CN108117582A - Big ring epoxy ketone peptides and preparation method thereof and medical usage - Google Patents

Abstract

The invention provides a novel macrocyclic epoxy ketone peptide derivative and its optical isomers or pharmaceutically acceptable salts or their solvates. It is obtained by using carboxyl-protected compounds as starting materials, condensing with amino-protected amino acids, deamination protection, olefin ring-closing metathesis, decarboxylation protection, and condensation with epoxy ketone fragments. The macrocyclic epoxy ketone peptide compound with a new skeleton of the present invention has good proteasome inhibitory activity, and is extremely effective against multiple myeloma such as RPMI 8226, MM.1S, NCI-H929 and other various solid tumor cell lines It has a strong inhibitory effect on proliferation in vitro, and at the same time, this type of compound also has a good oral effect. It can be used in the preparation of drugs against tumors and immune diseases. The raw materials required for the synthesis of the compounds of the present invention are readily available. The route design is reasonable, the reaction conditions are mild, the yield of each step is high, the operation is simple and convenient, and it is suitable for industrial production. Has general formula (I) structure:

Description

Macrocyclic epoxy ketone peptide compound and its preparation method and medical application

technical field

The present invention relates to the field of medicine, and relates to a class of macrocyclic peptide compounds, in particular to a macrocyclic epoxy ketone peptide derivative, in particular to macrocyclic peptide proteasome inhibitor derivatives, salts of the compound and the The compound or its salt is a drug with active ingredients, which can be used for treating tumor-related diseases such as multiple myeloma and mantle cell lymphoma.

Background technique

Malignant tumors are one of the major diseases that threaten human health. In 2008, there were about 12.7 million new cancer patients and 7.6 million cancer deaths worldwide. It is estimated that by 2020, there will be 15 million new cancer patients worldwide, and cancer deaths The number is also skyrocketing globally and could reach 13.2 million. With the continuous increase of the population and the aging trend of the population, coupled with the widespread existence of various unhealthy lifestyles including smoking and the pollution of the environment, this trend will become increasingly serious. important research topic.

Proteasome is a protein complex ubiquitously present in the cytoplasm and nucleus of eukaryotes and archaea, which controls the degradation of 80%-90% of proteins in cells, and is involved in cell cycle regulation, apoptosis, cell signal transduction, DNA repair, cell growth, development and many other physiological functions play an extremely important role. The proteasome plays a regulatory role in various life activities by regulating the level of key proteins (such as P53 and NF-κB) affecting cell signaling pathways. By inhibiting proteasome activity, it can affect the degradation of multiple cycle proteins in cells and promote cell apoptosis.

Since the discovery of the proteasome, small molecular compounds of various structural types have been found to inhibit the activity of the proteasome, and have shown good anti-tumor effects in hematological tumors. At present, the clinically applied small-molecule proteasome inhibitors are boronic acid peptide compounds Bortezomib, Ixazomib and epoxyketone peptide compound Carfilzomib.

Although Bortezomib and Carfilzomib have brought hope to the treatment of multiple myeloma, both are administered through intravenous injection. In order to improve the compliance and safety of patients' medication, the structural modification of the above two has been successful. Ixazomib is transformed from Bortezomib. Through prodrug design and structural adjustment, the absorption and stability of the compound are greatly improved. Due to its good oral effect, the compound was approved by the FDA in 2015 for the treatment of at least A treatment for multiple myeloma patients, however, the drug still cannot prevent peripheral neuropathy caused by boronic acid groups. Therefore, the design of orally active analogues of the epoxy ketone compound Carfilzomib has become the focus of research, among which Oprozomib transformed from Carfilzomib has attracted widespread attention, and the oral bioavailability of this compound in beagle dogs reaches 39%. , is currently in the second phase of clinical research, but according to relevant literature reports, the compound can cause serious dose-related gastrointestinal side effects during oral administration, and corresponding dosage forms are currently being developed to solve this problem.

Contents of the invention

The first object of the present invention is to provide a novel macrocyclic epoxy ketone peptide derivative and its salts or their solvates.

Specifically, the present invention provides a macrocyclic peptide derivative, which has a structure of general formula (I):

and its optical isomers or pharmaceutically acceptable salts or solvates thereof;

in:

R ₁ and R ₂ are each independently selected from H, -C _1-10 alkyl-D, C _1-10 hydroxyalkyl, -C _2-10 unsaturated alkyl-D, -halogenated C _1-10 alkane -D, -C _1-3 alkyl-SC _1-5 alkyl, C _1-10 alkoxyalkyl, halogenated C _1-10 alkoxyalkyl, C _3-10 unsaturated alkoxy Group, C _3-10 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl, aralkyl, heteroaryl, heteroaralkyl. Wherein, D is N(R _a )R _b or missing; R _a is selected from H, OH, C _1-6 alkyl, halogenated C _1-6 alkyl; R _b is selected from N-terminal protecting group;

R ₃ and R ₄ are each independently selected from H, C _1-10 alkyl, halogenated C _1-10 alkyl, aryl, and aralkyl;

_R is selected from H, C _1-6 alkyl, C _1-6 hydroxyalkyl, halogenated C _1-6 alkyl, C _1-6 alkoxyalkyl, halogenated C _1-6 alkoxy Alkyl;

R _is selected from H, C _1-10 alkyl, C _1-10 alkoxyalkyl, C _2-10 unsaturated alkyl, cycloalkyl;

X is O, S, NH, NC _1-6 alkyl;

Y is and Or missing, wherein R is selected from H, C _1-10 alkyl, halogenated C _1-10 alkyl;

Ar is selected from unsubstituted or substituted cycloalkyl, unsubstituted or substituted heterocycloalkyl, unsubstituted or substituted cycloalkenyl, unsubstituted or substituted heterocycloalkenyl, unsubstituted or substituted aryl, none Substituted or substituted aralkyl, unsubstituted or substituted heteroaryl, unsubstituted or substituted heteroaralkyl, optionally fused aryl;

L is

where _B1 is selected from

D ₁ and D ₂ are the same or different, independently selected from -C _1-8 alkyl-, -OC _1-8 alkyl-, -C _1-8 alkyl O-, -C _1-8 alkyl OC _1-8 Alkyl-, -SC _1-8 Alkyl-, -C _1-8 Alkyl S-, -C _1-8 Alkyl, SC _1-8 Alkyl-, -N(R _d )-, - N(R _d )C _1-8 alkyl-, -C _1-8 alkylN(R _d )-,-C _1-8 alkylN(R _d )C _1-8 alkyl-,-N( R _d )C(O)-, -N(R _d )C(O)C _1-8 alkyl-, -C _1-8 alkylN(R _d )C(O)-, -C _1-8 Alkyl N(R _d )C(O)C _1-8 alkyl-, -C(O)N(R _d )-, -C(O)N(R _d )C _1-8 alkyl-,- C _1-8 alkyl C(O)N(R _d )-, -C _1-8 alkyl C(O)N(R _d )C _1-8 alkyl-, -C(O)C _1-8 Alkyl-, -C(O)C _1-8 unsaturated alkyl-, -N(R _d )SO ₂ -, -N(R _d )SO ₂ C _1-8 alkyl-, -C _1-8 Alkyl N(R _d )SO ₂ -, -C _1-8 Alkyl N(R _d )SO ₂ C _1-8 Alkyl-, -OC(O)C _1-8 Alkyl-, -C _{1- 8} alkyl OC (O)-, -C _1-8 alkyl OC (O) C _1-8 alkyl-, -C (O) OC _1-8 alkyl-, -C _1-8 alkyl C ( O) O-, -C _1-8 alkyl C(O)OC _1-8 alkyl-; R _d is selected from H, C _1-4 alkyl, halogenated C _1-4 alkyl, cycloalkyl , heterocycloalkenyl, aryl, aralkyl, heteroaryl, heteroaralkyl.

The substituted substituent is optionally selected from halogen, nitro, amino, cyano, hydroxyl, C _1-6 alkyl, halogenated C _1-6 alkyl, C _1-6 alkoxy, C _1-6 Alkoxyalkyl, C _1-6 alkylamino, halogenated C _1-6 alkoxy, halogenated C _1-6 alkoxyalkyl, halogenated C _1-6 alkylamino, C _3-8 Cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl, aralkyl, heteroaryl, heteroaralkyl.

Preferably, the present invention provides compounds with the structure of formula (II):

and its optical isomers or pharmaceutically acceptable salts or solvates thereof, wherein:

X is preferably O;

Y preferred

Ring A is preferably the following structure:

Wherein, V ₁ , V ₂ , V ₃ , V ₄ , W ₁ , W ₂ and W ₃ are each independently selected from N and C;

R _e is selected from H, halogen, nitro, amino, cyano, hydroxyl, C _1-6 alkyl, halogenated C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkoxy Alkyl, C _1-6 alkylamino, halogenated C _1-6 alkoxy _, halogenated C _1-6 alkoxyalkyl, halogenated C _1-6 alkylamino, C _3-8 Cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl, aralkyl, heteroaryl, heteroaralkyl;

Z is selected from O, S, -N(R _f )-; wherein, R _f is present or missing, _selected from H, C _1-6 alkyl, halogenated C _1-6 alkyl, C _1-6 alkoxy Base, C _1-6 alkoxyalkyl, halogenated C _1-6 alkoxy, C _3-8 cycloalkyl, heterocycloalkyl, C _3-8 cycloalkenyl, heterocycloalkenyl, aromatic radical, aralkyl, heteroaryl, heteroaralkyl;

L, R ₁ , R ₂ , R ₃ , R ₄ and R ₅ are as defined in the general formula (I).

More specifically, the present invention provides the following compounds with the structure of formula (III), or pharmaceutically acceptable salts or solvates thereof:

in:

V ₁ , V ₂ , V ₃ , V ₄ and R _g are as defined in the general formula (II);

B ₁ , D ₁ , D ₂ , R ₁ and R ₂ are as defined in the general formula (I).

Further, preferred compounds of the present invention have the structure of general formula (IV):

and its optical isomers or pharmaceutically acceptable salts or solvates thereof, wherein:

V ₁ and V ₄ are each independently preferably C, N;

_B1 is preferably

D ₁ and D ₂ are the same or different, each independently preferably being -C _1-8 alkyl-, -OC _1-8 alkyl-, -C _1-8 alkyl O-, -C _1-8 alkyl OC _{1 -8} alkyl-;

_R is preferably methoxy-methyl, isobutyl, isopropyl, (N-morpholinyl)formyl-methyl, (N-morpholinyl)formyl-ethyl;

R ₂ is preferably benzyl, isobutyl;

R _g is preferably selected from H, halogen, C _1-6 alkyl, C _1-6 halogenated alkyl, C _1-6 alkoxy, C _1-6 alkoxyalkyl.

Wherein, unless otherwise specified, the amino acids involved are all L-type amino acids.

More specifically, the preferred compound of the general formula (IV) structure of the present invention is:

N-[(8S,11S)-11-(methoxymethyl)-10,13-dioxo-3,4,5,7,8,9,10,11,12,13-decahydro- 2H-1,6,9,12-Benzodioxadiazacyclopentadecyl-8-formyl]-Phe-epoxyketone

N-[(9S,12S)-12-(methoxymethyl)-11,14-dioxo-2,3,4,5,6,8,9,10,11,12,13,14 -Dodecahydro-1,7,10,13-benzodioxadiazacyclohexadeca-9-formyl]-Phe-epoxyketone

N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-3,4,5,6,7,9,10,11,12,13,14,15 -Dodecahydro-2H-1,8,11,14-benzodioxadiazacyclohexadeca-10-formyl]-Leu-epoxyketone

N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-3,4,5,6,7,9,10,11,12,13,14,15 -2H-Dodecahydro-1,8,11,14-benzodioxadiazacyclohexadeca-10-formyl]-Phe-epoxyketone

N-[(11S,14S)-14-(methoxymethyl)-13,16-dioxo-2,3,4,5,6,7,8,10,11,12,13,14 ,15,16-tetradetrahydro-1,9,12,15-benzodioxadiazacycloctadec-11-formyl]-Phe-epoxyketone

N-[(12S,15S)-15-(methoxymethyl)-14,17-dioxo-3,4,5,6,7,8,9,11,12,13,14,15 ,16,17-2H-Tetradetrahydro-2H-1,10,13,16-Benzodioxadiazacyclonadeca-12-formyl]-Phe-epoxyketone

N-[(13S,16S)-16-(methoxymethyl)-15,18-dioxo-3,4,5,6,7,8,9,10,13,14,15,16 ,17,18-tetradecahydro-2H,12H-benzo[i][1,11]dioxa[4,7]diazacycloeicos-13-formyl]-Phe-epoxyketone

N-[(7S,10S)-7-(methoxymethyl)-5,8-dioxo-6,7,8,9,10,11,13,14,15,16,17,18 -Dodecahydro-5H-pyrido[3,2-i][1,11]dioxa[4,7]diazacyclohexadeca-10-formyl]-Phe-epoxyketone

N-[(14S,17S)-17-(methoxymethyl)-16,19-dioxo-6,7,8,9,10,11,14,15,16,17,18,19 -Dodecahydro-13H-pyrido[2,3-i][1,11]dioxa[4,7]diazacyclohexadeca-14-formyl]-Phe-epoxyketone

N-[(9S,12S)-12-(methoxymethyl)-11,14-dioxo-3,4,5,6,7,8,9,10,11,12,13,14 -Dodecahydro-2H-benzo[b][1]oxa[5,8]-diazacyclohexadeca-9-formyl]-Phe-epoxyketone

N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,8,9,10,11,12,13 ,14,15-Tetradetrahydro-benzo[b][1]oxa[5,8]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone

N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 -Dodecahydro-9H-19-methoxy-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxy ketone

N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 -Dodecahydro-9H-19-fluoro-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone

N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 -Dodecahydro-9H-18-fluoro-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone

N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 -Dodecahydro-9H-18-chloro-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone

N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 -Dodecahydro-9H-18-methoxy-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxy ketone

N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 -Dodecahydro-9H-18-methyl-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone

N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 -Dodecahydro-9H-17-chloro-benzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-formyl]-Phe-epoxyketone

N-[(10S,13S)-13-(2-morpholino-2-oxoethyl)-12,15-dioxo-2,3,4,5,6,7,10,11, 12,13,14,15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-cyclo Oxyketone

N-[(10S,13S)-13-(3-morpholino-3-oxopropyl)-12,15-dioxo-2,3,4,5,6,7,10,11, 12,13,14,15-Dodecahydro-9Hbenzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxy ketone

N-[(9S,12S)-12-(methoxymethyl)-11,14-dioxo-2,3,5,6,9,10,11,12,13,14-decahydro- 8H-Benzo[o][1,4,7]-trioxa[10,13]-diazacyclohexadeca-9-formyl]-Phe-epoxyketone

N-[(12S,15S)-15-(methoxymethyl)-14,17-dioxo-2,3,5,6,8,9,12,13,14,15,16,17 -Dodecahydro-11H-benzo[r][1,4,7,10]tetraoxa[13,16]-diazacyclonadeca-12-formyl]-Phe-epoxyketone

N-[(10S,13S)-13-(2-(cyclopropylamino)-2-oxoethyl)-12,15-dioxo-2,3,4,5,6,7,10 ,11,12,13,14,15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-formyl]-Phe - epoxy ketone

N-[(10S,13S)-13-isobutyl-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-dodecahydro -9H-Benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone

N-[(10S,13S)-isopropyl-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-dodecahydro-9H -13-Benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone

and optical isomers of the above-mentioned compounds or pharmaceutically acceptable salts or solvates thereof.

More specifically, the present invention provides the following compounds with the structure of formula (V), or pharmaceutically acceptable salts or solvates thereof:

in:

_B1 is preferably

D ₁ and D ₂ are the same or different, and are independently preferably -OC _1-8 alkyl-, -C _1-8 alkyl O-, -C _1-8 alkylOC _1-8 alkyl-, -C _{1 -8} Alkyl OC _1-8 Alkyl-;

_Rh is preferably selected from H.

Wherein, unless otherwise specified, the amino acids involved are all L-type amino acids.

More specifically, the preferred compound of the general formula (V) structure of the present invention is:

N-[(13S,16S)-16-(methoxymethyl)-15,18-dioxo-2,11-dioxa-14,17-diazabicyclo[17.3.1]-di Tridecyl-1(23),19,21-triene-13-formyl]-Phe-epoxyketone

and optical isomers of the above-mentioned compounds or pharmaceutically acceptable salts or solvates thereof.

Still further, the preferred compound of the present invention has the structure of general formula (VI):

and its optical isomers or pharmaceutically acceptable salts or solvates thereof, wherein:

W ₁ and W ₂ are each preferably C, N;

Z is preferably N(C _1-6 alkyl);

_B1 is preferably

D ₁ and D ₂ are the same or different, each independently preferably being -C _1-8 alkyl-, -OC _1-8 alkyl-, -C _1-8 alkyl O-, -C _1-8 alkyl OC _{1 -8} alkyl-;

R _i is preferably H.

Wherein, unless otherwise specified, the amino acids involved are all L-type amino acids.

More specifically, the preferred compound of the general formula (VI) structure of the present invention is:

N-[(6S,9S)-6-(methoxymethyl)-4,7-dioxo-1,4,5,6,7,8,9,10,12,13,14,15 ,16,17-Dodecahydro-1-methyl-pyrazolo[4,3-i][1,11]dioxa[4,7]diazacyclohexadeca-9-formyl] -Phe-epoxyketone

N-[(6S,9S)-6-(methoxymethyl)-1-methyl-4,7-dioxo-4,5,6,7,8,9,10,12,13, 14,15,16,17,18-Tetradetrahydro-1H-pyrazolo[4,3-i][1]oxa[4,7]-diazacyclohexadeca-9-formyl] -Phe-epoxyketone

and optical isomers of the above-mentioned compounds or pharmaceutically acceptable salts or solvates thereof.

Preferably, the present invention also provides a compound having the structure of formula (VII), or a pharmaceutically acceptable salt or solvate thereof:

in:

W ₁ and W ₂ are each preferably C, N;

Z is preferably S;

_B1 is preferably

D ₁ and D ₂ are the same or different, each independently preferably being -C _1-8 alkyl-, -OC _1-8 alkyl-, -C _1-8 alkyl O-, -C _1-8 alkyl OC _{1 -8} alkyl-;

_Rj is preferably H, methyl.

Wherein, unless otherwise specified, the amino acids involved are all L-type amino acids.

More specifically, preferred compounds of the general formula (VII) structure of the present invention are:

N-[(13S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15,16,17,18 -Dodecahydro-12H-thieno[2,3-i][1,11]dioxa[4,7]diazacyclohexadeca-13-formyl]-Phe-epoxyketone

N-[(13S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15,16,17,18 -Dodecahydro-12H-2-methyl-thiazolo[4,5-i][1,11]dioxa[4,7]diazacyclohexadeca-13-formyl]-Phe- epoxy ketone

and optical isomers of the above-mentioned compounds or pharmaceutically acceptable salts or solvates thereof.

Preferably, the present invention also provides a compound having a structure of formula (VIII), or a pharmaceutically acceptable salt or solvate thereof:

in:

W ₁ , W ₂ and W ₃ are each preferably C, N;

Z is preferably N;

_B1 is preferably

D ₁ and D ₂ are the same or different, each independently preferably being -C _1-8 alkyl-, -OC _1-8 alkyl-, -C _1-8 alkyl O-, -C _1-8 alkyl OC _{1 -8} alkyl-;

_Rk is preferably H.

Wherein, unless otherwise specified, the amino acids involved are all L-type amino acids.

More specifically, the preferred compound of the general formula (VIII) structure of the present invention is:

N-(13S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15,16,17,18- Dodecahydro-12H-imidazo[2,1-i][1]oxa[4,7,10]-triazacyclohexadeca-13-formyl]-Phe-epoxyketone

N-(12S,15S)-15-(methoxymethyl)-14,17-dioxo-6,7,8,9,12,13,14,15,16,17-decahydro-5H ,11H-imidazo[2,1-i][1]oxa[4,7,10]-triazacyclopentadecan-12-formyl]-Phe-epoxyketone

N-(11S,14S)-14-(methoxymethyl)-13,16-dioxo-5,6,7,8,11,12,13,14,15,16-decahydro-10H -Imidazolo[2,1-i][1]oxa[4,7,10]-triazacyclotetradec-11-formyl]-Phe-epoxyketone

N-(3S,6S)-3-(methoxymethyl)-1,4-dioxo-2,3,4,5,6,7,9,10,11,12,13,14- Dodecahydro-1H-pyrrolo[2,1-i][1]oxa[4,7,10]-triazacyclohexadeca-6-formyl]-Phe-epoxyketone

N-(6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,12,13,14,15,16,17- Dodecahydro-4H-pyrazolo[5,1-i][1]oxa[4,7,10]-triazacyclohexadeca-9-formyl]-Phe-epoxyketone

N-(6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,13,14,15,16-decahydro-4H ,12H-pyrazolo[5,1-i][1]oxa[4,7,10]triazacyclopentadeca-9-formyl]-Phe-epoxyketone

N-(6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,12,13,14,15-decahydro-4H -pyrazolo[5,1-i][1]oxa[4,7,10]-triazacyclocarbo-9-formyl]-Phe-epoxyketone

and optical isomers of the above-mentioned compounds or pharmaceutically acceptable salts or solvates thereof.

Preferably, the present invention also provides a compound having a structure of formula (IX), or a pharmaceutically acceptable salt or solvate thereof:

in:

_B1 is preferably

_D2 is preferably -C _1-8 alkyl-, -C _1-8 alkylOC _1-8 alkyl-;

R _m is preferably H.

Wherein, unless otherwise specified, the amino acids involved are all L-type amino acids.

More specifically, the preferred compound of the general formula (IX) structure of the present invention is:

N-[(3S,6S,18aS)-3-(methoxymethyl)-1,4,14-trioxo-hexadecahydro-1H-pyrrolo[2,1-i][1]oxo Hetero[4,7,10]triazacyclohexadec-6-formyl]-Phe-epoxyketone

and optical isomers of the above-mentioned compounds or pharmaceutically acceptable salts or solvates thereof.

Another object of the present invention is to provide the preparation method of above-mentioned target compound, realize through the following steps:

(1) Compound 5 reacts with amino-protected amino acid under the action of condensation reagent to obtain compound 6. The selected condensation reagent has dicyclohexylcarbodiimide/4-dimethylaminopyridine, dicyclohexylcarbodiimide/1- Hydroxybenzotriazole, N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride/1-hydroxybenzotriazole, benzotriazole-N,N ,N',N'-tetramethyluronium hexafluorophosphate/1-hydroxybenzotriazole. The reaction temperature is 0-30°C, and the reaction time is 2-8 hours. The crude product can be directly used in the next reaction;

(2) Compound 6 removes the Boc protecting group under the action of trifluoroacetic acid, and the crude product is directly used for the next step reaction;

(3) Compound 7 reacts with compound 2 to obtain compound 8 under the action of a condensation reagent. The selected condensation reagent has dicyclohexylcarbodiimide/4-dimethylaminopyridine, dicyclohexylcarbodiimide/1-hydroxybenzene Triazole, N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride/1-hydroxybenzotriazole, benzotriazole-N,N,N ',N'-Tetramethyluronium hexafluorophosphate/1-hydroxybenzotriazole. The reaction temperature is 0-30°C, and the reaction time is 2-8 hours. The crude product can be directly used in the next reaction;

(4) Compound 8 is ring-merged under the effect of a metal catalyst or a condensing agent to remove the carboxyl protecting group to obtain compound 9. The metal catalyst selected is Grubbs second-generation catalyst, and the condensing agent is the same as in (1), and the metal-catalyzed reaction temperature is 30 -100°C, reaction time 0.5-3 hours, condensation reaction temperature 0-30°C, reaction time 3-8 hours, the obtained product is used for the next reaction;

(5) Compound 9 and Compound 11 were reacted under the action of a condensation reagent to obtain the product Compound 10. The condensation reagent was selected as in step (1), and the obtained crude product was separated by column chromatography to obtain a pure product.

Reaction formula:

in:

The definitions of the substituents of R ₁ -R ₄ , B ₁ , D ₁ , and D ₂ are the same as those of the general formula I.

The definitions of ring A substituents are the same as in general formula II.

For the synthesis of the raw material compound 11 used, refer to the literature J.Med.Chem.2009, 52, 3028.

Another object of the present invention is to provide the application of the macrocyclic epoxy ketone peptide derivatives in the preparation of antitumor drugs. The tumor is selected from blood tumors such as myeloma, lymphoma, leukemia, breast cancer, sarcoma, lung cancer, prostate cancer, colon cancer, rectal cancer, kidney cancer, pancreatic cancer, neuroblastoma, glioma, Head cancer, neck cancer, thyroid cancer, liver cancer, ovarian cancer, vulvar cancer, cervical cancer, endometrial cancer, testicular cancer, bladder cancer, esophagus cancer, stomach cancer, nasopharyngeal cancer, buccal cancer, oral cancer, gastrointestinal tract cancer Stromal tumors, skin cancer.

The macrocyclic epoxy ketone peptide derivatives include their salts or their solvates.

The medicine is prepared from macrocyclic epoxy ketone peptide derivatives and their salts or their solvates and pharmaceutical carriers.

Another object of the present invention is to provide the application of the macrocyclic epoxy ketone peptide derivatives in the preparation of drugs for immune diseases. The macrocyclic epoxy ketone peptide derivatives include their salts or their solvates.

The pharmaceutical dosage forms of the present invention include injections, freeze-dried powder injections, tablets, capsules, granules and the like. The administration route of the drug of the present invention is selected from injection administration, oral administration, inhalation or implantation administration and the like.

Experiments have proved that the macrocyclic epoxy ketone peptide compound with a new skeleton of the present invention has good proteasome inhibitory activity, and is effective against multiple myeloma such as RPMI 8226, MM.1S, NCI-H929 and other various solid tumor cells. The strain has a strong in vitro proliferation inhibitory effect, and the compound also has a good oral effect. The raw materials required for the synthesis of the compounds of the present invention are readily available. The route design is reasonable, the reaction conditions are mild, the yield of each step is high, the operation is simple and convenient, and it is suitable for industrial production.

Description of drawings

Fig. 1 Inhibitory activity of macrocyclic epoxy ketone peptides on proteasome in mouse PBMC after oral administration.

Detailed ways

The present invention is further described in conjunction with examples, and the following examples are only to illustrate the present invention, but not to limit the present invention in any way.

Preparation Example 1 Methyl 2-(allyloxy)benzoate (1a)

Methyl salicylate (4.2ml, 33mmol), potassium carbonate (13.8g, 99mmol) and 3-bromopropene (4.0ml, 46mmol) were placed in a 100mL three-necked flask, 50mL of acetone was added, and the reaction solution was refluxed The reaction was carried out for 18 hours. After the reaction was monitored by TLC, the reaction solution was cooled to room temperature, the solvent was removed under reduced pressure, 100ml of ethyl acetate was added, washed with water (2×100ml) and saturated brine (2×100ml) successively, dried over anhydrous sodium sulfate and then decompressed The solvent was removed and separated by column chromatography to obtain 5.9 g of a colorless oily liquid with a yield of 93%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.80 (dd, J=8.0, 2.0Hz, 1H), 7.44 (ddd, J=8.0, 7.5, 2.0Hz, 1H), 7.01–6.94 (m, 2H), 6.11–6.02(m,1H),5.54–5.48(m,1H),5.32–5.27(m,1H),4.63(dt,J=5.0,1.5Hz,2H),3.90(s,3H)ppm; -MS: m/z＝193[M+H] ⁺ .

Preparation Example 2 Methyl 2-(3-buten-1-yloxy)benzoate (1b).

Using 4-bromo-1-butene as raw material, the synthesis and post-treatment were the same as in Preparation Example 1 to obtain 4.0 g of a colorless oily liquid with a yield of 88%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.77 (dd, J=7.5, 1.5Hz, 1H), 7.47–7.38(m, 1H), 7.00–6.91(m, 2H), 6.00–5.88(m, 1H ), 5.22–5.07(m,2H), 4.08(t,J＝6.5Hz,2H), 3.88(s,3H), 2.62–2.54(m,2H)ppm; ESI-MS: m/z＝207[ M+H] ⁺ .

Preparation Example 3 Methyl 2-(4-penten-1-yloxy)benzoate (1c)

Using 5-bromo-1-pentene as raw material, the synthesis and post-treatment were the same as in Preparation Example 1 to obtain 5.8 g of a colorless oily liquid with a yield of 80%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.78 (dd, J=7.5, 1.5Hz, 1H), 7.44 (ddd, J=8.0, 7.5, 1.5Hz, 1H), 6.99–6.94 (m, 2H), 5.92–5.80(m,1H),5.09–5.04(m,1H),5.02–4.98(m,1H),4.05(t,J＝6.5Hz,2H),3.89(s,3H),2.34–2.25( m,2H),1.98–1.88(m,2H)ppm; ESI-MS: m/z=221[M+H] ⁺ .

Preparation Example 4 Methyl 2-(5-hexen-1-yloxy)benzoate (1d)

Using 6-bromo-1-hexene as the raw material, the synthesis and post-treatment were the same as in Preparation Example 1 to obtain 6.3 g of a colorless oily liquid with a yield of 81%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.78(d, J=8.0Hz, 1H), 7.47–7.41(m, 1H), 6.99–6.93(m, 2H), 5.88–5.78(m, 1H), 5.08–4.94(m,2H),4.04(t,J＝6.5Hz,2H),3.88(s,3H),2.17–2.10(m,2H),1.90–1.81(m,2H),1.65–1.57( m, 2H) ppm; ESI-MS: m/z = 235 [M+H] ⁺ .

Preparation Example 5 Methyl 2-(6-hepten-1-yloxy)benzoate (1e)

Using 7-bromo-1-heptene as a raw material, the synthesis and post-treatment were the same as in Preparation Example 1 to obtain 6.1 g of a colorless oily liquid with a yield of 75%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.78(d, J=8.0Hz, 1H), 7.46–7.41(m, 1H), 6.98–6.93(m, 2H), 5.89–5.76(m, 1H), 5.04–4.90(m,2H),4.03(t,J＝6.5Hz,2H),3.89(s,3H),2.13–2.06(m,2H),1.89–1.80(m,2H),1.56–1.42( m, 4H) ppm; ESI-MS: m/z = 249 [M+H] ⁺ .

Preparation Example 6, 2-(7-octen-1-yloxy)methyl benzoate (1f)

Using 8-bromo-1-octene as the raw material, the synthesis and post-treatment were the same as in Preparation Example 1 to obtain 6.1 g of a colorless oily liquid with a yield of 70%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.78(d, J=8.0Hz, 1H), 7.46–7.40(m, 1H), 6.99–6.92(m, 2H), 5.87–5.75(m, 1H), 5.05–4.90(m,2H),4.03(t,J＝6.5Hz,2H),3.89(s,3H),2.10–2.01(m,2H),1.88–1.79(m,2H),1.55–1.46( m,2H),1.46–1.34(m,4H)ppm; ESI-MS: m/z=263[M+H] ⁺ .

Preparation Example 7, 5-chloro-2-(4-penten-1-yloxy)methyl benzoate (1g)

Methyl 5-chloro-2-hydroxybenzoate (6.1g, 33mmol), potassium carbonate (9.1g, 66mmol) and 5-bromo-1-pentene (4.0ml, 46mmol) were placed in a 100mL three-necked flask, 50 mL of acetone was added, and the reaction solution was reacted under reflux for 18 hours. Cool the reaction solution to room temperature, remove the solvent under reduced pressure, add 100ml of ethyl acetate, wash with water (2×100ml) and saturated brine (2×100ml) successively, dry over anhydrous sodium sulfate, remove the solvent under reduced pressure, and perform column chromatography 6.9 g of a colorless oily liquid was isolated, with a yield of 82%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.75 (d, J=3.0Hz, 1H), 7.38 (dd, J=9.0, 3.0Hz, 1H), 6.89 (d, J=9.0Hz, 1H), 5.90 –5.79(m,1H),5.09–5.03(m,1H),5.02–4.97(m,1H),4.02(t,J＝6.5Hz,2H),3.89(s,3H),2.31–2.24(m ,2H),1.96–1.87(m,2H)ppm; ESI-MS: m/z＝255[M+H] ⁺ .

Preparation Example 8, methyl 4-chloro-2-(4-penten-1-yloxy)benzoate (1h)

Using methyl 4-chloro-2-hydroxybenzoate as the raw material, the synthesis and post-treatment were the same as in Preparation Example 7 to obtain 7.1 g of a colorless oily liquid with a yield of 85%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.77–7.72(m,1H),6.97–6.93(m,2H),5.90–5.80(m,1H),5.10–4.99(m,2H),4.03(t , J＝6.5Hz, 2H), 3.88(s, 3H), 2.32–2.25(m, 2H), 1.98–1.90(m, 2H)ppm; ESI-MS: m/z＝255[M+H] ⁺ .

Preparation Example 9, methyl 4-fluoro-2-(4-penten-1-yloxy)benzoate (1i)

Using methyl 4-fluoro-2-hydroxybenzoate as the raw material, the synthesis and post-treatment were the same as in Preparation Example 7 to obtain 6.4 g of a colorless oily liquid with a yield of 82%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.86–7.81(m,1H),6.68–6.63(m,2H),5.90–5.80(m,1H),5.10–5.04(m,1H),5.03–4.99 (m,1H),4.02(t,J=6.5Hz,2H),3.88(s,3H),2.32–2.26(m,2H),1.98–1.91(m,2H)ppm; ESI-MS:m/ z＝239[M+H] ⁺ .

