Camptothecin compound, preparation method therefor, and application thereof

Abstract

To provide camptothecin compounds and their conjugates with novel structures, improved efficacy and improved safety.SOLUTION: The present invention provides a compound having the structure of the upper formula in the figure, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotope-labeled product, metabolite or prodrug thereof. In the formula, R1 and R2 are each hydrogen, halogen, C1-6 alkyl, C1-6 alkoxyl, or the like, or are connected with adjacent carbon atoms to form a 5- to 6-membered oxygen-containing heterocyclic ring; R3 is hydrogen or is connected with an ortho-carbon atom of R1 to form a 6-membered carbocyclic ring; and A is selected from one of the lower two formulas in the figure.SELECTED DRAWING: None

Description

[Technical Field]

[0001] This application claims priority from Chinese Patent Application No. 202110159956.6, filed February 5, 2021; Chinese Patent Application No. 202110533304.4, filed May 17, 2021; Chinese Patent Application No. 202110718245.8, filed June 28, 2021; Chinese Patent Application No. 202110936768.X, filed August 16, 2021; and Chinese Patent Application No. 202111355330.9, filed November 16, 2021, the disclosures of which are incorporated herein in their entireties.

[0002] The present invention relates to camptothecin compounds and conjugates thereof having antitumor activity, as well as their preparation methods and applications in the medical field. [Background technology]

[0003] Camptothecin (CPT, Formula 1) was isolated from Camptotheca acuminata, a plant of the Botrytis family. It is a five-membered quinoline compound consisting of a quinoline ring AB, a pyrrole ring C, a pyridone ring D, and an α-hydroxylactone ring E, with the 20-position being in the S-configuration (see the structural formula below). In the early 1970s, camptothecin was used in clinical settings due to its excellent anticancer activity, but clinical trials were later terminated due to serious side effects such as diarrhea and hemorrhagic cystitis.

[0004] [ka]

[0005] Research data show that camptothecin can form a ternary complex with cellular DNA topoisomerase I, thereby inhibiting DNA unwinding, leading to blockage of DNA replication and cell death (Cancer Res. 1989, 49, 6365). Camptothecin and its derivatives have potent antitumor activity in animal models of lung, breast, colorectal, and ovarian cancers (Nature Review Cancer. 2006, 6, 789).

[0006] Currently, several camptothecin drugs are approved for marketing for the treatment of tumors (Med. Res. Rev. 2015, 35, 753). Irinotecan is used to treat colorectal cancer; topotecan is used to treat ovarian cancer; and belotecan is used to treat ovarian cancer and small cell lung cancer. Camptothecin derivatives further include exatecan, rubitecan, karenitecan, diflomotecan, lurtotecan, gimatecan, namitecan, cimitecan, siratecan, chimitecan, eromotecan, and the like.

[0007] Camptothecin drugs or their derivatives often have hematotoxicity induced by bone marrow suppression, such as neutropenia, leukopenia, thrombocytopenia, anemia, etc., as well as gastrointestinal side effects, such as nausea, vomiting, and diarrhea. Clinical studies have found that ways to improve the safety and efficacy of camptothecin compounds include improving their pharmacokinetic properties, adjusting activity, reducing dosage, or using their conjugates and antibodies to form antibody-conjugated drugs. Therefore, there is still a high clinical demand and application value for developing camptothecin compounds and their conjugates with novel structures, improved efficacy, and improved safety. Summary of the Invention

[0008] The present invention provides novel camptothecin compounds and their conjugates. The camptothecin compounds have good antitumor activity and are expected to be used in the treatment of tumor diseases; and the conjugates thereof have broad application prospects in ADC drugs. A first aspect of the present invention provides a compound, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitroxide, isotopically labeled product, metabolite, or prodrug thereof, wherein the compound has the structure shown below: [ka] (In the formula, R1 and R2 are each hydrogen, halogen, or C 1~6 Alkyl, C 1~6 Alkoxyl, C 1~6 Haloalkyl, hydroxyl, cyano and C 3~6 cycloalkyl; or R1 and R2 are joined to adjacent carbon atoms to form a 5-6 membered oxygen-containing heterocyclic ring; R3 is hydrogen or is joined to the ortho carbon atom of R1 to form a 6-membered carbocyclic ring; A is, [ka] is selected from one of R4 is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 3~6 selected from the group consisting of cycloalkyl and 3- to 6-membered heterocyclyl; R5 and R6 are each hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkylaminoalkyl, C 1~6 Alkoxyalkyl, C 2~6 Alkenyl, C 2~6 Alkynyl, C 3~6independently selected from the group consisting of cycloalkyl, 3- to 6-membered heterocyclyl, 3- to 6-membered heterocyclylalkyl, aryl, and heteroaryl; or R5 and R6 are joined to adjacent carbon atoms to form a 3- to 6-membered carbocyclic or heterocyclic ring; m=1 or 2) It has.

[0009] In some embodiments, the compound has the structure of Formula (I): [ka] (In formula (I), R x is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 3~6 selected from the group consisting of cycloalkyl and 3- to 6-membered heterocyclyl; R y and R z is not hydrogen at the same time, but hydrogen and C 1~6 Alkyl, C 1~6 Haloalkyl, C 2~6 Alkenyl, C 2~6 Alkynyl, C 1~6 Alkylaminoalkyl, C 1~6 independently selected from the group consisting of alkoxyalkyl, 3- to 6-membered heterocyclylalkyl, and 3- to 6-membered heterocyclyl It has.

[0010] In some embodiments, R x is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 3~6 selected from the group consisting of cycloalkyl and 3- to 6-membered heterocyclyl; R y and R z is not hydrogen at the same time, but hydrogen and C 2~6 Alkenyl, C 2~6 Alkynyl, C 1~6 Alkylaminoalkyl, C1~6 It is independently selected from the group consisting of alkoxyalkyl, 3- to 6-membered heterocyclylalkyl, and 3- to 6-membered heterocyclyl.

[0011] In some embodiments, in formula (I), R x is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 3~6 selected from the group consisting of cycloalkyl, 3- to 6-membered heterocyclyl; R y and R z is not hydrogen at the same time, but hydrogen and C 2~6 Alkenyl and C 2~6 alkynyl.

[0012] In some embodiments, R x is hydrogen or C 1~6 alkyl.

[0013] In some embodiments, R x is hydrogen.

[0014] In some embodiments, R y and R z is not simultaneously hydrogen, hydrogen, [ka] , dimethylaminomethylene, morpholinomethylene, and methoxymethylene.

[0015] In some embodiments, R y and R z is not simultaneously hydrogen, hydrogen, [ka] , dimethylaminomethylene, and methoxymethylene.

[0016] In some embodiments, R y is hydrogen and R z teeth, [ka] dimethylaminomethylene, morpholinomethylene, and methoxymethylene.

[0017] In some embodiments, R y is hydrogen and R z teeth, [ka] , dimethylaminomethylene and methoxymethylene.

[0018] In some embodiments, R x is hydrogen and R y is hydrogen and R z teeth, [ka] dimethylaminomethylene, morpholinomethylene, and methoxymethylene.

[0019] In some embodiments, R x is hydrogen and R y is hydrogen and R z teeth, [ka] , dimethylaminomethylene and methoxymethylene.

[0020] In some embodiments, R x is hydrogen and R y is hydrogen and R z teeth, [ka] and dimethylaminomethylene.

[0021] In some embodiments, in formula (I): [ka] teeth, [ka] It has the following arrangement.

[0022] In some embodiments, in formula (I): [ka] teeth, [ka] It has the following arrangement.

[0023] In some embodiments, in formula (I): [ka] teeth, [ka] It has the following arrangement.

[0024] In some embodiments, in formula (I): [ka] teeth, [ka] It has the following arrangement.

[0025] In some embodiments, in formula (I): [ka] teeth, [ka] It has the following arrangement.

[0026] In some embodiments, in formula (I): [ka] teeth, [ka] It has the following arrangement.

[0027] In some embodiments, in formula (I): [ka] teeth, [ka] It has the following arrangement.

[0028] In some embodiments, in formula (I): [ka] teeth, [ka] It has the following arrangement.

[0029] In some embodiments, the compound has the structure of Formula (II): [ka] (In formula (II), A' is [ka] is selected from one of R x’ is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 3~6 selected from the group consisting of cycloalkyl and 3- to 6-membered heterocyclyl; R y’ and R z’ is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 1~6 Alkylaminoalkyl, C 3~6 Cycloalkyl, 3- to 6-membered heterocyclyl, 3- to 6-membered heterocyclylalkyl, C 2~6 Alkenyl, C 2~6 independently selected from the group consisting of alkynyl, aryl, and heteroaryl, or R y’ and R z’ bonds to adjacent carbon atoms to form a 3- to 6-membered ring) It has.

[0030] In some embodiments, the structure of formula (II) is the following formula (II)-1: [ka] As shown in the figure.

[0031] In some embodiments, R x’ is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 3~6 selected from the group consisting of cycloalkyl, 3- to 6-membered heterocyclyl; R y’ and R z’is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 3~6 Cycloalkyl, 3-6 membered heterocyclyl, C 2~6 Alkenyl, C 2~6 independently selected from the group consisting of alkynyl, aryl, heteroaryl, or R y’ and R z’ is bonded to adjacent carbon atoms to form a 3- to 6-membered ring.

[0032] In some embodiments, the 3- to 6-membered ring is a 3- to 6-membered carbocyclic ring or a 3- to 6-membered heterocyclic ring.

[0033] In some embodiments, R x’ is hydrogen and C 1~6 alkyl.

[0034] In some embodiments, R x’ is selected from the group consisting of hydrogen and methyl.

[0035] In some embodiments, R y’ and R z’ is hydrogen, C 1~6 Alkyl, C 1~6 Alkoxyalkyl, C 1~6 Alkylaminoalkyl, C 3~6 Cycloalkyl and C 2~6 alkenyl, or R y’ and R z’ is linked to adjacent carbon atoms to form a 3- to 6-membered cycloalkyl.

[0036] In some embodiments, R y’ is hydrogen and C 1~6 alkyl; R z’ is hydrogen, C 1~6 Alkyl and C 3~6 cycloalkyl, or R y’ and R z’is bonded to adjacent carbon atoms to form a 3- to 6-membered ring.

[0037] In some embodiments, R y’ is selected from the group consisting of hydrogen and methyl; R z’ is selected from the group consisting of hydrogen, methyl and cyclopropyl, or R y’ and R z’ is linked to adjacent carbon atoms to form a three-membered carbocyclic ring.

[0038] In some embodiments, R x’ is selected from the group consisting of hydrogen and methyl; R y’ is selected from the group consisting of hydrogen and methyl; R z’ is selected from the group consisting of hydrogen, methyl and cyclopropyl, or R y’ and R z’ is linked to adjacent carbon atoms to form a three-membered carbocyclic ring.

