S)-2-(2-(6-oxa-3-azabicyclo[3.1.1]hept-3-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide.

[1138] MS (ESI) M / Z: 635 [M+H] + .

[1139] 1 H NMR (400MHz, CD3OD): δ7.43-7.36(m,2H),7.08-7.01(m,2H),6.79(t,J=74.3Hz,1H),5.44(s,1H),5. 06(d,J=4.5Hz,1H),4.67(dd,J=8.6,5.1Hz,1H),4.54(d,J=4.4Hz,1H),4.52-4.48(m,2H),4.45(q,J =7.0Hz,1H),3.29-3.20(m,2H),3.17-2.99(m,4H),2.92-2.77(m,3H),2.50(dd,J=14.2,5.0Hz,1H), 2.42(d,J=8.3Hz,1H),2.35-2.23(m,5H),1.85(p,J=7.5Hz,2H),1.42(s,3H),1.31(d,J=7.0Hz,3H).

[1140] Example 38:

[1141] (2S,3R)-2-((2S)-2-(2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide

[1142] Reaction route:

[1143] Steps:

[1144] Step A: Dissolve 8-oxa-3-azabicyclo[3.2.1]octane hydrochloride (2 g, 13.37 mmol) in acetonitrile (30 mL), add tert-butyl 2-bromoacetate (2.61 g, 13.37 mmol) and potassium carbonate (9.24 g, 66.85 mmol), and stir at 20°C for 4 hours.

[1145] After LC-MS monitoring showed that the reaction of the starting materials was complete, the mixture was diluted with ethyl acetate (20 mL), filtered, and the filtrate was concentrated under reduced pressure to obtain 3 g of crude tert-butyl 2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetate, which was used directly in the next step.

[1146] MS (ESI) M / Z: 228 [M+H] + .

[1147] Step B: Dissolve tert-butyl 2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetate (3 g, 13.20 mmol) in ethyl acetate (100 mL), add 4 M hydrochloric acid in ethyl acetate solution (16.5 mL, 66 mmol), and stir at 25°C for 16 hours.

[1148] After LC-MS monitoring showed that the raw material reaction was complete, the reaction solution was filtered through a pad of celite, and the filter cake was washed with ethyl acetate (50 mL) and dried to obtain 1.5 g of 2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetic acid hydrochloride, which was used directly in the next step.

[1149] MS (ESI) M / Z: 172 [M+H] + .

[1150] Step C: At 0°C, 2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetic acid hydrochloride (1 g, 4.82 mmol) was dissolved in N,N-dimethylformamide (5 mL), and L-alanine benzyl ester hydrochloride (0.66 g, 3.05 mmol) and O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (3.67 g, 9.64 mmol) and N,N-diisopropylethylamine (3.11 g, 24.1 mmol) were added and stirred at 25°C for 4 hours.

[1151] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was added to a saturated aqueous sodium bicarbonate solution (10 mL). The aqueous phase was extracted with ethyl acetate (20 mL x 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 1 / 1) to yield 1.36 g of N-(2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetyl)-L-alanine benzyl ester.

[1152] MS (ESI) M / Z: 333 [M+H] + .

[1153] Step D: Dissolve N-(2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetyl)-L-alanine benzyl ester (1.36 g, 4.09 mmol) in methanol (20 mL), add 10% wet palladium on carbon (44 mg), and stir under a hydrogen atmosphere (15 psi) at 25°C for 1 hour.

[1154] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction solution was filtered through a pad of celite, and the filtrate was concentrated under reduced pressure to obtain 0.72 g of crude N-(2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetyl)-L-alanine, which was used directly in the next step.

[1155] MS (ESI) M / Z: 243 [M+H] + .

[1156] Step E: Benzyl (2S,3R)-2-amino-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate (100 mg, 0.30 mmol) and N-(2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetyl)-L-alanine (72.68 mg, 0.30 mmol) were dissolved in tetrahydrofuran (5 mL). The reaction system was cooled to 0°C in an ice-water bath. 3-(diethoxyphosphoryloxy)-1,2,3-benzotriazine-4-one (98.74 mg, 0.33 mmol) and N,N-diisopropylethylamine (77.54 mg, 0.60 mmol) were added and stirring was continued at 25°C for 12 hours.