Preparation Example 10, methyl 4-methyl-2-(4-penten-1-yloxy)benzoate (1j)

Using methyl 4-methyl-2-hydroxybenzoate as raw material, the synthesis and post-treatment were the same as in Preparation Example 7 to obtain 6.1 g of a colorless oily liquid with a yield of 79%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.71(d, J=7.5Hz, 1H), 6.80–6.74(m, 2H), 5.92–5.82(m, 1H), 5.10–5.04(m, 1H), 5.02–4.97(m,1H),4.03(t,J＝6.4Hz,2H),3.87(s,3H),2.36(s,3H),2.32–2.26(m,2H),1.97–1.90(m, 2H) ppm; ESI-MS: m/z=235[M+H] ⁺ .

Preparation example 11, methyl 4-methoxy-2-(4-penten-1-yloxy)benzoate (1k)

Using methyl 4-methoxy-2-hydroxybenzoate as raw material, the synthesis and post-treatment were the same as in Preparation Example 7 to obtain 6.9 g of a colorless oily liquid with a yield of 84%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.84 (d, J=8.5Hz, 1H), 6.49 (dd, J=8.5, 2.5Hz, 1H), 6.46 (d, J=2.5Hz, 1H), 5.91 –5.81(m,1H),5.09–5.03(m,1H),5.02–4.97(m,1H),4.02(t,J＝6.5Hz,2H),3.85(s,3H),3.84(s,3H ), 2.33–2.26(m,2H), 1.98–1.91(m,2H)ppm; ESI-MS: m/z＝251[M+H] ⁺ .

Preparation Example 12, ethyl 2-methyl-4-(4-penten-1-yloxy)thiazole-5-carboxylate (1l)

Using ethyl 2-methyl-4-hydroxy-thiazole-5-carboxylate as raw material, the synthesis and post-treatment were the same as in Preparation Example 7 to obtain 6.6 g of a colorless oily liquid with a yield of 78%. ¹ H NMR (500MHz, CDCl ₃ )δ5.92–5.78(m,1H),5.09–5.03(m,1H),5.01–4.96(m,1H),4.45(t,J＝6.5Hz,2H), 4.28(q, J=7.0Hz, 2H), 2.61(s, 3H), 2.28–2.21(m, 2H), 1.96–1.87(m, 2H), 1.33(t, J=7.0Hz, 3H)ppm; ESI-MS: m/z＝256[M+H] ⁺ .

Preparation Example 13, 2-(4-penten-1-yloxy)thiophene-3-carboxylic acid methyl ester (1m)

Using methyl 2-hydroxythiophene-3-carboxylate as raw material, the synthesis and post-treatment were the same as in Preparation Example 7 to obtain 6.2 g of white solid with a yield of 83%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.39(d, J=5.5Hz, 1H), 6.83(d, J=5.5Hz, 1H), 5.91–5.80(m, 1H), 5.09–5.03(m, 1H),5.02–4.98(m,1H),4.14(t,J＝6.5Hz,2H),3.84(s,3H),2.30–2.24(m,2H),1.97–1.90(m,2H)ppm; ESI-MS: m/z＝227[M+H] ⁺ .

Preparation Example 14, 3-(4-penten-1-yloxy)picolinate methyl ester (1n)

Methyl 3-hydroxypicolinate (0.31g, 2mmol), potassium carbonate (0.61mg, 4.4mmol) and 5-bromo-1-pentene (0.28ml, 2.4mmol) were placed in a 25mL single-necked bottle, and 10ml DMF was added , and the reaction solution was reacted at 80° C. for 3 hours. Cool the reaction solution to room temperature, add 20ml of ethyl acetate and 20ml of water, separate the organic layer, then wash with water (20ml×2) and saturated brine (20ml×1) successively, dry over anhydrous sodium sulfate, and remove the solvent under reduced pressure , separated by column chromatography to obtain 0.39 g of a colorless oily liquid, with a yield of 89%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.25 (dd, J=4.5, 1.5Hz, 1H), 7.38 (dd, J=8.5, 4.5Hz, 1H), 7.33 (dd, J=8.5, 1.5Hz, 1H),5.89–5.78(m,1H),5.09–5.03(m,1H),5.02–4.97(m,1H),4.06(t,J＝6.5Hz,2H),3.96(s,3H),2.31 –2.23(m,2H),1.97–1.89(m,2H)ppm; ESI-MS: m/z=222[M+H] ⁺ .

Preparation Example 15, 4-penten-1-yl 2-(4-penten-1-yloxy)nicotinate (1o)

Add 4-penten-1-ol (1.45ml, 14.0mmol) into 24ml THF, cool to -8°C, slowly add 1.0N sodium bis(trimethylsilyl)amide (14.0ml, 14.0mmol) dropwise After 25 min, methyl 2-chloronicotinate (0.61 ml, 4.7 mmol) dissolved in 24 ml THF was added, and the reaction solution was reacted at room temperature for 2 hours. Quench the reaction with 50ml of saturated aqueous ammonium chloride solution, add dichloromethane (30ml×3) for extraction, combine the organic phases, wash with saturated brine (50ml×1), dry over anhydrous sodium sulfate, and remove the solvent under reduced pressure. Column chromatography separated 0.52 g of a colorless oily liquid with a yield of 40%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.28(dd, J=5.0, 2.0Hz, 1H), 8.14(dd, J=7.5, 2.0Hz, 1H), 6.92(dd, J=7.5, 5.0Hz, 1H),5.91–5.78(m,2H),5.09–5.07(m,1H),5.05–5.03(m,1H),5.03–4.96(m,2H),4.42(t,J＝6.5Hz,2H) ,4.32(t,J=6.5Hz,2H),2.29–2.19(m,4H),1.95–1.82(m,4H)ppm; ESI-MS: m/z=276[M+H] ⁺ .

Preparation Example 16, 1-methyl-5-(4-penten-1-yloxy)-1H-pyrazole-4-carboxylic acid mixed ester (1p)

Using ethyl 5-chloro-1-methyl-1H-pyrazole-4-carboxylate as raw material, the synthesis and post-treatment were the same as in Preparation Example 15 to obtain a colorless oily liquid mixture (methyl ester and 4-pentene-1- Phthalate) 0.51, yield 39%.

Preparation Example 17, methyl 3-(hept-6-en-1-yloxy)benzoate (1q)

Methyl 3-hydroxybenzoate (0.46g, 3mmol), potassium carbonate (0.91mg, 6.6mmol) and 7-bromo-1-heptene (0.55ml, 3.6mmol) were placed in a 25mL single-necked bottle, and 15ml DMF, and the reaction solution was reacted at 80°C for 3 hours. Cool the reaction solution to room temperature, add 30ml of ethyl acetate and 30ml of water, separate the organic layer, then wash with water (30ml×2) and saturated brine (30ml×1) successively, dry over anhydrous sodium sulfate, and remove the solvent under reduced pressure , separated by column chromatography to obtain 0.61 g of a colorless oily liquid, with a yield of 82%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.62 (d, J=7.5Hz, 1H), 7.57–7.52 (m, 1H), 7.36–7.30 (m, 1H), 7.09 (dd, J=8.0, 2.0 Hz,1H),5.87–5.76(m,1H),5.04–4.98(m,1H),4.98–4.93(m,1H),4.00(t,J＝6.5Hz,2H),3.91(s,3H) ,2.14–2.06(m,2H),1.83–1.77(m,2H),1.55–1.40(m,4H)ppm; ESI-MS: m/z=249[M+H] ⁺ .

Preparation Example 18, Methyl 3-fluoro-2-(4-penten-1-yloxy)benzoate (1r)

Using methyl 3-fluoro-2-hydroxybenzoate as the raw material, the synthesis and post-treatment were the same as in Preparation Example 7 to obtain 6.2 g of a colorless oily liquid with a yield of 80%. ppm; ESI-MS: m/z＝239[M+H] ⁺ .

Preparation example 19, methyl 3-methoxy-2-(pent-4-en-1-yloxy)benzoate (1s)

Using methyl 3-methoxy-2-hydroxybenzoate as the raw material, the synthesis and post-treatment were the same as in Preparation Example 7 to obtain 7.4 g of a colorless oily liquid with a yield of 90%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.31(dd, J=7.5, 2.0Hz, 1H), 7.09–7.02(m, 2H), 5.93–5.84(m, 1H), 5.09–5.03(m, 1H ),5.00–4.96(m,1H),4.04(t,J＝6.5Hz,2H),3.89(s,3H),3.86(s,3H),2.29–2.21(m,2H),1.93–1.84( m,2H)ppm; ESI-MS: m/z＝251[M+H] ⁺ .

Preparation Example 20, 5-allyl-1-methyl-1H-pyrazole-4-carboxylic acid ethyl ester (1t)

5-Bromo-1-methyl-1H-pyrazole-4-carboxylic acid ethyl ester (1.2g, 5.3mmol), allyltributyltin (1.8ml, 5.8mmol), Pd ₂ (dba) ₃ (0.24g , 0.27mmol), tri-tert-butylphosphine (0.27ml, 1.1mmol) and potassium fluoride (1.5g, 26mmol) were placed in a 50ml three-necked flask, and N ₂ was injected into anhydrous THF 25ml under protection, and refluxed for 5 hours. Rotate THF to dryness under reduced pressure, add 40ml of ethyl acetate and 30ml of water, separate the organic layer, wash with saturated brine (30ml×2), dry over anhydrous sodium sulfate, remove the solvent under reduced pressure, and separate by column chromatography to obtain Colorless oily liquid 0.82g, yield 80%. ¹ H NMR (500MHz, CDCl ₃ )δ7.85(s,1H),5.94–5.80(m,1H),5.16–5.06(m,1H),5.02–4.90(m,1H),4.28(q,J =7.0Hz, 2H), 3.85–3.73(m, 5H), 1.34(t, J=7.0Hz, 3H) ppm; ESI-MS: m/z=195[M+H] ⁺ .

Preparation Example 21, 1-(pent-4-en-1-yl)-1H-pyrazole-5-carboxylic acid methyl ester (1u)

Using methyl 3-pyrazolecarboxylate as the raw material, the synthesis and post-treatment were the same as in Preparation Example 14 to obtain 0.16 g of a colorless oily liquid with a yield of 40%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.48(d, J=2.0Hz, 1H), 6.83(d, J=2.0Hz, 1H), 5.86–5.76(m, 1H), 5.08–5.01(m, 1H),5.00–4.96(m,1H),4.62–4.54(m,2H),3.88(s,3H),2.12–2.04(m,2H),1.99–1.90(m,2H) ppm; :m/z＝195[M+H] ⁺ .

Preparation Example 22, ethyl 1-allyl-1H-imidazole-2-carboxylate (1v)

Using ethyl imidazole-2-carboxylate as raw material, the synthesis and post-treatment were the same as in Preparation Example 14 to obtain 0.21 g of a colorless oily liquid with a yield of 58%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.18(s,1H),7.08(s,1H),6.06–5.95(m,1H),5.28–5.19(m,1H),5.13–5.01(m,3H ), 4.40(q, J=7.0Hz, 2H), 1.42(t, J=7.0Hz, 3H) ppm; ESI-MS: m/z=

181[M+H] ⁺ .

Preparation Example 23, 1-(but-3-en-1-yl)-1H-imidazole-2-carboxylic acid ethyl ester (1w)

Using ethyl imidazole-2-carboxylate as the raw material, the synthesis and post-treatment were the same as in Preparation Example 14 to obtain 0.25 g of a colorless oily liquid with a yield of 69%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.14(d, J=1.0Hz, 1H), 7.05(d, J=1.0Hz, 1H), 5.80–5.70(m, 1H), 5.08–5.01(m, ESI -MS: m/z＝195[M+H] ⁺ .

Preparative Example 24, Ethyl 1-(pent-4-en-1-yl)-1H-imidazole-2-carboxylate (1x)

Using ethyl imidazole-2-carboxylate as raw material, the synthesis and post-treatment were the same as in Preparation Example 14 to obtain 0.34 g of a colorless oily liquid with a yield of 81%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.15(d, J=1.0Hz, 1H), 7.06(d, J=1.0Hz, 1H), 5.85–5.73(m, 1H), 5.08–4.99(m, 2H), 4.44–4.35(m,4H), 2.14–2.05(m,2H), 1.96–1.86(m,2H), 1.43(t,J=7.0Hz,3H)ppm; ESI-MS: m/z ＝209[M+H] ⁺ .

Preparation Example 25, Methyl 1-(pent-4-en-1-yl)-1H-pyrrole-2-carboxylate (1y)

Methyl 2-pyrrolecarboxylate (1.0g, 8.0mmol), potassium carbonate (2.2g, 16mmol), potassium iodide (0.13g, 0.8mmol) and 5-bromo-1-pentene (1.1ml, 9.6mmol) were placed in 10ml of DMF was added to a 50mL single-necked bottle, and the reaction solution was reacted at 80°C for 3 hours. Cool the reaction solution to room temperature, add 30ml of ethyl acetate and 20ml of water, separate the organic layer, then wash with water (20ml×2) and saturated brine (20ml×1) successively, dry over anhydrous sodium sulfate, and remove the solvent under reduced pressure , separated by column chromatography to obtain 0.46 g of a colorless oily liquid, with a yield of 30%. ¹ H NMR (500MHz, CDCl ₃ ) δ6.95 (dd, J=4.0, 2.0Hz, 1H), 6.84–6.82 (m, 1H), 6.12 (dd, J=4.0, 2.5Hz, 1H), 5.86– 5.76(m,1H),5.08–4.98(m,2H),4.34–4.28(m,2H),3.81(s,3H),2.10–2.03(m,2H),1.91–1.83(m,2H)ppm ;ESI-MS: m/z=194[M+H] ⁺ .

Preparation Example 26, Pent-4-enoyl-L-proline methyl ester (1z)

Starting material 4-pentenoic acid (1.0g, 10mmol) (2.0g, 12mmol) was dissolved in 30mL of dichloromethane, 1-hydroxybenzotriazole (1.6g, 12mmol) and N-(3-dimethylaminopropyl base)-N'-ethylcarbodiimide hydrochloride (3.5g, 18mmol), react at room temperature for 30 minutes. Cool to 0°C in an ice bath, add L-proline methyl ester hydrochloride (2.0 g, 12 mmol) and DIPEA (5.0 ml, 30 mmol), and react at room temperature for 3 hours. Add 50 mL of saturated sodium bicarbonate for dilution, separate the organic layer, wash with saturated brine (10 mL×1), dry over anhydrous sodium sulfate, evaporate the solvent, and separate by column chromatography to obtain 1.9 g of a colorless oily liquid with a yield of 92 %. ¹ H NMR (500MHz, CDCl ₃ )δ5.93–5.80(m,1H),5.09–4.96(m,2H),4.50(dd,J＝8.5,3.5Hz,1H),3.73(s,3H), 3.68–3.62(m,1H),3.54–3.46(m,1H),2.47–2.36(m,4H),2.21–2.14(m,1H),2.11–2.03(m,1H),2.03–1.94(m ,2H)ppm; ESI-MS: m/z=212[M+H] ⁺

Preparation Example 27, Allyl 2-Hydroxybenzoate (1za)

Place salicylic acid (4.1g, 30mmol), potassium carbonate (4.1mg, 36mmol) and 3-bromopropene (3.0ml, 36mmol) in a 100mL single-necked bottle, add 40ml of DMF, and react the reaction solution at room temperature for 3 hours . Add 80ml of ethyl acetate and 60ml of water, separate the organic layer, then wash with water (50ml×2) and saturated brine (50ml×1) successively, dry over anhydrous sodium sulfate, remove the solvent under reduced pressure, and separate by column chromatography. Color oily liquid 4.6g, yield 86%. ¹ H NMR (500MHz, CDCl ₃ ) δ10.74(s,1H),7.89(dd,J=8.0,1.5Hz,1H),7.49–7.43(m,1H),6.99(dd,J=8.5,1.0 Hz,1H),6.93–6.86(m,1H),6.04(ddt,J＝17.0,10.5,5.5Hz,1H),5.47–5.40(m,1H),5.36–5.30(m,1H),4.86( dt,J＝5.5,1.5Hz,2H)ppm; ESI-MS:m/z＝179[M+H] ⁺ .

Preparation Example 28, 2-(allyloxy)benzoic acid (2a)

The raw material 2-(allyloxy)benzoic acid methyl ester (6.0g, 31mmol) was dissolved in 20mL of methanol and 20ml of THF, and 50mL of 3N KOH aqueous solution was added dropwise, and heated to 50°C for 3 hours. Methanol and THF were evaporated from the reaction solution under reduced pressure, the pH of the aqueous layer was adjusted to 2-3 with 3N HCl, extracted with ethyl acetate (50 mL×3), the organic layers were combined, dried over anhydrous sodium sulfate, and the solvent was evaporated under reduced pressure. The resulting product was directly used in the next reaction.

Preparation Example 29, 2-(3-buten-1-yloxy)benzoic acid (2b)

Using methyl 2-(but-3-en-1-yloxy)benzoate as a raw material, the synthesis and post-treatment were the same as in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 30, 2-(4-penten-1-yloxy)benzoic acid (2c)

Using methyl 2-(pent-4-en-1-yloxy)benzoate as raw material, the synthesis and post-treatment were the same as in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 31, 2-(5-hexen-1-yloxy)benzoic acid (2d)

Using methyl 2-(hex-5-en-1-yloxy)benzoate as a raw material, the synthesis and post-treatment were the same as in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparative Example 32, 2-(6-hepten-1-yloxy)benzoic acid (2e)

Using methyl 2-(hept-6-en-1-yloxy)benzoate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 33, 2-(7-octen-1-yloxy)benzoic acid (2f)

Using methyl 2-(oct-7-en-1-yloxy)benzoate as a raw material, the synthesis and post-treatment were the same as in Preparation Example 28, and the obtained product was directly used in the next reaction.

PREPARATIVE EXAMPLE 34, 5-Chloro-2-(4-penten-1-yloxy)benzoic acid (2 g)

Using methyl 5-chloro-2-(4-penten-1-yloxy)benzoate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparative Example 35, 4-Chloro-2-(4-penten-1-yloxy)benzoic acid (2h)

Using methyl 4-chloro-2-(4-penten-1-yloxy)benzoate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 36, 4-fluoro-2-(4-penten-1-yloxy)benzoic acid (2i)

Using methyl 4-fluoro-2-(4-penten-1-yloxy)benzoate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 37, 4-methyl-2-(4-penten-1-yloxy)benzoic acid (2j)

Using methyl 4-methyl-2-(4-penten-1-yloxy)benzoate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparative Example 38, 4-methoxy-2-(4-penten-1-yloxy)benzoic acid (2k)

Using methyl 4-methoxy-2-(4-penten-1-yloxy)benzoate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 39, 2-Methyl-4-(4-penten-1-yloxy)thiazole-5-carboxylic acid (2l)

Using ethyl 2-methyl-4-(4-penten-1-yloxy)thiazole-5-carboxylate as raw material, the synthesis and post-treatment were the same as in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 40, 2-(4-penten-1-yloxy)thiophene-3-carboxylic acid (2m)

Using methyl 2-(4-penten-1-yloxy)thiophene-3-carboxylate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 41, 3-(4-penten-1-yloxy)picolinic acid (2n)

Using methyl 3-(4-penten-1-yloxy)picolinate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 42, 4-penten-1-yl 2-(4-penten-1-yloxy)nicotinic acid (2o)

Using 4-penten-1-yl 2-(4-penten-1-yloxy)nicotinate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 43, 1-Methyl-5-(4-penten-1-yloxy)-1H-pyrazole-4-carboxylic acid (2p)

Using 1-methyl-5-(4-penten-1-yloxy)-1H-pyrazole-4-carboxylic acid mixed ester as raw material, the synthesis and post-treatment are the same as in Preparation Example 28, and the obtained product is directly used in the following step response.

Preparation Example 44, 3-(hept-6-en-1-yloxy)benzoic acid (2q)

Using methyl 3-(hept-6-en-1-yloxy)benzoate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 45, 3-fluoro-2-(4-penten-1-yloxy)benzoic acid (2r)

Using methyl 3-fluoro-2-(4-penten-1-yloxy)benzoate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 46, 3-methoxy-2-(pent-4-en-1-yloxy)benzoic acid (2s)

Using methyl 3-methoxy-2-(pent-4-en-1-yloxy)benzoate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 47, 5-allyl-1-methyl-1H-pyrazole-4-carboxylic acid (2t)

Using ethyl 5-allyl-1-methyl-1H-pyrazole-4-carboxylate as raw material, the synthesis and post-treatment were the same as those in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 48, 1-(pent-4-en-1-yl)-1H-pyrazole-5-carboxylic acid (2u)

Using methyl 1-(pent-4-en-1-yl)-1H-pyrazole-5-carboxylate as a raw material, the synthesis and post-treatment were the same as in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 49, 1-allyl-1H-imidazole-2-carboxylic acid (2v)

The raw material 1-allyl-1H-imidazole-2-carboxylic acid ethyl ester (0.37g, 2.0mmol) was dissolved in 0.5mL of methanol and 0.5ml of THF, and 1.0mL of 3N KOH aqueous solution was added dropwise, heated to 50°C for 3 hours. The reaction solution was evaporated under reduced pressure to remove methanol and THF, the aqueous layer was adjusted to pH 2-3 with 3N HCl, extracted with dichloromethane: acetone = 3:1 (4mL×3), the organic layers were combined, dried over anhydrous sodium sulfate and then reduced pressure The solvent was distilled off to obtain 0.18 g of white colloidal liquid (10% decarboxylation product), yield 59%. ¹ H NMR (500MHz, DMSO) δ8.04 (d, J = 4.0Hz, 1H), 7.10 (s, 1H), 6.05–5.93 (m, 1H), 5.20 (dd, J = 10.0, 1.5Hz, 1H ), 5.11 (dd, J=17.0, 1.5Hz, 1H), 4.67 (d, J=6.0Hz, 2H) ppm; ESI-MS: m/z=153[M+H] ⁺ .

Preparation Example 50, 1-(but-3-en-1-yl)-1H-imidazole-2-carboxylic acid (2w)

Using ethyl 1-(but-3-en-1-yl)-1H-imidazole-2-carboxylate as raw material, the synthesis and post-treatment were the same as in Preparation Example 49 to obtain 0.23 g of a white colloidal liquid (12% decarboxylation product) , yield 69%. ¹ H NMR (500MHz,DMSO)δ8.26(s,1H),7.06(s,1H),5.55–5.43(m,1H),4.79–4.72(m,2H),3.92(t,J=7.0Hz ,2H),2.31–2.20(m,2H)ppm; ESI-MS: m/z＝167[M+H] ⁺ .

Preparative Example 51, 1-(pent-4-en-1-yl)-1H-imidazole-2-carboxylic acid (2x)

Using ethyl 1-(pent-4-en-1-yl)-1H-imidazole-2-carboxylate as raw material, the synthesis and post-treatment were the same as in Preparation Example 49 to obtain 0.26 g of white colloidal liquid (15% decarboxylation product) , yield 72%. ¹ H NMR (500MHz,DMSO)δ8.87(s,1H),8.09(s,1H),5.86–5.75(m,1H),5.10–4.95(m,2H),4.44–4.36(m,2H) ,2.07–2.00(m,2H),1.93–1.83(m,2H)ppm; ESI-MS: m/z=181[M+H] ⁺ .

Preparation Example 52, 1-(pent-4-en-1-yl)-1H-pyrrole-2-carboxylic acid (2y)

Using methyl 1-(pent-4-en-1-yl)-1H-pyrrole-2-carboxylate as raw material, the synthesis and post-treatment were the same as in Preparation Example 28, and the obtained product was directly used in the next reaction.

Preparation Example 53, Pent-4-enoyl-L-proline (2z)

The raw material pent-4-enoyl-L-proline methyl ester (0.47g, 2.2mmol) was dissolved in 1.6mL of methanol and 1.6ml of THF, and 3.3mL of 1N LiOH aqueous solution was added dropwise, and reacted at room temperature for 5 hours. Methanol and THF were evaporated from the reaction solution under reduced pressure, the pH of the aqueous layer was adjusted to 2-3 with 3N HCl, extracted with ethyl acetate (6 mL×3), the organic layers were combined, dried over anhydrous sodium sulfate, and the solvent was evaporated under reduced pressure. The resulting product was directly used in the next reaction.

Preparation Example 54, Boc-Ser-OBn(3a)

Boc-L-serine (4.0g, 19.5mmol) was dissolved in 500ml of DMF, potassium carbonate (3.2g, 23mmol) was added, cooled to 0°C in an ice bath, benzyl bromide (2.8ml, 23mmol) was dissolved in 100ml of DMF and then slowly The above reaction solution was slowly added dropwise, and reacted at room temperature for 10 hours. Add 100ml of ethyl acetate and 100ml of water to the reaction solution, separate the organic layer, then wash with 100ml of saturated aqueous sodium bicarbonate solution, 100ml of saturated brine, dry over anhydrous sodium sulfate, evaporate the solvent under reduced pressure, and pass column chromatography 5.2 g of white solid was isolated with a yield of 90%. mp:69-71℃; ¹ H NMR (500MHz, CDCl ₃ ) δ7.40–7.31(m,5H),5.45(s,1H),5.28–5.16(m,2H),4.48–4.38(m,1H ), 3.99 (dd, J = 11.0, 3.5Hz, 1H), 3.92 (dd, J = 11.0, 3.0Hz, 1H), 1.94 (br s, 1H), 1.45 (s, 9H) ppm; ESI-MS: m/z＝296[M+H] ⁺ .

Preparation Example 55, Boc-Ser-OMe(3b)

L-serine methyl ester hydrochloride (3.6g, 30.0mmol) was dissolved in 50ml of methanol, triethylamine (8.3ml, 60.0mmol) was added, cooled to 0°C in an ice bath, Boc anhydride (7.9g, 36mmol) was dissolved in After 20ml of methanol, slowly drop into the above reaction solution, and react at room temperature for 10 hours. Evaporate the solvent under reduced pressure, add 100ml of ethyl acetate and 100ml of water to the reaction solution, separate the organic layer, then successively wash with 100ml of saturated aqueous ammonium chloride solution, 100ml of saturated brine, dry over anhydrous sodium sulfate, and evaporate under reduced pressure The solvent was separated by column chromatography to obtain 5.9 g of light yellow oily liquid with a yield of 90%. ¹ H NMR (500MHz, CDCl ₃ ) δ5.47(s, 1H), 4.43–4.33(m, 1H), 3.96(dd, J=11.0, 3.5Hz, 1H), 3.90(dd, J=11.0, 3.5 Hz,1H),3.78(s,3H),2.27(br s,1H),1.45(s,9H)ppm; ESI-MS: m/z=220[M+H] ⁺ .

Preparation Example 56, O-allyl-Boc-Ser-benzyl ester (4a)

Reactant 3a (2.7g, 9.1mmol) and tetrakistriphenylphosphine palladium (0.53g, 0.46mmol) were dissolved in 40mL THF, and slowly injected into allyl methyl carbonate (1.5ml, 12.7mmol) under the protection of _N2 , heated to reflux for 5 hours. The reaction solution was evaporated to remove the solvent under reduced pressure, 50 ml of ethyl acetate and saturated aqueous sodium bicarbonate (50 ml x 1) were added, the organic layer was washed with saturated brine (30 ml x 1), dried over anhydrous sodium sulfate, and the solvent was evaporated under reduced pressure. Separated by column chromatography, 1.9 g of light yellow oily liquid was obtained, with a yield of 62%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.41–7.30(m,5H),5.86–5.68(m,1H),5.41(d,J＝8.5Hz,1H),5.32–5.17(m,2H), ESI -MS: m/z＝336[M+H] ⁺ .

Preparation Example 57, O-allyl-Boc-Ser-methyl ester (4b)

Using 3b as raw material, the synthesis and post-treatment were the same as those in Preparation Example 37, and 1.4 g of light yellow oily liquid was obtained with a yield of 59%. ¹ HNMR (500MHz, CDCl ₃ ) δ5.87–5.78(m,1H),5.38(d,J＝8.0Hz,1H),5.27–5.20(m,1H),5.20–5.15(m,1H),4.42 (dt,J=7.0,3.0Hz,1H),4.01–3.92(m,2H),3.85(dd,J=9.5,3.0Hz,1H),3.76(s,3H),3.64(dd,J=9.5 , 3.0 Hz, 1H), 1.45 (s, 9H) ppm; ESI-MS: m/z = 260 [M+H] ⁺ .

Preparation Example 58, (S)-2-((tert-butoxycarbonyl)amino)-5-hexenoic acid benzyl ester (4c)

Reactant 3a (3.0g, 10mmol) and triphenylphosphine (3.9g, 15mmol) were added to a 100ml three-necked flask, and 30ml of THF was added after nitrogen replacement and cooled to 0°C. At this temperature, pyridine (1.7ml, 20mmol) was added dropwise, and finally solid iodine (3.8g, 15mmol) was added three times, and reacted at room temperature for 4 hours. Diethyl ether (20ml x 3) was added to the reaction solution, washed successively with 1N HCl (20ml x 3), 1N Na ₂ S ₂ O ₃ (20ml x 2) and saturated brine (20ml x 1), dried over anhydrous sodium sulfate and reduced The solvent was evaporated under pressure and separated by column chromatography (EtOAc:petroleum ether=1:60 to 1:30) to obtain 3.6 g of white solid with a yield of 89%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.41–7.33 (m, 5H), 5.36 (d, J=7.0Hz, 1H), 5.21 (q, J=12.0Hz, 2H), 4.59–4.51 (m, 1H), 3.61(dd, J=10.0, 4.0Hz, 1H), 3.56(dd, J=10.0, 4.0Hz, 1H), 1.45(s, 9H)ppm; ESI-MS: m/z=406[M +H] ⁺ .

The activated zinc powder (0.33g, 4.9mmol) was placed in a 25ml three-neck flask, cooled to 0°C, and (R)-2-((tert-butoxycarbonyl) dissolved in 3ml DMF was added dropwise under nitrogen protection. Amino)-3-iodopropionic acid benzyl ester (0.5g, 1.2mmol), react at room temperature for 3h. Stop stirring, and wash the supernatant after the solid settles for later use.

Cuprous bromide (0.036g, 0.25mmol) and 3-bromopropene (0.22g, 1.9mmol) were added to 2ml of DMF, cooled to -15°C, injected dropwise into the supernatant, and reacted at room temperature overnight. Add 10ml of ethyl acetate to the reaction solution, wash with H ₂ O (10ml x 2), 1N Na ₂ S ₂ O ₃ (20ml x 2) and saturated brine (20ml x 1) successively, and dry over anhydrous sodium sulfate The solvent was distilled off under reduced pressure, and 0.23 g of a white solid was obtained by column chromatography (EtOAc:petroleum ether=1:15), with a yield of 60%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.41–7.31(m,5H),5.81–5.70(m,1H),5.24–5.10(m,2H),5.10–4.93(m,3H),4.42–4.32 (m,1H),2.15–2.02(m,2H),1.97–1.82(m,1H),1.78–1.67(m,1H),1.44(s,9H)ppm; ESI-MS: m/z=320 [M+H] ⁺ .

Preparation Example 59, N-(tert-butoxycarbonyl)-O-(hex-5-en-1-yl)-L-serine benzyl ester (4d)

Boc-Ser (2.0g, 10mmol) was dissolved in 25ml of DMF, cooled to 0°C in an ice bath, NaH (0.60g, 25mmol) was added slowly, reacted at this temperature for 30min, and 6-bromo-1-hexene (1.3ml , 10mmol), react overnight at room temperature. Slowly add 50ml of water and 50ml of ethyl acetate in an ice bath, separate the organic layer, add water (50ml x 2) and saturated brine (50ml x 2) to wash, dry over anhydrous sodium sulfate, evaporate the solvent under reduced pressure to obtain a colorless oil Liquid 1.6g, yield 56%. ¹ H NMR (500MHz, CDCl ₃ )δ5.84–5.72(m,1H),5.42(d,J=8.0Hz,1H),5.06–4.88(m,2H),4.47–4.37(m,1H), 3.87(dd, J=11.0, 3.0Hz, 2H), 3.65(dd, J=9.5, 3.5Hz, 1H), 3.45(t, J=6.5Hz, 2H), 2.08–2.02(m, 2H), 1.60 - 1.52 (m, 2H), 1.49 - 1.38 (m, 11H) ppm; ESI-MS: m/z = 288 [M+H] ⁺ .

The raw materials obtained above were dissolved in 15ml of DMF, anhydrous potassium carbonate (0.92g, 6.7mmol) and benzyl bromide (0.79ml, 6.7mmol) were added under ice cooling, and reacted at room temperature overnight. Add 30ml of ethyl acetate and 30ml of water to the reaction solution, separate the organic layer, then wash with water (30ml x 2) and saturated brine (30ml x 2) successively, dry over anhydrous sodium sulfate, evaporate the solvent under reduced pressure, pass Column chromatography separated 1.1 g of a colorless oily liquid with a yield of 52%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.41–7.29(m,5H),5.85–5.71(m,1H),5.38(d,J＝8.5Hz,1H),5.28–5.09(m,2H), 5.05–4.89(m,2H),4.52–4.39(m,1H),3.84(dd,J＝9.5,3.0Hz,1H),3.63(dd,J＝9.5,3.0Hz,1H),3.45–3.27( m, 2H), 2.07-1.99 (m, 2H), 1.55-1.33 (m, 13H) ppm; ESI-MS: m/z=378 [M+H] ⁺ .