[0039] In some embodiments, R x’ is hydrogen and R y’ is selected from the group consisting of hydrogen and methyl; R z’ is selected from the group consisting of hydrogen, methyl and cyclopropyl, or R y’ and R z’ is linked to adjacent carbon atoms to form a three-membered carbocyclic ring.

[0040] In some embodiments, in Formula (II), A' is [ka] is.

[0041] In some embodiments, in formula (II)-1, [ka] teeth, [ka] It has the following arrangement.

[0042] In some embodiments, in formula (II)-1, [ka] teeth, [ka] It has the following arrangement.

[0043] In some embodiments, in formula (II)-1, [ka] teeth, [ka] It has the following arrangement.

[0044] In some embodiments, in formula (II)-1, [ka] teeth, [ka] It has the following arrangement.

[0045] In some embodiments, in formula (II)-1, [ka] teeth, [ka] It has the following arrangement.

[0046] In some embodiments, in formula (II)-1, [ka] teeth, [ka] It has the following arrangement.

[0047] In some embodiments, in formula (II)-1, [ka] teeth, [ka] It has the following arrangement.

[0048] In some embodiments, in formula (II)-1, [ka] teeth, [ka] It has the following arrangement.

[0049] In some embodiments, the compound has the structure of Formula (III): [ka] (In formula (III), A″ is [ka] is selected from one of R x’’ is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 3~6selected from the group consisting of cycloalkyl and 3- to 6-membered heterocyclyl; R y’’ and R z’’ is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 1~6 Alkylaminoalkyl, C 3~6 Cycloalkyl, 3- to 6-membered heterocyclyl, 3- to 6-membered heterocyclylalkyl, 4- to 6-membered heterocyclyl, C 2~6 Alkenyl, C 2~6 independently selected from the group consisting of alkynyl, aryl, and heteroaryl, or R y’’ and R z’’ forms a 3- to 6-membered ring with adjacent carbon atoms) It has.

[0050] In some embodiments, the structure of the compound of formula (III) is the following formula (III)-1: [ka] As shown in the figure.

[0051] In some embodiments, R x’’ is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 3~6 selected from the group consisting of cycloalkyl, 3- to 6-membered heterocyclyl; R y’’ and R z’’ is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 3~6 Cycloalkyl, 4-6 membered heterocyclyl, C 2~6 Alkenyl, C 2~6 independently selected from the group consisting of alkynyl, aryl, heteroaryl, or R y’’ and R z’’ is bonded to adjacent carbon atoms to form a 3- to 6-membered ring.

[0052] In some embodiments, R x’’ is hydrogen and C 1~6 alkyl.

[0053] In some embodiments, R x’’ is hydrogen.

[0054] In some embodiments, the 3- to 6-membered ring is a 3- to 6-membered carbocyclic ring or a 3- to 6-membered heterocyclic ring.

[0055] In some embodiments, R y’’ and R z’’ are hydrogen and C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 1~6 Alkylaminoalkyl, C 3~6 independently selected from the group consisting of cycloalkyl and vinyl, or R y’’ and R z’’ is bonded to adjacent carbon atoms to form a 3- to 6-membered ring.

[0056] In some embodiments, R y’’ is hydrogen and R z’’ is hydrogen, C 1~6 Alkyl, C 3~6 cycloalkyl and vinyl, or R y’’ and R z’’ is bonded to adjacent carbon atoms to form a 3- to 6-membered carbocyclic ring.

[0057] In some embodiments, R y’’ is hydrogen and R z’’ is selected from the group consisting of hydrogen, methyl, cyclopropyl, and vinyl, or R y’’ and R z’’ is linked to adjacent carbon atoms to form a three-membered carbocyclic ring.

[0058] In some embodiments, R x’’ is hydrogen and R y’’is hydrogen and R z’’ is selected from the group consisting of hydrogen, methyl, cyclopropyl, and vinyl, or R y’’ and R z’’ is linked to adjacent carbon atoms to form a three-membered carbocyclic ring.

[0059] In some embodiments, in formula (III), A″ is [ka] is.

[0060] In some embodiments, in formula (III)-1, [ka] teeth, [ka] It has the following arrangement.

[0061] In some embodiments, in formula (III)-1, [ka] teeth, [ka] It has the following arrangement.

[0062] In some embodiments, in formula (III)-1, [ka] teeth, [ka] It has the following arrangement.

[0063] In some embodiments, in formula (III)-1, [ka] teeth, [ka] It has the following arrangement.

[0064] In some embodiments, in formula (III)-1, [ka] teeth, [ka] It has the following arrangement.

[0065] In some embodiments, in formula (III)-1, [ka] teeth, [ka] It has the following arrangement.

[0066] In some embodiments, in formula (III)-1, [ka] teeth, [ka] It has the following arrangement.

[0067] In some embodiments, in formula (III)-1, [ka] teeth, [ka] It has the following arrangement.

[0068] In some embodiments, the compound has the structure of Formula (IV): [ka] (In formula (IV), R a and R b is hydrogen, halogen, C 1~6 Alkyl, C 1~6 Hydroxyalkyl, C 1~6 Alkoxyalkyl, C 1~6 Alkoxy, C 1~6 independently selected from the group consisting of haloalkyl, hydroxyl, and cyano; or R a and R b is bonded to adjacent carbon atoms to form a 5- to 6-membered oxygen-containing heterocyclic ring; R c and R d is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 1~6 Alkylaminoalkyl, C 3~6 Cycloalkyl, 3- to 6-membered heterocyclyl, 3- to 6-membered heterocyclylalkyl, C 2~6 Alkenyl and C 2~6 alkynyl, or R c and R d combines with adjacent carbon atoms to form a 3- to 6-membered carbocyclic or heterocyclic ring; R e is hydrogen, C 1~6 Alkyl, C 3~6 Cycloalkyl, C 1~6 Haloalkyl, C 1~6 selected from the group consisting of alkoxyalkyl and C2-C5 heterocyclyl; q=0 or 1; When q=0, R c and R d cannot also be hydrogen) It has.

[0069] In some embodiments, R a and R b is hydrogen, halogen, C 1~6 Alkyl, C 1~6 Alkoxy, C 1~6 independently selected from the group consisting of haloalkyl, hydroxyl, and cyano; or R a and R b is bonded to adjacent carbon atoms to form a 5- to 6-membered oxygen-containing heterocyclic ring; R c and R d is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 1~6 Alkylaminoalkyl, C 3~6 Cycloalkyl, 3- to 6-membered heterocyclyl, 3- to 6-membered heterocyclylalkyl, C 2~6 Alkenyl and C 2~6 alkynyl, or R c and R d combines with adjacent carbon atoms to form a 3- to 6-membered carbocyclic or heterocyclic ring; R e is hydrogen, C 1~6 Alkyl, C 3~6 Cycloalkyl, C 1~6 Haloalkyl, C 1~6 selected from the group consisting of alkoxyalkyl and 4- to 6-membered heterocyclyl; q=0 or 1; When q=0, R c and R d cannot simultaneously be hydrogen.

[0070] In some embodiments, in formula (IV), R a and R b is hydrogen, halogen, C 1~6 Alkyl, C1~6 Alkoxy, C 1~6 independently selected from the group consisting of haloalkyl, hydroxyl, and cyano; or R a and R b is bonded to adjacent carbon atoms to form a 5- to 6-membered oxygen-containing heterocyclic ring; R c and R d is hydrogen, C 1~6 Alkyl, C 3~6 Cycloalkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, 4-6 membered heterocyclyl, C 2~6 Alkenyl, C 2~6 alkynyl, or R c and R d combines with adjacent carbon atoms to form a 3- to 6-membered carbocyclic or heterocyclic ring; R e is hydrogen, C 1~6 Alkyl, C 3~6 Cycloalkyl, C 1~6 Haloalkyl, C 1~6 selected from the group consisting of alkoxyalkyl and 4- to 6-membered heterocyclyl; q=0 or 1; When q=0, R c and R d cannot simultaneously be hydrogen.

[0071] In some embodiments, R a and R b is hydrogen, halogen and C 1~6 alkyl, or R a and R b is bonded to adjacent carbon atoms to form a 5- or 6-membered oxygen-containing heterocyclic ring.

[0072] In some embodiments, R a and R b are independently selected from the group consisting of hydrogen, fluorine, chlorine and methyl, or R a and R b together with the benzene rings attached to them, [ka] wherein Z is selected from the group consisting of -CH2-, -CD2-, -CH2CH2-, and -CF2-. Form.

[0073] In some embodiments, R a is methyl and R b is fluorine or R a and R b together with the benzene rings attached to them, [ka] Form.

[0074] In some embodiments, R c and R d is hydrogen, [ka] , C 1~6 Alkoxyalkyl and C 1~6 alkylaminoalkyl, or R c and R d is bonded to adjacent carbon atoms to form a 3- to 6-membered carbocyclic ring.

[0075] In some embodiments, R c is hydrogen and R d is hydrogen, [ka] , methoxyethyl, and cyclopropyl, or R c and R d is bonded to adjacent carbon atoms to form a 3- to 6-membered carbocyclic ring.

[0076] In some embodiments, R e is hydrogen and C1~6 alkyl.

[0077] In some embodiments, R e is selected from the group consisting of hydrogen and isopropyl.

[0078] In some embodiments, R a is methyl and R b is fluorine or R a and R b together with the benzene rings attached to them, [ka] Form R e is selected from the group consisting of hydrogen and isopropyl; R c is hydrogen and R d is hydrogen, [ka] , methoxyethyl, and cyclopropyl, or R c and R d is linked to adjacent carbon atoms to form a three-membered carbocyclic ring.

[0079] In some embodiments, R a is methyl and R b is fluorine or R a and R b together with the benzene rings attached to them, [ka] Form R e is selected from the group consisting of hydrogen and isopropyl; R c is hydrogen and R d is selected from the group consisting of hydrogen, methoxyethyl and cyclopropyl, or R c and R d is linked to adjacent carbon atoms to form a three-membered carbocyclic ring.

[0080] In some embodiments, in formula (IV): [ka] teeth, [ka] It has the following arrangement.

[0081] In some embodiments, in formula (IV): [ka] teeth, [ka] It has the following arrangement.

[0082] In some embodiments, the compound has the structure of Formula (V): [ka] (In formula (V), R is C 3~6 Cycloalkyl and C 1~6 selected from the group consisting of alkoxy; A''' is [ka] is selected from one of R x’’’ is hydrogen, C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 3~6 selected from the group consisting of cycloalkyl and 3- to 6-membered heterocyclyl; R y’’’ and R z’’’ is hydrogen, C 1~6 Alkyl, C 1~6Haloalkyl, C 1~6 Alkoxyalkyl, C 1~6 Alkylaminoalkyl, C 3~6 Cycloalkyl, 3- to 6-membered heterocyclyl, 3- to 6-membered heterocyclylalkyl, C 2~6 Alkenyl, C 2~6 independently selected from the group consisting of alkynyl, aryl, and heteroaryl, or R y’’’ and R z’’’ forms a 3- to 6-membered ring with adjacent carbon atoms) It has.