[1157] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was quenched with water (10 mL). The aqueous phase was extracted with dichloromethane (20 mL x 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: dichloromethane / methanol = 20 / 1) to yield 150 mg of benzyl (2S,3R)-2-((2S)-2-(2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate.

[1158] MS (ESI) M / Z: 562 [M+H] + .

[1159] Step F: Dissolve benzyl (2S,3R)-2-((2S)-2-(2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoate (150 mg, 0.27 mmol) in methanol (5 mL), add 10% wet palladium on carbon (29 mg), and stir under a hydrogen atmosphere (15 psi) at 25°C for 1 hour.

[1160] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was filtered through a pad of celite, and the filtrate was concentrated under reduced pressure to yield 125 mg of crude (2S,3R)-2-((2S)-2-(2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid, which was used directly in the next step.

[1161] MS (ESI) M / Z: 472 [M+H] + .

[1162] Step G: (2S,3R)-2-((2S)-2-(2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetylamino)propionylamino)-3-(4-difluoromethoxyphenyl)-3-hydroxypropanoic acid (125 mg, 0.27 mmol) and (S)-2-amino-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)propan-1 -Ketone p-toluenesulfonate (99.21 mg, 0.27 mmol) was dissolved in tetrahydrofuran (5 mL), and the reaction system was placed in an ice-water bath and cooled to 0 ° C. 3-(diethoxyphosphoryloxy)-1,2,3-benzotriazine-4-one (88.87 mg, 0.30 mmol) and N,N-diisopropylethylamine (69.79 mg, 0.54 mmol) were added, and stirring was continued at 25 ° C for 12 hours.

[1163] After LC-MS monitoring indicated complete reaction of the starting materials, the reaction mixture was quenched with water (10 mL). The aqueous phase was extracted with dichloromethane (20 mL x 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to yield 58 mg of (2S,3R)-2-((2S)-2-(2-(8-oxa-3-azabicyclo[3.2.1]octan-3-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide.

[1164] MS (ESI) M / Z: 649 [M+H] + .

[1165] 1 H NMR (400MHz, CD3OD): δ7.44-7.36(m,2H),7.07-7.01(m,2H),6.79(t,J=74.2Hz,1H),5.45(s,1H),5.06(d,J= 4.6Hz,1H),4.67(dd,J=8.6,5.0Hz,1H),4.57(d,J=4.5Hz,1H),4.44(q,J=7.0Hz,1H),4.30-4.21(m,2H),3.14 (d,J=5.1Hz,1H),2.98(d,J=16.1Hz,1H),2.92-2.83(m,2H),2.59(d,J=10.8Hz,1H),2.55-2.41(m,3H),2.40- 2.30(m,2H),2.27(t,J=7.6Hz,4H),2.07-1.96(m,2H),1.91-1.74(m,4H),1.42(s,3H),1.32(d,J=7.0Hz,3H).

[1166] Example 39:

[1167] (2S,3R)-2-((S)-2-(2-(1-oxa-7-azaspiro[3.5]non-7-yl)acetylamino)propionylamino)-N-((S)-3-(cyclopent-1-en-1-yl)-1-((R)-2-methyloxiran-2-yl)-1-oxopropan-2-yl)-3-(4-difluoromethoxyphenyl)-3-hydroxypropionamide

[1168] Reaction route:

[1169] Steps:

[1170] Step A: Dissolve 1-oxa-7-azaspiro[3.5]nonane (300.00 mg, 2.36 mmol) and benzyl 2-bromoacetate (540.33 mg, 2.36 mmol, 370.09 μL) in acetonitrile (5 mL), add potassium carbonate (978.03 mg, 7.08 mmol), and stir at 20°C for 3 hours.

[1171] After LC-MS monitoring indicated complete rea...