Preparation Example 60, (S)-2-((tert-butoxycarbonyl)amino)-4-morpholino-4-oxobutanoic acid benzyl ester (4e) raw material Boc-L-aspartic acid-1 -Benzyl ester (1.0 g, 3.0 mmol) was dissolved in 15 mL of dichloromethane, 1-hydroxybenzotriazole (0.49 g, 3.6 mmol) and N-(3-dimethylaminopropyl)-N'-ethyl Carbodiimide hydrochloride (1.0 g, 5.4 mmol) was reacted at room temperature for half an hour. Cool in an ice bath to 0°C, add morpholine (0.31g, 3.6mmol) and DIPEA (0.99ml, 6.0mmol), and react at room temperature for 3 hours. Add 20mL of saturated sodium bicarbonate solution for dilution, separate the organic layer, wash with saturated brine (10mL×1), dry over anhydrous sodium sulfate, evaporate the solvent under reduced pressure, and separate by column chromatography to obtain 1.1g of light yellow oily liquid with a yield of 93 %. ¹ H NMR (500MHz, CDCl ₃ ) δ7.37–7.29(m, 5H), 5.80(d, J=9.0Hz, 1H), 5.22(d, J=12.5Hz, 1H), 5.13(d, J= 12.5Hz, 1H), 4.65–4.56(m, 1H), 3.68–3.49(m, 6H), 3.39(t, J＝4.5Hz, 2H), 3.12(dd, J＝16.5, 4.0Hz, 1H), 2.76(dd, J=16.5,4.0Hz,1H), 1.41(s,9H)ppm; ESI-MS: m/z=393[M+H] ⁺ .

Preparation Example 61, (S)-2-((tert-butoxycarbonyl)amino)-5-morpholino-5-oxopentanoic acid benzyl ester (4f)

Using Boc-L-glutamic acid-1-benzyl ester as raw material, the synthesis and post-treatment were the same as those in Preparation Example 59 to obtain 1.1 g of white solid with a yield of 90%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.39–7.31(m, 5H), 5.31(d, J=7.0Hz, 1H), 5.22(d, J=12.0Hz, 1H), 5.13(d, J= 12.0Hz,1H),4.40–4.28(m,1H),3.68–3.53(m,6H),3.37–3.26(m,2H),2.41–2.15(m,3H),2.06–1.97(m,1H) , 1.43 (s, 9H) ppm; ESI-MS: m/z = 407 [M+H] ⁺ . Preparation Example 62, N ² -(tert-butoxycarbonyl)-N ⁴ -cyclopropyl-L-asparagine benzyl ester (4 g) using cyclopropylamine as raw material, the synthesis and post-treatment were the same as in Preparation Example 59, to obtain White solid 1.1g, yield 99%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.38–7.28(m,6H),5.24–5.12(m,3H),4.56–4.47(m,1H),2.83(dd,J＝15.5,4.5Hz,1H ),2.70–2.61(m,2H),1.42(s,9H),0.77–0.70(m,1H),0.48–0.42(m,2H)ppm; ESI-MS: m/z＝363[M+H ] ⁺ .

Preparation Example 63, O-allyl-Ser-OBn hydrochloride (5a)

Reactant 4a (3.4 g, 10 mmol) was dissolved in 15 mL CH ₂ Cl ₂ , cooled to 0° C. in an ice bath, 2N HCl-saturated ethyl acetate solution (20 mL) was added, and reacted at room temperature for 2 hours. The solvent was distilled off from the reaction solution under reduced pressure to obtain a light yellow solid, which was directly used in the next reaction.

Preparation Example 64, O-allyl-Ser-OMe hydrochloride (5b)

Using 4b as raw material, the synthesis and post-treatment were the same as those in Preparation Example 63, and the obtained product was directly used in the next reaction.

Preparation Example 65, (S)-2-amino-5-hexenoic acid benzyl ester hydrochloride (5c)

Using 4c as raw material, the synthesis and post-treatment were the same as those in Preparation Example 63, and the obtained product was directly used in the next reaction.

Preparation Example 66, O-(5-hexen-1-yl)-L-serine benzyl ester hydrochloride (5d)

Using 4d as raw material, the synthesis and post-treatment were the same as those in Preparation Example 63, and the obtained product was directly used in the next reaction.

PREPARATIVE EXAMPLE 67, (2-(2-(Benzyloxy)ethoxy)ethyl)-L-serine methyl ester hydrochloride (5e)

2-[2-(Benzyloxy)ethoxy]ethanol (5.7g, 29mmol), triethylamine (4.4ml, 32mmol) and DMAP (0.35g, 2.9mmol) were dissolved in 100ml of dichloromethane, ice bath Cool down to 0°C. Add p-toluenesulfonyl chloride (6.0 g, 32 mmol) dropwise, and react at room temperature for 8 hours. Add 50ml of dichloromethane and 100ml of 2N HCl, separate the organic layer, add water (100ml x 2) and saturated brine (80ml x 2) to wash, dry over anhydrous sodium sulfate, evaporate the solvent under reduced pressure, and separate by column chromatography Yellow oily liquid 6.4g, yield 63%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.82–7.77(m,2H),7.37–7.27(m,7H),4.53(s,2H),4.20–4.15(m,2H),3.72–3.68(m , 2H), 3.64–3.54 (m, 4H), 2.43 (s, 3H) ppm; ESI-MS: m/z=351 [M+H] ⁺ .

Boc-Ser (2.1g, 10mmol) was dissolved in 30ml DMF, cooled to 0°C in an ice bath, NaH (0.6g, 25mmol) was added slowly, reacted at this temperature for 30min, and 2-[2-(benzyloxy)ethane was added Oxy)ethyl]-4-methylbenzenesulfonic acid ethyl ester (3.5g, 10mmol), react at room temperature overnight. Slowly add 50ml of water and 50ml of ethyl acetate in an ice bath, separate the organic layer, add water (50ml x 2) and saturated brine (50ml x 2) to wash, dry over anhydrous sodium sulfate, evaporate the solvent under reduced pressure to obtain a colorless oil Liquid 2.0g, yield 52%. ESI-MS: m/z = 384 [M+H] ⁺ .

O-(2-(2-(benzyloxy)ethoxy)ethyl)-N-(tert-butoxycarbonyl)-L-serine (2.0g, 5.2mmol) was dissolved in 25ml of anhydrous methanol, ice bath Thionyl chloride (0.83ml, 11mmol) was slowly added dropwise under reflux for 5 hours, and the solvent was distilled off under reduced pressure to obtain a colorless oily liquid, which was directly used in the next reaction.

Preparation Example 68, (S)-13-Amino-1-phenyl-2,5,8,11-tetraoxatetradec-14-oic acid methyl ester hydrochloride (5f)

Triethylene glycol monobenzyl ether (1.5g, 6.2mmol), triethylamine (1.7ml, 12mmol) and DMAP (0.076g, 0.62mmol) were dissolved in 20ml of dichloromethane and cooled to 0°C in an ice bath. p-Toluenesulfonyl chloride (1.8 g, 9.4 mmol) was added dropwise and reacted at room temperature for 8 hours. Add 10ml of dichloromethane and 2N HCl (20ml), separate the organic layer, add water (20ml x 2) and saturated brine (20ml x 2), wash with anhydrous sodium sulfate, evaporate the solvent under reduced pressure, and separate by column chromatography 1.5 g of a light yellow oily liquid was obtained, with a yield of 61%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.81–7.77(m,2H),7.36–7.27(m,7H),4.56(s,2H),4.18–4.14(m,2H),3.71–3.67(m , 2H), 3.66–3.56 (m, 8H), 2.43 (s, 3H) ppm; ESI-MS: m/z=395 [M+H] ⁺ .

Boc-Ser (0.68g, 3.3mmol) was dissolved in 10ml DMF, cooled to 0°C in an ice bath, NaH (0.33g, 8.3mmol) was added slowly, reacted at this temperature for 30min, and 2-(2-(2-( Benzyloxy)ethoxy)ethoxy)ethyl)-4-methylbenzenesulfonic acid ethyl ester (1.3g, 3.3mmol), react at room temperature overnight. Slowly add 20ml of water and 30ml of ethyl acetate under an ice bath, separate the organic layer, add water (20ml x 2) and saturated brine (20ml x 2) to wash, dry over anhydrous sodium sulfate, evaporate the solvent under reduced pressure to obtain a colorless oil Liquid 1.0g, yield 71%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.38–7.27(m,5H),5.56(d,J=8.0Hz,1H),4.66–4.56(m,2H),4.45–4.40(m,1H), 3.94 (dd, J = 9.5, 3.0 Hz, 1H), 3.76 - 3.55 (m, 13H), 1.45 (s, 9H) ppm; ESI-MS: m/z = 428 [M+H] ⁺ .

(S)-13-((tert-butoxycarbonyl)amino)-1-phenyl-2,5,8,11-tetraoxatetradec-14-acid (1.0g, 2.3mmol) was dissolved in 10ml without In water and methanol, thionyl chloride (0.37ml, 5.1mmol) was slowly added dropwise under ice bath, and the reaction was carried out under reflux for 5 hours. The solvent was evaporated under reduced pressure to obtain a colorless oily liquid, which was directly used in the next reaction.

Preparation Example 69, N-(N-(tert-butoxycarbonyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester (6a)

Starting material Boc-O-methyl-Ser (2.2 g, 10 mmol) was dissolved in 30 mL of dichloromethane, 1-hydroxybenzotriazole (1.6 g, 12 mmol) and N-(3-dimethylaminopropyl)- N'-Ethylcarbodiimide hydrochloride (3.5 g, 18 mmol) was reacted at room temperature for half an hour. Cool in an ice bath to 0°C, add O-allyl-Ser-OBn hydrochloride (5a, 3.3g, 12mmol) and DIPEA (5.0ml, 30mmol), and react at room temperature for 3 hours. Add 50 mL of saturated sodium bicarbonate for dilution, separate the organic layer, wash with saturated brine (30 mL x 2), dry over anhydrous sodium sulfate, evaporate the solvent, and separate by column chromatography to obtain 2.7 g of a colorless oily liquid with a yield of 61%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.39(d, J=6.5Hz, 1H), 7.36–7.29(m, 5H), 5.80–5.70(m, 1H), 5.43(d, J=5.0Hz, 1H),5.26–5.21(m,1H),5.21–5.16(m,1H),5.16–5.11(m,2H),4.74(dt,J＝8.0,3.0Hz,1H),4.31–4.24(m, 1H), 3.94–3.87(m, 3H), 3.77(dd, J＝9.0, 4.0Hz, 1H), 3.65(dd, J＝9.5, 3.0Hz, 1H), 3.43(dd, J＝9.0, 7.0Hz ,1H), 3.33(s,3H), 1.44(s,9H)ppm; ESI-MS: m/z=437[M+H] ⁺ .

Preparation Example 70, N-(N-(tert-butoxycarbonyl)-O-methyl-L-seryl)-O-allyl-L-serine methyl ester (6b)

Using 5b (2.3g, 12mmol) as raw material, the synthesis and post-treatment were the same as those in Preparation Example 69, and 2.6g of light yellow oily liquid was obtained with a yield of 72%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.34 (d, J=3.5Hz, 1H), 5.87–5.76 (m, 1H), 5.42 (s, 1H), 5.26–5.15 (m, 2H), 4.70 ( dt,J=8.0,3.5Hz,1H), 4.31–4.24(m,1H),3.99–3.95(m,2H),3.88(dd,J=9.5,3.0Hz,1H),3.81(dd,J= 9.0,3.5Hz,1H),3.76(s,3H),3.65(dd,J=9.5,3.5Hz,1H),3.48(dd,J=9.0,7.0Hz,1H),3.39(s,3H), 1.45(s,9H)ppm; ESI-MS: m/z=361[M+H] ⁺ .

Preparation Example 71, (S)-2-((S)-2-((tert-butoxycarbonyl)amino)-3-methoxypropionamido)-5-hexenoic acid benzyl ester (6c)

Using 5c (3.1 g, 12 mmol) as raw material, the synthesis and post-treatment were the same as in Preparation Example 69, and 2.4 g of light yellow oily liquid was obtained with a yield of 57%. ¹ H NMR (500MHz, CDCl ₃ )δ7.40–7.31(m,5H),7.11(d,J=5.5Hz,1H),5.79–5.68(m,1H),5.41(d,J=3.5Hz, 1H), 5.17(q, J=12.0Hz, 2H), 5.01–4.94(m, 2H), 4.67(td, J=7.5, 4.5Hz, 1H), 4.25(s, 1H), 3.79(dd,J ＝9.0,4.0Hz,1H),3.46–3.40(m,1H),3.35(s,3H),2.11–1.92(m,3H),1.84–1.74(m,1H),1.45(s,9H)ppm ; ESI-MS: m/z=421[M+H] ⁺ .

Preparation Example 72, N-(N-(tert-butoxycarbonyl)-O-methyl-L-seryl)-O-(5-hexen-1-yl)-L-serine benzyl ester (6d )

Using 5d (3.8 g, 12 mmol) as raw material, the synthesis and post-treatment were the same as those in Preparation Example 69, and 2.8 g of light yellow oily liquid was obtained with a yield of 59%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.41–7.29(m,6H),5.83–5.72(m,1H),5.42(s,1H),5.26–5.11(m,2H),5.03–4.92(m ,2H),4.73(dt,J=8.0,3.0Hz,1H),4.28(s,1H),3.88(dd,J=9.5,3.0Hz,1H),3.77(dd,J=9.0,4.0Hz, 1H), 3.64(dd, J＝9.5, 3.0Hz, 1H), 3.46–3.32(m, 6H), 2.07–1.99(m, 2H), 1.56–1.48(m, 2H), 1.45(s, 9H) ,1.41–1.33(m,2H)ppm; ESI-MS: m/z=479[M+H] ⁺ .

Preparation Example 73, N-(N-(tert-butoxycarbonyl)-O-methyl-L-seryl)-O-(2-(2-(benzyloxy)ethoxy)ethyl)- L-serine methyl ester (6e)

Using 5e (4.0 g, 12 mmol) as the raw material, the synthesis and post-treatment were the same as those in Preparation Example 69 to obtain 2.5 g of light yellow oily liquid with a yield of 51%. ¹ H NMR (500MHz, CDCl ₃ )δ7.39–7.27(m,6H),5.50(d,J=4.5Hz,1H),4.74–4.66(m,1H),4.57(s,2H),4.34– 4.25(m,1H),3.96(dd,J＝10.0,3.5Hz,1H),3.79(dd,J＝9.0,4.0Hz,1H),3.75–3.70(m,4H),3.67–3.59(m, 8H), 3.48(dd, J＝9.0, 6.5Hz, 1H), 3.37(s, 3H), 1.46(s, 9H)ppm; ESI-MS: m/z＝499[M+H] ⁺ .

Preparation Example 74, (S)-13-((S)-2-((tert-butoxycarbonyl)amino)-3-methoxypropionylamino)-1-phenyl-2,5,8,11 -tetraoxatetradecane-14-oic acid methyl ester (6f)

Using 5f (4.5 g, 12 mmol) as the raw material, the synthesis and post-treatment were the same as those in Preparation Example 69, and 2.3 g of light yellow oily liquid was obtained with a yield of 43%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.40–7.31(m,5H),7.31–7.27(m,1H),5.48(d,J=4.0Hz,1H),4.70(dt,J=7.0,3.0 Hz,1H),4.57(s,2H),4.33–4.25(m,1H),3.94(dd,J=10.0,3.5Hz,1H),3.80(dd,J=9.5,3.5Hz,1H),3.74 (s,3H),3.71(dd,J=9.5,3.0Hz,1H),3.69–3.58(m,12H),3.49(dd,J=9.2,6.6Hz,1H),3.38(s,3H), 1.46(s,9H)ppm; ESI-MS: m/z=543[M+H] ⁺ .

Preparation Example 75, N-((S)-2-((tert-butoxycarbonyl)amino)-4-morpholino-4-oxobutanoyl)-O-allyl-L-serine benzyl ester (6g)

Reactant 4e (0.80g, 2.0mmol) was dissolved in 4ml of methanol, Pd/C (0.08g) was added, and reacted at room temperature for 3h after hydrogen replacement. Pd/C was removed by suction filtration, the solvent was evaporated under reduced pressure, and the obtained product was directly used in the next reaction.

The raw material (S)-2-((tert-butoxycarbonyl)amino)-4-morpholino-4-oxobutanoic acid (0.19g, 0.63mmol) was dissolved in 5mL of dichloromethane, and 1-hydroxy Benzotriazole (0.1 g, 0.76 mmol) and N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (0.22 g, 1.1 mmol) were reacted at room temperature for half an hour. Cool in an ice bath to 0°C, add O-allyl-Ser-OBn hydrochloride (5a, 0.21g, 0.76mmol) and DIPEA (0.31ml, 1.9mmol), and react at room temperature for 3 hours. Add 5 mL of saturated sodium bicarbonate for dilution, separate the organic layer, wash with saturated brine (5 mL x 2), dry over anhydrous sodium sulfate, evaporate the solvent, and separate by column chromatography to obtain 0.22 g of a colorless oily liquid with a yield of 67%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.65(d, J=8.0Hz, 1H), 7.38–7.29(m, 5H), 6.02(d, J=8.0Hz, 1H), 5.84–5.74(m, 1H),5.28–5.10(m,4H),4.71(dt,J＝8.5,3.5Hz,1H),4.68–4.62(m,1H),4.01–3.95(m,1H),3.95–3.88(m, 2H),3.71–3.60(m,5H),3.60–3.53(m,2H),3.49–3.36(m,2H),3.13(dd,J＝16.5,2.0Hz,1H),2.57(dd,J＝ 16.5,6.0Hz,1H), 1.45(s,9H)ppm; ESI-MS: m/z=520[M+H] ⁺ .

Preparation Example 76, N-((S)-2-((tert-butoxycarbonyl)amino)-5-morpholino-5-oxopentanoyl)-O-allyl-L-serine benzyl ester (6h)

Reactant 4f (0.80g, 2.0mmol) was dissolved in 4ml of methanol, Pd/C (0.08g) was added, and reacted at room temperature for 3h after hydrogen replacement. Pd/C was removed by suction filtration, the solvent was evaporated under reduced pressure, and the obtained product was directly used in the next reaction.

The raw material (S)-2-((tert-butoxycarbonyl)amino)-5-morpholino-5-oxopentanoic acid (0.20 g, 0.63 mmol) in the previous step was dissolved in 5 mL of dichloromethane, and 1-hydroxy Benzotriazole (0.10 g, 0.76 mmol) and N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (0.22 g, 1.1 mmol) were reacted at room temperature for half an hour. Cool in an ice bath to 0°C, add O-allyl-Ser-OBn hydrochloride (5a, 0.21g, 0.76mmol) and DIPEA (0.31ml, 1.9mmol), and react at room temperature for 3 hours. Add 5 mL of saturated sodium bicarbonate for dilution, separate the organic layer, wash with saturated brine (5 mL x 2), dry over anhydrous sodium sulfate, evaporate the solvent, and separate by column chromatography to obtain 0.33 g of a colorless oily liquid with a yield of 98%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.56(d, J=7.0Hz, 1H), 7.40–7.29(m, 5H), 5.85–5.73(m, 1H), 5.56(d, J=6.3Hz, 1H),5.28–5.09(m,4H),4.79–4.73(m,1H),4.27–4.18(m,1H),4.01–3.88(m,3H),3.78–3.55(m,7H),3.49– 3.38(m,2H),2.61–2.51(m,1H),2.50–2.41(m,1H),2.19–2.10(m,1H),1.98–1.89(m,1H),1.43(s,9H)ppm ; ESI-MS: m/z=534[M+H] ⁺ .

Preparation Example 77, N-(N ² -(tert-butoxycarbonyl)-N ⁴ -cyclopropyl-L-asparaginyl)-O-allyl-L-serine benzyl ester (6i)

4g of the reactant (0.72g, 2.0mmol) was dissolved in 4ml of methanol, Pd/C (0.08g) was added, hydrogen was replaced and reacted at room temperature for 3h. Pd/C was removed by suction filtration, the solvent was evaporated under reduced pressure, and the obtained product was directly used in the next reaction.

The raw material N ² -(tert-butoxycarbonyl)-N ⁴ -cyclopropyl-L-asparagine (0.27g, 1.0mmol) was dissolved in 5mL of dichloromethane, and 1-hydroxybenzotriazole ( 0.16g, 1.2mmol) and N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (0.35g, 1.8mmol), react at room temperature for half an hour. Cool in an ice bath to 0°C, add O-allyl-Ser-OBn hydrochloride (5a, 0.30g, 1.1mmol) and DIPEA (0.50ml, 3.0mmol), and react at room temperature for 3 hours. Add 5 mL of saturated sodium bicarbonate for dilution, separate the organic layer, wash with saturated brine (5 mL x 2), dry over anhydrous sodium sulfate, evaporate the solvent under reduced pressure, and separate by column chromatography to obtain 0.35 g of white solid with a yield of 71%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.60(d, J=7.0Hz, 1H), 7.40–7.29(m, 6H), 6.15(d, J=5.5Hz, 1H), 5.83–5.72(m, 1H),5.27–5.09(m,4H),4.75–4.66(m,1H),4.56–4.44(m,1H),4.01–3.86(m,3H),3.63(dd,J＝9.5,3.5Hz, 1H),2.80–2.61(m,2H),2.49(dd,J＝15.0,6.0Hz,1H),1.44(s,9H),0.77–0.68(m,2H),0.53–0.44(m,2H) ppm; ESI-MS: m/z=499[M+H] ⁺ .

Preparation Example 78, N-((tert-butoxycarbonyl)-L-leucyl)-O-allyl-L-serine benzyl ester (6j)

Starting material Boc-leucine (0.28g, 1.2mmol) was dissolved in 5mL of dichloromethane, 1-hydroxybenzotriazole (0.19g, 1.4mmol) and N-(3-dimethylaminopropyl)-N '-Ethylcarbodiimide hydrochloride (0.42g, 2.2mmol), react at room temperature for half an hour. Cool in an ice bath to 0°C, add O-allyl-Ser-OBn hydrochloride (5a, 0.39g, 1.4mmol) and DIPEA (0.59ml, 3.6mmol), and react at room temperature for 3 hours. Add 5 mL of saturated sodium bicarbonate for dilution, separate the organic layer, wash with saturated brine (5 mL x 2), dry over anhydrous sodium sulfate, evaporate the solvent under reduced pressure, and separate by column chromatography to obtain 0.45 g of a colorless oily liquid with a yield of 83%. . ¹ H NMR (500MHz, CDCl ₃ ) δ7.39–7.30(m,5H),6.73(d,J=8.0Hz,1H),5.82–5.71(m,1H),5.31–5.11(m,4H), 4.91(d,J=7.0Hz,1H),4.77(dt,J=8.5,3.0Hz,1H),4.20–4.13(m,1H),3.99–3.86(m,3H),3.64(dd,J= 9.5,3.0Hz,1H),1.68–1.59(m,2H),1.51–1.39(m,10H),0.96–0.89(m,6H)ppm; ESI-MS: m/z=449[M+H] ⁺ .

Preparation Example 79, N-((tert-butoxycarbonyl)-L-valyl)-O-allyl-L-serine benzyl ester (6k)

Using Boc-valine (0.29g, 1.2mmol) as raw material, the synthesis and post-treatment were the same as those in Preparation Example 78, and 0.48g of light yellow oily liquid was obtained, with a yield of 92%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.40–7.30(m,5H),6.59(d,J=7.0Hz,1H),5.81–5.71(m,1H),5.30–5.07(m,5H), 4.79(dt, J＝8.0, 3.0Hz, 1H), 4.04–3.86(m, 4H), 3.64(dd, J＝9.5, 3.0Hz, 1H), 2.18–2.07(m, 1H), 1.44(s, 9H), 0.97(d, J=6.5Hz, 3H), 0.91(d, J=6.5Hz, 3H) ppm; ESI-MS: m/z=435[M+H] ⁺ .

Preparation Example 80, N-(O-methyl-L-seryl)-O-allyl-L-serine benzyl ester hydrochloride (7a)

Reactant 6a (2.2 g, 5.0 mmol) was dissolved in 5 mL CH ₂ Cl ₂ , cooled to 0° C. in an ice bath, 2N HCl-saturated ethyl acetate solution (10 mL) was added, and reacted at room temperature for 2 hours. The solvent was distilled off from the reaction solution under reduced pressure to obtain a light yellow oily liquid, which was directly used in the next reaction.

Preparation Example 81, N-(O-methyl-L-seryl)-O-allyl-L-serine benzyl ester hydrochloride (7b)

Using 6b as raw material, the synthesis and post-treatment were the same as those in Preparation Example 80, and the obtained product was directly used in the next reaction.

Preparation Example 82, (S)-2-((S)-2-Amino-3-methoxypropionamido)-5-hexenoic acid benzyl ester hydrochloride (7c)

Using 6c as raw material, the synthesis and post-treatment were the same as those in Preparation Example 80, and the obtained product was directly used in the next reaction.

Preparation Example 83, N-(O-methyl-L-seryl)-O-(hex-5-en-1-yl)-L-serine benzyl ester hydrochloride (7d)

Using 6d as raw material, the synthesis and post-treatment were the same as those in Preparation Example 80, and the obtained product was directly used in the next reaction.

Preparation Example 84, N-(N-(tert-butoxycarbonyl)-O-methyl-L-seryl)-O-(2-(2-hydroxyethoxy)ethyl)-L-serine Methyl ester (7e)

The reactant 6e (0.72g, 1.4mmol) was dissolved in 4ml of methanol and 2ml of ethyl acetate, added Pd/C (0.12g), and reacted at 50°C for 3h after hydrogen replacement. Pd/C was removed by suction filtration, and the solvent was distilled off under reduced pressure to obtain 0.59 g of light yellow oily liquid with a yield of 100%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.43 (d, J=8.0Hz, 1H), 5.64 (d, J=4.0Hz, 1H), 4.73 (dt, J=8.0, 3.0Hz, 1H), 4.35 –4.28(m,1H),3.98(dd,J＝10.0,3.0Hz,1H),3.81(dd,J＝9.0,4.0Hz,1H),3.78–3.70(m,6H),3.66–3.57(m ,6H), 3.50(dd,J＝9.5,6.0Hz,1H), 3.38(s,3H), 1.45(s,9H)ppm; ESI-MS: m/z＝409[M+H] ⁺ .

Preparation Example 85, N-(N-(tert-butoxycarbonyl)-O-methyl-L-seryl)-O-(2-(2-(2-hydroxyethoxy)ethoxy) Ethyl)-L-serine methyl ester (7f)

Using 6f (0.78g, 1.4mmol) as the raw material, the synthesis and post-treatment were the same as in Preparation Example 84, and 0.60g of light yellow oily liquid was obtained with a yield of 95%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.83(d, J=7.0Hz, 1H), 5.68(d, J=6.0Hz, 1H), 4.79–4.70(m, 1H), 4.42–4.32(m, ESI -MS: m/z＝453[M+H] ⁺ .

Preparation Example 86, N-((S)-2-amino-4-morpholino-4-oxobutanoyl)-O-allyl-L-serine benzyl ester hydrochloride (7 g)

Using 6 g as raw material, the synthesis and post-treatment were the same as those in Preparation Example 80, and the obtained product was directly used in the next reaction.

Preparation Example 87, N-((S)-2-amino-5-morpholino-5-oxopentanoyl)-O-allyl-L-serine benzyl ester hydrochloride (7h)

Using 6h as the raw material, the synthesis and post-treatment were the same as those in Preparation Example 80, and the obtained product was directly used in the next reaction.

Preparation Example 88, N-(N ⁴ -cyclopropyl-L-asparaginyl)-O-allyl-L-serine benzyl ester hydrochloride (7i)

Using 6i as raw material, the synthesis and post-treatment were the same as those in Preparation Example 80, and the obtained product was directly used in the next reaction.

Preparation Example 89, N-(L-leucyl)-O-allyl-L-serine benzyl ester hydrochloride (7j)

Using 6j as the raw material, the synthesis and post-treatment were the same as those in Preparation Example 80, and the obtained product was directly used in the next reaction.

Preparation Example 90, N-(L-valyl)-O-allyl-L-serine benzyl ester hydrochloride (7k)

Using 6k as raw material, the synthesis and post-treatment were the same as those in Preparation Example 80, and the obtained product was directly used in the next reaction.

Preparation Example 91, N-(N-(2-(allyloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester (8a)

The raw material 2-(allyloxy)benzoic acid (2a, 0.18g, 1.0mmol) was placed in a 10ml reaction flask, and 4mL of anhydrous CH ₂ Cl ₂ was added to dissolve it, followed by the addition of 1-hydroxybenzotriazole (0.16g , 1.2mmol) and N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (0.35g, 1.8mmol), react at room temperature for half an hour. Subsequently, cool to 0°C, add O-methyl-Ser-O-allyl-Ser-OBn hydrochloride (7a, 52.0mg, 1.2mmol) and DIPEA (0.50ml, 3.0mmol), and react at room temperature for 3 hours . The reaction solution was diluted by adding 5 mL of saturated aqueous sodium bicarbonate solution, the organic layer was separated, washed with saturated brine (5 mL×2), dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure. Column chromatography separated to obtain 0.39 g of a colorless oily liquid, yield 78%.

¹ H NMR (500MHz, CDCl ₃ ) δ8.80(d, J=6.5Hz, 1H), 8.19(d, J=8.0Hz, 1H), 7.48–7.40(m, 2H), 7.39–7.29(m, 5H),7.11–7.04(m,1H),6.97(d,J＝8.3Hz,1H),6.24–6.11(m,1H),5.80–5.70(m,1H),5.48–5.30(m,2H) ,5.27–5.08(m,4H),4.87–4.77(m,2H),4.75–4.67(m,2H),3.97–3.87(m,4H),3.71–3.65(m,1H),3.56–3.49( m,1H), 3.38(s,3H)ppm; ESI-MS: m/z＝497[M+H] ⁺ .

Preparation Example 92, N-(N-(2-(3-buten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L- Benzyl Serine (8b)

Using 2-(3-buten-1-yloxy)benzoic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.38 g of a colorless oily liquid with a yield of 77%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.79(d, J=6.5Hz, 1H), 8.20(d, J=7.5Hz, 1H), 7.49–7.39(m, 2H), 7.37–7.28(m, 5H),7.11–7.04(m,1H),6.97(d,J＝8.5Hz,1H),5.96–5.85(m,1H),5.80–5.68(m,1H),5.27–5.07(m,6H) ,4.90–4.82(m,1H),4.82–4.77(m,1H),4.18(t,J＝6.5Hz,2H),3.95–3.87(m,4H),3.72–3.66(m,1H),3.57 –3.50(m,1H),3.39(s,3H),2.77–2.63(m,2H)ppm; ESI-MS: m/z=511[M+H] ⁺ .

Preparation Example 93, N-(N-(2-(4-penten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L- Benzyl Serine (8c)

Using 2-(4-penten-1-yloxy)benzoic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.32 g of a colorless oily liquid with a yield of 61%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.85(d, J=6.5Hz, 1H), 8.21(d, J=8.0Hz, 1H), 7.48–7.42(m, 1H), 7.40(d, J= 8.0Hz, 1H), 7.36–7.29(m, 5H), 7.09–7.03(m, 1H), 6.96(d, J＝8.0Hz, 1H), 5.89–5.79(m, 1H), 5.79–5.69(m ,1H),5.27–4.98(m,6H),4.88–4.82(m,1H),4.81–4.77(m,1H),4.16–4.09(m,2H),3.97–3.87(m,4H),3.70 –3.65(m,1H),3.55–3.49(m,1H),3.38(s,3H),2.30–2.22(m,2H),2.11–1.97(m,2H)ppm; ESI-MS: m/z ＝525[M+H] ⁺ .

Preparation Example 94, N-(N-(2-(5-hexen-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L- Benzyl serine (8d)

Using 2-(5-hexen-1-yloxy)benzoic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.35 g of a colorless oily liquid with a yield of 65%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.85 (d, J=6.5Hz, 1H), 8.20 (dd, J=8.0, 2.0Hz, 1H), 7.44 (ddd, J=8.5, 7.5, 2.0Hz, 1H), 7.40(d, J＝8.0Hz, 1H), 7.37–7.28(m, 5H), 7.09–7.03(m, 1H), 6.96(d, J＝8.0Hz, 1H), 5.88–5.70(m ,2H),5.26–5.13(m,3H),5.12–5.09(m,1H),5.05–4.99(m,1H),4.99–4.93(m,1H),4.84(td,J＝7.0,4.0Hz ,1H),4.79(dt,J＝8.0,3.0Hz,1H),4.17–4.08(m,2H),3.97–3.87(m,4H),3.67(dd,J＝9.5,3.0Hz,1H), ESI -MS: m/z＝539[M+H] ⁺ .

Preparation Example 95, N-(N-(2-(6-hepten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L- Benzyl Serine (8e)

Using 2-(6-hepten-1-yloxy)benzoic acid as a raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.33 g of a colorless oily liquid with a yield of 60%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.86(d, J=6.5Hz, 1H), 8.20(dd, J=8.0, 2.0Hz, 1H), 7.47–7.42(m, 1H), 7.40(d, J=8.0Hz,1H),7.38–7.28(m,5H),7.09–7.02(m,1H),6.96(d,J=8.0Hz,1H),5.85–5.69(m,2H),5.27–5.13 (m,3H),5.13–5.08(m,1H),5.03–4.97(m,1H),4.97–4.92(m,1H),4.84(td,J＝7.0,4.0Hz,1H),4.79(dt ,J=8.0,3.0Hz,1H),4.15–4.09(m,2H),3.96–3.87(m,4H),3.67(dd,J=9.5,3.5Hz,1H),3.53(dd,J=9.0 ,7.0Hz,1H),3.39(s,3H),2.11-2.05(m,2H),1.99–1.91(m,2H),1.53–1.43(m,4H)ppm; ESI-MS: m/z＝ 553[M+H] ⁺ .