[0083] In some embodiments, R is selected from the group consisting of methoxy and cyclopropyl.

[0084] In some embodiments, the structure of the compound of formula (V) is represented by the following formula (V)-1: [ka] As shown in the figure.

[0085] In some embodiments, the 3- to 6-membered ring is a 3- to 6-membered carbocyclic ring or a 3- to 6-membered heterocyclic ring.

[0086] In some embodiments, R y’’’ and R z’’’ are hydrogen and C 1~6 Alkyl, C 1~6 Haloalkyl, C 1~6 Alkoxyalkyl, C 1~6 Alkylaminoalkyl, C 3~6 independently selected from the group consisting of cycloalkyl and vinyl, or R y’’’ and R z’’’ is linked to adjacent carbon atoms to form a 3- to 6-membered carbocyclic ring.

[0087] In some embodiments, R y’’’ and R z’’’ is hydrogen or R y’’’ and R z’’’is linked to adjacent carbon atoms to form a 3- to 6-membered carbocyclic ring.

[0088] In some embodiments, R x’’’ is hydrogen and C 1~6 alkyl.

[0089] In some embodiments, R x’’’ is hydrogen.

[0090] In some embodiments, A''' is [ka] is.

[0091] In some embodiments, R is selected from the group consisting of methoxy and cyclopropyl; x’’’ is hydrogen and R y’’’ and R z’’’ are both hydrogen, or R y’’’ and R z’’’ is linked to adjacent carbon atoms to form a three-membered carbocyclic ring.

[0092] In some embodiments, in formula (V)-1, [ka] teeth, [ka] It has the following arrangement.

[0093] In some embodiments, in formula (V)-1, [ka] teeth, [ka] It has the following arrangement.

[0094] In some embodiments, in formula (V)-1, [ka] teeth, [ka] It has the following arrangement.

[0095] In some embodiments, in formula (V)-1, [ka] teeth, [ka] It has the following arrangement.

[0096] In some embodiments, in formula (V)-1, [ka] teeth, [ka] It has the following arrangement.

[0097] In some embodiments, in formula (V)-1, [ka] teeth, [ka] It has the following arrangement.

[0098] In some embodiments, in formula (V)-1, [ka] teeth, [ka] It has the following arrangement.

[0099] In some embodiments, in formula (V)-1, [ka] teeth, [ka] It has the following arrangement.

[0100] In some embodiments, the present invention provides the following compound: [ka] TIFF2025160317000101.tif226149 TIFF2025160317000102.tif226149 TIFF2025160317000103.tif226149 TIFF2025160317000104.tif226149 to provide.

[0101] On the other hand, the present invention also provides a compound of formula (VI), or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite or prodrug thereof: MLED Formula (VI) (In the formula, M is a linker moiety of an antibody or antigen-binding fragment thereof; L is a linker connecting linker moieties M and E; E is a structural fragment connecting L and D; D is a structural fragment of a cytotoxic drug to provide.

[0102] In some embodiments, M has the following structure: [ka] is selected from the group consisting of:

[0103] In some embodiments, M has the following structure: [ka] is selected from the group consisting of:

[0104] In some embodiments, L is C 1~6 Alkylene, -N(R')-, Carbonyl, -O-, Val, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn, Val-Cit, Val-Ala, Val-Lys, Val-Lys(Ac), Phe-Lys, Phe-Lys( Ac), D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn, Ala-Ala-Ala, Val-Lys-Ala, Gly-Gly-Gly, Gly-Gly-Phe-Gly, Gly-Gly-Gly-Gly-Gly, [ka] (Wherein R' is hydrogen, C 1~6 Alkyl or -(CH2CH2O) r -containing alkyl; r is an integer selected from 1 to 10; and s is an integer selected from 1 to 10. It is a bivalent structure composed of one or more selected from the group consisting of:

[0105] In some embodiments, L has the following structure: [ka] is selected from.

[0106] In some embodiments, L has the following structure: [ka] is selected from.

[0107] In some embodiments, E is a single bond, —NH—CH—, [ka] is selected from the group consisting of:

[0108] In some embodiments, E is —NH—CH 2 —.

[0109] In some embodiments, the cytotoxic agent is selected from a compound according to any one of the items of the first aspect of the present invention.

[0110] In some embodiments, the cytotoxic agent is selected from the group consisting of compounds of the present invention: 1-1 to 1-15; 2-1 to 2-27; 3-1 to 3-26; 4-1 to 4-15; or 5-1 to 5-36.

[0111] In some embodiments, D is selected from structures formed by removing a hydrogen atom from a compound of the invention.

[0112] In some embodiments, D is selected from structures formed by removing a hydrogen atom from compounds of the invention: 1-1 through 1-15; 2-1 through 2-27; 3-1 through 3-26; 4-1 through 4-15; or 5-1 through 5-36.

[0113] In some embodiments, D has the following structure: [ka] is selected from the group consisting of:

[0114] In some embodiments, D has the following structure: [ka] is selected from the group consisting of:

[0115] In some embodiments, the MLED is the following compound: [ka] TIFF2025160317000114.tif218149 TIFF2025160317000115.tif221149 TIFF2025160317000116.tif199149 TIFF2025160317000117.tif90149 is selected from the group consisting of:

[0116] In some embodiments, the MLED is the following compound: [ka] is selected from the group consisting of: DETAILED DESCRIPTION OF THE INVENTION

[0117] definition Unless otherwise defined herein below, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art. References to technology used herein are intended to refer to technology as commonly understood in the art, including modifications of the technology or equivalent technical substitutions that would be obvious to those skilled in the art. Although the following terms are believed to be well understood by those skilled in the art, the following definitions are provided to better explain the present invention.

[0118] As used herein, the terms "comprising," "including," "having," "containing," or "involving," and variations thereof, are inclusive or open-ended and do not exclude other, unrecited elements or method steps.

[0119] As used herein, the marking of "*" in a structural formula of a compound indicates that the marked carbon atom is a chiral carbon atom, and the present invention includes the pair of enantiomers formed by the chiral carbon atom. When a compound contains two different chiral carbon atoms, the present invention includes the four optical isomers formed by the chiral carbon atoms.

[0120] As used herein, " [ka] " indicates a bond that may be stereospecific ((R) or (S)) or non-stereospecific.

[0121] The term "alkyl" is defined as a saturated, straight-chain or branched aliphatic hydrocarbon group. In some embodiments, alkyl has 1 to 12, e.g., 1 to 6, carbon atoms. For example, as used herein, "C 1~6 The term "alkyl" refers to a straight-chain or branched alkyl group having 1 to 6 carbon atoms (e.g., methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, and n-hexyl), optionally substituted by one or more (e.g., 1, 2, or 3) suitable substituents.

[0122] The term "alkenyl" refers to a straight or branched chain hydrocarbon group containing at least one carbon-carbon double bond, e.g., "C 2~6 alkenyl," "C 2~4Examples include, but are not limited to, vinyl, 1-propenyl, 2-propenyl, 1-butenyl, 2-butenyl, 1,3-butadienyl, 1-pentenyl, 2-pentenyl, 3-pentenyl, 1,3-pentadienyl, 1,4-pentadienyl, 1-hexenyl, 2-hexenyl, 3-hexenyl, 1,4-hexdienyl, and the like.

[0123] The term "alkynyl" refers to a straight or branched chain hydrocarbon group containing at least one carbon-carbon triple bond, e.g., "C 2~6 alkynyl", "C 4~6 Examples include, but are not limited to, ethynyl, 1-propynyl, 2-propynyl, 1-butynyl, 2-butynyl, 1,3-butadiynyl, 1-pentynyl, 2-pentynyl, 3-pentynyl, 1,3-pentadiynyl, 1,4-pentadiynyl, 1-hexynyl, 2-hexynyl, 3-hexynyl, 1,4-hexadiynyl, and the like.

[0124] The term "cycloalkyl" refers to saturated cyclic hydrocarbon groups, including, but not limited to, monocycloalkyls and bicycloalkyls (e.g., spiro-cycloalkyls, fused cycloalkyls, and bridged cycloalkyls). 3~6 The term "cycloalkyl" refers to a cycloalkyl having 3 to 6 ring-forming carbon atoms, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and the like, which may be optionally substituted with one or more (e.g., 1, 2, or 3) suitable substituents, for example, methyl-substituted cyclopropyl.

[0125] The term "carbocyclic ring" or "carbocyclyl" refers to a saturated or partially unsaturated hydrocarbon group having a non-aromatic monocyclic or polycyclic structure, which is attached to the rest of the compound through a ring-forming carbon atom. Examples include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloheptyl, and cyclooctyl.

[0126] The term "carbocyclic ring" refers to a saturated or unsaturated non-aromatic monocyclic or polycyclic (e.g., bicyclic) hydrocarbon ring (e.g., a monocyclic ring such as cyclopropane, cyclobutane, cyclopentane, cyclohexane, cycloheptane, cyclooctane, or cyclononane, or a spirocyclic, fused, or bridged ring system (e.g., a bicyclic ring including a bicyclo[1.1.1]pentane, bicyclo[2.2.1]heptane, bicyclo[3.2.1]octane, or bicyclo[5.2.0]nonane ring, decahydronaphthalene, or the like), which may be optionally substituted with one or more (e.g., one, two, or three) suitable substituents. The term "3- to 6-membered carbocyclic ring" refers to a carbocyclic ring containing 3, 4, 5, or 6 ring-forming carbon atoms.

[0127] The term "heterocyclyl" or "heterocyclic ring" refers to a saturated or partially saturated monocyclic or polycyclic (e.g., bicyclic) non-aromatic ring structure whose ring-forming atoms consist of carbon atoms and at least one (e.g., 1, 2, or 3) heteroatom selected from the group consisting of nitrogen, oxygen, and sulfur. A heterocyclyl can be attached to the remainder of the molecule through any ring atom as long as valence requirements are met. A heterocyclyl of the present invention is preferably a 3- to 6-membered heterocyclyl. The term "3- to 6-membered heterocyclyl" herein refers to a heterocyclic group having 3 to 6 ring atoms, and includes 3-, 4-, 5-, and 6-membered heterocyclyls, including nitrogen- and oxygen-containing heterocyclyls, such as 4- to 6-membered heterocyclyl, 4- to 6-membered nitrogen-containing heterocyclyl, and 4- to 6-membered oxygen-containing heterocyclyl. In general, heterocyclic groups include, but are not limited to, azetidinyl, oxetanyl, tetrahydrofuryl, pyrrolidinyl, pyrrolidinonyl, imidazolidinyl, pyrazolidinyl, tetrahydropyranyl, piperidinyl, piperazinyl, and morpholinyl. Heterocyclyls of the present invention can be optionally substituted with one or more substituents described herein. Heterocyclyls of the present invention can be optionally fused with one or more aromatic or non-aromatic rings.