Claims

1. A compound represented by formula (I-1), its stereoisomers, isotopic derivatives, solvates, nitrogen oxides, prodrugs or pharmaceutically acceptable salts thereof: in, Ring A is selected from a 4-10 membered heterocyclic group, and the 4-10 membered heterocyclic group contains 1-3 heteroatoms selected from N, O or S, and Ring A may be optionally further substituted with one or more selected from =O, OH, halogen, C 1-4 Alkyl or C 1-4 substituted by an alkoxy substituent; Y 1 , Y 2 and Y 3 Each independently selected from N or CR y ; R y Selected from H, OH, halogen, C 1-4 Alkyl or -OR 2a ; R 1 Selected from C 1-4 Alkyl, 3-6 membered heterocyclic group, the C 1-4 The alkyl group may be further optionally substituted with one or more alkyl radicals selected from OH, CN, halogen or -OC 3-6 Cycloalkyl, C 1-4 Alkoxy, -O-halogenated C 1-4 substituted by an alkyl substituent; R 2 Selected from -SR 2a , C 2-4 Alkynyl, C 2-4 Alkenyl, C 1-4 Alkyl, -OR 2a , C 3-6 Cycloalkyl, -C(=O)-R 2d or a 4-7 membered heterocyclic group, wherein the C 1-4 Alkyl, C 3-6 The cycloalkyl or 4-7 membered heterocyclic group may be optionally further substituted with one or more selected from OH, CN, halogen, C 1-4 Alkyl, C 1-4 Alkoxy or NR 2b R 2c substituted by a substituent; R 2a Selected from C 3-6 Cycloalkyl, C 1-4 Alkyl or halogenated C 1-4 alkyl; Or, R 2 With Y 2 and its adjacent carbon atoms together form a 5-8 membered heterocyclic ring; Or, R 2 With Y 3 and its adjacent carbon atoms together form a 5-8 membered heterocyclic ring; R 2b and R 2c Each independently selected from H or C 1-4 alkyl; R 2d Selected from H, OH, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Alkoxy or halogenated C 1-4 alkyl, and the C 1-4 The alkyl group may be further substituted with one or more selected from deuterium, halogen or C 1-4 substituted by an alkoxy substituent; R 3 Selected from H or OH; R 4 Select from H or C 1-4 alkyl; R 5 Selected from C 2-4 Alkenyl, C 4-6 Cycloalkenyl or C 3-6 Cycloalkyl, and the C 2-4 Alkenyl, C 4-6 Cycloalkenyl or C 3-6 The cycloalkyl group may be further optionally substituted with C 2-4 Alkenyl or C 1-4 Alkyl substituted; n is selected from 0 or 1.

2. The compound according to claim 1, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein ring A is selected from 3. The compound according to claim 1 or 2, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein R 1 Selected from methyl, -CH2OH, 4. The compound according to any one of claims 1 to 3, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein R 2 Selected from -OCH3, -CH2OH, -CHF2, -CH2NHCH3, 5. The compound according to any one of claims 1 to 4, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein: Y 1 is selected from N, CH or -CF; Y 2 is selected from CH, -CF, -CO-CHF2; Y 3 is selected from N or CH.

6. The compound according to any one of claims 1 to 5, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein: Y 3 With R 2 Together we form 7. The compound according to any one of claims 1 to 6, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein the structural unit Selected from 8. The compound according to any one of claims 1 to 7, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein: R 3 is selected from H or OH; preferably, R 3 For OH.

9. The compound according to any one of claims 1 to 8, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein: R 4 is selected from H or methyl; preferably, R 4 For H.

10. The compound according to any one of claims 1 to 9, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein: R 5 Selected from Preferably, R 5 Selected from 11. A compound according to any one of claims 1 to 10, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein the compound is selected from the compound represented by general formula (I): in, Ring A is selected from a 6-7 membered heterocyclic group, wherein the 6-7 membered heterocyclic group contains 1-3 heteroatoms selected from N, O or S, and Ring A may be further optionally substituted with one or more selected from OH, halogen, C 1-4 Alkyl or C 1-4 substituted by an alkoxy substituent; R 1 Selected from C 1-4 Alkyl, 3-6 membered heterocyclic group, the C 1-4 The alkyl group may be further optionally substituted with one or more alkyl radicals selected from OH, CN, halogen or -OC 3-6 Substituted by a cycloalkyl substituent; R 2 Selected from C 1-4 Alkyl, -OR 2a , C 3-6 Cycloalkyl, -C(=O)-R 2d or a 4-7 membered heterocyclic group, wherein the C 1-4 Alkyl, C 3-6 The cycloalkyl or 4-7 membered heterocyclic group may be optionally further substituted with one or more selected from OH, CN, halogen, C 1-4 Alkyl, C 1-4 Alkoxy or NR 2b R 2c substituted by a substituent; R 2a Selected from C 3-6 Cycloalkyl, C 1-4 Alkyl or halogenated C 1-4 alkyl; R 2b and R 2c Each independently selected from H or C 1-4 alkyl; R 2d Selected from H, OH, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Alkyl or halogenated C 1-4 alkyl; R 3 Selected from H or OH; n is selected from 0 or 1.