Preparation Example 96, N-(N-(2-(7-octen-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L- Benzyl Serine (8f)

Using 2-(7-octen-1-yloxy)benzoic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.31 g of a colorless oily liquid with a yield of 55%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.85 (d, J=7.0Hz, 1H), 8.20 (dd, J=8.0, 2.0Hz, 1H), 7.44 (ddd, J=8.0, 7.5, 2.0Hz, 1H), 7.39(d, J＝8.0Hz, 1H), 7.36–7.30(m, 6H), 7.08–7.03(m, 1H), 6.96(d, J＝8.0Hz, 1H), 5.84–5.69(m ,2H),5.26–5.13(m,4H),5.12–5.08(m,1H),5.02–4.96(m,1H),4.95–4.91(m,1H),4.84(td,J＝7.0,4.0Hz ,1H),4.79(dt,J=8.0,3.0Hz,1H),3.95–3.88(m,4H),3.67(dd,J=9.5,3.5Hz,1H),3.53(dd,J=9.0,7.0 ESI -MS: m/z＝567[M+H] ⁺ .

Preparation Example 97, N-(N-(5-chloro-2-(4-penten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl Benzyl-L-serine benzyl ester (8g)

Using 5-chloro-2-(4-penten-1-yloxy)benzoic acid as a raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.41 g of a colorless oily liquid with a yield of 73%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.80(d, J=6.5Hz, 1H), 8.17(d, J=3.0Hz, 1H), 7.40–7.30(m, 7H), 6.90(d, J= 9.0Hz,1H),5.89–5.69(m,2H),5.26–5.22(m,1H),5.21–5.09(m,3H),5.09–5.03(m,1H),5.03–4.98(m,1H) ,4.84–4.76(m,2H),4.13–4.09(m,2H),3.95–3.87(m,4H),3.67(dd,J＝9.5,3.5Hz,1H),3.52(dd,J＝9.0, 7.0Hz,1H),3.37(s,3H),2.28–2.22(m,2H),2.07–2.00(m,2H)ppm; ESI-MS: m/z=559[M+H] ⁺ .

Preparation Example 98, N-(N-(4-chloro-2-(4-penten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl Benzyl-L-serine benzyl ester (8h)

Using 4-chloro-2-(4-penten-1-yloxy)benzoic acid as a raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.39 g of a colorless oily liquid with a yield of 70%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.70 (d, J=7.0Hz, 1H), 8.21 (dd, J=8.5, 7.0Hz, 1H), 7.37 (d, J=9.0Hz, 1H), 7.35 –7.31(m,5H),6.76(ddd,J＝9.0,7.5,2.5Hz,1H),6.67(dd,J＝10.5,2.5Hz,1H),5.88–5.70(m,2H),5.26–5.05 (m,5H),5.04–5.00(m,1H),4.84–4.76(m,2H),4.12–4.08(m,2H),3.96–3.87(m,4H),3.67(dd,J＝9.5, 3.5Hz,1H),3.52(dd,J＝9.0,7.0Hz,1H),3.38(s,3H),2.29–2.23(m,2H),2.10–2.01(m,2H)ppm; ESI-MS: m/z＝559[M+H] ⁺ .

Preparation Example 99, N-(N-(4-fluoro-2-(4-penten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl Benzyl-L-serine benzyl ester (8i)

Using 4-fluoro-2-(4-penten-1-yloxy)benzoic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.40 g of a colorless oily liquid with a yield of 74%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.72 (d, J = 6.5Hz, 1H), 8.13 (d, J = 8.5Hz, 1H), 7.37 (d, J = 8.5Hz, 1H), 7.35–7.31 (m,5H),7.04(dd,J＝8.5,2.0Hz,1H),6.95(d,J＝2.0Hz,1H),5.88–5.70(m,2H),5.26–5.05(m,5H), 5.04–5.00(m,1H),4.84–4.76(m,2H),4.11(t,J＝6.5Hz,2H),3.97–3.87(m,4H),3.67(dd,J＝9.5,3.0Hz, 1H), 3.52(dd, J=9.0, 7.0Hz, 1H), 3.38(s, 3H), 2.29–2.23(m, 2H), 2.09–2.01(m, 2H)ppm; ESI-MS: m/z ＝543[M+H] ⁺ .

Preparation Example 100, N-(N-(4-methyl-2-(4-penten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-ene Propyl-L-serine benzyl ester (8j)

Using 4-methyl-2-(4-penten-1-yloxy)benzoic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.44 g of a colorless oily liquid with a yield of 82%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.80 (d, J=7.0Hz, 1H), 8.08 (d, J=8.0Hz, 1H), 7.39 (d, J=8.0Hz, 1H), 7.36–7.29 (m,5H),6.87(d,J＝8.0Hz,1H),6.76(s,1H),5.90–5.79(m,1H),5.79–5.69(m,1H),5.26–5.04(m,5H ),5.03–4.98(m,1H),4.84(td,J＝7.0,4.0Hz,1H),4.79(dt,J＝8.0,3.0Hz,1H),4.15–4.09(m,2H),3.96– 3.87(m,4H),3.67(dd,J=9.5,3.0Hz,1H),3.52(dd,J=9.0,6.5Hz,1H),3.37(s,3H),2.38(s,3H),2.29 –2.23(m,2H),2.06–2.02(m,2H)ppm; ESI-MS: m/z=539[M+H] ⁺ .

Preparation Example 101, N-(N-(4-methoxy-2-(4-penten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O- Allyl-L-serine benzyl ester (8k)

Using 4-methoxy-2-(4-penten-1-yloxy)benzoic acid as a raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.41 g of a colorless oily liquid with a yield of 76%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.71(d, J=6.5Hz, 1H), 8.16(d, J=8.5Hz, 1H), 7.40(d, J=8.0Hz, 1H), 7.37–7.28 (m,5H),6.58(dd,J=9.0,2.0Hz,1H),6.46(d,J=2.5Hz,1H),5.88–5.69(m,2H),5.26–5.03(m,5H), 5.02–4.98(m,1H),4.83(td,J=7.0,4.0Hz,1H),4.78(dt,J=8.0,3.0Hz,1H),4.09(t,J=6.5Hz,2H),3.96 –3.87(m,4H),3.84(s,3H),3.67(dd,J=9.5,3.0Hz,1H),3.52(dd,J=9.5,6.5Hz,1H),3.37(s,3H), 2.30–2.20(m,2H),2.09–1.98(m,2H)ppm; ESI-MS: m/z=555[M+H] ⁺ .

Preparation Example 102, N-(N-(2-methyl-4-(4-penten-1-yloxy)thiazole-5-formyl)-O-methyl-L-seryl)- O-allyl-L-serine methyl ester (8l)

The raw material 2-methyl-4-(4-penten-1-yloxy)thiazole-5-carboxylic acid (2l, 0.16g, 0.70mmol) was placed in _a 10ml reaction flask, and 3mL of anhydrous _CH2Cl2 was added to dissolve it , followed by the addition of 1-hydroxybenzotriazole (0.11 g, 0.84 mmol) and N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (0.24 g, 1.3 mmol) , half an hour at room temperature. Subsequently, cooled to 0°C, O-methyl-Ser-O-allyl-Ser-OMe hydrochloride (7b, 0.25mg, 0.84mmol) and DIPEA (0.35ml, 2.1mmol) were added, and reacted at room temperature for 3 hours . The reaction solution was diluted by adding 5 mL of saturated aqueous sodium bicarbonate solution, the organic layer was separated, washed with saturated brine (5 mL×2), dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure. Column chromatography separated to obtain 0.25 g of a colorless oily liquid, yield 76%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.93(d, J=7.0Hz, 1H), 7.34(d, J=8.0Hz, 1H), 5.89–5.76(m, 2H), 5.25–5.17(m, 1H),5.17–5.12(m,1H),5.10–5.04(m,1H),5.02–4.97(m,1H),4.76–4.68(m,2H),4.53–4.44(m,2H),3.99– 3.95(m,2H),3.93(dd,J=9.0,4.0Hz,1H),3.89(dd,J=9.5,3.0Hz,1H),3.76(s,3H),3.66(dd,J=9.5, 3.0Hz,1H),3.54(dd,J＝9.0,7.0Hz,1H),3.43(s,3H),2.61(s,3H),2.28–2.21(m,2H),1.97–1.90(m,2H )ppm; ESI-MS: m/z=470[M+H] ⁺ .

Preparation Example 103, N-(O-methyl-N-(2-(4-penten-1-yloxy)thiophene-3-formyl)-L-seryl)-O-allyl -L-serine methyl ester (8m)

Using 2-(4-penten-1-yloxy)thiophene-3-carboxylic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 102 to obtain 0.27 g of a colorless oily liquid with a yield of 84%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.13(d, J=7.0Hz, 1H), 7.39(d, J=5.5Hz, 1H), 7.36(d, J=8.5Hz, 1H), 6.83(d ,J＝5.5Hz,1H),5.88–5.76(m,2H),5.24–5.18(m,1H),5.16–5.12(m,1H),5.10–5.04(m,1H),5.03–4.98(m ,1H),4.79–4.71(m,2H),4.18(td,J＝6.5,2.0Hz,2H),3.98–3.94(m,3H),3.89(dd,J＝9.5,3.0Hz,1H), 3.75(s,3H),3.66(dd,J=9.5,3.5Hz,1H),3.55(dd,J=9.0,7.0Hz,1H),3.43(s,3H),2.30–2.24(m,2H) ,2.01–1.93(m,2H)ppm; ESI-MS: m/z=455[M+H] ⁺ .

Preparation Example 104, N-(O-methyl-N-(3-(4-penten-1-yloxy)pyridinecarbonyl)-L-seryl)-O-allyl-L- Benzyl serine (8n)

Using 3-(4-penten-1-yloxy)picolinic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.50 g of a colorless oily liquid with a yield of 95%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.58 (d, J=7.0Hz, 1H), 8.29 (dd, J=4.0, 1.5Hz, 1H), 7.42 (d, J=8.0Hz, 1H), 7.40 –7.28(m,7H),5.88–5.71(m,2H),5.26–5.10(m,4H),5.10–4.97(m,2H),4.83(td,J＝7.0,4.0Hz,1H),4.78 (dt,J=8.0,3.5Hz,1H),4.10(t,J=6.5Hz,2H),3.95–3.89(m,4H),3.68(dd,J=9.5,3.5Hz,1H),3.52( dd,J＝9.0,7.5Hz,1H), 3.37(s,3H), 2.31–2.24(m,2H), 2.06–1.99(m,2H)ppm; ESI-MS: m/z＝526[M+ H] ⁺ .

Preparation Example 105, N-(O-methyl-N-(2-(4-penten-1-yloxy)nicotinic acid formyl)-L-seryl)-O-allyl-L - Benzyl serine (8o)

Using 4-penten-1-yl 2-(4-penten-1-yloxy)nicotinic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.31 g of a colorless oily liquid with a yield of 59% . ¹ H NMR (500MHz, CDCl ₃ ) δ8.87 (d, J=6.5Hz, 1H), 8.50 (dd, J=7.5, 2.0Hz, 1H), 8.27 (dd, J=5.0, 2.0Hz, 1H) ,7.39–7.31(m,6H),7.04(dd,J＝7.5,5.0Hz,1H),5.91–5.81(m,1H),5.79–5.70(m,1H),5.27–5.04(m,5H) ,5.01–4.97(m,1H),4.84–4.78(m,2H),4.58–4.46(m,2H),3.96–3.87(m,4H),3.68(dd,J＝9.5,3.5Hz,1H) ,3.53(dd,J=9.0,7.0Hz,1H),3.39(s,3H),2.29–2.22(m,2H),2.05–1.98(m,2H)ppm; ESI-MS: m/z=526 [M+H] ⁺ .

Preparation Example 106, N-(O-methyl-N-(1-methyl-5-(4-penten-1-yloxy)-1H-pyrazole-4-formyl)-L-silk Aminoacyl)-O-allyl-L-serine benzyl ester (8p)

Using 1-methyl-5-(4-penten-1-yloxy)-1H-pyrazole-4-carboxylic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.36 g of a colorless oily liquid. Yield 68%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.74(s, 1H), 7.42(d, J=8.0Hz, 1H), 7.36–7.32(m, 5H), 7.02(d, J=6.5Hz, 1H) ,5.87–5.70(m,2H),5.28–5.12(m,4H),5.09–5.00(m,2H),4.78–4.69(m,2H),4.27–4.18(m,2H),3.94–3.85( m,5H),3.70(s,3H),3.67(dd,J＝9.5,3.0Hz,1H),3.47(dd,J＝9.5,7.5Hz,1H),3.37(s,2H),2.28–2.21 (m,2H),1.99–1.91(m,2H)ppm; ESI-MS: m/z=529[M+H] ⁺ .

Preparation Example 107, N-(N-(3-(hept-6-en-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L - Benzyl serine (8q)

Using 3-(hept-6-en-1-yloxy)benzoic acid as a raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.50 g of a colorless oily liquid with a yield of 90%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.51(d, J=8.0Hz, 1H), 7.39–7.30(m, 8H), 7.12(d, J=6.0Hz, 1H), 7.06–7.02(m, 1H),5.87–5.72(m,2H),5.29–5.12(m,4H),5.05–4.99(m,1H),4.97–4.93(m,1H),4.80–4.72(m,2H),4.00( t,J=6.5Hz,2H),3.96–3.88(m,4H),3.68(dd,J=9.5,3.0Hz,1H),3.49(t,J=8.5Hz,1H),3.39(s,3H ), 2.12–2.06(m,2H), 1.83–1.76(m,2H), 1.52–1.43(m,4H)ppm; ESI-MS: m/z＝553[M+H] ⁺ .

Preparation Example 108, N-(N-(3-fluoro-2-(pent-4-en-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-ene Propyl-L-serine benzyl ester (8r)

Using 3-fluoro-2-(4-penten-1-yloxy)benzoic acid as a raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.33 g of a colorless oily liquid with a yield of 61%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.78 (d, J=6.5Hz, 1H), 7.90 (dt, J=8.0, 1.5Hz, 1H), 7.37–7.29 (m, 6H), 7.25–7.19( m,1H),7.10(td,J＝8.0,4.5Hz,1H),5.88–5.70(m,2H),5.27–5.10(m,4H),5.08–5.02(m,1H),5.01–4.97( m,1H),4.84–4.76(m,2H),4.28–4.16(m,2H),3.97–3.88(m,4H),3.68(dd,J＝9.5,3.0Hz,1H),3.54(dd, J=9.0,7.0Hz,1H),3.38(s,3H),2.26–2.20(m,2H),2.01–1.94(m,2H)ppm; ESI-MS: m/z=543[M+H] ⁺ .

Preparation Example 109, N-(N-(3-methoxy-2-(pent-4-en-1-yloxy)benzoyl)-O-methyl-L-seryl)-O -Allyl-L-serine benzyl ester (8s)

Using 3-methoxy-2-(pent-4-en-1-yloxy)benzoic acid as raw material, the synthesis and post-treatment were the same as those in Preparation Example 91 to obtain 0.36 g of a colorless oily liquid with a yield of 65%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.93(d, J=7.0Hz, 1H), 7.70(dd, J=8.0, 1.5Hz, 1H), 7.39–7.28(m, 6H), 7.13(t, J＝8.0Hz, 1H), 7.05(dd, J＝8.0, 1.5Hz, 1H), 5.90–5.80(m, 1H), 5.78–5.69(m, 1H), 5.26–5.13(m, 3H), 5.13 –5.08(m,1H),5.08–5.02(m,1H),5.00–4.96(m,1H),4.85–4.76(m,2H),4.11–4.04(m,2H),3.97–3.86(m, 7H), 3.67(dd, J＝9.5, 3.5Hz, 1H), 3.54(dd, J＝9.0, 6.5Hz, 1H), 3.37(s, 3H), 2.26–2.20(m, 2H), 1.99–1.91 (m,2H)ppm; ESI-MS: m/z=555[M+H] ⁺ .

Preparation Example 110, N-(N-(5-allyl-1-methyl-1H-pyrazole-4-carbonyl)-O-methyl-L-seryl)-O-(5-hexyl En-1-yl)-L-serine benzyl ester (8t)

Using 5-allyl-1-methyl-1H-pyrazole-4-carboxylic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.36 g of a colorless oily liquid with a yield of 68%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.72(s, 1H), 7.47(d, J=8.0Hz, 1H), 7.38–7.29(m, 5H), 6.72(d, J=6.0Hz, 1H) ,5.89(ddt,J＝17.0,10.0,6.0Hz,1H),5.81–5.72(m,1H),5.26–5.21(m,1H),5.18–5.09(m,2H),5.02–4.92(m, 3H), 4.74(dt, J＝8.0, 3.0Hz, 1H), 4.69(ddd, J＝8.0, 6.5, 4.0Hz, 1H), 3.89(dd, J＝9.5, 3.0Hz, 1H), 3.85(dd ,J＝9.0,4.0Hz,1H),3.81–3.77(m,5H),3.65(dd,J＝9.5,3.5Hz,1H),3.48–3.43(m,1H),3.42–3.34(m,5H ), 2.06–1.97(m,2H), 1.54–1.45(m,2H), 1.42–1.31(m,2H)ppm; ESI-MS: m/z＝527[M+H] ⁺ .

Preparation Example 111, N-(N-(1-(pent-4-en-1-yl)-1H-pyrazole-5-carbonyl)-O-methyl-L-seryl)-O-ene Propyl-L-serine benzyl ester (8u)

Using 1-(pent-4-en-1-yl)-1H-pyrazole-5-carboxylic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.33 g of a colorless oily liquid with a yield of 66%. ¹ H NMR (500MHz, CDCl ₃ )δ7.50–7.45(m,2H),7.39–7.31(m,5H),6.96(d,J=6.5Hz,1H),6.59(d,J=2.0Hz, 1H),5.85–5.72(m,2H),5.29–5.12(m,4H),5.06–4.94(m,2H),4.76(dt,J＝8.5,3.0Hz,1H),4.67(ddd,J＝ 8.0,6.0,4.0Hz,1H),4.60–4.52(m,2H),3.99–3.90(m,3H),3.86(dd,J＝9.0,4.0Hz,1H),3.68(dd,J＝9.5, 3.5Hz,1H),3.47(t,J＝8.5Hz,1H),3.39(s,3H),2.10–2.05(m,2H),1.99–1.92(m,2H)ppm; ESI-MS:m/ z＝499[M+H] ⁺ .

Preparation Example 112, N-(N-(1-allyl-1H-imidazole-2-carbonyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester ( 8v)

Using 1-allyl-1H-imidazole-2-carboxylic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.28 g of a colorless oily liquid with a yield of 60%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.08 (d, J=7.5Hz, 1H), 7.42–7.29 (m, 6H), 7.10–7.01 (m, 2H), 6.07–5.94 (m, 1H), 5.79–5.69(m,1H),5.28–5.07(m,8H),4.78(dt,J＝8.0,3.0Hz,1H),4.73–4.65(m,1H),3.95–3.83(m,4H), 3.66(dd, J=9.5,3.5Hz,1H), 3.55(dd,J=9.5,6.5Hz,1H), 3.37(s,3H)ppm; ESI-MS: m/z=471[M+H] ⁺ .

Preparation Example 113, N-(N-(1-(but-3-en-1-yl)-1H-imidazole-2-carbonyl)-O-methyl-L-seryl)-O-allyl Benzyl-L-serine benzyl ester (8w)

Using 1-(but-3-en-1-yl)-1H-imidazole-2-carboxylic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.31 g of a colorless oily liquid with a yield of 64%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.09 (d, J=6.5Hz, 1H), 7.32–7.22 (m, 6H), 7.00–6.92 (m, 2H), 5.75–5.61 (m, 2H), 5.20–4.93(m,6H),4.72(dt,J＝8.0,3.0Hz,1H),4.62(td,J＝7.0,4.0Hz,1H),4.51–4.37(m,2H),3.90–3.78( m, 4H), 3.60(dd, J=9.5, 3.0Hz, 1H), 3.50(dd, J=9.0, 6.5Hz, 1H), 3.31(s, 3H), 2.53–2.45(m, 2H)ppm; ESI-MS: m/z＝485[M+H] ⁺ .

Preparation Example 114, N-(N-(1-(pent-4-en-1-yl)-1H-imidazole-2-carbonyl)-O-methyl-L-seryl)-O-allyl Benzyl-L-serine benzyl ester (8x)

Using 1-(pent-4-en-1-yl)-1H-imidazole-2-carboxylic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.35 g of a colorless oily liquid with a yield of 70%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.19(d, J=4.0Hz, 1H), 7.38–7.29(m, 6H), 7.07(d, J=1.0Hz, 1H), 7.03(d, J= 1.0Hz,1H),5.85–5.70(m,2H),5.26–5.22(m,1H),5.21–5.10(m,3H),5.08–4.99(m,2H),4.79(dt,J＝8.0, 3.0Hz, 1H), 4.69(td, J＝7.0, 4.5Hz, 1H), 4.48–4.42(m, 2H), 3.98–3.85(m, 4H), 3.67(dd, J＝9.5, 3.0Hz, 1H ),3.57(dd,J＝9.0,6.5Hz,1H),3.38(s,3H),2.81(s,3H),2.12–2.06(m,2H),1.97–1.88(m,2H)ppm; ESI -MS: m/z＝499[M+H] ⁺ .

Preparation Example 115, N-(N-(1-(pent-4-en-1-yl)-1H-pyrrole-2-carbonyl)-O-methyl-L-seryl)-O-allyl Benzyl-L-serine benzyl ester (8y)

Using 1-(pent-4-en-1-yl)-1H-pyrrole-2-carboxylic acid as the raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.25 g of a colorless oily liquid with a yield of 50%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.48 (d, J=8.0Hz, 1H), 7.39–7.30 (m, 5H), 6.80–6.75 (m, 2H), 6.67 (dd, J=4.0, 2.0 Hz,1H),6.10(dd,J＝4.0,2.5Hz,1H),5.84–5.71(m,2H),5.28–5.24(m,1H),5.22–5.12(m,3H),5.06–4.95( m,2H),4.77(dt,J＝8.0,3.0Hz,1H),4.70–4.65(m,1H),4.40–4.26(m,2H),3.97–3.90(m,3H),3.87(dd, J=9.0,4.0Hz,1H),3.68(dd,J=9.5,3.0Hz,1H),3.46(dd,J=9.0,8.0Hz,1H),3.38(s,3H),2.08–2.02(m ,2H),1.91–1.83(m,2H)ppm; ESI-MS: m/z=498[M+H] ⁺ .

Preparation Example 116, N-(N-(4-pentenoyl-L-prolyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester (8z)

Using pent-4-enoyl-L-proline as raw material, the synthesis and post-treatment were the same as in Preparation Example 91 to obtain 0.36 g of a colorless oily liquid with a yield of 70%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.40–7.29(m,6H),7.26–7.24(m,1H),5.92–5.72(m,2H),5.29–5.11(m,4H),5.09–4.95 (m,2H),4.78–4.70(m,1H),4.60–4.48(m,2H),3.98–3.91(m,2H),3.89(dd,J＝9.5,3.5Hz,1H),3.79(dd ,J=9.5,4.0Hz,1H),3.67(dd,J=9.5,3.5Hz,1H),3.62–3.56(m,1H),3.46(dd,J=9.0,7.0Hz,2H),3.33( s,3H),2.50–2.35(m,4H),2.32–2.23(m,1H),2.15–2.04(m,1H),2.02–1.93(m,2H)ppm; ESI-MS: m/z＝ 516[M+H] ⁺ .

Preparation Example 117, N-(N-(tert-butoxycarbonyl)-O-methyl-L-seryl)-O-(2-((2-allyloxycarbonyl)phenoxyethoxy Base) ethyl) -L-serine methyl ester (8za)

Reactant N-(N-(tert-butoxycarbonyl)-O-methyl-L-seryl)-O-(2-(2-(2-hydroxyethoxy)ethoxy)ethyl) -Methyl L-serine (7f, 0.61g, 1.5mmol), allyl 2-hydroxybenzoate (0.25g, 1.4mmol) and triphenylphosphine (0.39g, 1.5mmol) were placed in a 10ml three-necked flask , Added 3mL of anhydrous THF under the protection of N ₂ to dissolve, then cooled to 0°C, slowly added DIAD (0.35g, 1.7mmol) dropwise, and reacted overnight at room temperature. 1ml of ethyl acetate and 5ml of petroleum ether were added to the reaction solution, stirred at room temperature for 1 hour, filtered with suction, the filtrate was desolventized under reduced pressure, and separated by column chromatography to obtain 0.52g of a colorless oily liquid with a yield of 64%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.81 (dd, J=8.0, 1.5Hz, 1H), 7.57–7.51 (m, 1H), 7.35 (d, J=5.5Hz, 1H), 7.03–6.97( m,2H),6.08–5.97(m,1H),5.49–5.38(m,2H),5.30–5.24(m,1H),4.82–4.77(m,2H),4.74–4.66(m,1H), 4.31–4.25(m,1H),4.22–4.16(m,2H),3.95(dd,J=10.0,3.0Hz,1H),3.91–3.84(m,2H),3.79(dd,J=9.5,4.0 Hz,1H),3.76–3.67(m,6H),3.66–3.60(m,2H),3.48(dd,J＝9.5,6.5Hz,1H),3.37(s,3H),1.45(s,9H) ppm; ESI-MS: m/z=569[M+H] ⁺ .

Preparation Example 118, N-(N-(tert-butoxycarbonyl)-O-methyl-L-seryl)-O-(2-(2-((2-allyloxycarbonyl)phenoxy Ethoxy)ethoxy)ethyl)-L-serine methyl ester (8zb)

With N-(N-(tert-butoxycarbonyl)-O-methyl-L-seryl)-O-(2-(2-(2-hydroxyethoxy)ethoxy)ethyl)- L-serine methyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 117 to obtain 0.55 g of a colorless oily liquid with a yield of 60%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.80 (dd, J=8.0, 2.0Hz, 1H), 7.49–7.42 (m, 1H), 7.37 (d, J=5.5Hz, 1H), 7.02–6.96 ( m,2H),6.07–5.97(m,1H),5.49(d,J＝2.5Hz,1H),5.45–5.38(m,1H),5.29–5.23(m,1H),4.79(dt,J＝ 5.5,1.5Hz,2H),4.69(dt,J＝8.0,3.5Hz,1H),4.33–4.25(m,1H),4.23–4.18(m,2H),3.94(dd,J＝10.0,3.5Hz ,1H),3.92–3.87(m,2H),3.79(dd,J＝9.0,4.0Hz,1H),3.76–3.68(m,6H),3.66–3.57(m,6H),3.48(dd,J =9.0,6.5Hz,1H),3.37(s,3H),1.45(s,9H)ppm; ESI-MS: m/z=613[M+H] ⁺ .

Preparation Example 119, N-((S)-4-morpholino-4-oxo-2-(2-(4-penten-1-yloxy)benzamido)butyryl)-O -Allyl-L-serine benzyl ester (8zc)

The raw material 2-(4-penten-1-yloxy)benzoic acid (2c, 0.068g, _0.33mmol ) was placed in a 5ml reaction flask, and 2mL of anhydrous _CH2Cl2 was added to dissolve it, followed by the addition of 1-hydroxybenzo Triazole (0.053g, 0.40mmol) and N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (0.11g, 0.59mmol) were reacted at room temperature for half an hour. Subsequently, cooled to 0°C, N-((S)-2-amino-4-morpholino-4-oxobutanoyl)-O-allyl-L-serine benzyl ester hydrochloride (7 g, 0.18g, 0.40mmol) and DIPEA (0.16ml, 0.99mmol), react at room temperature for 3 hours. The reaction solution was diluted by adding 2 mL of saturated aqueous sodium bicarbonate solution, and the organic layer was separated, washed with saturated brine (2 mL×2), dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure, separated by column chromatography to obtain 0.13 g of a white solid, with a yield of 65%. . ¹ H NMR (500MHz, CDCl ₃ ) δ9.17 (d, J=7.5Hz, 1H), 8.18 (dd, J=7.5, 1.5Hz, 1H), 7.93 (d, J=8.0Hz, 1H), 7.46 –7.41(m,1H),7.37–7.28(m,5H),7.05(t,J＝7.5Hz,1H),6.96(d,J＝8.0Hz,1H),5.90–5.80(m,1H), 5.80–5.70(m,1H),5.27–5.03(m,6H),5.03–4.96(m,1H),4.75(dt,J＝8.0,3.0Hz,1H),4.17–4.08(m,2H), 3.98–3.88(m,3H),3.71–3.53(m,7H),3.47–3.33(m,2H),3.21(dd,J＝16.5,3.0Hz,1H),2.65(dd,J＝16.5,7.0 Hz,1H), 2.29–2.21(m,2H), 2.12–2.05(m,2H)ppm; ESI-MS: m/z=608[M+H] ⁺ .

Preparation Example 120, N-((S)-5-morpholino-5-oxo-2-(2-(4-penten-1-yloxy)benzamido)pentanoyl)-O -Allyl-L-serine benzyl ester (8zd)

Using N-((S)-2-amino-5-morpholino-5-oxopentanoyl)-O-allyl-L-serine benzyl ester hydrochloride as raw material, the synthesis and post-treatment are carried out in the same way as the preparation Example 119, 0.21 g of colorless oily liquid was obtained, yield 76%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.83 (d, J=7.0Hz, 1H), 8.15 (dd, J=8.0, 2.0Hz, 1H), 7.89 (d, J=8.5Hz, 1H), 7.42 (ddd, J=8.5,7.5,2.0Hz,1H),7.37–7.29(m,5H),7.07–7.01(m,1H),6.95(d,J=8.0Hz,1H),5.90–5.72(m ,2H),5.29–5.18(m,2H),5.15–5.04(m,3H),5.03–4.97(m,1H),4.83–4.73(m,2H),4.17–4.09(m,2H),4.00 –3.88(m,3H),3.74–3.61(m,6H),3.60–3.53(m,1H),3.53–3.40(m,2H),2.79–2.69(m,1H),2.58–2.49(m, 1H), 2.32–2.23(m,3H), 2.12–1.99(m,3H)ppm; ESI-MS: m/z＝622[M+H] ⁺ .

Preparation Example 121, N-(N ⁴ -cyclopropyl-N ² -(2-(4-penten-1-yloxy)benzoyl)-L-asparagineyl)-O-allyl Benzyl-L-serine benzyl ester (8ze)

Using N-(N ⁴ -cyclopropyl-L-asparaginyl)-O-allyl-L-serine benzyl ester hydrochloride as raw material, the synthesis and post-treatment were the same as in Preparation Example 119 to obtain a white solid 0.11 g, yield 58%. ¹ H NMR (500MHz, CDCl ₃ ) δ9.19(d, J=6.5Hz, 1H), 8.14(dd, J=8.0, 2.0Hz, 1H), 8.01(d, J=8.0Hz, 1H), 7.43 (ddd,J=8.5,7.5,2.0Hz,1H),7.38–7.29(m,5H),7.07–7.01(m,1H),6.96(d,J=8.0Hz,1H),6.20(d,J ＝3.5Hz,1H),5.90–5.70(m,2H),5.28–4.96(m,7H),4.78–4.72(m,1H),4.15(t,J＝6.5Hz,2H),3.99–3.87( m,3H),3.66(dd,J=9.5,3.5Hz,1H),2.82(dd,J=15.0,3.5Hz,1H),2.73–2.66(m,1H),2.61(dd,J=15.0, 7.0Hz,1H),2.29–2.18(m,2H),2.12–2.01(m,2H),0.78–0.69(m,2H),0.57–0.44(m,2H)ppm; ESI-MS: m/z ＝578[M+H] ⁺ .

Preparation Example 122, N-((2-(pent-4-en-1-yloxy)benzoyl)-L-leucyl)-O-allyl-L-serine benzyl ester (8zf)

Using N-(L-leucyl)-O-allyl-L-serine benzyl ester hydrochloride as raw material, the synthesis and post-treatment were the same as in Preparation Example 119 to obtain 0.12 g of a colorless oily liquid with a yield of 68% . ¹ H NMR (500MHz, CDCl ₃ ) δ8.42 (d, J=8.0Hz, 1H), 8.23–8.17 (m, 1H), 7.46–7.40 (m, 1H), 7.38–7.30 (m, 5H), 7.09–7.03(m,1H),6.95(d,J=8.5Hz,1H),6.90(d,J=8.0Hz,1H),5.90–5.80(m,1H),5.79–5.70(m,1H) ,5.29–5.23(m,1H),5.20–5.01(m,5H),4.81–4.73(m,2H),4.18–4.10(m,2H),3.97–3.85(m,3H),3.62(dd, J=9.5,3.5Hz,1H),2.29(q,J=7.5Hz,2H),2.07–1.98(m,2H),1.82–1.73(m,2H),1.68–1.59(m,1H),0.95 (d, J=6.0Hz,6H)ppm; ESI-MS: m/z=537[M+H] ⁺ .