[0128] The term "oxygen-containing heterocyclic ring" refers to a heterocyclic ring as described above in which one or more (e.g., 1, 2, or 3) ring atoms are oxygen atoms, for example, a 5- or 6-membered oxygen-containing heterocyclic ring, and specific examples include, but are not limited to, an oxirane ring, a tetrahydrofuran ring, a furan ring, a tetrahydropyran ring, a pyran ring, etc. In the present invention, the term "nitrogen-containing heterocyclic ring" refers to a heterocyclic ring as described above in which one or more (e.g., 1, 2, or 3) ring atoms are nitrogen atoms.

[0129] The term "haloalkyl" refers to an alkyl group substituted by one or more (e.g., 1, 2, or 3) of the same or different halogen atoms, where alkyl is as defined above. For example, "C 1~6The term "haloalkyl" refers to a haloalkyl having 1 to 6 carbon atoms. Common haloalkyls include, but are not limited to, -CHF, -CHF, -CF, -CHCF, -CFCF, -CHCHCF, -CHCl, and the like. Haloalkyls of the present invention can be optionally substituted with one or more substituents described herein.

[0130] The term "aryl" refers to a group obtained by removing a hydrogen atom from a carbon atom of an aromatic nucleus of an aromatic hydrocarbon molecule, e.g., a 6- to 14-membered aryl, and examples include, but are not limited to, phenyl, naphthyl, anthracenyl, and the like.

[0131] The term "heteroaryl" refers to an aromatic ring group containing at least one ring member selected from the group consisting of N, O, and S. Specific examples include, but are not limited to, 5- to 6-membered heteroaryls, 5- to 6-membered nitrogen-containing heteroaryls, and 5- to 6-membered oxygen-containing heteroaryls, such as furyl, thienyl, pyrrolyl, thiazolyl, isothiazolyl, thiadiazolyl, oxazolyl, isoxazolyl, oxadiazolyl, imidazolyl, pyrazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, pyridyl, pyrimidinyl, pyridazinyl, pyrazinyl, 1,2,3-triazinyl, 1,3,5-triazinyl, and 1,2,4,5-tetrazinyl.

[0132] The term "alkoxy" refers to a group having the structure "alkyl-O-", where alkyl is as defined above. Examples include C 1~6 Alkoxy, C 1~4 Alkoxy, C 1~3 Alkoxy or C 1~2Common alkoxy groups include (but are not limited to) methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, pentyloxy, hexyloxy, etc. Alkoxy groups in the present invention are optionally substituted with one or more substituents described herein.

[0133] The term "alkoxyalkyl" refers to an alkyl substituted with one or more (e.g., 1, 2, 3, or 4) alkoxy groups, where alkoxy and alkyl are as defined above. For example, the term "C 1~6 The term "alkoxyalkyl" refers to an alkyl having 1 to 6 carbon atoms and substituted with one or more (e.g., 1, 2, 3, or 4) alkoxy groups. Common alkoxyalkyls include (but are not limited to) CHO-CH-, CH-O-CH-, CH-O-CHCH-, and the like.

[0134] The term "halo" or "halogen" group is defined to include F, Cl, Br, or I.

[0135] The term "nitrogen oxide" refers to an oxide (e.g., mono- or di-oxide) of at least one nitrogen atom in the structure of a compound of the present application. The nitrogen mono-oxide may exist as a single positional isomer or a mixture of positional isomers.

[0136] The term "substituted" means the selective replacement of one or more (e.g., 1, 2, 3, or 4) hydrogens on the designated atom with the indicated group, provided that the specified atom's normal valence in the present environment is not exceeded and the substitution results in a stable compound. Combinations of substituents and / or variables are permissible only if such combinations result in stable compounds.

[0137] When a substituent is described as "optionally substituted," the substituent may be (1) unsubstituted or (2) substituted. When a carbon atom of a substituent is described as being optionally substituted with one or more substituents from a list of substituents, one or more hydrogens on the carbon atom (up to any hydrogens present) may be individually and / or simultaneously replaced with any individually selected substituents. When a nitrogen atom of a substituent is described as being optionally substituted with one or more of the listed substituents, one or more hydrogens on the nitrogen atom (up to any hydrogens present) may each independently be replaced with any selected substituent.

[0138] When substituents are described as being "independently selected from" a group, each substituent is selected independently of the other. Thus, each substituent can be the same or different from another (other) substituent.

[0139] As used herein, the term "one or more" means one or more than one, for example, two, three, four, five or ten, under appropriate circumstances.

[0140] As used herein, unless otherwise indicated, the site of attachment of a substituent may be at any suitable position on the substituent.

[0141] The term "stereoisomer" refers to an isomer formed as a result of at least one asymmetric center. In compounds possessing one or more (e.g., one, two, three, or four) asymmetric centers, racemic mixtures, single enantiomers, diastereomeric mixtures, and individual diastereoisomers may be produced. Certain individual molecules may also exist as geometric isomers (cis / trans). Similarly, compounds of the present invention may exist as mixtures of two or more structurally distinct forms (commonly referred to as tautomers) in rapid equilibrium. Representative examples of tautomers include keto-enol tautomers, phenol-keto tautomers, nitroso-oxime tautomers, imine-enamine tautomers, and the like. It will be understood that the scope of the present invention encompasses all such isomers or mixtures thereof in any proportion (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%).

[0142] Carbon-carbon bonds in the compounds of the present invention are represented herein by solid lines ( [ka] ), solid wedge ( [ka] ) or dashed wedge ( [ka] ) A solid line used to depict bonds to asymmetric carbon atoms is intended to indicate that all possible stereoisomers of that carbon atom (e.g., a specific enantiomer, a racemic mixture, etc.) are included. A solid or dashed wedge used to depict bonds to asymmetric carbon atoms is intended to represent the stereoisomer as shown. When a racemic mixture is depicted, the solid or dashed wedge is intended to define relative stereochemistry, not absolute stereochemistry. Unless otherwise indicated, compounds of the present invention may exist in stereoisomeric forms, including cis and trans isomers, optical isomers (e.g., R and S enantiomers), diastereomers, geometric isomers, rotamers, conformational isomers, atropisomers, and mixtures thereof. Compounds of the present invention may exhibit and consist of more than one type of isomerism, such as racemic mixtures and diastereoisomeric pairs.

[0143] The present invention includes all possible crystalline forms or polymorphs of the compounds of the present invention, which may be a single polymorph or a mixture of more than one polymorph in any proportion.

[0144] It is also understood that certain compounds of the present invention may exist in free form for therapeutic use, or, where appropriate, in the form of a pharmaceutically acceptable derivative thereof. In the present invention, a pharmaceutically acceptable derivative includes, but is not limited to, a pharmaceutically acceptable salt, ester, solvate, metabolite, or prodrug, which, after administration to a patient, can directly or indirectly provide the compound of the present invention or its metabolite or residue. Thus, when reference is made herein to a "compound of the present invention," it is also intended to encompass various derivative forms of the compound described above.

[0145] Pharmaceutically acceptable salts of the compounds of the present invention include the acid addition and base salts thereof.

[0146] Suitable acid addition salts can be formed from acids capable of forming pharmaceutically acceptable salts, including aspartate, fumarate, glucoheptonate, gluconate, glucuronate, hexafluorophosphate, and the like.

[0147] Suitable base addition salts can be formed from bases capable of forming pharmaceutically acceptable salts including aluminum salts, alginate salts, choline salts, diethylamine salts and the like.

[0148] For a review of suitable salts, see Stahl and Wermuth, "Handbook of Pharmaceutical Salts: Properties, Selection, and Use" (Wiley-VCH, 2002). Methods for preparing pharmaceutically acceptable salts of the compounds of the invention are known to those skilled in the art.

[0149] The term "ester" refers to an ester derived from each of the compounds of the formulae herein, and includes physiologically hydrolyzable esters (capable of being hydrolyzed under physiological conditions to release a compound of the invention in the form of a free acid or alcohol). The compounds of the invention themselves may also be esters.

[0150] The compounds of the present invention may exist in the form of solvates, preferably hydrates, which contain polar solvents (e.g., water, methanol or ethanol, among others) as structural elements of the crystalline lattice of the compounds. The amount of polar solvent, especially water, may be present in a stoichiometric or non-stoichiometric ratio.

[0151] The scope of the present invention further includes metabolites of the compounds of the invention, i.e., substances formed in vivo when the compounds of the invention are administered. Such substances may result, for example, from oxidation, reduction, hydrolysis, amidation, deamidation, esterification, delipidation, enzymatic degradation, etc. of the administered compound. Thus, the present invention includes metabolites of the compounds of the invention, including compounds produced by contact of a compound of the invention with a mammal for a time sufficient to produce a metabolite thereof.

[0152] The scope of the present invention may further include prodrugs of the compounds of the present invention. Typically, such prodrugs are functional derivatives of the compounds, which are easily converted in vivo into the desired therapeutically active compounds. Therefore, in these cases, the term "administration" used in the therapeutic methods of the present invention includes treating various diseases or conditions using one or more prodrugs of the compounds of the present invention, where the prodrugs are converted into the aforementioned compounds in vivo after administration to a subject. For example, "Design of Prodrug", edited by H. Bundgaard, Elsevier, 1985, describes general methods for selecting and preparing suitable prodrug derivatives.

[0153] The scope of the present invention further includes isotopically labeled products of the compounds of the present invention, which are identical to the compounds of the present invention except that at least one atom thereof is replaced by an atom having the same atomic number but an atomic mass or mass number different from the atomic mass or mass number of the atom predominant in nature.

[0154] The present invention also encompasses compounds of the present invention that contain protecting groups. During any method for preparing compounds of the present invention, it may be necessary / and / or desirable to protect sensitive or reactive groups on any molecule involved, thereby forming a chemically protected form of the compound of the present invention. This can be achieved by conventional protecting groups, such as those described in Protective Groups in Organic Chemistry, edited by J.F.W.M. Commie, Plenum Press, 1973; and T.W. Greene & P.G.W. Muts, Protective Groups in Organic Synthesis, John Wiley & Sons, 1991. These references are incorporated herein by reference. Protecting groups may be removed at a suitable subsequent stage using methods known in the art.

[0155] Pharmaceutical Composition In a third aspect, the present invention provides a pharmaceutical composition comprising a compound according to the first or second aspect of the invention, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite or prodrug thereof, and one or more pharmaceutically acceptable carriers.