12. A compound of formula (I) according to any one of claims 1 to 11, a stereoisomer, an isotopic derivative, a solvate, a nitrogen oxide, a prodrug or a pharmaceutically acceptable salt thereof, wherein the compound is selected from the compound of general formula (I): in, Ring A is selected from a 6-7 membered heterocyclic group, wherein the 6-7 membered heterocyclic group contains 1-3 heteroatoms selected from N, O or S, and Ring A may be further optionally substituted with one or more selected from OH, halogen, C 1-4 Alkyl, C 1-4 substituted by an alkoxy substituent; R 1 Selected from C 1-4 Alkyl, 3-6 membered heterocyclic group, the C 1-4 The alkyl group may be further optionally substituted with one or more alkyl radicals selected from OH, CN, halogen, -OC 3-6 Substituted by a cycloalkyl substituent; R 2 Selected from C 1-4 Alkyl, -OR 2a , C 3-6 Cycloalkyl, 4-7 membered heterocyclic group, the C 1-4 Alkyl, C 3-6 The cycloalkyl or 4-7 membered heterocyclic group may be optionally further substituted with one or more selected from OH, CN, halogen, C 1-4 Alkyl, C 1-4 Alkoxy, NR 2b R 2c substituted by a substituent; R 2a Selected from C 3-6 Cycloalkyl, C 1-4 Alkyl, halogenated C 1-4 alkyl; R 2b and R 2c Each independently selected from H or C 1-4 alkyl; R 3 Selected from H or OH; n is selected from 0 or 1.

13. A compound according to any one of claims 1 to 12, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein the compound is selected from the compound described by general formula (II): in, X is selected from N or CR x ; R x Selected from H, OH, halogen or C 1-4 Alkoxy; Y is selected from O, NH or NCH3; m is selected from 1 or 2; n is selected from 0 or 1; R 1 , R 2 , R 3 As defined in claim 1.

14. A compound according to any one of claims 1 to 13, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein the compound is selected from the compound represented by general formula (IIIA): R 2 Selected from -SR 2a , C 2-4 Alkynyl, C 2-4 Alkenyl, C 1-4 Alkyl, -OR 2a , C 3-6 Cycloalkyl, -C(=O)-R 2d or a 4-7 membered heterocyclic group, wherein the C 1-4 Alkyl, C 3-6 The cycloalkyl or 4-7 membered heterocyclic group may be optionally further substituted with one or more selected from OH, CN, halogen, C 1-4 Alkyl, C 1-4 Alkoxy or NR 2b R 2c substituted by a substituent; R 2a Selected from C 3-6 Cycloalkyl, C 1-4 Alkyl or halogenated C 1-4 alkyl; R 2b and R 2c Each independently selected from H or C 1-4 alkyl; R 2d Selected from H, OH, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Alkoxy or halogenated C 1-4 alkyl, and the C 1-4 The alkyl group may be further substituted with one or more selected from deuterium, halogen or C 1-4 substituted by an alkoxy substituent; R 3 Selected from H or OH.

15. The compound according to claim 14, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, in, R 2 Selected from C 1-4 Alkyl, -OR 2a , C 3-6 Cycloalkyl, 4-7 membered heterocyclic group; the C 1-4 The alkyl group is further substituted by OH, C 1-4 Alkoxy, NR 2b R 2c Replacement; said C 3-6 The cycloalkyl or 4-7 membered heterocyclic group may be further optionally substituted with one or more selected from OH, C 1-4 substituted by an alkyl substituent; R 2a Selected from C 3-6 Cycloalkyl or halogenated C 1-4 alkyl; R 2b and R 2c Each independently selected from H or C 1-4 alkyl; R 3 Selected from H or OH.