Preparation Example 123, N-((2-(pent-4-en-1-yloxy)benzoyl)-L-valyl)-O-allyl-L-serine benzyl ester (8zg)

Using N-(L-valyl)-O-allyl-L-serine benzyl ester hydrochloride as raw material, the synthesis and post-treatment were the same as in Preparation Example 119 to obtain 0.088 g of a colorless oily liquid with a yield of 51% . ¹ H NMR (500MHz, CDCl ₃ ) δ8.56 (d, J=8.5Hz, 1H), 8.20 (dd, J=8.0, 2.0Hz, 1H), 7.46–7.40 (m, 1H), 7.39–7.29( m,5H),7.09–7.02(m,1H),6.97(d,J=8.5Hz,1H),6.68(d,J=8.0Hz,1H),5.90–5.69(m,2H),5.30–5.23 (m,1H),5.20–4.99(m,5H),4.81(dt,J＝8.0,3.0Hz,1H),4.62(dd,J＝8.5,6.0Hz,1H),4.20–4.09(m,2H ),3.97–3.84(m,3H),3.62(dd,J＝9.5,3.0Hz,1H),2.36–2.16(m,3H),2.09–2.01(m,2H),1.06–0.99(m,6H )ppm; ESI-MS: m/z＝523[M+H] ⁺ .

Preparation Example 124, (S)-2-(N-(2-(pent-4-en-1-yloxy)benzoyl)-O-methyl-L-seryl)aminohexan-5 -Benzyl enoate (8zh)

Using (S)-2-((S)-2-amino-3-methoxypropionamido)-5-hexenoic acid benzyl ester hydrochloride as raw material, the synthesis and post-treatment were the same as in Preparation Example 119, to obtain Colorless oily liquid 0.15g, yield 88%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.83 (d, J=7.0Hz, 1H), 8.21 (dd, J=8.0, 1.5Hz, 1H), 7.50–7.42 (m, 1H), 7.39–7.29( m,5H),7.14(d,J=8.0Hz,1H),7.07(t,J=7.5Hz,1H),6.97(d,J=8.5Hz,1H),5.88–5.67(m,2H), 5.21–5.10(m,2H),5.09–4.90(m,4H),4.81(td,J=6.5,4.0Hz,1H),4.69(td,J=7.5,4.5Hz,1H),4.16–4.12( m,2H),3.96(dd,J=9.0,3.5Hz,1H),3.51(dd,J=9.0,6.0Hz,1H),3.39(s,3H),2.29–2.22(m,2H),2.11 –1.93(m,5H),1.84–1.73(m,1H)ppm; ESI-MS: m/z=509[M+H] ⁺ .

Preparation Example 125, (S)-2-(N-(2-(hex-5-en-1-yloxy)benzoyl)-O-methyl-L-seryl)aminohexan-5 -Benzyl enoate (8zi)

The raw material 2-(5-hexen-1-yloxy)benzoic acid (2d, 0.15g, 0.69mmol) was placed in a 5ml reaction flask, and 3mL of anhydrous THF was added to dissolve it, and then 1-hydroxybenzotriazole was added (0.11g, 0.80mmol), benzotriazole-N,N,N',N'-tetramethyluronium hexafluorophosphate (0.30g, 0.80mmol) and (S)-2-((S) - Benzyl 2-amino-3-methoxypropionamido)-5-hexenoate hydrochloride (0.29 g, 0.80 mmol). Subsequently, it was cooled to 0°C, DIPEA (0.23ml, 1.4mmol) was added, and reacted at room temperature for 3 hours. The solvent was evaporated under reduced pressure, 5 mL of ethyl acetate and 4 mL of water were added, the organic layer was separated, washed successively with saturated aqueous sodium bicarbonate (4 mL×2) and saturated brine (4 mL×2), dried over anhydrous sodium sulfate, and reduced The solvent was removed under pressure and separated by column chromatography to obtain 0.31 g of a colorless oily liquid with a yield of 86%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.83 (d, J=6.5Hz, 1H), 8.21 (dd, J=8.0, 1.5Hz, 1H), 7.45 (ddd, J=8.5, 7.0, 2.0Hz, 1H),7.38–7.29(m,5H),7.15(d,J＝8.0Hz,1H),7.10–7.04(m,1H),6.97(d,J＝8.0Hz,1H),5.86–5.67(m ,2H),5.21–5.10(m,2H),5.06–4.90(m,4H),4.81(td,J＝6.5,3.5Hz,1H),4.68(td,J＝7.5,4.5Hz,1H), 4.16–4.11(m,2H),3.94(dd,J＝9.0,3.5Hz,1H),3.51(dd,J＝9.0,6.5Hz,1H),3.40(s,3H),2.17–2.10(m, 2H), 2.09–1.90(m,5H), 1.84–1.75(m,1H), 1.63–1.54(m,2H)ppm; ESI-MS: m/z＝523[M+H] ⁺ .

Preparation Example 126, (8S,11S)-11-(methoxymethyl)-10,13-dioxo-3,4,5,7,8,9,10,11,12,13-deca Hydrogen-2H-1,6,9,12-benzodioxadiazacyclopentadec-8-carboxylic acid (9a)

Starting material N-(N-(2-(allyloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester (8a, 0.10 g, 0.20 mmol) was placed in a 100ml two-necked bottle, dissolved in 60mL of toluene, and heated to 100°C under the protection of _N2 . Subsequently, Grubbs second-generation catalyst was dissolved in 7ml of toluene and slowly injected into the reaction system, and reacted at this temperature for 1 hour. The solvent was removed under reduced pressure, and 80 mg of gray oily liquid (including cis-trans isomers) was obtained by column chromatography with a yield of 85%. The obtained product was directly used in the next reaction.

The above raw materials were dissolved in 3ml of methanol, 10% Pd/C (8mg) was added, reacted at room temperature for 3h under the protection of _H2 , the catalyst was removed by filtration with diatomaceous earth, and the solvent was distilled off under reduced pressure to obtain 58mg of a white foamy solid, which was collected The rate is 89%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.48(d, J=9.0Hz, 1H), 8.09(d, J=7.0Hz, 1H), 7.50–7.43(m, 1H), 7.35(d, J= 7.0Hz, 1H), 7.12–7.06(m, 1H), 6.99(d, J＝8.5Hz, 1H), 5.04–4.97(m, 1H), 4.74–4.68(m, 1H), 4.23–4.03(m ,4H),3.74–3.66(m,2H),3.51(dd,J＝9.0,3.5Hz,1H),3.45–3.40(m,1H),3.35(s,3H),2.06–1.97(m,1H ), 1.87–1.75(m,2H), 1.69–1.60(m,1H)ppm; ESI-MS: m/z＝381[M+H] ⁺ .

Preparation Example 127, (9S,12S)-12-(methoxymethyl)-11,14-dioxo-2,3,4,5,6,8,9,10,11,12,13 ,14-Dodecahydro-1,7,10,13-benzodioxadiazacyclohexadeca-9-carboxylic acid (9b)

Take N-(N-(2-(3-buten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester as The raw materials, synthesis and post-treatment were the same as those in Preparation Example 126 to obtain 66 mg of white foamy solid, with a two-step yield of 84%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.93(d, J=9.0Hz, 1H), 8.19(d, J=7.0Hz, 1H), 7.50–7.41(m, 1H), 7.28(d, J= 9.0Hz, 1H), 7.11–7.03(m, 1H), 6.98(d, J＝8.5Hz, 1H), 5.00–4.91(m, 1H), 4.80–4.71(m, 1H), 4.27–4.20(m ,1H),4.17–4.08(m,2H),4.06–3.96(m,1H),3.79–3.73(m,1H),3.67–3.61(m,1H),3.59–3.53(m,1H),3.50 (dd,J=9.0,3.5Hz,1H),3.37(s,3H),1.93–1.73(m,3H),1.60–1.48(m,3H)ppm; ESI-MS: m/z=395[M +H] ⁺ .

Preparation Example 128, (10S,13S)-13-(methoxymethyl)-12,15-dioxo-3,4,5,6,7,9,10,11,12,13,14 ,15-2H-Dodecahydro-1,8,11,14-benzodioxadiazacyclohexadeca-10-carboxylic acid (9c)

Take N-(N-(2-(4-penten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester as The raw materials, synthesis and post-treatment were the same as those in Preparation Example 126 to obtain 62 mg of white foamy solid, with a two-step yield of 76%. ¹ H NMR (500MHz, CDCl ₃ ) δ9.00(d, J=8.0Hz, 1H), 8.18(d, J=7.0Hz, 1H), 7.47–7.38(m, 1H), 7.23(d, J= 6.5Hz, 1H), 7.06–6.99(m, 1H), 6.94(d, J＝8.5Hz, 1H), 4.92–4.82(m, 1H), 4.65–4.55(m, 1H), 4.23–4.16(m ,1H),4.14–4.05(m,2H),3.80–3.67(m,2H),3.56–3.47(m,2H),3.45–3.40(m,1H),3.36(s,3H),1.98–1.88 (m,1H),1.82–1.69(m,2H),1.63–1.36(m,5H)ppm; ESI-MS: m/z=409[M+H] ⁺ .

Preparation Example 129, (11S,14S)-14-(methoxymethyl)-13,16-dioxo-2,3,4,5,6,7,8,10,11,12,13 ,14,15,16-tetrahydro-1,9,12,15-benzodioxadiazacycloctadec-11-carboxylic acid (9d)

N-(N-(2-(5-hexen-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester as The raw materials, synthesis and post-treatment were the same as those in Preparation Example 126 to obtain 68 mg of white foamy solid, with a two-step yield of 80%. ¹ H NMR (500MHz, CDCl ₃ ) δ9.00(d, J=7.5Hz, 1H), 8.18(d, J=7.5Hz, 1H), 7.50–7.40(m, 1H), 7.23(d, J= 6.5Hz, 1H), 7.10–7.01(m, 1H), 6.98(d, J＝8.5Hz, 1H), 4.87–4.81(m, 1H), 4.75–4.69(m, 1H), 4.28–4.21(m ,1H),4.15–4.03(m,2H),3.86(dd,J=10.5,4.5Hz,1H),3.76(dd,J=10.5,2.5Hz,1H),3.56(dd,J=9.0,4.5 Hz,1H),3.53–3.47(m,1H),3.45–3.35(m,4H),2.03–1.94(m,1H),1.88–1.77(m,1H),1.64–1.32(m,8H)ppm ;ESI-MS: m/z=423[M+H] ⁺ .

Preparative Example 130, (12S,15S)-15-(methoxymethyl)-14,17-dioxo-3,4,5,6,7,8,9,11,12,13,14 ,15,16,17-tetrahydro-2H-1,10,13,16-benzodioxane nonadecane-12-carboxylic acid (9e)

Take N-(N-(2-(6-hepten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester as The raw materials, synthesis and post-treatment were the same as those in Preparation Example 126 to obtain 67 mg of white foamy solid, and the two-step yield was 77%. ¹ H NMR (500MHz, CDCl ₃ ) δ9.03(d, J=6.0Hz, 1H), 8.20(d, J=7.5Hz, 1H), 7.49–7.39(m, 1H), 7.28(d, J= 7.5Hz, 1H), 7.07–7.00(m, 1H), 6.97(d, J＝8.5Hz, 1H), 4.80–4.75(m, 1H), 4.74–4.68(m, 1H), 4.22–4.12(m ,2H),3.97(dd,J＝9.5,4.0Hz,1H),3.80–3.70(m,2H),3.66(dd,J＝9.5,5.0Hz,1H),3.54–3.48(m,1H), 3.46–3.36(m,4H),2.03–1.96(m,1H),1.91–1.82(m,1H),1.60–1.28(m,10H)ppm; ESI-MS: m/z=437[M+H ] ⁺ .

Preparation Example 131, (13S,16S)-16-(methoxymethyl)-15,18-dioxo-3,4,5,6,7,8,9,10,13,14,15 ,16,17,18-tetrahydro-2H,12H-benzo[i][1,11]dioxa[4,7]diazacycloeicos-13-carboxylic acid (9f)

Take N-(N-(2-(7-octen-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester as The raw materials, synthesis and post-treatment were the same as those in Preparation Example 126 to obtain 67 mg of white foamy solid, and the two-step yield was 77%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.97(d, J=6.5Hz, 1H), 8.21(d, J=7.5Hz, 1H), 7.49–7.41(m, 1H), 7.22(d, J= 6.0Hz, 1H), 7.07–7.01(m, 1H), 6.97(d, J＝8.5Hz, 1H), 4.82–4.75(m, 1H), 4.68–4.61(m, 1H), 4.23–4.18(m ,1H),4.17–4.12(m,1H),4.01(dd,J＝9.5,3.0Hz,1H),3.80–3.72(m,2H),3.63(dd,J＝9.5,3.5Hz,1H), 3.55–3.48(m,1H),3.41–3.29(m,4H),2.01–1.89(m,1H),1.83–1.72(m,1H),1.67–1.29(m,12H)ppm; ESI-MS: m/z＝451[M+H] ⁺ .

Preparation Example 132, (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14 ,15-Dodecahydro-9H-17-chlorobenzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-carboxylic acid (9g)

With N-(N-(5-chloro-2-(4-penten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L- Serine benzyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 61 mg of white foamy solid, with a two-step yield of 69%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.95 (d, J = 8.5Hz, 1H), 8.18 (d, J = 3.0Hz, 1H), 7.39 (dd, J = 8.5, 3.0Hz, 1H), 7.13 (d,J=7.0Hz,1H),6.91(d,J=8.5Hz,1H),4.93–4.86(m,1H),4.75–4.66(m,1H),4.27–4.18(m,1H), 4.14–4.07(m,2H),3.81(dd,J＝10.0,4.5Hz,1H),3.75(dd,J＝10.0,3.5Hz,1H),3.58–3.52(m,2H),3.42–3.34( m,4H),2.00–1.90(m,1H),1.85–1.72(m,2H),1.68–1.38(m,5H)ppm; ESI-MS: m/z＝443[M+H] ⁺ .

Preparation Example 133, (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14 ,15-Dodecahydro-9H-18-chlorobenzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-carboxylic acid (9h)

With N-(N-(4-chloro-2-(4-penten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L- Serine benzyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 66 mg of off-white foamy solid, with a two-step yield of 75%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.86(d, J=8.0Hz, 1H), 8.26–8.18(m, 1H), 7.15(d, J=6.5Hz, 1H), 6.79–6.72(m, 1H),6.69(d,J＝10.5Hz,1H),4.93–4.86(m,1H),4.72–4.64(m,1H),4.23–4.16(m,1H),4.14–4.04(m,2H) ,3.82–3.69(m,2H),3.57–3.49(m,2H),3.41–3.32(m,4H),1.99–1.89(m,1H),1.82–1.73(m,2H),1.66–1.36( m,5H)ppm; ESI-MS: m/z=443[M+H] ⁺ .

Preparative Example 134, (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14 ,15-Dodecahydro-9H-18-fluorobenzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-carboxylic acid (9i)

With N-(N-(4-fluoro-2-(4-penten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L- Serine benzyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 59 mg of off-white foamy solid, with a two-step yield of 69%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.87(d, J=8.5Hz, 1H), 8.15(d, J=8.5Hz, 1H), 7.15(d, J=7.0Hz, 1H), 7.05(d ,J＝8.5Hz,1H),6.98(s,1H),4.93–4.86(m,1H),4.72–4.64(m,1H),4.24–4.18(m,1H),4.13–4.08(m,2H ),3.82–3.70(m,2H),3.56–3.49(m,2H),3.41–3.32(m,5H),2.00–1.89(m,1H),1.83–1.72(m,2H),1.67–1.35 (m,5H)ppm; ESI-MS: m/z=427[M+H] ⁺ .

Preparative Example 135, (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14 ,15-Dodecahydro-9H-18-methylbenzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-carboxylic acid (9j)

With N-(N-(4-methyl-2-(4-penten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L - Serine benzyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 63 mg of off-white foamy solid, with a two-step yield of 75%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.98(d, J=8.5Hz, 1H), 8.09(d, J=8.0Hz, 1H), 7.16(d, J=7.0Hz, 1H), 6.87(d ,J＝8.0Hz,1H),6.77(s,1H),4.96–4.89(m,1H),4.72–4.64(m,1H),4.23–4.15(m,1H),4.15–4.08(m,2H ),3.78(dd,J＝10.0,4.5Hz,1H),3.73(dd,J＝10.0,3.5Hz,1H),3.56–3.47(m,2H),3.43–3.33(m,4H),2.38( s,3H),1.99–1.89(m,1H),1.82–1.72(m,2H),1.66–1.36(m,5H)ppm; ESI-MS: m/z＝423[M+H] ⁺ .

Preparative Example 136, (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14 ,15-Dodecahydro-9H-18-methoxybenzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-carboxylic acid (9k)

With N-(N-(4-methoxy-2-(4-penten-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl- L-serine benzyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 59 mg of off-white foamy solid, with a two-step yield of 69%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.87(d, J=8.0Hz, 1H), 8.16(d, J=8.5Hz, 1H), 7.18(d, J=6.5Hz, 1H), 6.58(d ,J＝8.5Hz,1H),6.47(s,1H),4.93–4.84(m,1H),4.69–4.59(m,1H),4.22–4.16(m,1H),4.14–4.05(m,2H ),3.85(s,3H),3.79(dd,J＝9.5,4.5Hz,1H),3.73(dd,J＝9.5,3.0Hz,1H),3.57–3.48(m,2H),3.43–3.31( m,4H),2.00–1.88(m,1H),1.84–1.71(m,2H),1.69–1.36(m,5H)ppm; ESI-MS: m/z＝439[M+H] ⁺ .

Preparation Example 137, (3S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15,16,17 ,18-Dodecahydro-12H-2-methylthiazolo[5,4-i][1,11]dioxa[4,7]diazacyclohexadeca-13-carboxylic acid (9l)

Starting material N-(N-(2-methyl-4-(4-penten-1-yloxy)thiazole-5-formyl)-O-methyl-L-seryl)-O-allyl Base-L-serine methyl ester (8l, 0.12g, 0.26mmol) was placed in a 250ml two-neck flask, dissolved in 65mL of toluene, and heated to 100°C under nitrogen protection. Subsequently, the Grubbs second-generation catalyst was dissolved in 8 ml of toluene and slowly injected into the reaction system, and reacted at this temperature for 1 hour. The solvent was removed under reduced pressure, and 0.10 g of a colorless oily liquid (including cis and trans isomers) was obtained by column chromatography with a yield of 88%. The obtained product was directly used in the next reaction.

Dissolve the above raw materials in 3ml of methanol, add 10% Pd/C (30mg), react at room temperature for 3h under the protection of H ₂ , remove the catalyst by filtration with diatomaceous earth, evaporate the solvent under reduced pressure, and obtain a colorless Oily liquid 86mg, yield 85%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.95 (d, J = 9.5Hz, 1H), 7.21 (d, J = 7.0Hz, 1H), 4.89–4.83 (m, 1H), 4.69–4.63 (m, 1H ),4.59(td,J＝10.0,2.5Hz,1H),4.51–4.45(m,1H),4.16–4.09(m,2H),3.84(dd,J＝10.0,3.0Hz,1H),3.72( s,3H),3.68(dd,J＝10.0,3.0Hz,1H),3.56–3.49(m,2H),3.38(s,3H),2.64(s,3H),1.89–1.73(m,2H) , 1.69 - 1.50 (m, 5H), 1.48 - 1.38 (m, 1H) ppm; ESI-MS: m/z = 444 [M+H] ⁺ .

Reactant (3S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15,16,17,18- Dodecahydro-12H-2-methylthiazolo[5,4-i][1,11]dioxa[4,7]diazacyclohexadeca-13-carboxylic acid methyl ester (60mg, 0.14 mmol) was dissolved in 0.68 mL of acetone, cooled to 0° C. under an ice bath, 0.67 mL of 0.3 N LiOH aqueous solution was added dropwise, and reacted at this temperature for 2 hours. Acetone was removed under reduced pressure, the pH of the aqueous layer was adjusted to 2-3 with 3N HCl, extracted with ethyl acetate (2 mL×3), the organic layers were combined, dried over anhydrous sodium sulfate, and the solvent was evaporated under reduced pressure. The resulting product was directly used in the next reaction.

Preparation Example 138, (6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,12,13,14,15,16 ,17-Dodecahydro-4H-thieno[3,2-i][1,11]dioxa[4,7]diazacyclohexadeca-9-carboxylic acid (9m)

N-(O-methyl-N-(2-(4-penten-1-yloxy)thiophene-3-formyl)-L-seryl)-O-allyl-L-serine Methyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 135 to obtain 76 mg of a colorless oily liquid, with a two-step yield of 68%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.14(d, J=9.0Hz, 1H), 7.44(d, J=5.5Hz, 1H), 7.24(d, J=7.0Hz, 1H), 6.87(d ,J＝5.5Hz,1H),4.96–4.91(m,1H),4.71–4.66(m,1H),4.33–4.27(m,1H),4.21–4.13(m,2H),3.82(dd,J ＝10.0,3.5Hz,1H),3.73–3.68(m,4H),3.57–3.50(m,2H),3.42–3.36(m,4H),1.95–1.85(m,1H),1.80–1.69(m ,2H), 1.63–1.54(m,4H), 1.48–1.40(m,1H)ppm; ESI-MS: m/z＝429[M+H] ⁺ .

Reactant (6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,12,13,14,15,16,17- Dodecahydro-4H-thieno[3,2-i][1,11]dioxa[4,7]diazacyclohexadeca-9-carboxylate (60mg, 0.14mmol) was dissolved in 0.70 mL of acetone was cooled to 0°C under an ice bath, 0.70 mL of 0.3N LiOH aqueous solution was added dropwise, and the reaction was carried out at this temperature for 2 hours. Acetone was removed under reduced pressure, the pH of the aqueous layer was adjusted to 2-3 with 3N HCl, extracted with ethyl acetate (2 mL x 3), the organic layers were combined, dried over anhydrous sodium sulfate, and the solvent was evaporated under reduced pressure. The resulting product was directly used in the next reaction.

Preparation Example 139, (14S,17S)-17-(methoxymethyl)-16,19-dioxo-6,7,8,9,10,11,14,15,16,17,18 ,19-Dodecahydro-13H-pyrido[3-i][1,11]dioxa[4,7]diazacyclohexadeca-14-carboxylic acid (9n)

N-(O-methyl-N-(3-(4-penten-1-yloxy)pyridineformyl)-L-seryl)-O-allyl-L-serine benzyl ester as The raw materials, synthesis and post-treatment were the same as in Preparation Example 126 to obtain 45 mg of a light red foamy solid, with a two-step yield of 55%. ESI-MS: m/z=410[M+H] ⁺ .

Preparative Example 140, (7S,10S)-7-(methoxymethyl)-5,8-dioxo-6,7,8,9,10,11,13,14,15,16,17 ,18-Dodecahydro-5H-pyrido[3,2-i][1,11]dioxa[1,11]diazacyclohexadeca-10-carboxylic acid (9o)

N-(O-methyl-N-(2-(4-penten-1-yloxy)nicotinylformyl)-L-seryl)-O-allyl-L-serine benzyl ester As the raw material, the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 50 mg of a light red foamy solid, and the two-step yield was 61%. ESI-MS: m/z=410[M+H] ⁺ .

Preparation Example 141, (6S,9S)-6-(methoxymethyl)-4,7-dioxo-1,4,5,6,7,8,9,10,12,13,14 ,15,16,17-Tetradetrahydro-1-methyl-pyrazolo[4,3-i][1,11]dioxa[4,7]diazacyclohexadeca-9-carboxylic acid (9p)

With N-(O-methyl-N-(1-methyl-5-(4-penten-1-yloxy)-1H-pyrazole-4-formyl)-L-seryl)- O-allyl-L-serine benzyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 68 mg of off-white foamy solid with a two-step yield of 82%. ESI-MS: m/z＝413[M+H] ⁺ .

Preparation Example 142, (13S,16S)-16-(methoxymethyl)-15,18-dioxo-2,11-dioxa-14,17-diazabicyclo[17.3.1] - Tricos-1(23),19,21-triene-13-carboxylic acid (9q)

N-(N-(3-(hept-6-en-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester As the raw material, the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 61 mg of off-white foamy solid, with a two-step yield of 70%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.42(d, J=8.0Hz, 1H), 7.39–7.31(m, 3H), 7.09–7.00(m, 2H), 4.92–4.84(m, 1H), 4.74–4.68(m,1H),4.19–4.09(m,2H),4.07(dd,J=10.0,3.5Hz,1H),4.03–3.93(m,1H),3.88(dd,J=10.0,3.5 Hz, 1H), 3.67(dd, J＝10.0, 3.5Hz, 1H), 3.62(dd, J＝9.5, 4.0Hz, 1H), 3.44–3.36(m, 4H), 1.86–1.69(m, 3H) ,1.55–1.40(m,5H),1.40–1.29(m,4H)ppm; ESI-MS: m/z=437[M+H] ⁺ .

Preparation Example 143, (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14 ,15-Dodecahydro-9H-19-fluoro-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-carboxylic acid (9r)

With N-(N-(3-fluoro-2-(pent-4-en-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl-L - Serine benzyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 70 mg of off-white foamy solid, with a two-step yield of 82%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.96(d, J=8.0Hz, 1H), 7.92(d, J=8.0Hz, 1H), 7.25–7.20(m, 2H), 7.13–7.02(m, 1H),4.88–4.79(m,1H),4.68–4.60(m,1H),4.42–4.34(m,1H),4.32–4.24(m,1H),4.13–4.06(m,1H),3.80( dd,J＝9.5,3.5Hz,1H),3.75(dd,J＝9.5,2.5Hz,1H),3.57–3.49(m,2H),3.39–3.34(m,4H),2.01–1.91(m, 1H), 1.77–1.68(m,1H), 1.64–1.36(m,6H)ppm; ESI-MS: m/z=427[M+H] ⁺ .

Preparation Example 144, (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14 ,15-Dodecahydro-9H-19-methoxy-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-carboxylic acid (9s)

N-(N-(3-methoxy-2-(pent-4-en-1-yloxy)benzoyl)-O-methyl-L-seryl)-O-allyl - L-serine benzyl ester as raw material, the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 60 mg of off-white foamy solid, and the two-step yield was 68%. ¹ H NMR (500MHz, CDCl ₃ ) δ9.14(d, J=7.5Hz, 1H), 7.72(d, J=7.5Hz, 1H), 7.23(d, J=7.0Hz, 1H), 7.15–7.10 (m,1H),7.10–7.04(m,1H),4.85–4.78(m,1H),4.69–4.61(m,1H),4.11–4.01(m,2H),3.91–3.81(m,5H) ,3.75(dd,J=9.5,2.5Hz,1H),3.54(dd,J=9.0,3.5Hz,2H),3.42–3.32(m,4H),1.78–1.67(m,1H),1.64–1.55 (m,1H),1.53–1.18(m,6H)ppm; ESI-MS: m/z=439[M+H] ⁺ .

Preparation Example 145, (6S,9S)-6-(methoxymethyl)-4,7-dioxo-4,5,6,7,8,9,10,12,13,14,15 ,16,17,18-Tetradetrahydro-1H-1-methyl-pyrazolo[4,3-i][1]oxa[4,7]-diazacyclohexadeca-9-carboxy Acid (9t)

With N-(N-(5-allyl-1-methyl-1H-pyrazole-4-carbonyl)-O-methyl-L-seryl)-O-(5-hexene-1- Base)-L-serine benzyl ester as raw material, the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 71 mg of off-white foamy solid, and the two-step yield was 87%. ¹ H NMR (500MHz, CDCl ₃ ) δ9.14(d, J=7.5Hz, 1H), 7.72(d, J=7.5Hz, 1H), 7.23(d, J=7.0Hz, 1H), 7.15–7.10 (m,1H),7.10–7.04(m,1H),4.85–4.78(m,1H),4.69–4.61(m,1H),4.11–4.01(m,2H),3.91–3.81(m,5H) ,3.75(dd,J=9.5,2.5Hz,1H),3.54(dd,J=9.0,3.5Hz,2H),3.42–3.32(m,4H),1.78–1.67(m,1H),1.64–1.55 (m,1H),1.53–1.18(m,6H)ppm; ESI-MS: m/z=411[M+H] ⁺ .

Preparation Example 146, (6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,12,13,14,15,16 ,17-Dodecahydro-4H-pyrazolo[5,1-i][1]oxa[4,7,10]-triazacyclohexadeca-9-carboxylic acid (9u)

With N-(N-(1-(pent-4-en-1-yl)-1H-pyrazole-5-carbonyl)-O-methyl-L-seryl)-O-allyl-L - Serine benzyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 57 mg of off-white foamy solid, with a two-step yield of 75%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.47(d, J=2.0Hz, 1H), 6.60(d, J=2.0Hz, 1H), 5.15–5.06(m, 1H), 4.75–4.69(m, 1H), 4.58–4.52(m, 1H), 4.19(dt, J=13.5, 5.0Hz, 1H), 3.90(dd, J=10.0, 5.5Hz, 1H), 3.75(dd, J=10.0, 4.0Hz ,1H),3.70–3.62(m,2H),3.48–3.28(m,5H),2.01–1.90(m,1H),1.76–1.66(m,1H),1.51–1.29(m,3H),1.28 –0.99(m,3H)ppm; ESI-MS: m/z=383[M+H] ⁺ .

Preparation Example 147, (11S,14S)-14-(methoxymethyl)-13,16-dioxo-5,6,7,8,11,12,13,14,15,16-deca Hydrogen-10H-imidazo[2,1-i][1]oxa[4,7,10]-triazacyclotetradecan-11-carboxylic acid (9v)

Using N-(N-(1-allyl-1H-imidazole-2-carbonyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester as raw material, synthesis and The post-treatment was the same as that of Preparation Example 126 to obtain 47 mg of a light red solid, and the two-step yield was 66%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.27(d, J=5.5Hz, 1H), 7.61(d, J=6.5Hz, 1H), 7.09–6.94(m, 2H), 4.69–4.47(m, 3H),4.34–4.24(m,1H),3.98(dd,J＝10.0,6.5Hz,1H),3.93–3.80(m,2H),3.69(dd,J＝9.0,2.5Hz,1H),3.62 –3.56(m,1H),3.44–3.28(m,4H),1.90–1.80(m,1H),1.76–1.67(m,1H),1.54–1.39(m,2H)ppm; ESI-MS:m /z＝355[M+H] ⁺ .

Preparation Example 148, (12S,15S)-15-(methoxymethyl)-14,17-dioxo-6,7,8,9,12,13,14,15,16,17-deca Hydrogen-5H,11H-imidazo[2,1-i][1]oxa[4,7,10]-triazacyclopentadeca-12-carboxylic acid (9w)

With N-(N-(1-(but-3-en-1-yl)-1H-imidazole-2-carbonyl)-O-methyl-L-seryl)-O-allyl-L- Serine benzyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 44 mg of a light red solid with a two-step yield of 60%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.13(d, J=7.5Hz, 1H), 7.34(d, J=7.5Hz, 1H), 7.10–6.97(m, 2H), 4.76–4.69(m, 1H), 4.56–4.47(m,1H), 4.08–3.95(m,3H), 3.73(dd,J＝9.5,4.0Hz,1H), 3.61–3.52(m,2H), 3.49(s,1H) ,3.42(s,3H),3.26–3.18(m,1H),1.86–1.75(m,1H),1.68–1.55(m,2H),1.43–1.26(m,3H)ppm; ESI-MS:m /z＝369[M+H] ⁺ .

Preparation Example 149, (13S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15,16,17 ,18-Dodecahydro-12H-imidazo[2,1-i][1]oxa[4,7,10]-triazacyclohexadeca-13-carboxylic acid (9x)

With N-(N-(1-(pent-4-en-1-yl)-1H-imidazole-2-carbonyl)-O-methyl-L-seryl)-O-allyl-L- Serine benzyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 45 mg of a light red solid with a two-step yield of 59%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.13(d, J=6.7Hz, 1H), 7.46(d, J=7.9Hz, 1H), 7.10–7.04(m, 2H), 4.68–4.62(m, 1H),4.56–4.50(m,1H),4.03–3.94(m,2H),3.80(dd,J＝9.5,3.0Hz,1H),3.74(dd,J＝10.0,4.5Hz,1H),3.63 (dd,J=9.5,3.0Hz,1H),3.49–3.31(m,6H),1.87–1.75(m,2H),1.61–1.45(m,2H),1.43–1.32(m,2H),1.30 –1.21(m,2H)ppm; ESI-MS: m/z=383[M+H] ⁺ .

Preparative Example 150, (3S,6S)-3-(methoxymethyl)-1,4-dioxo-2,3,4,5,6,7,9,10,11,12,13 ,14-Dodecahydro-1H-pyrrolo[2,1-i][1]oxa[4,7,10]-triazacyclohexadeca-6-carboxylic acid (9y)

With N-(N-(1-(pent-4-en-1-yl)-1H-pyrrole-2-carbonyl)-O-methyl-L-seryl)-O-allyl-L- Serine benzyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 58 mg of off-white foamy solid, with a two-step yield of 76%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.40(d, J=8.0Hz, 1H), 6.91–6.76(m, 2H), 6.66(d, J=2.5Hz, 1H), 6.17(dd, J= 3.5,2.5Hz,1H),5.22–5.09(m,1H),4.62(dt,J＝8.0,2.5Hz,1H),4.59–4.52(m,1H),3.99(dd,J＝10.0,5.0Hz ,1H),3.93(dd,J＝9.5,2.0Hz,2H),3.76–3.65(m,2H),3.51–3.36(m,5H),1.78–1.59(m,2H),1.60–1.49(m ,1H), 1.47–1.36(m,2H), 1.33–1.19(m,3H)ppm; ESI-MS: m/z=382[M+H] ⁺ .