[0156] The term "pharmaceutical composition" refers to a composition that can be used as a medicine, which comprises an active pharmaceutical ingredient (API) (or therapeutic agent) and, optionally, one or more pharmaceutically acceptable carriers. The term "pharmaceutically acceptable carrier" refers to an excipient that is suitable for administration with a therapeutic agent, to come into contact with the tissues of human beings and / or other animals without causing undue toxicity, irritation, anaphylaxis, or other problem or complication, and that is consistent with sound medical judgment and with a reasonable benefit / risk ratio.

[0157] The pharmaceutical compositions described above can act systemically and / or locally, which can be achieved by suitable dosage forms, including but not limited to tablets, capsules, lozenges, hard troches, powders, sprays, creams, ointments, suppositories, gels, pastes, lotions, salves, aqueous suspensions, injectable solutions, elixirs, and syrups.

[0158] The pharmaceutical compositions described above may contain 0.01 mg to 1000 mg of at least one compound of the present invention, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof.

[0159] The present invention also provides a method for preparing the above-mentioned pharmaceutical composition or its corresponding dosage form, comprising combining at least one compound of the present invention, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, with one or more pharmaceutically acceptable carriers.

[0160] Kit Products In a fourth aspect, the present invention provides a method for producing a composition comprising: a) as a first therapeutic agent, at least one compound described in the first or second aspect of the invention, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite or prodrug thereof, or as a first pharmaceutical composition, a pharmaceutical composition described in the third aspect; b) optionally, as a second therapeutic agent, at least one additional therapeutic agent, or as a second pharmaceutical composition, a pharmaceutical composition comprising an additional therapeutic agent; c) optionally, packaging and / or instructions for use; A kit product comprising:

[0161] The kit product may comprise 0.01 mg to 1000 mg of at least one compound of the present invention, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitroxide, isotopically labeled product, metabolite, or prodrug thereof.

[0162] The present invention also provides a method for preparing the above-described kit, comprising combining at least one compound of the present invention, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitroxide, isotopically labeled product, metabolite, or prodrug thereof, or the above-described pharmaceutical composition, with optionally at least one additional therapeutic agent or a pharmaceutical composition comprising the additional therapeutic agent, and optional packaging and / or instructions for use.

[0163] medical use The compounds of the present invention may exhibit potent effects on inhibiting abnormal cell proliferation.

[0164] Accordingly, the present application provides a compound of the present invention, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, or a pharmaceutical composition thereof, for use in treating a disease associated with abnormal cell proliferation.

[0165] Additionally, the present application also provides the use of a compound of the present invention, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, or a pharmaceutical composition as described above, in the manufacture of a medicament for treating a disease associated with abnormal cell proliferation.

[0166] In some embodiments, diseases associated with abnormal cell proliferation include, but are not limited to, tumors, for example, advanced solid tumors.

[0167] The present application also provides the use of a compound of the present invention, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitroxide, isotopically labeled product, metabolite, or prodrug thereof, or the aforementioned pharmaceutical composition, in the manufacture of a preparation, wherein the preparation is used to inhibit tumor cell growth. In certain embodiments, the preparation is used in vivo or in vitro. For example, the preparation may be administered to a subject to inhibit tumor cell growth in the subject; or the preparation may be applied to cells in vitro (e.g., cell lines or cells derived from a subject) to inhibit tumor cell growth in vitro.

[0168] Tumors of the present invention include (but are not limited to) brain tumors, lung cancer, squamous cell carcinoma, bladder cancer, gastric cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colorectal cancer, liver cancer, kidney cancer, esophageal adenocarcinoma, esophageal squamous cell carcinoma, prostate cancer, cancer of the female reproductive system, carcinoma in situ, lymphoma, neurofibroma, thyroid cancer, bone cancer, skin cancer, brain cancer, colon cancer, testicular cancer, gastrointestinal stromal tumor, prostate tumor, mast cell tumor, multiple myeloma, melanoma, glioma, or sarcoma.

[0169] Treatment method In another aspect, the present invention provides a method for treating a disease associated with abnormal cell proliferation, comprising administering to an individual in need thereof a therapeutically effective amount of a compound of the present invention, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitroxide, isotopically labeled product, metabolite, or prodrug thereof, or a pharmaceutical composition as described above.

[0170] The term "effective amount" refers to a dose that is capable of eliciting a biological or medical response in a cell, tissue, organ or organism (e.g., an individual) and is sufficient to achieve the desired prophylactic and / or therapeutic effect.

[0171] The dosage regimen may be adjusted to obtain the optimum desired response. For example, it may be administered as a single dose or as divided doses over time, or the dose may be proportionally reduced or increased according to the actual situation. It is recognized that for any particular individual, the specific dosage regimen will be adjusted according to the needs and professional judgment of the person administering or supervising the administration of the composition.

[0172] The dosage of the compounds of the present invention will depend on the individual's condition, the severity of the disease or condition, the rate of administration, the nature of the compound, and the judgment of the prescribing physician. Generally, an effective amount is about 0.001 to 10,000 mg / kg of subject body weight / day. Where appropriate, an effective amount is about 0.01 to 1,000 mg / kg of subject body weight / day. About 0.01 to 1,000 mg / kg of subject body weight, usually about 0.1 to 500 mg / kg of subject body weight, can be administered daily, every two days, or every three days. Exemplary dosage regimens are one or more times daily, one or more times weekly, or one or more times monthly. In the case of multiple administrations, the interval between single doses can generally be daily, weekly, monthly, or yearly. Alternatively, it can be administered as a sustained-release preparation, in which case less frequent administration is required. Dosage and frequency of administration may vary depending on the half-life of the drug in the subject and whether the use is for prophylaxis or treatment. For prophylactic use, relatively low doses are administered at relatively infrequent intervals over a long period of time; for therapeutic use, relatively higher doses may need to be administered at shorter intervals until disease progression is slowed or halted, preferably until the individual shows partial or complete alleviation of disease symptoms, after which prophylactic use may be employed.

[0173] The term "treating" means alleviating or eliminating the target disease or condition. When a subject receives a therapeutic amount of a compound of the present invention or a pharmaceutically acceptable form thereof, or a pharmaceutical composition of the present invention, at least one indicator and symptom of the subject shows observable and / or detectable remission and / or improvement, indicating that the subject has been successfully "treated." It is understood that treatment includes not only complete cure, but also the achievement of some biologically or medically relevant result that is less than a complete cure.

[0174] The term "administer / administering / administration" refers to the process of applying an active pharmaceutical ingredient (e.g., a compound of the present invention) or a pharmaceutical composition containing an active pharmaceutical ingredient (e.g., a pharmaceutical composition of the present invention) to an individual or its cells, tissues, organs, biological fluids, etc., so that the active pharmaceutical ingredient or pharmaceutical composition comes into contact with the individual or its cells, tissues, organs, biological fluids, etc. Common modes of administration include, but are not limited to, oral, subcutaneous, intramuscular, subperitoneal, ocular, nasal, sublingual, rectal, vaginal, etc.

[0175] The term "in need thereof" refers to a physician or other caregiver's determination that an individual needs or will benefit from a preventative and / or therapeutic procedure based on a variety of factors within the physician's or other caregiver's own area of ​​expertise.

[0176] The term "individual" (or subject) refers to a human or a non-human animal. Individuals of the present invention include individuals (patients) with a disease and / or condition, as well as normal individuals. Non-human animals of the present invention include all vertebrates, e.g., non-mammals, such as birds, amphibians, reptiles, etc., and mammals, e.g., non-human primates, domestic and / or farm animals (e.g., sheep, dogs, cats, cows, pigs, etc.).

[0177] Preparation method A fourth aspect of the present invention provides a method for synthesizing a compound.

[0178] The compounds of formula (I) of the present invention can be synthesized by the following synthetic route.

[0179] [ka] (In the formula, R x , R y and R zhas the meaning as defined above; LG is a leaving group selected from the group consisting of methylsulfonyl, trifluoromethylsulfonyloxy and halogen, preferably trifluoromethylsulfonyloxy or iodine).

[0180] Step 1: The compound of formula (I)-IM1 can be obtained by a substitution reaction between the compound of formula (I)-SM1 and the compound of formula (I)-SM2.

[0181] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 50°C.

[0182] In some embodiments, this step is carried out in a suitable organic solvent, preferably acetonitrile, selected from the group consisting of halogenated hydrocarbons (e.g., dichloromethane (DCM), chloroform (TCM), 1,2-dichloroethane (1,2-DCE), etc.), nitriles (e.g., acetonitrile (AN)), N-methylpyrrolidone (NMP), N,N-dimethylformamide (DMF), N,N-dimethylacetamide (DMA), tetrahydrofuran (THF), 1,4-dioxane (dioxane), dimethyl sulfoxide (DMSO), and any combination thereof.

[0183] In some embodiments, this step is carried out in the presence of a suitable base, wherein the base comprises an organic base or an inorganic base; the organic base may be selected from the group consisting of N,N-diisopropylethylamine (DIPEA), triethylamine (TEA), potassium tert-butoxide (t-BuOK), and pyridine (Py); and the inorganic base may be selected from the group consisting of potassium phosphate (KPO), sodium hydride (NaH), potassium carbonate (KCO), sodium carbonate (NaCO), sodium bicarbonate (NaHCO), cesium carbonate (CsCO), and NaOH, preferably NaCO or NaHCO.

[0184] Step 2: The compound of formula (I) can be obtained by a condensation reaction between a compound of formula (I)-IM1 and a compound of formula (I)-SM3.

[0185] In some embodiments, this step is carried out in the presence of a suitable condensing reagent selected from the group consisting of HATU, HBTU, EDCI, DCC and HOBT, preferably HATU.

[0186] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 25°C.

[0187] In some embodiments, this step is carried out in a suitable organic solvent selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably N,N-dimethylformamide.

[0188] In some embodiments, this step is carried out in the presence of a suitable base, which comprises an organic base or an inorganic base, where the organic base may be selected from the group consisting of DIPEA, TEA, t-BuOK and Py, and the inorganic base may be selected from the group consisting of K3PO4, NaH, K2CO3, Na2CO3, Cs2CO3 and NaOH, preferably DIPEA.

[0189] The compounds of formula (II)-1 of the present invention may be synthesized via the following synthetic routes.

[0190] [ka] (In the formula, R x‘ , R y’ and R z‘has the meaning as defined above; LG is a leaving group selected from the group consisting of methylsulfonyl, trifluoromethylsulfonyloxy and halogen, preferably trifluoromethylsulfonyloxy or iodine; PG is [ka] (wherein the protecting group is selected from the group consisting of:

[0191] Step 1 The compound of formula (II)-IM1 can be obtained by a substitution reaction of the compound of formula (II)-SM1.

[0192] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 50°C, 60°C, 100°C, preferably 50°C.

[0193] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably n-heptane.

[0194] Step 2 The compound of formula (II)-IM2 can be obtained by reduction of the compound of formula (II)-IM1.