16. The compound according to any one of claims 1 to 15, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein R 2 Selected from -OCH3, or CHF2.

17. A compound according to any one of claims 1 to 13, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein the compound is selected from the compound represented by general formula (IIIB): in, X is selected from N or CR x ; R x Selected from OH, halogen or C 1-4 Alkoxy; Y is O; m is selected from 1 or 2; n is 1; R 1 Selected from C 1-4 alkyl, and the C 1-4 The alkyl group may be further optionally substituted with one or more selected from -OC 3-6 Substituted by a cycloalkyl substituent; R 3 Selected from H or OH.

18. The compound according to claim 13 or 17 or a pharmaceutically acceptable salt thereof, wherein: X is selected from N or CR x ; R x Selected from OH, F, methoxy.

19. The compound according to any one of claims 1 to 13, 17 to 18, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein R 1 Selected from methyl or 20. A compound according to any one of claims 1 to 11, a stereoisomer, an isotopic derivative, a solvate, a nitrogen oxide, a prodrug or a pharmaceutically acceptable salt thereof, wherein the compound is selected from the compounds represented by the general formula (IIIC) or (IIID): X is selected from N or CR x ; R x Selected from H, OH, halogen or C 1-4 Alkoxy; Y is selected from O, NH or NCH3; m is selected from 1 or 2; R 1 Selected from C 1-4 Alkyl, 3-6 membered heterocyclic group, the C 1-4 The alkyl group may be further optionally substituted with one or more alkyl radicals selected from OH, CN, halogen, -OC 3-6 Substituted by a cycloalkyl substituent; R 2d Selected from C 1-4 Alkyl or halogenated C 1-4 alkyl; R 3 Selected from H or OH.

21. The compound according to claim 20, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein X is selected from N or CH.

22. The compound according to claim 19 or 21, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein R 1 Selected from methyl, -CH2OH, -CH2CN or 23. The compound according to any one of claims 20 to 22, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein R 3 For H.

24. The compound according to any one of claims 20 to 23, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein R 2d Selected from C 1-4 Alkyl or halogenated C 1-4 Alkyl; preferably, R 2d Selected from methyl, ethyl or fluoroethyl.

25. A compound according to any one of claims 20 to 24, a stereoisomer, an isotopic derivative, a solvate, a nitrogen oxide, a prodrug or a pharmaceutically acceptable salt thereof, wherein the compound is selected from the compound represented by the general formula (IIID-1): R 2d Selected from methyl, ethyl or fluoroethyl.

26. A compound according to any one of claims 1 to 12, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, wherein the compound is selected from the compound represented by general formula (IIIE): in, X is N, Y is O, m is selected from 0 or 1, R 2 for R 3 Selected from H or OH, R 4 For H.

27. A compound, a stereoisomer, an isotopic derivative, a solvate, a nitrogen oxide, a prodrug or a pharmaceutically acceptable salt thereof, wherein the compound is selected from:

28. A pharmaceutical composition comprising the compound of any one of claims 1 to 27, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

29. Use of the compound according to any one of claims 1 to 27, its stereoisomer, isotopic derivative, solvate, nitrogen oxide, prodrug or pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 28 for the preparation of a medicament for treating a disease mediated by a selective proteasome inhibitor.

30. The use according to claim 29, wherein the disease is a tumor, an infectious disease, an autoimmune disease or a central nervous system disease, preferably an autoimmune disease.

31. The use according to claim 30, wherein the autoimmune disease is selected from systemic lupus erythematosus, arthritis, myasthenia gravis, polymyositis, dermatomyositis, autoimmune gastritis, autoimmune hepatitis, lupus nephritis, Goodpasture's hemorrhage-nephritis syndrome, IgA nephropathy, IgG4-related nephropathy, Crohn's disease, ulcerative colitis, psoriasis or multiple sclerosis.

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