Preparation Example 151, (3S,6S,18aS)-3-(methoxymethyl)-1,4,14-trioxo-hexadecahydro-1H-pyrrolo[2,1-i][1 ]oxa[4,7,10]triazacyclohexadeca-6-carboxylic acid (9z)

Using N-(N-(4-pentenoyl-L-prolyl)-O-methyl-L-seryl)-O-allyl-L-serine benzyl ester as raw material, synthesis and post-processing The same as in Preparation Example 126, 68 mg of off-white oily liquid was obtained, and the two-step yield was 85%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.68(d, J=7.0Hz, 1H), 6.65(d, J=8.0Hz, 1H), 4.79–4.69(m, 1H), 4.59–4.44(m, 2H), 3.98(dd, J＝10.0, 2.5Hz, 1H), 3.90–3.77(m, 2H), 3.76–3.67(m, 1H), 3.65–3.49(m, 4H), 3.32(s, 3H) ,2.47–2.39(m,2H),2.36–2.30(m,1H),2.10–1.94(m,4H),1.72–1.63(m,1H),1.55–1.39(m,3H),1.35–1.28( m,1H)ppm; ESI-MS: m/z=400[M+H] ⁺ .

Preparation Example 152, (9S,12S)-12-(methoxymethyl)-11,14-dioxo-2,3,5,6,9,10,11,12,13,14-deca Hydrogen-8H-benzo[o][1,4,7]-trioxa[10,13]-diazacyclohexadeca-9-carboxylic acid (9za)

Raw material N-(N-(tert-butoxycarbonyl)-O-methyl-L-seryl)-O-(2-((2-allyloxycarbonyl)phenoxyethoxy)ethyl )-L-serine methyl ester (8za, 0.68g, 1.2mmol) was placed in a 25ml round bottom flask, tetrakistriphenylphosphinepalladium (0.069g, 0.06mmol) and 1,3-dimethylbarbituric acid were added (0.19g, 1.2mmol), injected 8ml of dichloromethane under the protection of N ₂ , and reacted overnight at room temperature. Add 8ml of saturated sodium bicarbonate solution, stir for 1 hour, separate the organic layer, adjust the pH of the aqueous layer to 2-3 with 3N HCl, extract with ethyl acetate (10ml x 2), wash with saturated brine (8ml x 2), anhydrous After drying over sodium sulfate, the solvent was removed under reduced pressure to obtain 0.37 g of a colorless oily liquid with a yield of 58%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.14(d, J=7.5Hz, 1H), 7.57–7.51(m, 1H), 7.40(d, J=4.0Hz, 1H), 7.17–7.10(m, 1H),7.04(d,J=8.0Hz,1H),5.55(d,J=6.5Hz,1H),4.76–4.70(m,1H),4.39–4.30(m,3H),3.94(dd,J ＝9.5,2.5Hz,1H),3.91–3.86(m,2H),3.81(dd,J＝9.5,4.0Hz,1H),3.77–3.71(m,4H),3.70–3.63(m,4H), 3.55–3.48(m,1H), 3.38(s,3H), 1.44(s,9H)ppm; ESI-MS: m/z=529[M+H] ⁺ .

Reactant 2-(((6S,9S)-9-(methoxycarbonyl)-6-(methoxymethyl)-2,2-dimethyl-4,7-dioxo 3,11, 14 Trioxa-5,8-diazahexadec-16-oxy)benzoic acid (0.37g, 0.69mmol) was dissolved in 2.0mL CH ₂ Cl ₂ , cooled to 0°C in an ice bath, and 1.0mL Tris Fluoroacetic acid was reacted at room temperature for 2 hours. The solvent was evaporated from the reaction solution under reduced pressure to obtain a light yellow solid, which was directly used in the next reaction.

2-(((4S,7S)-4-amino-7-(methoxycarbonyl)-5-oxo-2,9,12-trioxa-6-azatetradecyl-14-yl) oxy)benzoic acid trifluoroacetate (0.69 mmol), 1-hydroxybenzotriazole (0.11 g, 0.83 mmol) and benzotriazole-N,N,N',N'-tetramethyl Urea hexafluorophosphate (0.31g, 0.83mmol) was dissolved in 138ml of anhydrous THF, cooled to 0°C, DIPEA (0.27mL, 1.7mmol) was added dropwise, and reacted at room temperature for 5 hours. The solvent was removed under reduced pressure, 10 ml of ethyl acetate and 8 ml of water were added, and the organic layer was separated. Subsequently, washed successively with saturated sodium bicarbonate solution (10mL x 2) and saturated brine (8mL x2), dried over anhydrous sodium sulfate and removed the solvent under reduced pressure, separated by column chromatography to obtain 0.15 g of a colorless oily liquid, collected in two steps rate 53%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.90(d, J=9.5Hz, 1H), 8.25(dd, J=8.0, 2.0Hz, 1H), 7.51–7.45(m, 1H), 7.38(d, J＝8.5Hz, 1H), 7.14–7.08(m, 1H), 6.97(d, J＝8.0Hz, 1H), 5.13-5.07(m, 1H), 4.84(dt, J＝8.5, 3.0Hz, 1H ),4.42–4.35(m,1H),4.31–4.24(m,1H),4.16(dd,J＝9.0,2.0Hz,1H),4.07(dd,J＝10.0,2.0Hz,1H),3.96– 3.89(m,2H),3.86–3.79(m,1H),3.75–3.66(m,4H),3.62(dd,J＝10.0,3.0Hz,1H),3.59–3.51(m,2H),3.50– 3.45(m,1H), 3.39(s,3H)ppm; ESI-MS: m/z=411[M+H] ⁺ .

The above raw material (9S,12S)-12-(methoxymethyl)-11,14-dioxo-2,3,5,6,9,10,11,12,13,14-decahydro-8H -Benzo[o][1,4,7]-trioxa[10,13]-diazacyclohexadec-9-carboxylic acid methyl ester (0.14 g, 0.34 mmol) was dissolved in 1.3 mL of acetone, Cool to 0° C. in an ice bath, add 1.7 ml of 0.3 N lithium hydroxide aqueous solution dropwise, and react at this temperature for 30 minutes. Slowly add 3N HCl to adjust the pH to 2-3, remove acetone under reduced pressure, extract with ethyl acetate (3mL x 2), wash with saturated brine (2mL x2), dry over anhydrous sodium sulfate, and remove the solvent under reduced pressure to obtain a colorless oily liquid 0.11 mg, yield 82%. The product obtained is directly used in the next step reaction.

Preparation Example 153, N-[(12S,15S)-15-(methoxymethyl)-14,17-dioxo-2,3,5,6,8,9,12,13,14, 15,16,17-Dodecahydro-11H-benzo[r][1,4,7,10]tetraoxa[13,16]-diazacyclinedecane-12-carboxylic acid (9zb)

Starting material N-(N-(tert-butoxycarbonyl)-O-methyl-L-seryl)-O-(2-(2–((2-allyloxycarbonyl)phenoxyethoxy )ethoxy)ethyl)-L-serine methyl ester (8zb, 0.12g, 0.20mmol) was placed in a 10ml round bottom flask, tetrakistriphenylphosphine palladium (0.011g, 0.01mmol) and 1,3- Dimethylbarbituric acid (0.03g, 0.20mmol), injected 4ml of dichloromethane under _N2 protection, reacted overnight at room temperature. Add 5ml of saturated sodium bicarbonate solution, stir for 1 hour and separate the organic layer, adjust the pH of the aqueous layer to 2-3 with 3N HCl, extract with ethyl acetate (5ml x 2), wash with saturated brine (4ml x 2), and remove Dry over sodium sulfate, and remove the solvent under reduced pressure to obtain 0.098 g of a colorless oily liquid, with a yield of 86%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.15(d, J=7.5Hz, 1H), 7.57–7.51(m, 1H), 7.37(d, J=5.5Hz, 1H), 7.17–7.11(m, 1H), 7.05(d, J＝8.0Hz, 1H), 5.48(d, J＝5.0Hz, 1H), 4.72–4.66(m, 1H), 4.41–4.36(m, 2H), 4.33–4.25(m ,1H),3.96–3.90(m,3H),3.79(dd,J＝9.0,4.0Hz,1H),3.76–3.64(m,9H),3.62–3.58(m,4H),3.49(dd,J =8.5,7.0Hz,1H),3.38(s,3H),1.45(s,9H)ppm; ESI-MS: m/z=573[M+H] ⁺ .

Reactant 2-(((6S,9S)-9-(methoxycarbonyl)-6-(methoxymethyl)-2,2-dimethyl-4,7-dioxo-3,11 ,14,17-tetraoxa-5,8-diazanonadecan-19-yl)oxy)benzoic acid (0.16g, 0.28mmol) was dissolved in 0.6mL CH ₂ Cl ₂ , cooled to 0 in an ice bath ℃, add 0.3mL trifluoroacetic acid, react at room temperature for 2 hours. The solvent was distilled off from the reaction solution under reduced pressure to obtain a light yellow solid, which was directly used in the next reaction.

2–(((4S,7S)-4-amino-7-(methoxycarbonyl)-5-oxo-2,9,12,15-tetraoxa-6-azaheptadecane-17- base)oxy)benzoic acid trifluoroacetate (0.16g, 0.28mmol), 1-hydroxybenzotriazole (0.045g, 0.34mmol) and benzotriazole-N,N,N',N '-Tetramethylurea hexafluorophosphate (0.13g, 0.34mmol) was dissolved in 93ml of anhydrous THF, cooled to 0°C, DIPEA (92μL, 0.56mmol) was added dropwise, and reacted at room temperature for 5 hours. The solvent was removed under reduced pressure, 5 ml of ethyl acetate and 4 ml of water were added, and the organic layer was separated. Subsequently, washed successively with saturated sodium bicarbonate solution (5mL x 2) and saturated brine (4mL x 2), dried over anhydrous sodium sulfate and removed the solvent under reduced pressure, separated by column chromatography to obtain 59mg of a colorless oily liquid, collected in two steps rate 46%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.77(d, J=7.5Hz, 1H), 8.23(d, J=8.0Hz, 1H), 7.49–7.42(m, 1H), 7.30(d, J= 7.0Hz, 1H), 7.13–7.05(m, 1H), 6.97(d, J＝8.5Hz, 1H), 4.95–4.88(m, 1H), 4.77–4.70(m, 1H), 4.42–4.35(m ,1H),4.28–4.20(m,1H),4.11–4.01(m,2H),3.91–3.82(m,3H),3.73–3.64(m,8H),3.64–3.58(m,2H),3.57 –3.47(m,2H), 3.41(s,3H)ppm; ESI-MS: m/z=455[M+H] ⁺ .

The above raw materials (12S,15S)-15-(methoxymethyl)-14,17-dioxo-2,3,5,6,8,9,12,13,14,15,16,17- Dodecahydro-11H-benzo[r][1,4,7,10]tetraoxa[13,16]-diazacyclic nonadecane-12-carboxylic acid methyl ester (65mg, 0.14mmol) In 0.5 mL of acetone, cooled to 0° C. under an ice bath, 0.7 ml of 0.3 N LiOH aqueous solution was added dropwise, and the reaction was carried out at this temperature for 30 minutes. Slowly add 3N HCl to adjust the pH to 2-3, remove acetone under reduced pressure, extract with ethyl acetate (1mL x 2), wash with saturated brine (1mL x 2), dry over anhydrous sodium sulfate, remove the solvent under reduced pressure to obtain a colorless oil Liquid 53mg, yield 86%. The product obtained is directly used in the next step reaction.

PREPARATIVE EXAMPLE 154, (10S,13S)-13-(2-Morpholino-2-oxoethyl)-12,15-dioxo-2,3,4,5,6,7,10, 11,12,13,14,15-Dodecahydro-9Hbenzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-carboxylic acid (9zc)

N-((S)-4-morpholino-4-oxo-2-(2-(4-penten-1-yloxy)benzamido)butyryl)-O-allyl - L-serine benzyl ester as the raw material, the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 62 mg of off-white foamy solid, and the two-step yield was 63%. ¹ H NMR (500MHz, CDCl ₃ ) δ9.40(d, J=9.5Hz, 1H), 8.24(dd, J=8.0, 2.0Hz, 1H), 7.49–7.41(m, 1H), 7.20(d, J＝7.0Hz, 1H), 7.09–7.02(m, 1H), 6.97(d, J＝8.0Hz, 1H), 5.36–5.28(m, 1H), 4.78–4.71(m, 1H), 4.25–4.12 (m,2H),3.83–3.73(m,2H),3.69–3.61(m,5H),3.55–3.46(m,4H),3.43–3.37(m,2H),2.56(dd,J＝16.5, 4.5Hz,1H),2.19–2.09(m,1H),1.89–1.79(m,1H),1.75–1.61(m,2H),1.58–1.44(m,4H)ppm; ESI-MS: m/z ＝492[M+H] ⁺ .

Preparative Example 155, (10S,13S)-13-(3-morpholino-3-oxopropyl)-12,15-dioxo-2,3,4,5,6,7,10, 11,12,13,14,15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-carboxylic acid (9zd)

N-((S)-5-morpholino-5-oxo-2-(2-(4-penten-1-yloxy)benzamido)pentanoyl)-O-allyl -L-serine benzyl ester as the raw material, the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 70 mg of off-white solid, and the two-step yield was 69%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.67(d, J=7.5Hz, 1H), 8.16–8.11(m, 1H), 7.48–7.42(m, 1H), 7.29(d, J=8.0Hz, 1H),7.09–7.03(m,1H),6.96(d,J＝8.5Hz,1H),4.87–4.80(m,1H),4.29–4.23(m,1H),4.17–4.08(m,2H) ,3.81(dd,J=10.0,4.0Hz,1H),3.71(dd,J=10.0,6.5Hz,1H),3.68–3.39(m,10H),2.56–2.48(m,1H),2.47–2.40 (m,1H),2.38–2.29(m,1H),2.23–2.14(m,1H),2.06–1.95(m,2H),1.86–1.78(m,1H),1.69–1.59(m,2H) ,1.55–1.42(m,3H)ppm; ESI-MS: m/z=506[M+H] ⁺ .

Preparation Example 156, (10S,13S)-13-(2-(cyclopropylamino)-2-oxoethyl)-12,15-dioxo-2,3,4,5,6,7 ,10,11,12,13,14,15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-carboxylic acid ( 9ze)

With N-(N ⁴ -cyclopropyl-N ² -(2-(4-penten-1-yloxy)benzoyl)-L-asparagyl)-O-allyl-L- Serine benzyl ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 70 mg of off-white oily liquid with a two-step yield of 76%. ESI-MS: m/z＝462[M+H] ⁺ .

Preparation Example 157, (10S,13S)-13-isobutyl-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-deca Dihydro-9H-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-carboxylic acid (9zf)

Using N-((2-(pent-4-en-1-yloxy)benzoyl)-L-leucyl)-O-allyl-L-serine benzyl ester as raw material, synthesis and post-treatment The same as in Preparation Example 126, 79 mg of off-white solid was obtained, and the two-step yield was 94%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.47 (d, J=8.5Hz, 1H), 8.25 (dd, J=8.0, 1.5Hz, 1H), 7.51–7.44 (m, 1H), 7.16–7.06 ( m,2H),7.01(d,J=8.0Hz,1H),4.84(dd,J=9.0,4.0Hz,1H),4.76–4.70(m,1H),4.33–4.26(m,1H),4.19 –4.09(m,1H),3.76(d,J＝4.0Hz,2H),3.57–3.50(m,1H),3.42–3.35(m,1H),1.98–1.88(m,1H),1.85–1.76 (m,2H),1.69–1.32(m,6H),1.30–1.23(m,2H),0.98(d,J=7.0Hz,3H),0.95(d,J=7.0Hz,3H) ppm; ESI -MS: m/z＝421[M+H] ⁺ .

Preparation Example 158, (10S,13S)-13-isopropyl-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-deca Dihydro-9H-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-carboxylic acid (9zg)

Using N-((2-(pent-4-en-1-yloxy)benzoyl)-L-valyl)-O-allyl-L-serine benzyl ester as raw material, synthesis and post-treatment In the same manner as in Preparation Example 126, 67 mg of off-white solid was obtained, and the two-step yield was 82%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.50(d, J=8.5Hz, 1H), 8.22(dd, J=8.0, 1.0Hz, 1H), 7.51–7.44(m, 1H), 7.17(d, J＝7.0Hz, 1H), 7.11–7.06(m, 1H), 6.98(d, J＝8.5Hz, 1H), 4.88(td, J＝9.6, 4.5Hz, 1H), 4.79–4.73(m, 1H ),4.30–4.24(m,1H),4.19–4.10(m,1H),3.78–3.69(m,2H),3.52–3.41(m,2H),2.01–1.93(m,1H),1.90–1.82 (m,2H),1.75–1.66(m,2H),1.65–1.43(m,4H),1.00–0.91(m,6H)ppm; ESI-MS: m/z＝407[M+H] ⁺ .

Preparative Example 159, (9S,12S)-12-(methoxymethyl)-11,14-dioxo-3,4,5,6,7,8,9,10,11,12,13 ,14-Dodecahydro-2H-benzo[b][1]oxa[5,8]-diazacyclohexadeca-9-carboxylic acid (9zh)

(S)-2-(N-(2-(pent-4-en-1-yloxy)benzoyl)-O-methyl-L-seryl)aminohex-5-enoic acid benzyl The ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 62 mg of off-white solid, with a two-step yield of 79%. ¹ HNMR (500MHz, CDCl ₃ ) δ9.02(d, J＝8.5Hz, 1H), 8.24(dd, J＝8.0, 1.5Hz, 1H), 7.51–7.44(m, 1H), 7.11–7.05(m ,1H),7.00–6.92(m,2H),4.97–4.91(m,1H),4.66–4.59(m,1H),4.25–4.19(m,1H),4.18–4.08(m,2H),3.52 (dd,J=9.0,4.0Hz,1H),3.41(s,3H),2.11–1.89(m,3H),1.83–1.74(m,1H),1.69–1.62(m,1H),1.60–1.52 (m,1H),1.50–1.29(m,6H)ppm; ESI-MS: m/z=393[M+H] ⁺ .

Preparative Example 160, (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,8,9,10,11,12 ,13,14,15-tetrahydro-benzo[b][1]oxa[5,8]-diazacyclohexadeca-10-carboxylic acid (9zi)

Benzyl (S)-2-(N-(2-(hex-5-en-1-yloxy)benzoyl)-O-methyl-L-seryl)aminohex-5-enoate The ester was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 126 to obtain 66 mg of off-white solid, with a two-step yield of 81%. ¹ HNMR (500MHz, CDCl ₃ ) δ8.70(d, J＝8.0Hz, 1H), 8.13(dd, J＝8.0, 1.5Hz, 1H), 7.45–7.38(m, 1H), 7.06–7.00(m ,1H),6.94(d,J=8.0Hz,1H),6.89(d,J=8.0Hz,1H),4.94–4.85(m,1H),4.64(td,J=8.5,3.5Hz,1H) ,4.25–4.18(m,1H),4.06–3.98(m,2H),3.46(dd,J＝9.0,5.5Hz,1H),3.35(s,3H),2.02–1.91(m,2H),1.71 –1.44(m,4H),1.41–1.11(m,8H)ppm; ESI-MS: m/z=407[M+H] ⁺ .

Preparation Example 161, N-[(8S,11S)-11-(methoxymethyl)-10,13-dioxo-3,4,5,7,8,9,10,11,12, 13-Decahydro-2H-1,6,9,12-benzodioxadiazacyclopentadecyl-8-formyl]-Phe-epoxyketone (10a)

Starting material (8S,11S)-11-(methoxymethyl)-10,13-dioxo-3,4,5,7,8,9,10,11,12,13-decahydro-2H- 1,6,9,12-Benzodioxadiazacyclopentadeca-8-carboxylic acid (9a, 38 mg, 0.10 mmol) was dissolved in 2 mL of anhydrous THF, and 1-hydroxybenzotriazole ( 16mg, 0.12mmol), benzotriazole-N,N,N',N'-tetramethyluronium hexafluorophosphate (45mg, 0.12mmol) and Phe-epoxyketone trifluoroacetate (38mg, 0.12 mmol), cooled to 0°C in an ice bath. Subsequently, DIPEA (0.033 mL, 0.20 mmol) was added and reacted at room temperature for 3 hours. The solvent was removed under reduced pressure, 5 mL of ethyl acetate and 4 mL of water were added, the organic layer was separated, washed successively with saturated sodium bicarbonate solution (3 mL x 2) and saturated brine (3 mL x 2), dried over anhydrous sodium sulfate, and decompressed The solvent was distilled off, and 42 mg of white solid was separated by column chromatography, with a yield of 74%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.03–7.97 (m, 2H), 7.49 (ddd, J=8.5, 7.5, 2.0Hz, 1H), 7.15–7.10 (m, 2H), 7.07–6.94 (m ,7H),4.84–4.78(m,1H),4.78–4.72(m,1H),4.56–4.51(m,1H),4.20–4.15(m,1H),4.12–4.06(m,1H),4.04 (dd,J=9.0,2.5Hz,1H),3.76(dd,J=9.5,4.5Hz,1H),3.56(dd,J=9.5,3.5Hz,1H),3.53–3.44(m,2H), 3.37–3.30(m,5H),3.06(dd,J=14.0,4.0Hz,1H),2.87(d,J=5.0Hz,1H),2.70(dd,J=14.0,9.5Hz,1H),1.99 –1.81(m,2H),1.71–1.60(m,1H),1.51–1.41(m,4H)ppm; ESI-MS: m/z=568[M+H] ⁺ .

Preparation Example 162, N-[(9S,12S)-12-(methoxymethyl)-11,14-dioxo-2,3,4,5,6,8,9,10,11, 12,13,14-Dodecahydro-1,7,10,13-benzodioxadiazacyclohexadeca-9-formyl]-Phe-epoxyketone (10b)

With (9S,12S)-12-(methoxymethyl)-11,14-dioxo-2,3,4,5,6,8,9,10,11,12,13,14-deca Dihydro-1,7,10,13-benzodioxadiazacyclohexadecane-9-carboxylic acid was used as raw material, and the synthesis and post-treatment were the same as in Preparation Example 161 to obtain 45 mg of white solid with a yield of 79% . ¹ H NMR (500MHz, CDCl ₃ ) δ8.78(d, J=5.0Hz, 1H), 8.19(d, J=7.5Hz, 1H), 7.54–7.44(m, 1H), 7.10–6.92(m, 9H),4.80–4.67(m,2H),4.66–4.58(m,1H),4.39–4.20(m,2H),4.07–3.96(m,2H),3.68–3.58(m,1H),3.56– 3.50(m,1H),3.40(s,3H),3.34(d,J＝5.0Hz,1H),3.31–3.27(m,1H),3.22–3.15(m,1H),3.04(dd,J＝ 13.5,4.5Hz,1H),2.82(d,J＝4.0Hz,1H),2.73(dd,J＝13.5,9.0Hz,1H),1.93–1.87(m,1H),1.83–1.76(m,1H ), 1.65–1.54(m,2H), 1.48–1.39(m,5H)ppm; ESI-MS: m/z＝582[M+H] ⁺ .

Preparation Example 163, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-3,4,5,6,7,9,10,11,12, 13,14,15-2H-Dodecahydro-1,8,11,14-benzodioxadiazacyclohexadeca-10-formyl]-Phe-epoxyketone (10c)

(10S,13S)-13-(methoxymethyl)-12,15-dioxo-3,4,5,6,7,9,10,11,12,13,14,15-2H -Dodecahydro-1,8,11,14-benzodioxadiazacyclohexadecan-10-carboxylic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 161 to obtain 42mg of white solid, the yield 71%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.90 (d, J=6.5Hz, 1H), 8.23 (dd, J=8.0, 2.0Hz, 1H), 7.52–7.46 (m, 1H), 7.13–6.96 ( m,8H),6.87(d,J=8.0Hz,1H),4.78–4.72(m,1H),4.70–4.65(m,1H),4.58(dt,J=7.5,4.5Hz,1H),4.31 –4.27(m,1H),4.15–4.12(m,1H),4.04(dd,J＝9.5,2.0Hz,1H),3.64(dd,J＝9.5,4.0Hz,1H),3.60(dd,J =9.5,5.0Hz,1H),3.50(dd,J=10.0,4.0Hz,1H),3.41(s,3H),3.35–3.28(m,3H),3.04(dd,J=14.0,4.5Hz, 1H), 2.83(d, J=5.0Hz, 1H), 2.70(dd, J=14.0, 9.0Hz, 1H), 1.99–1.91(m,1), 1.88–1.80(m,1H), 1.64–1.56 (m,2H),1.51–1.45(m,2H),1.44–1.40(m,5H)ppm; ESI-MS: m/z=596[M+H] ⁺ .

Preparation Example 164, N-[(11S,14S)-14-(methoxymethyl)-13,16-dioxo-2,3,4,5,6,7,8,10,11, 12,13,14,15,16-tetradetrahydro-1,9,12,15-benzodioxadiazacycloctadec-11-formyl]-Phe-epoxyketone (10d)

(11S,14S)-14-(methoxymethyl)-13,16-dioxo-2,3,4,5,6,7,8,10,11,12,13,14,15 , 16-Tetrahydro-1,9,12,15-benzodioxadiazacycloctadec-11-carboxylic acid as raw material, the synthesis and post-treatment were the same as Preparation Example 161 to obtain 46 mg of white solid, Yield 71%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.89(d, J=5.5Hz, 1H), 8.19(dd, J=8.0, 1.5Hz, 1H), 7.49–7.43(m, 1H), 7.21(d, J＝8.0Hz, 1H), 7.15–6.98(m, 7H), 6.78(d, J＝8.0Hz, 1H), 4.82(td, J＝8.5,5.5Hz, 1H), 4.72–4.66(m, 2H ),4.35–4.27(m,1H),4.11–4.05(m,1H),3.98(dd,J＝9.5,3.5Hz,1H),3.72–3.61(m,2H),3.52(dd,J＝10.0 ,4.0Hz,1H),3.47–3.39(m,4H),3.36–3.31(m,2H),3.08(dd,J＝14.0,5.5Hz,1H),2.83(d,J＝5.0Hz,1H) ,2.78(dd,J＝14.0,9.0Hz,1H),2.02–1.96(m,1H),1.88–1.79(m,1H),1.61–1.52(m,1H),1.50–1.31(m,10H) ppm; ESI-MS: m/z=610[M+H] ⁺ .

Preparation Example 165, N-[(12S,15S)-15-(methoxymethyl)-14,17-dioxo-3,4,5,6,7,8,9,11,12, 13,14,15,16,17-2H-Tetradetrahydro-2H-1,10,13,16-Benzodioxadiazacyclonadeca-12-formyl]-Phe-epoxyketone (10e)

(12S,15S)-15-(methoxymethyl)-14,17-dioxo-3,4,5,6,7,8,9,11,12,13,14,15,16 , 17-2H-tetradecylhydro-2H-1,10,13,16-benzodioxadiazacycline nineadecane-12-carboxylic acid as raw material, the synthesis and post-treatment are the same as in Preparation Example 161, to obtain White solid 46mg, yield 71%. ¹ H NMR (500MHz, CDCl ₃ ) δ9.00(d, J=4.0Hz, 1H), 8.27(dd, J=8.0, 2.0Hz, 1H), 7.52–7.45(m, 1H), 7.42(d, J＝8.0Hz, 1H), 7.18–6.99(m, 7H), 6.80(d, J＝8.0Hz, 1H), 4.82(td, J＝8.5,5.5Hz, 1H), 4.67–4.60(m, 2H ),4.28–4.17(m,2H),3.95(dd,J＝9.5,4.5Hz,1H),3.70(dd,J＝9.5,4.5Hz,1H),3.63(dd,J＝9.5,5.0Hz, 1H), 3.51(dd, J=9.5, 4.5Hz, 1H), 3.46–3.28(m, 6H), 3.11(dd, J=14.0, 5.5Hz, 1H), 2.88(dd, J=14.0, 8.5Hz ,1H),2.83(d,J＝5.0Hz,1H),2.03–1.97(m,1H),1.95–1.77(m,2H),1.66–1.53(m,2H),1.49–1.29(m,10H )ppm; ESI-MS: m/z＝624[M+H] ⁺ .

Preparation Example 166, N-[(13S,16S)-16-(methoxymethyl)-15,18-dioxo-3,4,5,6,7,8,9,10,13, 14,15,16,17,18-Tetrahydro-2H,12H-benzo[i][1,11]dioxa[4,7]diazacycloeicos-13-formyl]-Phe - Epoxyketone (10f)

With (13S,16S)-16-(methoxymethyl)-15,18-dioxo-3,4,5,6,7,8,9,10,13,14,15,16,17 , 18-tetrahydro-2H, 12H-benzo[i][1,11]dioxa[4,7]diazacyclineicos-13-carboxylic acid as raw material, the synthesis and post-treatment are the same as the preparation Example 161, 42mg of white solid was obtained, the yield was 66%. ¹ H NMR (500MHz, CDCl ₃ ) δ9.02 (d, J=4.5Hz, 1H), 8.22 (dd, J=8.0, 2.0Hz, 1H), 7.49–7.41 (m, 2H), 7.25–7.11( m,5H),7.06–6.97(m,3H),4.83(td,J＝8.0,5.5Hz,1H),4.70–4.64(m,1H),4.58–4.50(m,1H),4.24–4.06( m,2H),3.89(dd,J=9.5,4.5Hz,1H),3.71(dd,J=9.5,5.0Hz,1H),3.59(dd,J=9.5,6.5Hz,1H),3.50–3.34 (m,7H),3.13(dd,J＝14.0,5.0Hz,1H),2.91–2.82(m,2H),2.12–2.02(m,1H),1.95–1.85(m,1H),1.64–1.23 (m,15H)ppm; ESI-MS: m/z＝638[M+H] ⁺ .

Preparation Example 167, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12, 13,14,15-Dodecahydro-9H-17-chlorobenzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-formyl]-Phe-cyclo Oxyketone (10g)

With (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-deca Dihydro-9H-17-chlorobenzo[i][1,11]dioxa[4,7]diazacyclohexadecan-10-carboxylic acid as raw material, the synthesis and post-treatment are the same as in Preparation Example 161 , to obtain 49 mg of white solid, yield 78%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.82 (d, J=6.5Hz, 1H), 8.19 (d, J=3.0Hz, 1H), 7.41 (dd, J=9.0, 2.5Hz, 1H), 7.14 –7.02(m,6H),6.93(d,J=9.0Hz,1H),6.78(d,J=8.0Hz,1H),4.76(td,J=8.5,5.0Hz,1H),4.73–4.69( m,1H),4.59(dt,J＝7.5,5.0Hz,1H),4.29–4.22(m,1H),4.13–4.03(m,2H),3.63–3.57(m,2H),3.52(dd, J=10.0,4.0Hz,1H),3.42–3.29(m,6H),3.06(dd,J=14.0,4.5Hz,1H),2.83(d,J=5.0Hz,1H),2.79–2.74(m ,1H),2.01–1.90(m,1H),1.88–1.80(m,1H),1.66–1.56(m,2H),1.55–1.39(m,7H)ppm; ESI-MS: m/z＝631 [M+H] ⁺ .

Preparation Example 168, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12, 13,14,15-Dodecahydro-9H-18-chlorobenzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-formyl]-Phe-cyclo Oxyketone (10h)

With (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-deca Dihydro-9H-18-chlorobenzo[i][1,11]dioxa[4,7]diazacyclohexadecan-10-carboxylic acid as raw material, the synthesis and post-treatment are the same as in Preparation Example 161 , to obtain 44 mg of white solid, yield 70%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.73(d, J=7.0Hz, 1H), 8.16(d, J=8.5Hz, 1H), 7.12–7.07(m, 4H), 7.06–7.02(m, 3H), 6.99(d, J=1.5Hz, 1H), 6.80(d, J=8.0Hz, 1H), 4.78–4.68(m, 2H), 4.58(dt, J=7.5, 5.5Hz, 1H), 4.30–4.24(m,1H),4.15–4.08(m,1H),4.05(dd,J＝9.0,2.0Hz,1H),3.64–3.57(m,2H),3.51(dd,J＝9.5,4.0 Hz,1H),3.39(s,3H),3.38–3.29(m,3H),3.04(dd,J＝14.0,5.0Hz,1H),2.82(d,J＝5.0Hz,1H),2.75(dd ,J＝14.0,8.5Hz,1H),2.00–1.91(m,1H),1.89–1.81(m,1H),1.67–1.57(m,2H),1.54–1.40(m,7H)ppm; ESI- MS: m/z＝631[M+H] ⁺ .

Preparation Example 169, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12, 13,14,15-Dodecahydro-9H-18-fluorobenzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-formyl]-Phe-cyclo Oxyketone (10i)

With (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-deca Dihydro-9H-18-fluorobenzo[i][1,11]dioxa[4,7]diazacyclohexadecan-10-carboxylic acid as raw material, the synthesis and post-treatment are the same as in Preparation Example 161 , to obtain 42mg of white solid, yield 68%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.72 (d, J=6.5Hz, 1H), 8.28–8.21 (m, 1H), 7.12–7.02 (m, 6H), 6.84–6.74 (m, 2H), 6.71(dd,J=10.5,2.5Hz,1H),4.79–4.70(m,2H),4.59(dt,J=8.0,5.5Hz,1H),4.30–4.22(m,1H),4.13–4.03( m,2H),3.64–3.56(m,2H),3.51(dd,J＝9.9,4.0Hz,1H),3.40(s,3H),3.37–3.31(m,3H),3.05(dd,J＝ 14.0,5.0Hz,1H),2.83(d,J=5.0Hz,1H),2.75(dd,J=14.0,9.0Hz,1H),2.02–1.93(m,1H),1.90–1.81(m,1H ), 1.69–1.58(m,2H), 1.57–1.40(m,7H)ppm; ESI-MS: m/z＝614[M+H] ⁺ .