[0195] In some embodiments, this step is carried out in the presence of a suitable reducing agent, which may be selected from the group consisting of a palladium catalyst, a platinum catalyst, a rhodium catalyst, preferably a platinum catalyst.

[0196] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 50°C, 60°C, 100°C, preferably 60°C.

[0197] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably ethyl acetate.

[0198] Step 3 The compound of formula (II)-IM3 can be obtained by a substitution reaction of the compound of formula (II)-IM2.

[0199] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 50°C, 60°C, 100°C, preferably 20°C.

[0200] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably ethyl acetate.

[0201] In some embodiments, this step is carried out under basic conditions, and reagents that provide basic conditions include organic and inorganic bases, including but not limited to triethylamine, pyridine, N,N-diisopropylethylamine, n-butyllithium, lithium diisopropylamide, lithium bistrimethylsilylamide, and sodium bistrimethylsilylamide; and inorganic bases, including but not limited to potassium carbonate, sodium carbonate, sodium bicarbonate, potassium tert-butoxide, sodium hydride, sodium hydroxide, and potassium hydroxide, preferably triethylamine.

[0202] Step 4 The compound of formula (II)-IM4 can be obtained by coupling reaction of the compound of formula (II)-IM3.

[0203] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 50°C, 60°C, 70°C, 100°C, preferably 70°C.

[0204] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, water, preferably a mixture of tetrahydrofuran and water.

[0205] In some embodiments, this step is carried out under basic conditions, and reagents that provide basic conditions include organic and inorganic bases, including but not limited to triethylamine, pyridine, N,N-diisopropylethylamine, n-butyllithium, lithium diisopropylamide, lithium bistrimethylsilylamide, and sodium bistrimethylsilylamide; and inorganic bases, including but not limited to potassium carbonate, sodium carbonate, sodium bicarbonate, potassium tert-butoxide, sodium hydride, sodium hydroxide, and potassium hydroxide, preferably N,N-diisopropylethylamine.

[0206] Step 5 The compound of formula (II)-IM5 can be obtained by reduction of the compound of formula (II)-IM4.

[0207] In some embodiments, this step is carried out in the presence of a suitable reducing agent, which may be selected from the group consisting of a palladium catalyst, a platinum catalyst, a rhodium catalyst, preferably a platinum catalyst.

[0208] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 40°C.

[0209] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably tetrahydrofuran.

[0210] Step 6 The compound of formula (II)-IM6 can be obtained by ring closure reaction of the compound of formula (II)-IM5.

[0211] In some embodiments, this step is carried out at a suitable temperature, which may be 5°C, 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 5°C.

[0212] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of trifluoroacetic acid, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, tert-butanol, preferably a mixed solvent of tetrahydrofuran and tert-butanol.

[0213] Step 7 The compound of formula (II)-IM7 can be obtained by a substitution reaction of the compound of formula (II)-IM6.

[0214] In some embodiments, this step is carried out at a suitable temperature, which may be 5°C, 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 5°C.

[0215] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of trifluoroacetic acid, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably trifluoroacetic acid.

[0216] In some embodiments, this step is carried out under basic conditions, and reagents that provide basic conditions include organic and inorganic bases, where organic bases include, but are not limited to, triethylamine, pyridine, N,N-diisopropylethylamine, n-butyllithium, lithium diisopropylamide, lithium bistrimethylsilylamide, and sodium bistrimethylsilylamide; and inorganic bases include, but are not limited to, potassium carbonate, sodium carbonate, sodium bicarbonate, potassium tert-butoxide, sodium hydride, sodium hydroxide, and potassium hydroxide, preferably potassium tert-butoxide.

[0217] Step 8 The compound of formula (II)-IM8 can be obtained by reduction of the compound of formula (II)-IM7.

[0218] In some embodiments, this step is carried out in the presence of a suitable reducing agent, which may be selected from the group consisting of a palladium catalyst, a platinum catalyst, a rhodium catalyst, preferably a palladium catalyst.

[0219] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 20°C.

[0220] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably methanol.

[0221] Step 9 The compound of formula (II)-IM9 can be obtained by a substitution reaction of the compound of formula (II)-IM8.

[0222] In some embodiments, this step is carried out at a suitable temperature, which may be 5°C, 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 20°C.

[0223] In some embodiments, this step is carried out under basic conditions, and reagents that provide basic conditions include organic and inorganic bases, including but not limited to triethylamine, pyridine, N,N-diisopropylethylamine, n-butyllithium, lithium diisopropylamide, lithium bistrimethylsilylamide, and sodium bistrimethylsilylamide; and inorganic bases, including but not limited to potassium carbonate, sodium carbonate, sodium bicarbonate, potassium tert-butoxide, sodium hydride, sodium hydroxide, and potassium hydroxide, preferably pyridine.

[0224] Step 10 The compound of formula (II)-IM10 can be obtained by hydrolysis of the compound of formula (II)-IM9.

[0225] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 60°C.

[0226] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably methanol.

[0227] In some embodiments, the reaction is carried out under acidic conditions, and reagents that provide acidic conditions include hydrochloric acid, trifluoroacetic acid, formic acid, sulfuric acid, methanesulfonic acid, preferably hydrochloric acid.

[0228] Step 11 The compound of formula (II)-IM11 can be obtained by ring closure reaction of the compound of formula (II)-IM10 with (S)-4-ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-F]indolizine-3,6,10(4H)-trione.

[0229] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 140°C.

[0230] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of toluene, methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably toluene.

[0231] In some embodiments, the reaction is carried out under acidic conditions, and reagents that provide acidic conditions include p-toluenesulfonic acid, hydrochloric acid, trifluoroacetic acid, formic acid, sulfuric acid, methanesulfonic acid, preferably p-toluenesulfonic acid.

[0232] Step 12 The compound of formula (II)-IM12 can be obtained by hydrolysis of the compound of formula (II)-IM11.

[0233] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 100°C.

[0234] In some embodiments, the reaction is carried out under acidic conditions, and reagents that provide acidic conditions include p-toluenesulfonic acid, hydrochloric acid, trifluoroacetic acid, formic acid, sulfuric acid, methanesulfonic acid, preferably hydrochloric acid.

[0235] Step 13 The compound of formula (II)-IM13 can be obtained by a substitution reaction between the compound of formula (II)-IM12 and the compound of formula (II)-SM2.

[0236] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 50°C.

[0237] In some embodiments, this step is carried out in a suitable organic solvent, preferably acetonitrile, selected from the group consisting of halogenated hydrocarbons (e.g., dichloromethane (DCM), chloroform (TCM), 1,2-dichloroethane (1,2-DCE), etc.), nitriles (e.g., acetonitrile (AN)), N-methylpyrrolidone (NMP), N,N-dimethylformamide (DMF), N,N-dimethylacetamide (DMA), tetrahydrofuran (THF), 1,4-dioxane (Diox), dimethyl sulfoxide (DMSO), and any combination thereof.

[0238] In some embodiments, this step is carried out in the presence of a suitable base, which comprises an organic base or an inorganic base; the organic base may be selected from the group consisting of N,N-diisopropylethylamine (DIPEA), triethylamine (TEA), potassium tert-butoxide (t-BuOK) and pyridine (Py); the inorganic base may be selected from the group consisting of potassium phosphate (KPO), sodium hydride (NaH), potassium carbonate (KCO), sodium carbonate (NaCO), sodium bicarbonate (NaHCO), cesium carbonate (CsCO) and NaOH, preferably NaCO or NaHCO.

[0239] Step 14 The compound of formula (II)-IM14 can be obtained by condensation reaction of the compound of formula (II)-IM13 with the compound of formula (II)-SM3.

[0240] In some embodiments, this step is carried out in the presence of a suitable condensing reagent selected from the group consisting of HATU, HBTU, EDCI, DCC and HOBT, preferably HATU.

[0241] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 25°C.

[0242] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethylsulfoxide, n-heptane, n-hexane, ethyl acetate, preferably N,N-dimethylformamide.

[0243] In some embodiments, this step is carried out in the presence of a suitable base, including organic and inorganic bases, where the organic base may be selected from the group consisting of DIPEA, TEA, t-BuOK, and Py, and the inorganic base may be selected from the group consisting of KPO, NaH, KCO, NaCO, CsCO, and NaOH, preferably DIPEA.

[0244] Step 15 The compound of formula (II)-1 can be obtained by acid hydrolysis of the compound of formula (II)-IM14.

[0245] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 25°C.

[0246] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, and mixtures thereof, preferably a mixture of dichloromethane and methanol (volume ratio 2:1).

[0247] In some embodiments, the reaction is carried out under acidic conditions, and reagents that provide acidic conditions include p-toluenesulfonic acid, hydrochloric acid, trifluoroacetic acid, formic acid, sulfuric acid, methanesulfonic acid, preferably hydrochloric acid.

[0248] The synthesis method of formula (II)-SM3 is as follows: PG, [ka] If: [ka]

[0249] Step 1 The compound of formula (II)-SM3-3 can be obtained by a substitution reaction between the compound of formula (II)-SM3-1 and the compound of formula (II)-SM3-2.

[0250] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 0-25°C.

[0251] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, and mixtures thereof, preferably tetrahydrofuran.

[0252] In some embodiments, this step is carried out in the presence of a suitable base, including organic and inorganic bases, where the organic base may be selected from the group consisting of DIPEA, TEA, t-BuOK, and Py, and the inorganic base may be selected from the group consisting of K3PO4, NaH, K2CO3, Na2CO3, Cs2CO3, and NaOH, preferably K2CO3.

[0253] Step 2 The compound of formula (II)-SM3 can be obtained by hydrogenation of the compound of formula (II)-SM3-3.

[0254] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 25°C.

[0255] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, and mixtures thereof, preferably methanol.

[0256] In some embodiments, this step is carried out in the presence of a suitable reducing agent, which may be selected from the group consisting of a palladium catalyst, a platinum catalyst, a rhodium catalyst, preferably a palladium catalyst.

[0257] PG, [ka] and R y’ and R z’ If is hydrogen: [ka]

[0258] Step 1 The compound of formula (II)-SM3 can be obtained by a condensation reaction between the compound of formula (II)-SM3-4 and the compound of formula (II)-SM3-5.

[0259] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 25°C.

[0260] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, and mixtures thereof, preferably N,N-dimethylformamide.

[0261] Alternatively, compounds of formula (II)-1 can be synthesized by the following synthetic route: [ka]

[0262] Step 1 The compound of formula (II)-IM15 can be obtained by condensation reaction of the compound of formula (II)-IM13 with the compound of formula (II)-SM4.

[0263] In some embodiments, this step is carried out in the presence of a suitable condensing reagent selected from the group consisting of HATU, HBTU, EDCI, DCC and HOBT, preferably HATU.

[0264] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 25°C.

[0265] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethylsulfoxide, n-heptane, n-hexane, ethyl acetate, preferably N,N-dimethylformamide.

[0266] In some embodiments, this step is carried out in the presence of a suitable base, which comprises an organic base or an inorganic base, where the organic base may be selected from the group consisting of DIPEA, TEA, t-BuOK and Py, and the inorganic base may be selected from the group consisting of K3PO4, NaH, K2CO3, Na2CO3, Cs2CO3 and NaOH, preferably DIPEA.

[0267] Step 2 The compound of formula (II)-1 can be obtained by removing the silicon protecting group of the compound of formula (II)-IM15.

[0268] Alternatively, compounds of formula (II) may be synthesized by the following synthetic route: [ka]

[0269] Step 1 The compound of formula (II) can be obtained by a condensation reaction between a compound of formula (II)-IM13 and a compound of formula (II)-SM5.

[0270] In some embodiments, this step is carried out in the presence of a suitable condensing reagent selected from the group consisting of HATU, HBTU, EDCI, DCC and HOBT, preferably HATU.

[0271] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 25°C.

[0272] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethylsulfoxide, n-heptane, n-hexane, ethyl acetate, preferably N,N-dimethylformamide.

[0273] In some embodiments, this step is carried out in the presence of a suitable base, which comprises an organic base or an inorganic base, where the organic base may be selected from the group consisting of DIPEA, TEA, t-BuOK and Py, and the inorganic base may be selected from the group consisting of K3PO4, NaH, K2CO3, Na2CO3, Cs2CO3 and NaOH, preferably DIPEA.

[0274] The compound of formula (III)-1 of the present invention can be synthesized by the following synthetic route.

[0275] [ka] (In the formula, R x’’ , R y’’ and R z’’ is as defined above; LG is a leaving group selected from the group consisting of methylsulfonyl, trifluoromethylsulfonyloxy and halogen, preferably trifluoromethylsulfonyl or chlorine; PG is [ka] (wherein the protecting group is selected from the group consisting of:

[0276] Step 1 The compound of formula (III)-IM1 can be obtained by a substitution reaction of the compound of formula (III)-SM1.

[0277] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 50°C, 60°C, 100°C, preferably 50°C.

[0278] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably n-heptane.

[0279] Step 2 The compound of formula (III)-IM2 can be obtained by reduction of the compound of formula (III)-IM1.

[0280] In some embodiments, this step is carried out in the presence of a suitable reducing agent, which may be selected from the group consisting of a palladium catalyst, a platinum catalyst, a rhodium catalyst, preferably a platinum catalyst.

[0281] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 50°C, 60°C, 100°C, preferably 60°C.

[0282] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably ethyl acetate.

[0283] Step 3 The compound of formula (III)-IM3 can be obtained by a substitution reaction of the compound of formula (III)-IM2.

[0284] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 50°C, 60°C, 100°C, preferably 20°C.

[0285] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably ethyl acetate.

[0286] In some embodiments, this step is carried out under basic conditions, and reagents that provide basic conditions include organic and inorganic bases, including but not limited to triethylamine, pyridine, N,N-diisopropylethylamine, n-butyllithium, lithium diisopropylamide, lithium bistrimethylsilylamide, and sodium bistrimethylsilylamide; and inorganic bases, including but not limited to potassium carbonate, sodium carbonate, sodium bicarbonate, potassium tert-butoxide, sodium hydride, sodium hydroxide, and potassium hydroxide, preferably triethylamine.

[0287] Step 4 The compound of formula (III)-IM4 can be obtained by coupling reaction of the compound of formula (III)-IM3.

[0288] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 50°C, 60°C, 70°C, 100°C, preferably 70°C.

[0289] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, water, preferably a mixture of tetrahydrofuran and water.

[0290] In some embodiments, this step is carried out under basic conditions, and reagents that provide basic conditions include organic and inorganic bases, including but not limited to triethylamine, pyridine, N,N-diisopropylethylamine, n-butyllithium, lithium diisopropylamide, lithium bistrimethylsilylamide, and sodium bistrimethylsilylamide; and inorganic bases, including but not limited to potassium carbonate, sodium carbonate, sodium bicarbonate, potassium tert-butoxide, sodium hydride, sodium hydroxide, and potassium hydroxide, preferably N,N-diisopropylethylamine.

[0291] Step 5 The compound of formula (III)-IM5 can be obtained by reduction of the compound of formula (III)-IM4.

[0292] In some embodiments, this step is carried out in the presence of a suitable reducing agent, which may be selected from the group consisting of a palladium catalyst, a platinum catalyst, a rhodium catalyst, preferably a platinum catalyst.

[0293] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 40°C.

[0294] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably tetrahydrofuran.

[0295] Step 6 The compound of formula (III)-IM6 can be obtained by ring closure reaction of the compound of formula (III)-IM5.

[0296] In some embodiments, this step is carried out at a suitable temperature, which may be 5°C, 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 5°C.

[0297] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of trifluoroacetic acid, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, tert-butanol, preferably a mixed solvent of tetrahydrofuran and tert-butanol.

[0298] Step 7 The compound of formula (III)-IM7 can be obtained by a substitution reaction of the compound of formula (III)-IM6.

[0299] In some embodiments, this step is carried out at a suitable temperature, which may be 5°C, 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 5°C.

[0300] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of trifluoroacetic acid, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably trifluoroacetic acid.

[0301] In some embodiments, this step is carried out under basic conditions, and reagents that provide basic conditions include organic and inorganic bases, where organic bases include, but are not limited to, triethylamine, pyridine, N,N-diisopropylethylamine, n-butyllithium, lithium diisopropylamide, lithium bistrimethylsilylamide, and sodium bistrimethylsilylamide; and inorganic bases include, but are not limited to, potassium carbonate, sodium carbonate, sodium bicarbonate, potassium tert-butoxide, sodium hydride, sodium hydroxide, and potassium hydroxide, preferably potassium tert-butoxide.

[0302] Step 8 The compound of formula (III)-IM8 can be obtained by reduction of the compound of formula (III)-IM7.

[0303] In some embodiments, this step is carried out in the presence of a suitable reducing agent, which may be selected from the group consisting of a palladium catalyst, a platinum catalyst, a rhodium catalyst, preferably a palladium catalyst.

[0304] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 20°C.

[0305] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably methanol.

[0306] Step 9 The compound of formula (III)-IM9 can be obtained by a substitution reaction of the compound of formula (III)-IM8.

[0307] In some embodiments, this step is carried out at a suitable temperature, which may be 5°C, 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 20°C.

[0308] In some embodiments, this step is carried out under basic conditions, and reagents that provide basic conditions include organic and inorganic bases, including but not limited to triethylamine, pyridine, N,N-diisopropylethylamine, n-butyllithium, lithium diisopropylamide, lithium bistrimethylsilylamide, and sodium bistrimethylsilylamide; and inorganic bases, including but not limited to potassium carbonate, sodium carbonate, sodium bicarbonate, potassium tert-butoxide, sodium hydride, sodium hydroxide, and potassium hydroxide, preferably pyridine.

[0309] Step 10 The compound of formula (III)-IM10 can be obtained by hydrolysis of the compound of formula (III)-IM9.

[0310] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 60°C.

[0311] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably methanol.

[0312] In some embodiments, the reaction is carried out under acidic conditions, and reagents that provide acidic conditions include hydrochloric acid, trifluoroacetic acid, formic acid, sulfuric acid, methanesulfonic acid, preferably hydrochloric acid.

[0313] Step 11 The compound of formula (III)-IM11 can be obtained by ring closure reaction of the compound of formula (III)-IM10 with (S)-4-ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-F]indolizine-3,6,10(4H)-trione.

[0314] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 140°C.

[0315] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of toluene, methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably toluene.

[0316] In some embodiments, the reaction is carried out under acidic conditions, and reagents that provide acidic conditions include p-toluenesulfonic acid, hydrochloric acid, trifluoroacetic acid, formic acid, sulfuric acid, methanesulfonic acid, preferably p-toluenesulfonic acid.

[0317] Step 12 The compound of formula (III)-IM12 can be obtained by hydrolysis of the compound of formula (III)-IM11.

[0318] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 100°C.

[0319] In some embodiments, the reaction is carried out under acidic conditions, and reagents that provide acidic conditions include p-toluenesulfonic acid, hydrochloric acid, trifluoroacetic acid, formic acid, sulfuric acid, methanesulfonic acid, preferably hydrochloric acid.

[0320] Step 13 The compound of formula (III)-IM13 can be obtained by a substitution reaction between the compound of formula (III)-IM12 and the compound of formula (III)-SM2.

[0321] A substitution reaction is carried out to give a compound of formula (II)-IM13.

[0322] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 50°C.

[0323] In some embodiments, this step is carried out in a suitable organic solvent, preferably acetonitrile, selected from the group consisting of halogenated hydrocarbons (e.g., dichloromethane (DCM), chloroform (TCM), 1,2-dichloroethane (1,2-DCE), etc.), nitriles (e.g., acetonitrile (AN)), N-methylpyrrolidone (NMP), N,N-dimethylformamide (DMF), N,N-dimethylacetamide (DMA), tetrahydrofuran (THF), 1,4-dioxane (dioxane), dimethyl sulfoxide (DMSO), and any combination thereof.

[0324] In some embodiments, this step is carried out in the presence of a suitable base, which comprises an organic base or an inorganic base; the organic base may be selected from the group consisting of N,N-diisopropylethylamine (DIPEA), triethylamine (TEA), potassium tert-butoxide (t-BuOK) and pyridine (Py); the inorganic base may be selected from the group consisting of potassium phosphate (KPO), sodium hydride (NaH), potassium carbonate (KCO), sodium carbonate (NaCO), sodium bicarbonate (NaHCO), cesium carbonate (CsCO) and NaOH, preferably NaCO or NaHCO.

[0325] Step 14 The compound of formula (III)-IM14 can be obtained by a condensation reaction of the compound of formula (III)-IM13 and the compound of formula (III)-SM3.

[0326] In some embodiments, this step is carried out in the presence of a suitable condensing reagent selected from the group consisting of HATU, HBTU, EDCI, DCC and HOBT, preferably HATU.

[0327] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 25°C.

[0328] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethylsulfoxide, n-heptane, n-hexane, ethyl acetate, preferably N,N-dimethylformamide.

[0329] In some embodiments, this step is carried out in the presence of a suitable base, which comprises an organic base or an inorganic base, where the organic base may be selected from the group consisting of DIPEA, TEA, t-BuOK and Py, and the inorganic base may be selected from the group consisting of K3PO4, NaH, K2CO3, Na2CO3, Cs2CO3 and NaOH, preferably DIPEA.

[0330] Step 15 The compound of formula (III)-1 can be obtained by acid hydrolysis of the compound of formula (III)-IM14.