Preparation Example 170, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12, 13,14,15-Dodecahydro-9H-18-methylbenzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-formyl]-Phe- Epoxy ketone (10j)

With (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-deca Dihydro-9H-18-methylbenzo[i][1,11]dioxa[4,7]diazacyclohexadecane-10-carboxylic acid as raw material, the synthesis and post-treatment are the same as the preparation examples 161, 48mg of white solid was obtained, the yield was 79%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.84 (d, J = 6.5Hz, 1H), 8.12 (d, J = 8.0Hz, 1H), 7.18 (d, J = 8.1Hz, 1H), 7.11–7.01 (m,5H),6.87(d,J=8.0Hz,1H),6.83–6.76(m,2H),4.79–4.69(m,2H),4.58(dt,J=8.0,5.5Hz,1H), 4.31–4.22(m,1H),4.17–4.09(m,1H),4.06(dd,J＝9.5,2.0Hz,1H),3.61(dd,J＝9.0,3.5Hz,1H),3.57(dd, J=9.5,5.5Hz,1H), 3.49(dd,J=9.5,4.0Hz,1H),3.39(s,3H),3.36–3.28(m,3H),3.04(dd,J=14.0,5.0Hz ,1H),2.82(d,J=5.0Hz,1H),2.75(dd,J=14.0,9.0Hz,1H),2.39(s,3H),2.00–1.91(m,1H),1.88–1.79( m,1H),1.67–1.56(m,2H),1.53–1.40(m,7H)ppm; ESI-MS: m/z＝610[M+H] ⁺ .

Preparation Example 171, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12, 13,14,15-Dodecahydro-9H-18-methoxybenzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-formyl]-Phe - Epoxyketone (10k)

With (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-deca Dihydro-9H-18-methoxybenzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-carboxylic acid as raw material, the synthesis and post-treatment are carried out in the same way as the preparation Example 161, 50 mg of white solid was obtained, the yield was 80%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.74(d, J=6.5Hz, 1H), 8.19(d, J=9.0Hz, 1H), 7.18(d, J=8.0Hz, 1H), 7.11–7.03 (m,5H),6.82(d,J=8.0Hz,1H),6.57(dd,J=8.8,2.5Hz,1H),6.50(d,J=2.5Hz,1H),4.78–4.70(m, 2H), 4.58(dt, J＝8.0, 5.5Hz, 1H), 4.28–4.21(m, 1H), 4.13–4.02(m, 2H), 3.85(s, 3H), 3.63–3.55(m, 2H) ,3.49(dd,J＝10.0,4.0Hz,1H),3.39(s,3H),3.36–3.30(m,3H),3.04(dd,J＝14.0,5.0Hz,1H),2.82(d,J =5.0Hz,1H),2.76(dd,J=14.0,9.0Hz,1H),2.00–1.91(m,1H),1.88–1.79(m,1H),1.65–1.37(m,9H)ppm; ESI -MS: m/z＝626[M+H] ⁺ .

Preparation Example 172, N-[(3S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15, 16,17,18-Dodecahydro-12H-2-methylthiazolo[5,4-i][1,11]dioxa[4,7]diazacyclohexadeca-13-formyl ]-Phe-epoxyketone (10l)

With (3S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15,16,17,18-deca Dihydro-12H-2-methylthiazolo[5,4-i][1,11]dioxa[4,7]diazacyclohexadeca-13-carboxylic acid as raw material, synthesis and post-treatment The same as in Preparation Example 161, 46 mg of white solid was obtained with a yield of 75%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.96(d, J=7.5Hz, 1H), 7.21–7.13(m, 3H), 7.11–7.07(m, 2H), 7.04(d, J=8.0Hz, 1H), 6.86(d, J=7.5Hz, 1H), 4.78–4.69(m, 2H), 4.66–4.60(m, 1H), 4.54(dt, J=9.0, 5.0Hz, 1H), 4.45(dt ,J=10.5,4.0Hz,1H),4.06(dd,J=9.0,2.0Hz,1H),3.71(dd,J=10.0,5.0Hz,1H),3.56(dd,J=9.0,3.5Hz, 1H), 3.47(dd, J＝10.0, 4.0Hz, 1H), 3.44–3.37(m, 4H), 3.34–3.27(m, 2H), 3.05(dd, J＝14.0, 5.0Hz, 1H), 2.83 (d,J=5.0Hz,1H),2.81–2.75(m,1H),2.61(s,3H),1.88–1.80(m,2H),1.58–1.45(m,6H),1.42(s,3H )ppm; ESI-MS: m/z＝617[M+H] ⁺ .

Preparation Example 173, N-[(6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,12,13,14, 15,16,17-Dodecahydro-4H-thieno[3,2-i][1,11]dioxa[4,7]diazacyclohexadeca-9-formyl]-Phe- Epoxy ketone (10m)

With (6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,12,13,14,15,16,17-deca Dihydro-4H-thieno[3,2-i][1,11]dioxa[4,7]diazacyclohexadecan-9-carboxylic acid as raw material, the synthesis and post-treatment are the same as the preparation examples 161, 40 mg of white solid was obtained, the yield was 66%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.15(d, J=7.5Hz, 1H), 7.44(d, J=5.5Hz, 1H), 7.20–7.08(m, 5H), 7.05(d, J= 8.0Hz,1H),6.89–6.83(m,2H),4.81–4.70(m,2H),4.60–4.51(m,1H),4.39–4.31(m,1H),4.20–4.13(m,1H) ,4.11–4.04(m,1H),3.67(dd,J＝10.0,6.0Hz,1H),3.57(dd,J＝9.0,3.0Hz,1H),3.48(dd,J＝10.0,3.5Hz,1H ),3.42–3.27(m,6H),3.06(dd,J＝14.0,5.0Hz,1H),2.82(d,J＝5.0Hz,1H),2.77(dd,J＝13.5,8.5Hz,1H) ,1.94–1.78(m,2H),1.63–1.44(m,6H),1.42(s,3H)ppm; ESI-MS: m/z=602[M+H] ⁺ .

Preparation Example 174, N-[(14S,17S)-17-(methoxymethyl)-16,19-dioxo-6,7,8,9,10,11,14,15,16, 17,18,19-Dodecahydro-13H-pyrido[3-i][1,11]dioxa[4,7]diazacyclohexadeca-14-formyl]-Phe-epoxy Ketone (10n)

With (14S,17S)-17-(methoxymethyl)-16,19-dioxo-6,7,8,9,10,11,14,15,16,17,18,19-deca Dihydro-13H-pyrido[3-i][1,11]dioxa[4,7]diazacyclohexadecan-14-carboxylic acid as raw material, the synthesis and post-treatment are the same as in Preparation Example 161, 39 mg of white solid was obtained, and the yield was 65%. ESI-MS: m/z＝597[M+H] ⁺ .

Preparation Example 175, N-[(7S,10S)-7-(methoxymethyl)-5,8-dioxo-6,7,8,9,10,11,13,14,15, 16,17,18-Dodecahydro-5H-pyrido[3,2-i][1,11]dioxa[1,11]diazacyclohexadeca-10-formyl]-Phe- Epoxy ketone (10o)

(7S,10S)-7-(methoxymethyl)-5,8-dioxo-6,7,8,9,10,11,13,14,15,16,17,18-dodeca Hydrogen-5H-pyrido[3,2-i][1,11]dioxa[1,11]diazacyclohexadecan-10-carboxylic acid as raw material, the synthesis and post-treatment are the same as in Preparation Example 161 , 40mg of white solid was obtained, the yield was 67%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.90(d, J=7.0Hz, 1H), 8.50(dd, J=7.6, 2.0Hz, 1H), 8.29(dd, J=4.8, 2.0Hz, 1H) ,7.12–6.98(m,7H),6.81(d,J＝7.8Hz,1H),4.78–4.69(m,3H),4.60–4.53(m,1H),4.41–4.35(m,1H),4.07 (dd,J=9.3,2.1Hz,1H),3.65–3.58(m,2H),3.50(dd,J=9.8,4.0Hz,1H),3.41(s,3H),3.37–3.27(m,3H ),3.04(dd,J=13.9,4.9Hz,1H),2.82(d,J=5.0Hz,1H),2.75(dd,J=13.9,8.6Hz,1H),1.93–1.86(m,2H) ,1.63–1.39(m,9H)ppm; ESI-MS: m/z=597[M+H] ⁺ .

Preparation Example 176, N-[(6S,9S)-6-(methoxymethyl)-4,7-dioxo-1,4,5,6,7,8,9,10,12, 13,14,15,16,17-Tetradetrahydro-1-methyl-pyrazolo[4,3-i][1,11]dioxa[4,7]diazacyclohexadeca- 9-Formyl]-Phe-epoxyketone (10p)

With (6S,9S)-6-(methoxymethyl)-4,7-dioxo-1,4,5,6,7,8,9,10,12,13,14,15,16 ,17-Tetradetrahydro-1-methyl-pyrazolo[4,3-i][1,11]dioxa[4,7]diazacyclohexadecan-9-carboxylic acid as raw material, The synthesis and post-treatment were the same as in Preparation Example 161 to obtain 50 mg of white solid with a yield of 83%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.75(s, 1H), 7.23–7.09(m, 6H), 7.06(d, J=8.0Hz, 1H), 6.90(d, J=6.0Hz, 1H) ,4.87(td,J=8.0,5.5Hz,1H),4.69–4.61(m,1H),4.55–4.50(m,1H),4.38–4.0(m,1H),4.18(ddd,J=10.0, 8.0,5.5Hz,1H),4.00(dd,J＝9.5,3.5Hz,1H),3.76–3.69(m,4H),3.61(dd,J＝9.5,4.5Hz,1H),3.54(dd,J ＝9.5,3.5Hz,1H),3.45–3.27(m,6H),3.10(dd,J＝14.0,5.5Hz,1H),2.90–2.81(m,2H),1.90–1.81(m,1H), 1.79–1.71(m,1H),1.49–1.34(m,9H)ppm; ESI-MS: m/z=600[M+H] ⁺ .

Preparation Example 177, N-[(13S,16S)-16-(methoxymethyl)-15,18-dioxo-2,11-dioxa-14,17-diazabicyclo[17.3 .1] -tricosyl-1(23),19,21-triene-13-formyl]-Phe-epoxyketone (10q)

With (13S,16S)-16-(methoxymethyl)-15,18-dioxo-2,11-dioxa-14,17-diazabicyclo[17.3.1]-23 Carbon-1(23), 19,21-triene-13-carboxylic acid was used as raw material, and the synthesis and post-treatment were the same as in Preparation Example 161 to obtain 50 mg of a white solid with a yield of 83%. ¹ H NMR (500MHz, CDCl ₃ )δ7.44–7.38(m,2H),7.32–7.29(m,1H),7.25(d,J=8.0Hz,1H),7.16–7.12(m,2H), 7.11–7.03(m,5H),6.97(d,J=5.5Hz,1H),4.82(td,J=8.5,4.5Hz,1H),4.73–4.68(m,1H),4.46–4.41(m, 1H), 4.22–4.10(m, 2H), 4.03(dd, J=10.0, 4.0Hz, 1H), 3.81(dd, J=9.5, 2.5Hz, 1H), 3.69(dd, J=9.5, 4.0Hz ,1H),3.45(dd,J＝9.5,4.0Hz,1H),3.40(s,3H),3.37(d,J＝5.0Hz,1H),3.20–3.15(m,2H),3.11(dd, ( m,6H)ppm; ESI-MS: m/z=624[M+H] ⁺ .

Preparation Example 178, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12, 13,14,15-Dodecahydro-9H-19-fluoro-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe - Epoxyketone (10r)

With (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-deca Dihydro-9H-19-fluoro-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-carboxylic acid as raw material, the synthesis and post-treatment are carried out in the same way as the preparation Example 161, 43mg of white solid was obtained, the yield was 70%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.84 (d, J=6.0Hz, 1H), 8.01–7.95 (m, 1H), 7.32–7.20 (m, 3H), 7.12–7.01 (m, 6H), 6.85(d,J=8.5Hz,1H),4.80(td,J=9.0,5.0Hz,1H),4.67–4.57(m,2H),4.56–4.49(m,1H),4.41–4.33(m, 1H), 4.03(dd, J＝9.5, 2.5Hz, 1H), 3.70–3.61(m, 2H), 3.52(dd, J＝9.5, 4.0Hz, 1H), 3.44–3.38(m, 4H), 3.35 (d,J=5.0Hz,1H),3.33–3.26(m,1H),3.08(dd,J=14.0,5.0Hz,1H),2.87–2.77(m,2H),2.03–1.92(m,1H ),1.91–1.82(m,1H),1.82–1.72(m,1H),1.63–1.48(m,2H),1.47–1.37(m,6H)ppm; ESI-MS: m/z＝614[M +H] ⁺ .

Preparation Example 179, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12, 13,14,15-Dodecahydro-9H-19-fluoro-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe - Epoxyketone (10s)

With (10S,13S)-19-methoxy-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13 ,14,15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-carboxylic acid as raw material, the synthesis and post-treatment are the same Example 161 was prepared to obtain 45 mg of a white solid with a yield of 72%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.99 (d, J=5.5Hz, 1H), 7.78 (dd, J=7.5, 2.0Hz, 1H), 7.30–7.27 (m, 1H), 7.13–7.04( m,7H),6.86(d,J=8.5Hz,1H),4.81(td,J=8.5,5.0Hz,1H),4.65–4.58(m,2H),4.30(dt,J=10.0,6.5Hz ,1H),4.13(dt,J=10.0,6.5Hz,1H),4.00(dd,J=9.5,3.0Hz,1H),3.91(s,3H),3.72(dd,J=9.5,3.5Hz, 1H), 3.63(dd, J＝9.5, 4.0Hz, 1H), 3.49(dd, J＝9.5, 4.0Hz, 1H), 3.44–3.35(m, 5H), 3.28–3.22(m, 1H), 3.08 (dd,J=14.0,5.0Hz,1H),2.86–2.80(m,2H),1.86–1.73(m,2H),1.57–1.50(m,1H),1.44(s,3H),1.41–1.30 (m,5H)ppm; ESI-MS: m/z=626[M+H] ⁺ .

Preparation Example 180, N-[(6S,9S)-6-(methoxymethyl)-4,7-dioxo-4,5,6,7,8,9,10,12,13, 14,15,16,17,18-Tetradetrahydro-1H-1-methyl-pyrazolo[4,3-i][1]oxa[4,7]-diazacyclohexadeca- 9-Formyl]-Phe-epoxyketone (10t)

With (6S,9S)-6-(methoxymethyl)-1-methyl-4,7-dioxo-4,5,6,7,8,9,10,12,13,14, 15,16,17,18-Tetradetrahydro-1H-pyrazolo[4,3-i][1]oxa[4,7]-diazacyclohexadeca-9-carboxylic acid as raw material, The synthesis and post-treatment were the same as in Preparation Example 161 to obtain 40 mg of white solid with a yield of 67%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.70(s, 1H), 7.46(d, J=8.0Hz, 1H), 7.30–7.14(m, 6H), 6.66(d, J=6.0Hz, 1H) ,5.00–4.93(m,1H),4.62–4.56(m,1H),4.42(dt,J＝7.5,3.0Hz,1H),3.97(dd,J＝10.0,5.0Hz,1H),3.83–3.76 (m,4H),3.68(dd,J＝10.0,5.0Hz,1H),3.51(dd,J＝9.0,3.5Hz,1H),3.47–3.37(mHu8219Hu8219,4H),3.35–3.24(m,3H ),3.10(dd,J=13.5,7.5Hz,1H),2.94(dd,J=14.0,6.5Hz,1H),2.74(d,J=5.0Hz,1H),2.60–2.50(m,1H) ,1.82–1.70(m,1H),1.48–1.18(m,11H),1.17–1.07(m,1H)ppm; ESI-MS: m/z=598[M+H] ⁺ .

Preparation Example 181, N-[(6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,12,13,14, 15,16,17-Dodecahydro-4H-pyrazolo[5,1-i][1]oxa[4,7,10]-triazacyclohexadeca-9-formyl]-Phe - Epoxyketone (10u)

With (6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,12,13,14,15,16,17-deca Dihydro-4H-pyrazolo[5,1-i][1]oxa[4,7,10]-triazacyclohexadecane-9-carboxylic acid as raw material, the synthesis and post-treatment are carried out in the same way as the preparation Example 161, 42 mg of white solid was obtained, yield 74%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.52(d, J=2.0Hz, 1H), 7.47(d, J=8.5Hz, 1H), 7.32–7.27(m, 2H), 7.25–7.20(m, 3H), 7.14(d, J=8.0Hz, 1H), 6.93(d, J=5.5Hz, 1H), 6.59(d, J=2.0Hz, 1H), 5.10(ddd, J=14.0, 12.0, 4.5 Hz,1H),5.03(td,J=8.0,6.0Hz,1H),4.59(dd,J=10.0,4.5Hz,1H),4.43(ddd,J=6.0,3.5,2.5Hz,1H),4.14 (ddd, J=13.5,5.0,3.5Hz,1H),4.03(dd,J=10.0,4.5Hz,1H),3.82(dd,J=9.5,2.5Hz,1H),3.72(dd,J=10.0 ,4.5Hz,1H),3.53(dd,J＝9.5,4.0Hz,1H),3.45–3.38(m,4H),3.37–3.27(m,2H),3.15(dd,J＝14.0,6.0Hz, 1H), 2.95(dd, J＝14.0, 8.0Hz, 1H), 2.82(d, J＝4.9Hz, 1H), 2.02–1.90(m, 1H), 1.61–1.50(m, 1H), 1.50–1.41 (m,1H),1.39–1.27(m,5H),1.20–1.06(m,2H),1.03–0.90(m,1H)ppm; ESI-MS: m/z＝626[M+H] ⁺ .

Preparation Example 182, N-[(11S,14S)-14-(methoxymethyl)-13,16-dioxo-5,6,7,8,11,12,13,14,15, 16-Decahydro-10H-imidazo[2,1-i][1]oxa[4,7,10]-triazacyclotetradecan-11-formyl]-Phe-epoxyketone (10v )

With (11S,14S)-14-(methoxymethyl)-13,16-dioxo-5,6,7,8,11,12,13,14,15,16-decahydro-10H- Imidazo[2,1-i][1]oxa[4,7,10]-triazacyclotetradec-11-carboxylic acid was used as raw material, the synthesis and post-treatment were the same as in Preparation Example 161, and a white solid was obtained 39 mg, yield 72%. ¹ H NMR (500MHz, CDCl ₃ ) δ9.38(s, 1H), 7.33(d, J=7.0Hz, 1H), 7.30–7.25(m, 3H), 7.24–7.17(m, 4H), 7.13( s,1H),4.91(td,J＝8.0,5.0Hz,1H),4.84–4.75(m,1H),4.56–4.49(m,1H),4.42–4.33(m,1H),4.01–3.90( m,2H),3.77–3.68(m,2H),3.58(dd,J＝9.5,3.5Hz,1H),3.53–3.46(m,1H),3.38(s,3H),3.34–3.26(m, 2H), 3.16(dd, J=14.0, 4.5Hz, 1H), 2.88(d, J=5.0Hz, 1H), 2.84(dd, J=14.4, 8.6Hz, 1H), 1.90–1.77(m, 1H ), 1.77–1.66(m,1H), 1.55–1.35(m,5H)ppm; ESI-MS: m/z＝542[M+H] ⁺ .

Preparation Example 183, N-[(12S,15S)-15-(methoxymethyl)-14,17-dioxo-6,7,8,9,12,13,14,15,16, 17-Decahydro-5H,11H-imidazo[2,1-i][1]oxa[4,7,10]-triazacyclopentadeca-12-formyl]-Phe-epoxyketone (10w)

With (12S,15S)-15-(methoxymethyl)-14,17-dioxo-6,7,8,9,12,13,14,15,16,17-decahydro-5H, 11H-imidazo[2,1-i][1]oxa[4,7,10]-triazacyclopentadecan-12-carboxylic acid as raw material, the synthesis and post-treatment are the same as in Preparation Example 161, to obtain White solid 37mg, yield 67%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.10(d, J=6.5Hz, 1H), 7.34(d, J=8.5Hz, 1H), 7.30–7.18(m, 5H), 7.13(d, J= 7.5Hz, 1H), 7.06(d, J=1.0Hz, 1H), 6.99(d, J=1.0Hz, 1H), 5.16–5.08(m, 1H), 5.01(td, J=8.5, 6.0Hz, 1H),4.64–4.58(m,1H),4.31(ddd,J＝7.5,3.5,2.0Hz,1H),4.14–4.09(m,1H),3.95–3.88(m,2H),3.71(dd, J=9.5,4.0Hz,1H), 3.43(dd,J=10.0,4.0Hz,1H),3.40(s,3H),3.37–3.33(m,1H),3.31(d,J=4.5Hz,1H ),3.20–3.12(m,2H),2.92(dd,J＝14.0,8.0Hz,1H),2.83(d,J＝4.5Hz,1H),1.82–1.71(m,1H),1.54–1.44( m,1H), 1.43–1.34(m,4H), 1.30–1.21(m,1H), 1.20–1.09(m,2H)ppm; ESI-MS: m/z=556[M+H] ⁺ . Preparation Example 184, N-[(13S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15,16 ,17,18-Dodecahydro-12H-imidazo[2,1-i][1]oxa[4,7,10]-triazacyclohexadeca-13-formyl]-Phe-cyclo Oxyketone (10x)

With (13S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15,16,17,18-deca Dihydro-12H-imidazo[2,1-i][1]oxa[4,7,10]-triazacyclohexadecane-13-carboxylic acid as raw material, the synthesis and post-treatment are the same as the preparation examples 161, 41 mg of white solid was obtained, the yield was 72%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.13(d, J=5.0Hz, 1H), 7.62(d, J=8.0Hz, 1H), 7.31–7.15(m, 6H), 7.10(s, 1H) ,7.07(s,1H),5.40(td,J＝13.0,3.5Hz,1H),5.08–5.01(m,1H),4.53(dd,J＝9.0,4.5Hz,1H),4.41–4.34(m ,1H),4.02(dd,J=9.5,4.5Hz,1H),3.94(dt,J=13.5,3.5Hz,1H),3.79(dd,J=9.5,2.0Hz,1H),3.74(dd, J=9.5,4.0Hz,1H), 3.56(dd,J=9.5,3.5Hz,1H),3.40(s,3H),3.38–3.32(m,2H),3.29(d,J=5.0Hz,1H ),3.14(dd,J=14.0,6.0Hz,1H),2.96(dd,J=14.0,7.5Hz,1H),2.80(d,J=5.0Hz,1H),1.76–1.66(m,1H) ,1.66–1.56(m,1H),1.48–1.38(m,2H),1.36(s,3H),1.33–1.26(m,1H),1.17–1.09(m,1H),1.08–1.00(m, 1H),0.98–0.89(m,1H)ppm; ESI-MS: m/z=570[M+H] ⁺ .

Preparation Example 185, N-[(3S,6S)-3-(methoxymethyl)-1,4-dioxo-2,3,4,5,6,7,9,10,11, 12,13,14-Dodecahydro-1H-pyrrolo[2,1-i][1]oxa[4,7,10]-triazacyclohexadeca-6-formyl]-Phe- Epoxyketone (10y)

(3S,6S)-2,3,4,5,6,7,9,10,11,12,13,14-dodecahydro-1H-3-(methoxymethyl)-1,4 -Dioxo-pyrrolo[2,1-i][1]oxa[4,7,10]-triazacyclohexadecane-6-carboxylic acid as raw material, the synthesis and post-treatment are the same as the preparation examples 161, 41 mg of white solid was obtained, the yield was 72%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.85(d, J=8.5Hz, 1H), 7.31–7.18(m, 6H), 6.86–6.81(m, 1H), 6.74(d, J=5.0Hz, 1H), 6.62(dd, J＝3.9, 1.4Hz, 1H), 6.17–6.12(m, 1H), 5.12–5.03(m, 2H), 4.51(q, J＝4.5Hz, 1H), 4.45–4.40 (m,1H),4.03(dd,J=9.5,4.5Hz,1H),3.87–3.77(m,2H),3.72(dd,J=9.5,4.5Hz,1H),3.54(dd,J=9.5 ,3.5Hz,1H),3.44–3.38(m,4H),3.37–3.29(m,2H),3.15(dd,J＝14.0,6.5Hz,1H),2.98(dd,J＝14.2,7.8Hz, 1H), 2.78(d, J＝5.0Hz, 1H), 1.66–1.57(m, 1H), 1.55–1.41(m, 3H), 1.37–1.29(m, 5H), 1.24–1.11(m, 2H) ppm; ESI-MS: m/z=569[M+H] ⁺ .

Preparation Example 186, N-[(3S,6S,18aS)-3-(methoxymethyl)-1,4,14-trioxo-hexadecahydro-1H-pyrrolo[2,1-i ][1]oxa[4,7,10]triazacyclohexadec-6-formyl]-Phe-epoxyketone (10z)

With (3S,6S,18aS)-3-(methoxymethyl)-1,4,14-trioxo-hexadecahydro-1H-pyrrolo[2,1-i][1]oxa[ 4,7,10] Triazacyclohexadeca-6-carboxylic acid was used as the raw material, and the synthesis and post-treatment were the same as in Preparation Example 161 to obtain 45 mg of white solid with a yield of 77%. ¹ H NMR (500MHz, CDCl ₃ ) δ7.66(d, J=7.0Hz, 1H), 7.33–7.13(m, 6H), 6.56(d, J=7.0Hz, 1H), 4.77(td, J= 9.0,5.0Hz,1H),4.61–4.53(m,1H),4.44–4.37(m,1H),4.37–4.32(m,1H),3.90(dd,J＝10.5,2.5Hz,1H),3.75 –3.69(m,1H),3.67(dd,J＝9.0,4.0Hz,1H),3.62(dd,J＝10.0,4.0Hz,1H),3.60–3.55(m,1H),3.55–3.48(m ,2H),3.45–3.39(m,1H),3.32–3.26(m,4H),3.04(dd,J＝14.0,5.0Hz,1H),2.86–2.78(m,2H),2.41–2.32(m ,1H),2.12–1.87(m,5H),1.71–1.61(m,1H),1.57–1.45(m,1H),1.44–1.31(m,5H),1.31–1.24(m,2H)ppm; ESI-MS: m/z＝587[M+H] ⁺ .

Preparation Example 187, N-[(9S,12S)-12-(methoxymethyl)-11,14-dioxo-2,3,5,6,9,10,11,12,13, 14-Decahydro-8H-benzo[o][1,4,7]-trioxa[10,13]-diazacyclohexadeca-9-formyl]-Phe-epoxyketone (10za )

With (9S,12S)-12-(methoxymethyl)-11,14-dioxo-2,3,5,6,9,10,11,12,13,14-decahydro-8H- Benzo[o][1,4,7]-trioxa[10,13]-diazacyclohexadecane-9-carboxylic acid was used as raw material, and the synthesis and post-treatment were the same as in Preparation Example 161 to obtain a white solid 43 mg, yield 77%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.82 (d, J=7.0Hz, 1H), 8.18 (dd, J=7.5, 2.5Hz, 1H), 7.47 (ddd, J=8.5, 7.5, 1.5Hz, 1H), 7.12–6.95(m, 9H), 4.78(ddd, J=7.0, 3.0, 2.5Hz, 1H), 4.70(td, J=8.5, 5.5Hz, 1H), 4.64(dt, J=8.5, 3.5Hz, 1H), 4.43–4.32(m, 2H), 4.03(dd, J＝9.5, 2.5Hz, 1H), 3.99(dd, J＝9.0, 3.5Hz, 1H), 3.95–3.89(m, 1H ),3.78–3.72(m,1H),3.67–3.61(m,2H),3.55(t,J＝8.0Hz,1H),3.43–3.38(m,6H),3.33(d,J＝5.0Hz, 1H), 3.00(dd, J=13.5, 5.0Hz, 1H), 2.79(d, J=5.0Hz, 1H), 2.75(dd, J=13.5, 8.5Hz, 1H), 1.40(s, 3H)ppm ;ESI-MS: m/z=584[M+H] ⁺ .

Preparation Example 188, N-[(12S,15S)-15-(methoxymethyl)-14,17-dioxo-2,3,5,6,8,9,12,13,14, 15,16,17-Dodecahydro-11H-benzo[r][1,4,7,10]tetraoxa[13,16]-diazacyclonadeca-12-formyl]-Phe - Epoxyketone (10zb)

With (12S,15S)-15-(methoxymethyl)-14,17-dioxo-2,3,5,6,8,9,12,13,14,15,16,17-deca Dihydro-11H-benzo[r][1,4,7,10]tetraoxa[13,16]-diazacyclinedecane-12-carboxylic acid as raw material, the synthesis and post-treatment are the same as the preparation Example 161, 42mg of white solid was obtained, the yield was 67%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.96 (d, J=4.5Hz, 1H), 8.26 (dd, J=7.5, 1.5Hz, 1H), 7.56–7.50 (m, 1H), 7.49–7.42( m,2H),7.24–7.08(m,5H),7.07–7.02(m,1H),6.97(d,J＝8.5Hz,1H),4.77(td,J＝8.5,5.5Hz,1H),4.71 –4.61(m,2H),4.41(td,J=10.0,2.0Hz,1H),4.30–4.24(m,1H),4.09–4.02(m,1H),3.97(dd,J=10.0,6.5Hz ,1H),3.85–3.79(m,1H),3.75(dd,J＝10.5,4.0Hz,1H),3.73–3.68(m,1H),3.68–3.51(m,6H),3.50–3.43(m ,3H),3.42(s,3H),3.35(d,J＝5.0Hz,1H),3.06(dd,J＝14.0,5.5Hz,1H),2.88–2.78(m,2H),1.40(s, 3H) ppm; ESI-MS: m/z=628[M+H] ⁺ .

Preparation Example 189, N-[(10S,13S)-13-(2-morpholino-2-oxoethyl)-12,15-dioxo-2,3,4,5,6,7 ,10,11,12,13,14,15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl ]-Phe-epoxyketone (10zc)

With (10S,13S)-13-(2-morpholino-2-oxoethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12, 13,14,15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-carboxylic acid as raw material, synthesis and post-treatment Same as Preparation Example 161, 37 mg of white solid was obtained, yield 55%. ¹ H NMR (500MHz, CDCl ₃ ) δ9.24 (d, J=8.5Hz, 1H), 8.24 (dd, J=8.0, 1.5Hz, 1H), 7.50–7.41 (m, 1H), 7.20–7.08( m,6H),7.08–7.02(m,1H),6.98(d,J＝8.5Hz,1H),6.91(d,J＝7.5Hz,1H),5.16–5.10(m,1H),4.75(td ,J＝8.5,4.5Hz,1H),4.64–4.57(m,1H),4.30–4.15(m,2H),3.68–3.62(m,4H),3.61–3.56(m,1H),3.55–3.30 (m,9H),3.07(dd,J=13.5,4.5Hz,1H),2.84(d,J=5.0Hz,1H),2.77(dd,J=14.0,9.0Hz,1H),2.62(dd, J=16.5, 5.0Hz, 1H), 2.12–2.04(m, 1H), 1.81–1.73(m, 1H), 1.72–1.58(m, 2H), 1.52–1.37(m, 7H) ppm; ESI-MS :m/z＝679[M+H] ⁺ .

Preparation Example 190, N-[(10S,13S)-13-(3-morpholino-3-oxopropyl)-12,15-dioxo-2,3,4,5,6,7 ,10,11,12,13,14,15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl ]-Phe-epoxyketone (10zd)

With (10S,13S)-13-(3-morpholino-3-oxopropyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12, 13,14,15-Dodecahydro-9Hbenzo[i][1,11]dioxa[4,7]-diazacyclohexadecyl-10-carboxylic acid as raw material, synthesis and post-treatment The same as in Preparation Example 161, 40 mg of white solid was obtained with a yield of 58%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.72(d, J=5.5Hz, 1H), 8.10(dd, J=7.5, 2.0Hz, 1H), 7.48–7.41(m, 1H), 7.34(d, J＝8.5Hz, 1H), 7.11–6.91(m, 7H), 4.76(td, J＝9.0, 4.5Hz, 1H), 4.62–4.46(m, 2H), 4.37–4.28(m, 1H), 4.18 (td,J=9.0,2.0Hz,1H),3.66–3.39(m,9H),3.37–3.29(m,2H),3.04(dd,J=13.5,4.5Hz,1H),2.83(d,J ＝5.0Hz,1H),2.73(dd,J＝13.5,9.0Hz,1H),2.63–2.42(m,2H),2.30–2.20(m,2H),2.02–1.96(m,1H),1.90– 1.84(m,1H),1.59–1.35(m,8H),1.31–1.21(m,3H)ppm; ESI-MS: m/z=693[M+H] ⁺ .

Preparation Example 191, N-[(10S,13S)-13-(2-(cyclopropylamino)-2-oxoethyl)-12,15-dioxo-2,3,4,5, 6,7,10,11,12,13,14,15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10- Formyl]-Phe-epoxyketone (10ze)

With (10S,13S)-13-(2-(cyclopropylamino)-2-oxoethyl)-12,15-dioxo-2,3,4,5,6,7,10,11 ,12,13,14,15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-carboxylic acid as raw material, synthesized and The post-treatment was the same as that of Preparation Example 161 to obtain 41 mg of white solid with a yield of 63%. ¹ H NMR (500MHz, CDCl ₃ ) δ9.43(d, J=6.5Hz, 1H), 8.17(dd, J=7.5, 1.5Hz, 1H), 7.51–7.42(m, 1H), 7.38(d, J＝8.0Hz, 1H), 7.16–6.99(m, 8H), 6.88(d, J＝3.0Hz, 1H), 4.97–4.87(m, 1H), 4.77(td, J＝9.0, 4.5Hz, 1H ),4.62–4.54(m,1H),4.49–4.40(m,1H),4.31–4.18(m,1H),3.64–3.54(m,1H),3.50(dd,J＝9.5,3.5Hz,1H ),3.42–3.27(m,2H),3.07(dd,J＝14.0,4.5Hz,1H),2.90(dd,J＝15.0,5.5Hz,1H),2.87–2.77(m,2H),2.75– 2.67(m,1H),2.63(dd,J＝15.0,5.0Hz,2H),2.24–2.12(m,1H),1.91–1.80(m,1H),1.71–1.54(m,2H),1.53– 1.33(m,7H),0.75–0.63(m,2H),0.57–0.45(m,2H)ppm; ESI-MS: m/z=649[M+H] ⁺ .

Preparation Example 192, N-[(10S,13S)-13-isobutyl-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14, 15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone (10zf)

(10S,13S)-13-isobutyl-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-dodecahydro-9H -Benzo[i][1,11]dioxa[4,7]-diazacyclohexadecan-10-carboxylic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 161 to obtain 45 mg of white solid, Yield 74%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.29(d, J=6.0Hz, 1H), 8.23(dd, J=8.0, 1.5Hz, 1H), 7.52–7.45(m, 1H), 7.21(d, J＝8.0Hz, 1H), 7.11–6.99(m, 7H), 6.81(d, J＝8.0Hz, 1H), 4.76(td, J＝8.5,5.0Hz, 1H), 4.63–4.52(m, 2H ),4.35–4.30(m,1H),4.19(td,J＝10.0,2.0Hz,1H),3.60(dd,J＝10.0,5.5Hz,1H),3.48(dd,J＝10.0,4.5Hz, 1H), 3.36–3.27(m, 3H), 3.05(dd, J＝14.0, 5.0Hz, 1H), 2.82(d, J＝5.0Hz, 1H), 2.76(dd, J＝13.5, 9.0Hz, 1H ),1.98–1.88(m,2H),1.87–1.70(m,2H),1.63–1.49(m,4H),1.47–1.40(m,6H),0.99(d,J＝6.5Hz,3H), 0.95(d, J=6.5Hz,3H)ppm; ESI-MS: m/z=608[M+H] ⁺ .

Preparation Example 193, N-[(10S,13S)-13-isopropyl-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14, 15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone (10zg)

(10S,13S)-13-isopropyl-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-dodecahydro-9H -Benzo[i][1,11]dioxa[4,7]-diazacyclohexadecan-10-carboxylic acid as raw material, the synthesis and post-treatment were the same as in Preparation Example 161 to obtain 46 mg of white solid, Yield 77%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.26–8.19 (m, 2H), 7.52–7.46 (m, 1H), 7.21 (d, J=8.0Hz, 1H), 7.11–6.99 (m, 7H), 6.78(d,J=8.5Hz,1H),4.77(td,J=9.0,5.0Hz,1H),4.62–4.55(m,2H),4.37–4.31(m,1H),4.22(td,J= 9.5,3.5Hz,1H),3.62(dd,J＝9.5,5.0Hz,1H),3.47(dd,J＝9.5,4.0Hz,1H),3.37–3.26(m,3H),3.05(dd,J =14.0,4.5Hz,1H),2.82(d,J=5.0Hz,1H),2.76(dd,J=14.0,9.0Hz,1H),1.96–1.81(m,2H),1.67–1.53(m, 2H), 1.49–1.36(m,7H), 1.30–1.23(m,1H), 1.06–0.97(m,6H)ppm; ESI-MS: m/z＝594[M+H] ⁺ .

Preparation Example 194, N-[(9S,12S)-12-(methoxymethyl)-11,14-dioxo-3,4,5,6,7,8,9,10,11, 12,13,14-Dodecahydro-2H-benzo[b][1]oxa[5,8]-diazacyclohexadeca-9-formyl]-Phe-epoxyketone (10zh)

With (9S,12S)-12-(methoxymethyl)-11,14-dioxo-3,4,5,6,7,8,9,10,11,12,13,14-deca Dihydro-2H-benzo[b][1]oxa[5,8]-diazacyclohexadecane-9-carboxylic acid was used as raw material, and the synthesis and post-treatment were the same as in Preparation Example 161 to obtain 35 mg of white solid , yield 60%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.87 (d, J=7.5Hz, 1H), 8.27 (dd, J=8.0, 2.0Hz, 1H), 7.52–7.45 (m, 1H), 7.23–7.15( m,3H),7.14–7.08(m,3H),7.00(d,J=8.5Hz,1H),6.73(d,J=7.5Hz,1H),6.58(d,J=8.0Hz,1H), 4.85–4.73(m,2H),4.46–4.38(m,1H),4.26–4.19(m,1H),4.18–4.08(m,2H),3.53(dd,J＝9.0,4.0Hz,1H), 3.41(s,3H),3.32(d,J=5.0Hz,1H),3.10(dd,J=14.0,5.0Hz,1H),2.86(d,J=5.0Hz,1H),2.75(dd,J =14.0,9.0Hz,1H), 2.03–1.75(m,4H),1.56–1.26(m,11H)ppm; ESI-MS: m/z=580[M+H] ⁺ .

Preparation Example 195, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,8,9,10, 11,12,13,14,15-tetrahydro-benzo[b][1]oxa[5,8]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone ( 10zi)

With (10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,8,9,10,11,12,13,14 , 15-Tetradecylhydro-benzo[b][1]oxa[5,8]-diazacyclohexadecan-10-carboxylic acid as raw material, the synthesis and aftertreatment are the same as in Preparation Example 161 to obtain white Solid 34mg, yield 57%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.86 (d, J=8.0Hz, 1H), 8.17 (dd, J=8.0, 1.5Hz, 1H), 7.48–7.40 (m, 1H), 7.19–7.00 ( m,7H),6.95(d,J=8.5Hz,1H),6.84(d,J=8.0Hz,1H),4.80–4.74(m,1H),4.73–4.67(m,1H),4.39–4.33 (m,1H),4.20–4.14(m,1H),4.11–4.01(m,2H),3.48(dd,J＝9.0,4.0Hz,1H),3.34(s,3H),3.25(d,J =5.0Hz,1H),3.02(dd,J=14.0,4.5Hz,1H),2.80(d,J=5.0Hz,1H),2.69(dd,J=14.0,9.0Hz,1H),1.93–1.83 (m,1H),1.82–1.70(m,3H),1.52–1.19(m,13H)ppm; ESI-MS: m/z=594[M+H] ⁺ .

Preparation Example 196, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-3,4,5,6,7,9,10,11,12, 13,14,15-Dodecahydro-2H-1,8-diaza-11,14-benzodioxadiazacyclohexadeca-10-formyl]-Leu-epoxyketone (10zj )

With (10S,13S)-13-(methoxymethyl)-12,15-dioxo-3,4,5,6,7,9,10,11,12,13,14,15-deca Synthetic and The post-treatment was the same as that of Preparation Example 161 to obtain 42 mg of white solid with a yield of 75%. ¹ H NMR (500MHz, CDCl ₃ ) δ8.81(d, J=6.5Hz, 1H), 8.15(d, J=8.0Hz, 1H), 7.49–7.40(m, 1H), 7.07–7.01(m, 1H),7.00–6.94(m,2H),6.86(d,J＝8.0Hz,1H),4.67–4.62(m,2H),4.48–4.42(m,1H),4.28–4.24(m,1H) ,4.12–4.07(m,1H),4.03–3.99(m,1H),3.81(dd,J＝9.5,5.0Hz,1H),3.61(dd,J＝9.5,3.5Hz,1H),3.54–3.44 (m,2H),3.39(s,3H),3.30(d,J＝5.0Hz,1H),2.81–2.76(m,2H),1.99–1.90(m,1H),1.86–1.77(m,1H ),1.71–1.57(m,2H),1.55–1.37(m,10H),0.74(d,J=6.0Hz,3H),0.67(d,J=6.0Hz,3H) ¹ ppm; ESI-MS: m/z＝562[M+H] ⁺ .

Test Example 1, Proteasome Inhibitory Activity Test of Macrocyclic Epoxy Ketone Peptide Compounds

Experimental method: The fluorescent substrate Suc-Leu-Leu-Val-Tyr-AMC was used to detect the activity, and the inhibition of enzyme activity by different compounds was observed, so as to preliminarily evaluate the inhibitory effect of the compound. Human proteasome chymotrypsin-likeprotease hydrolyzes the Tyr-AMC sequence in the substrate to release AMC. Under the conditions of excitation light 355nm and emission light 460nm, the fluorescence absorption value of the hydrolyzed product AMC can be detected to observe the inhibition of the compound on the enzyme activity . See Table 1 for the results.

Table 1 Compounds have inhibitory activity on proteasome CT-L and multiple myeloma cells (English in the table is changed to Chinese)

Note: NT - not tested

Experimental Example 2: Multiple Myeloma Cell Proliferation Inhibitory Activity Test of Macrocyclic Epoxy Ketone Peptide Compounds

Experimental method: use the MTT method to detect the cell survival rate, the cells that are about to grow in the logarithmic growth phase are digested with 0.01% trypsin, counted, and seeded in 100ml of a 96-well plate at a cell density of 2.0×10 ³ /well. Incubate overnight at 37°C in a 5% CO ₂ incubator. Six concentration gradients were set up for each compound, and three replicate wells were set up for each concentration. Each concentration was added to corresponding wells, cultured in a 5% CO ₂ 37° C. incubator for 72 hours, and 20 ml of 5 mg/ml MTT was added. After incubating at 37°C for 3 hours, discard the supernatant, add 100ml of DMSO to dissolve, use SpectraMAX 340 to measure the light absorption value at 550nm (L1), refer to the wavelength of 690nm (L2), and calculate the (L1-L2) value for different concentrations of inhibitors. Figure, IC ₅₀ was obtained by formula fitting. See Table 1 for the results.

Experimental Example 3, Macrocyclic Epoxy Ketone Peptide Compounds' Proliferation Inhibitory Activity Test on Various Tumor Cells For the experimental method, refer to Experimental Example 2, except that the RPMI8226 and MM.1S cell lines were replaced with corresponding tumor cells. See Table 2 for the results.

Table 2. Proliferation inhibitory activity of compounds on various tumor cells

Experimental Example 4: Test of Proteasome Inhibitory Activity in Mouse PBMCs After Oral Administration of Macrocyclic Epoxy Ketone Peptide Compounds

Experimental method: select compounds 10a, 10b and 10c, and divide male BABL/C mice into 4 groups, one of which is a negative control group, with 3 mice in each group, and each is orally administered 30 mg/kg, 100 mg/kg according to the group kg, samples were taken 24 hours after the administration, and the changes in proteasome activity in PBMC after the compound was administered once to BABL/C mice were investigated.

The compounds of the present invention are all efficient inhibitors of proteasome, and the activity test results show (see Figure 1), most of the compounds have obvious inhibitory activity on proteasome, and the proteasome inhibitory activity of 19 compounds is equivalent to that of positive control Oprozzomib, Some compounds are superior to Oprozzomib and comparable to the marketed compound Carfilzomib. At the same time, we can also see that most of the compounds have moderate to strong in vitro proliferation inhibitory activity on multiple myeloma cells and other nine tumor cell lines; 10c inhibited the proteasome in mouse PBMC to varying degrees. In conclusion, this type of compound has potent anti-tumor activity in vitro and good oral activity, and has the in vitro and in vivo activity levels optimized to become drug candidates. Therefore, the macrocyclic epoxy ketone peptide derivatives that can be used as proteasome inhibitors involved in the present invention have broad anti-tumor application prospects.

Claims (18)

1. A macrocyclic epoxy ketone peptide derivative and a salt thereof, characterized in that it has a general formula (I) structure: and its optical isomers or solvates thereof, in: R ₁ and R ₂ are each independently selected from H, -C _1-10 alkyl-D, C _1-10 hydroxyalkyl, -C _2-10 unsaturated alkyl-D, -halogenated C _1-10 alkane -D, -C _1-3 alkyl-SC _1-5 alkyl, C _1-10 alkoxyalkyl, halogenated C _1-10 alkoxyalkyl, C _3-10 unsaturated alkoxy base, C _3-10 cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl, aralkyl, heteroaryl, heteroarylalkyl; wherein, D is N(R _a )R _b or missing; R _a is selected from H, OH, C _1-6 alkyl, halogenated C _1-6 alkyl; R _b is selected from N-terminal protecting groups; R ₃ and R ₄ are each independently selected from H, C _1-10 alkyl, halogenated C _1-10 alkyl, aryl, and aralkyl; _R is selected from H, C _1-6 alkyl, C _1-6 hydroxyalkyl, halogenated C _1-6 alkyl, C _1-6 alkoxyalkyl, halogenated C _1-6 alkoxy Alkyl; R _is selected from H, C _1-10 alkyl, C _1-10 alkoxyalkyl, C _2-10 unsaturated alkyl, cycloalkyl; X is O, S, NH, NC _1-6 alkyl; Y is Or missing, wherein R is selected from H, C _1-10 alkyl, halogenated C _1-10 alkyl; Ar is selected from unsubstituted or substituted cycloalkyl, unsubstituted or substituted heterocycloalkyl, unsubstituted or substituted cycloalkenyl, unsubstituted or substituted heterocycloalkenyl, unsubstituted or substituted aryl, none Substituted or substituted aralkyl, unsubstituted or substituted heteroaryl, unsubstituted or substituted heteroaralkyl, optionally fused aryl; L is where _B1 is selected from D ₁ and D ₂ are the same or different, independently selected from -C _1-8 alkyl-, -OC _1-8 alkyl-, -C _1-8 alkyl O-, -C _1-8 alkyl OC _{1 -8} Alkyl-, -SC _1-8 Alkyl-, -C _1-8 Alkyl S-, -C _1-8 Alkyl, SC _1-8 Alkyl-, -N(R _d )-, -N (R _d )C _1-8 alkyl-, -C _1-8 alkylN(R _d )-, -C _1-8 alkylN(R _d )C _1-8 alkyl-, -N(R _d ) C(O)-, -N(R _d )C(O)C _1-8 alkyl-, -C _1-8 alkyl N(R _d )C(O)-, -C _1-8 alkane N(R _d )C(O)C _1-8 alkyl-, -C(O)N(R _d )-, -C(O)N(R _d )C _1-8 alkyl-, -C _1-8 Alkyl C(O)N(R _d )-, -C _1-8 Alkyl C(O)N(R _d )C _1-8 Alkyl-, -C(O)C _1-8 Alkane -, -C(O)C _1-8 unsaturated alkyl-, -N(R _d )SO ₂ -, -N(R _d )SO ₂ C _1-8 alkyl-, -C _1-8 alkane N(R _d )SO ₂ -, -C _1-8 alkylN(R _d )SO ₂ C _1-8 alkyl-, -OC(O)C _1-8 alkyl-, -C _1-8 AlkylOC(O)-, -C _1-8alkylOC (O)C _1-8alkyl- , -C(O)OC _1-8alkyl- , -C _1-8alkylC (O )O-, -C _1-8 alkyl C(O)OC _1-8 alkyl-; R _d is selected from H, C _1-4 alkyl, halogenated C _1-4 alkyl, cycloalkyl, Heterocycloalkenyl, aryl, aralkyl, heteroaryl, heteroaralkyl; The substituted substituent is selected from halogen, nitro, amino, cyano, hydroxyl, C _1-6 alkyl, halogenated C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkane Oxyalkyl, C _1-6 alkylamino, halogenated C _1-6 alkoxy, halogenated C _1-6 alkoxyalkyl, halogenated C _1-6 alkylamino, C _{3 -8} Cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl, aralkyl, heteroaryl, heteroaralkyl. 2. The compound according to claim 1, characterized in that, the compound has the general formula (II) structure: and its optical isomers or pharmaceutically acceptable salts or solvates, wherein: X is selected as O; Y choice Ring A is selected from the following structures: Wherein, V ₁ , V ₂ , V ₃ , V ₄ , W ₁ , W ₂ and W ₃ are each independently selected from N and C; R _e is selected from H, halogen, nitro, amino, cyano, hydroxyl, _{C 1-6} alkyl, halogenated C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkoxy Alkyl, C _1-6 alkylamino, halogenated C _1-6 alkoxy, halogenated C _1-6 alkoxyalkyl, halogenated C _1-6 alkylamino, C _3-8 Cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl, aralkyl, heteroaryl, heteroaralkyl; Z is selected from O, S, -N(R _f )-; wherein, R _f is present or missing, selected from H, C _1-6 alkyl, halogenated C _1-6 alkyl, C _1-6 alkoxy Base, C _1-6 alkoxyalkyl, halogenated C _1-6 alkoxy, C _3-8 cycloalkyl, heterocycloalkyl, C _3-8 cycloalkenyl, heterocycloalkenyl, aromatic radical, aralkyl, heteroaryl, heteroaralkyl; L, R ₁ , R ₂ , R ₃ , R ₄ and R ₅ are as defined in the structure of the general formula (I) in claim 1. 3. The compound according to claim 2, characterized in that, the compound has the general formula (III) structure: and its optical isomers or pharmaceutically acceptable salts or solvates, wherein: V ₁ , V ₂ , V ₃ , V ₄ and R _g are as defined in the general formula (II); B ₁ , D ₁ , D ₂ , R ₁ and R ₂ are as defined in the general formula (I) structure of claim 1. 4. the compound according to claim 3, is characterized in that, described compound has general formula (IV) structure: and its optical isomers or pharmaceutically acceptable salts or solvates thereof, wherein: V _and V are _each independently selected from C, N; B ₁ selected as D ₁ and D ₂ are the same or different, independently selected from -C _1-8 alkyl-, -OC _1-8 alkyl-, -C _1-8 alkyl O-, -C _1-8 alkyl OC _{1 -8} alkyl; _R is selected from methoxy-methyl, isobutyl, isopropyl, (N-morpholinyl) formyl-methyl, (N-morpholinyl) formyl-ethyl; R ₂ is selected from benzyl, isobutyl; R _g is selected from H, halogen, C _1-6 alkyl, C _1-6 halogenated alkyl, C _1-6 alkoxy, C _1-6 alkoxyalkyl; Wherein, unless otherwise specified, the amino acids involved are all L-type amino acids. 5. the compound according to claim 4, is characterized in that, described compound is selected from: N-[(8S,11S)-11-(methoxymethyl)-10,13-dioxo-3,4,5,7,8,9,10,11,12,13-decahydro- 2H-1,6,9,12-Benzodioxadiazacyclopentadecyl-8-formyl]-Phe-epoxyketone, N-[(9S,12S)-12-(methoxymethyl)-11,14-dioxo-2,3,4,5,6,8,9,10,11,12,13,14 - dodecahydro-1,7,10,13-benzodioxadiazacyclohexadeca-9-formyl]-Phe-epoxyketone, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-3,4,5,6,7,9,10,11,12,13,14,15 - dodecahydro-2H-1,8,11,14-benzodioxadiazacyclohexadeca-10-formyl]-Leu-epoxyketone, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-3,4,5,6,7,9,10,11,12,13,14,15 -2H-Dodecahydro-1,8,11,14-benzodioxadiazacyclohexadeca-10-formyl]-Phe-epoxyketone, N-[(11S,14S)-14-(methoxymethyl)-13,16-dioxo-2,3,4,5,6,7,8,10,11,12,13,14 ,15,16-tetradetrahydro-1,9,12,15-benzodioxadiazacycloctadec-11-formyl]-Phe-epoxyketone, N-[(12S,15S)-15-(methoxymethyl)-14,17-dioxo-3,4,5,6,7,8,9,11,12,13,14,15 ,16,17-2H-Tetradetrahydro-2H-1,10,13,16-Benzodioxadiazacyclonadeca-12-formyl]-Phe-epoxyketone, N-[(13S,16S)-16-(methoxymethyl)-15,18-dioxo-3,4,5,6,7,8,9,10,13,14,15,16 ,17,18-tetradecahydro-2H,12H-benzo[i][1,11]dioxa[4,7]diazacycloeicos-13-formyl]-Phe-epoxyketone , N-[(7S,10S)-7-(methoxymethyl)-5,8-dioxo-6,7,8,9,10,11,13,14,15,16,17,18 - dodecahydro-5H-pyrido[3,2-i][1,11]dioxa[4,7]diazacyclohexadeca-10-formyl]-Phe-epoxyketone, N-[(14S,17S)-17-(methoxymethyl)-16,19-dioxo-6,7,8,9,10,11,14,15,16,17,18,19 - dodecahydro-13H-pyrido[2,3-i][1,11]dioxa[4,7]diazacyclohexadeca-14-formyl]-Phe-epoxyketone, N-[(9S,12S)-12-(methoxymethyl)-11,14-dioxo-3,4,5,6,7,8,9,10,11,12,13,14 - dodecahydro-2H-benzo[b][1]oxa[5,8]-diazacyclohexadeca-9-formyl]-Phe-epoxyketone, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,8,9,10,11,12,13 ,14,15-Tetradetrahydro-benzo[b][1]oxa[5,8]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 -Dodecahydro-9H-19-methoxy-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxy ketone, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 - dodecahydro-9H-19-fluoro-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 - dodecahydro-9H-18-fluoro-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 - dodecahydro-9H-18-chloro-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 -Dodecahydro-9H-18-methoxy-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxy ketone, N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 -Dodecahydro-9H-18-methyl-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone , N-[(10S,13S)-13-(methoxymethyl)-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15 - dodecahydro-9H-17-chloro-benzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-formyl]-Phe-epoxyketone, N-[(10S,13S)-13-(2-morpholino-2-oxoethyl)-12,15-dioxo-2,3,4,5,6,7,10,11, 12,13,14,15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-cyclo Oxyketone, N-[(10S,13S)-13-(3-morpholino-3-oxopropyl)-12,15-dioxo-2,3,4,5,6,7,10,11, 12,13,14,15-Dodecahydro-9Hbenzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxy ketone, N-[(9S,12S)-12-(methoxymethyl)-11,14-dioxo-2,3,5,6,9,10,11,12,13,14-decahydro- 8H-Benzo[o][1,4,7]-trioxa[10,13]-diazacyclohexadec-9-formyl]-Phe-epoxyketone, N-[(12S,15S)-15-(methoxymethyl)-14,17-dioxo-2,3,5,6,8,9,12,13,14,15,16,17 - dodecahydro-11H-benzo[r][1,4,7,10]tetraoxa[13,16]-diazacyclonadeca-12-formyl]-Phe-epoxyketone, N-[(10S,13S)-13-(2-(cyclopropylamino)-2-oxoethyl)-12,15-dioxo-2,3,4,5,6,7,10 ,11,12,13,14,15-Dodecahydro-9H-benzo[i][1,11]dioxa[4,7]diazacyclohexadeca-10-formyl]-Phe - epoxyketone, N-[(10S,13S)-13-isobutyl-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-dodecahydro -9H-Benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone, N-[(10S,13S)-isopropyl-12,15-dioxo-2,3,4,5,6,7,10,11,12,13,14,15-dodecahydro-9H -13-Benzo[i][1,11]dioxa[4,7]-diazacyclohexadeca-10-formyl]-Phe-epoxyketone, and optical isomers of the above-mentioned compounds or pharmaceutically acceptable salts or solvates thereof. 6. the compound according to claim 2, is characterized in that, described compound has general formula (V) structure: and its optical isomers or pharmaceutically acceptable salts or solvates thereof, wherein: B ₁ selected as D ₁ and D ₂ are the same or different, independently selected from -C _1-8 alkyl-, -OC _1-8 alkyl-, -C _1-8 alkyl O-, -C _1-8 alkyl OC _{1 -8} alkyl-; _Rh is selected from H; Wherein, unless otherwise specified, the amino acids involved are all L-type amino acids. 7. The compound according to claim 6, characterized in that, the compound is selected from: N-[(13S,16S)-16-(methoxymethyl)-15,18-dioxo-2,11-dioxa-14,17-diazabicyclo[17.3.1]-di Tridecyl-1(23),19,21-triene-13-formyl]-Phe-epoxyketone, and optical isomers of the above-mentioned compounds or pharmaceutically acceptable salts or solvates thereof. 8. The compound according to claim 2, characterized in that the compound has the general formula (VI) structure: and its optical isomers or pharmaceutically acceptable salts or solvates thereof, wherein: W ₁ and W ₂ are each selected as C, N; Z is selected as N(C _1-6 alkyl); B ₁ selected as D ₁ and D ₂ are the same or different, independently selected from -C _1-8 alkyl-, -OC _1-8 alkyl-, -C _1-8 alkyl O-, -C _1-8 alkyl OC _{1 -8} alkyl-; R _i is selected as H; Wherein, unless otherwise specified, the amino acids involved are all L-type amino acids. 9. The compound according to claim 8, characterized in that, the compound is selected from: N-[(6S,9S)-6-(methoxymethyl)-4,7-dioxo-1,4,5,6,7,8,9,10,12,13,14,15 ,16,17-Dodecahydro-1-methyl-pyrazolo[4,3-i][1,11]dioxa[4,7]diazacyclohexadeca-9-formyl] -Phe-epoxyketone N-[(6S,9S)-6-(methoxymethyl)-1-methyl-4,7-dioxo-4,5,6,7,8,9,10,12,13, 14,15,16,17,18-Tetradetrahydro-1H-pyrazolo[4,3-i][1]oxa[4,7]-diazacyclohexadeca-9-formyl] -Phe-epoxyketone, and optical isomers of the above-mentioned compounds or pharmaceutically acceptable salts or solvates thereof. 10. the compound according to claim 2, is characterized in that, described compound has general formula (VII) structure: and its optical isomers or pharmaceutically acceptable salts or solvates thereof, wherein: W ₁ and W ₂ are each selected as C, N; Z is chosen as S; B ₁ selected as D ₁ and D ₂ are the same or different, and are independently selected from -C _1-8 alkyl-, -OC _1-8 alkyl-, -C _1-8 alkyl O-; R _j is selected from H, methyl; Wherein, unless otherwise specified, the amino acids involved are all L-type amino acids. 11. The compound according to claim 10, characterized in that, the compound is selected from: N-[(13S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15,16,17,18 - dodecahydro-12H-thieno[2,3-i][1,11]dioxa[4,7]diazacyclohexadeca-13-formyl]-Phe-epoxyketone, N-[(13S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15,16,17,18 -Dodecahydro-12H-2-methyl-thiazolo[4,5-i][1,11]dioxa[4,7]diazacyclohexadeca-13-formyl]-Phe- epoxy ketone, and optical isomers of the above-mentioned compounds or pharmaceutically acceptable salts or solvates thereof. 12. The compound according to claim 2, characterized in that, the compound has the general formula (Ⅷ) structure: and its optical isomers or pharmaceutically acceptable salts or solvates thereof, wherein: W ₁ , W ₂ and W ₃ are each selected as C, N; Z is selected as N; B ₁ selected as D ₁ and D ₂ are the same or different, independently selected from -C _1-8 alkyl-, -OC _1-8 alkyl-, -C _1-8 alkyl O-, -C _1-8 alkyl OC _{1 -8} alkyl-; R _k is selected as H; Wherein, unless otherwise specified, the amino acids involved are all L-type amino acids. 13. The compound according to claim 12, characterized in that said compound is selected from: NN-(13S,16S)-16-(methoxymethyl)-15,18-dioxo-5,6,7,8,9,10,13,14,15,16,17,18- dodecahydro-12H-imidazo[2,1-i][1]oxa[4,7,10]-triazacyclohexadeca-13-formyl]-Phe-epoxyketone, N-(12S,15S)-15-(methoxymethyl)-14,17-dioxo-6,7,8,9,12,13,14,15,16,17-decahydro-5H ,11H-imidazo[2,1-i][1]oxa[4,7,10]-triazacyclopentadecan-12-formyl]-Phe-epoxyketone, N-(11S,14S)-14-(methoxymethyl)-13,16-dioxo-5,6,7,8,11,12,13,14,15,16-decahydro-10H -imidazo[2,1-i][1]oxa[4,7,10]-triazacyclotetradec-11-formyl]-Phe-epoxyketone, N-(3S,6S)-3-(methoxymethyl)-1,4-dioxo-2,3,4,5,6,7,9,10,11,12,13,14- Dodecahydro-1H-pyrrolo[2,1-i][1]oxa[4,7,10]-triazacyclohexadeca-6-formyl]-Phe-epoxyketone, N-(6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,12,13,14,15,16,17- Dodecahydro-4H-pyrazolo[5,1-i][1]oxa[4,7,10]-triazacyclohexadeca-9-formyl]-Phe-epoxyketone, N-(6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,13,14,15,16-decahydro-4H ,12H-pyrazolo[5,1-i][1]oxa[4,7,10]triazacyclopentadeca-9-formyl]-Phe-epoxyketone, N-(6S,9S)-6-(methoxymethyl)-4,7-dioxo-5,6,7,8,9,10,12,13,14,15-decahydro-4H -pyrazolo[5,1-i][1]oxa[4,7,10]-triazacyclocarbo-9-formyl]-Phe-epoxyketone, and optical isomers of the above-mentioned compounds or pharmaceutically acceptable salts or solvates thereof. 14. The compound according to claim 2, characterized in that, the compound has a general formula (IX) structure: and its optical isomers or pharmaceutically acceptable salts or solvates thereof, wherein: B ₁ selected as _D2 is selected from -C _1-8 alkyl-, -C _1-8 alkylOC _1-8 alkyl-, R _m is selected as H; Wherein, unless otherwise specified, the amino acids involved are all L-type amino acids. 15. The compound according to claim 14, characterized in that, the compound is selected from the group consisting of: N-[(3S,6S,18aS)-3-(methoxymethyl)-1,4,14-trioxo-hexadecahydro-1H-pyrrolo[2,1-i][1]oxo Hetero[4,7,10]triazacyclohexadec-6-formyl]-Phe-epoxyketone, and optical isomers of the above-mentioned compounds or pharmaceutically acceptable salts or solvates thereof. 16. A method for preparing a macrocyclic peptide compound, characterized in that, it is realized through the following steps: (1) Compound 5 reacts with amino-protected amino acid under the action of condensation reagent to obtain compound 6. The selected condensation reagent has dicyclohexylcarbodiimide/4-dimethylaminopyridine, dicyclohexylcarbodiimide/1- Hydroxybenzotriazole, N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride/1-hydroxybenzotriazole, benzotriazole-N,N ,N',N'-Tetramethyluronium hexafluorophosphate/1-hydroxybenzotriazole, the reaction temperature is 0-30°C, the reaction time is 2-8 hours, the crude product can be directly used in the next reaction; (2) Compound 6 removes the Boc protecting group under the action of trifluoroacetic acid, and the crude product is directly used for the next step reaction; (3) Compound 7 reacts with compound 2 to obtain compound 8 under the action of a condensation reagent. The selected condensation reagent has dicyclohexylcarbodiimide/4-dimethylaminopyridine, dicyclohexylcarbodiimide/1-hydroxybenzene Triazole, N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride/1-hydroxybenzotriazole, benzotriazole-N,N,N ', N'-Tetramethyluronium hexafluorophosphate/1-hydroxybenzotriazole, the reaction temperature is 0-30°C, the reaction time is 2-8 hours, the crude product can be directly used in the next reaction; (4) Compound 8 is ring-merged under the effect of metal catalyst or condensing agent to remove carboxyl protecting group to obtain compound 9. The metal catalyst selected is Grubbs second-generation catalyst, and the condensing agent is the same as in step (1), and the metal catalyzed reaction temperature 0-100°C, reaction time 0.5-3 hours, condensation reaction temperature 0-30°C, reaction time 3-8 hours, the obtained product is used for the next reaction; (5) compound 9 and compound 11 were reacted under the action of condensation reagent to obtain product 10, and the condensation reagent was selected as in step (1), and the obtained crude product was separated by column chromatography to obtain pure product; Reaction formula: in: The definitions of R ₁ -R ₄ , B ₁ , D ₁ , and D ₂ substituents are the same as the general formula (I) of claim 1; The definition of the ring A substituent is the same as that of the general formula (II) in claim 2. 17. The application of the macrocyclic epoxy ketone peptide derivatives according to claim 1 in the preparation of anti-tumor and anti-immune disease drugs, characterized in that, the macrocyclic epoxy ketone peptide derivatives include its Optical isomers or pharmaceutically acceptable salts or their solvates. 18. The application according to claim 17, wherein the tumor is selected from blood tumors such as myeloma, lymphoma, leukemia, and breast cancer, sarcoma, lung cancer, prostate cancer, colon cancer, rectal cancer, kidney cancer, etc. Cancer, pancreatic cancer, neuroblastoma, glioma, head cancer, neck cancer, thyroid cancer, liver cancer, ovarian cancer, vulvar cancer, cervical cancer, endometrial cancer, testicular cancer, bladder cancer, esophageal cancer, Gastric cancer, nasopharyngeal cancer, buccal cancer, oral cancer, gastrointestinal stromal tumor, skin cancer.