[0331] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 25°C.

[0332] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, and mixtures thereof, preferably a mixture of dichloromethane and methanol (volume ratio 2:1).

[0333] In some embodiments, the reaction is carried out under acidic conditions, and reagents that provide acidic conditions include p-toluenesulfonic acid, hydrochloric acid, trifluoroacetic acid, formic acid, sulfuric acid, methanesulfonic acid, preferably hydrochloric acid.

[0334] The synthesis method of formula (III)-SM3 is as follows: PG, [ka] If: [ka]

[0335] Step 1: The compound of formula (III)-SM3-3 can be obtained by a substitution reaction between the compound of formula (III)-SM3-1 and the compound of formula (II)-SM3-2.

[0336] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 0-25°C.

[0337] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, and mixtures thereof, preferably tetrahydrofuran.

[0338] In some embodiments, this step is carried out in the presence of a suitable base, which comprises an organic base or an inorganic base, where the organic base may be selected from the group consisting of DIPEA, TEA, t-BuOK, and Py, and the inorganic base may be selected from the group consisting of K3PO4, NaH, K2CO3, Na2CO3, Cs2CO3, and NaOH, preferably K2CO3.

[0339] Step 2: The compound of formula (III)-SM3 can be obtained by hydrogenation of the compound of formula (III)-SM3-3.

[0340] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 25°C.

[0341] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, and mixtures thereof, preferably methanol.

[0342] In some embodiments, this step is carried out in the presence of a suitable reducing agent, which may be selected from the group consisting of a palladium catalyst, a platinum catalyst, a rhodium catalyst, preferably a palladium catalyst.

[0343] PG, [ka] If R y’ and R z’ is hydrogen: [ka]

[0344] Step 1: The compound of formula (III)-SM3 can be obtained by a condensation reaction between the compound of formula (III)-SM3-4 and the compound of formula (III)-SM3-5.

[0345] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 25°C.

[0346] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, and mixtures thereof, preferably N,N-dimethylformamide.

[0347] Alternatively, compounds of formula (III)-1 may be synthesized by the following synthetic route: [ka]

[0348] Step 1: The compound of formula (III)-IM15 can be obtained by a condensation reaction between the compound of formula (III)-IM13 and the compound of formula (III)-SM4.

[0349] In some embodiments, this step is carried out in the presence of a suitable condensing reagent selected from the group consisting of HATU, HBTU, EDCI, DCC and HOBT, preferably HATU.

[0350] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 25°C.

[0351] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethylsulfoxide, n-heptane, n-hexane, ethyl acetate, preferably N,N-dimethylformamide.

[0352] In some embodiments, this step is carried out in the presence of a suitable base, which comprises an organic base or an inorganic base, where the organic base may be selected from the group consisting of DIPEA, TEA, t-BuOK and Py, and the inorganic base may be selected from the group consisting of K3PO4, NaH, K2CO3, Na2CO3, Cs2CO3 and NaOH, preferably DIPEA.

[0353] Step 2: The compound of formula (III)-1 can be obtained by removing the silicon protecting group of the compound of formula (III)-IM15.

[0354] Alternatively, compounds of formula (III)-1 can be synthesized by the following synthetic route: [ka]

[0355] Step 1: The compound of formula (III)-1 can be obtained by a condensation reaction of the compound of formula (III)-IM13 and the compound of formula (III)-SM5.

[0356] In some embodiments, this step is carried out in the presence of a suitable condensing reagent selected from the group consisting of HATU, HBTU, EDCI, DCC and HOBT, preferably HATU.

[0357] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 25°C.

[0358] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethylsulfoxide, n-heptane, n-hexane, ethyl acetate, preferably N,N-dimethylformamide.

[0359] In some embodiments, this step is carried out in the presence of a suitable base, which comprises an organic base or an inorganic base, where the organic base may be selected from the group consisting of DIPEA, TEA, t-BuOK and Py, and the inorganic base may be selected from the group consisting of K3PO4, NaH, K2CO3, Na2CO3, Cs2CO3 and NaOH, preferably DIPEA.

[0360] The compounds of formula (IV) of the present invention can be synthesized and prepared by the following synthetic routes: (In the formula, R a , R b , R c , R d and R e The meaning of is as above) When q=1, [ka]

[0361] Step 1 The compound of formula (IV)-IM1 can be obtained by nitration of the compound of formula (IV)-SM1.

[0362] In some embodiments, this step is carried out at a suitable temperature, which is 5°C, 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 25°C.

[0363] Step 2 The compound of formula (IV)-IM2 can be obtained by hydrogenation of the compound of formula (IV)-IM1.

[0364] In some embodiments, this step is carried out in the presence of a suitable reducing agent, which may be selected from the group consisting of a palladium catalyst, a platinum catalyst, a rhodium catalyst, preferably a palladium catalyst.

[0365] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 25°C.

[0366] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably ethyl acetate.

[0367] Step 3 The compound of formula (IV)-IM3 can be obtained by acylation of the compound of formula (IV)-IM2.

[0368] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 50°C, 60°C, 100°C, preferably 25°C.

[0369] Step 4 The compound of formula (IV)-IM4 can be obtained by reacting the compound of formula (IV)-IM3 with DMF-DMA.

[0370] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 120°C, preferably 120°C.

[0371] Step 5 The compound of formula (IV)-IM5 can be obtained by a substitution reaction between the compound of formula (IV)-IM4 and the compound of formula (IV)-SM2.

[0372] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 50°C.

[0373] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, ethanol, N-methylpyrrolidone, dimethyl sulfoxide, preferably ethanol.

[0374] Step 6 The compound of formula (IV)-IM6 can be obtained by reduction of the compound of formula (IV)-IM5.

[0375] In some embodiments, this step is carried out in the presence of a suitable reducing agent, which is preferably sodium borohydride.

[0376] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 0-25°C.

[0377] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of tetrahydrofuran, glacial acetic acid, methanol, and mixtures thereof, preferably glacial acetic acid.

[0378] Step 7 The compound of formula (IV)-IM7 can be obtained by protecting the amino group of the compound of formula (IV)-IM6 with Fmoc.

[0379] Step 8 The compound of formula (IV)-IM8 can be obtained by removing the acetyl protecting group of the amino group of the compound of formula (IV)-IM7.

[0380] Step 9 The compound of formula (IV)-IM9 can be obtained by ring closure reaction of the compound of formula (IV)-IM8 under acidic conditions.

[0381] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 120°C, preferably 120°C.

[0382] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of toluene, xylene, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, preferably toluene and xylene.

[0383] In some embodiments, this step is carried out under acidic conditions, and reagents that provide acidic conditions include p-toluenesulfonic acid, hydrochloric acid, trifluoroacetic acid, formic acid, sulfuric acid, methanesulfonic acid, preferably p-toluenesulfonic acid.

[0384] Step 10 The compound of formula (IV)-IM10 can be obtained by removing the Fmoc protecting group of the compound of formula (IV)-IM9.

[0385] Step 11 The compound of formula (IV) can be obtained by a condensation reaction between the compound of formula (IV)-IM10 and the compound of formula (IV)-SM4.

[0386] In some embodiments, this step is carried out in the presence of a suitable condensing reagent selected from the group consisting of HATU, HBTU, EDCI, DCC and HOBT, preferably HBTU.

[0387] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 25°C.

[0388] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethylsulfoxide, n-heptane, n-hexane, ethyl acetate, preferably N,N-dimethylformamide.

[0389] In some embodiments, this step is carried out in the presence of a suitable base, which comprises an organic base or an inorganic base, where the organic base may be selected from the group consisting of DIPEA, TEA, t-BuOK and Py, and the inorganic base may be selected from the group consisting of K3PO4, NaH, K2CO3, Na2CO3, Cs2CO3 and NaOH, preferably DIPEA.

[0390] Or, if q=0, [ka]

[0391] LG is a leaving group selected from the group consisting of methylsulfonyl, trifluoromethylsulfonyloxy and halogen, preferably trifluoromethylsulfonyloxy or iodine.

[0392] Step 1 The compound of formula (IV)-IM11 can be obtained by reduction of the compound of formula (IV)-SM5.

[0393] In some embodiments, this step is carried out in the presence of a suitable reducing agent, which may be selected from the group consisting of a palladium catalyst, a platinum catalyst, a rhodium catalyst, preferably a platinum catalyst.

[0394] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 50°C, 60°C, 100°C, preferably 25°C.

[0395] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, preferably ethyl acetate and tetrahydrofuran.

[0396] Step 2 The compound of formula (IV)-IM12 can be obtained by Friedel-Crafts acylation of the compound of formula (IV)-IM11.

[0397] In some embodiments, this step is carried out at a suitable temperature, which is 5°C, 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 25°C.

[0398] Step 3 The compound of formula (IV)-IM13 can be obtained by ring closure reaction of formula (IV)-IM12 under acidic conditions.

[0399] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 120°C, preferably 120°C.

[0400] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of toluene, xylene, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, preferably toluene and xylene.

[0401] In some embodiments, this step is carried out under acidic conditions, and reagents that provide acidic conditions include p-toluenesulfonic acid, hydrochloric acid, trifluoroacetic acid, formic acid, sulfuric acid, methanesulfonic acid, preferably p-toluenesulfonic acid.

[0402] Step 4 The compound of formula (IV)-IM14 can be obtained by a substitution reaction of formula (IV)-IM13.

[0403] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 25°C.

[0404] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, ethanol, N,N-methylpyrrolidone, dimethyl sulfoxide, preferably dimethyl sulfoxide.

[0405] Step 5 The compound of formula (IV)-IM15 can be obtained by reduction of the compound of formula (IV)-IM14.

[0406] In some embodiments, this step is carried out in the presence of a suitable reducing agent selected from the group consisting of a palladium catalyst, a platinum catalyst, a rhodium catalyst, triphenylphosphine, triethylphosphite, preferably triethylphosphite.

[0407] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 50°C, 60°C, 80°C, 100°C, preferably 80°C.

[0408] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, n-heptane, n-hexane, ethyl acetate, toluene, and mixtures thereof, preferably a mixture of methanol and toluene.

[0409] Step 6 The compound of formula (IV)-IM16 can be obtained by a substitution reaction between the compound of formula (IV)-IM15 and the compound of formula (IV)-SM6.

[0410] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, 140°C, preferably 50°C.

[0411] In some embodiments, this step is carried out in a suitable organic solvent, preferably acetonitrile, selected from the group consisting of halogenated hydrocarbons (e.g., dichloromethane (DCM), chloroform (TCM), 1,2-dichloroethane (1,2-DCE), etc.), nitriles (e.g., acetonitrile (AN)), N-methylpyrrolidone (NMP), N,N-dimethylformamide (DMF), N,N-dimethylacetamide (DMA), tetrahydrofuran (THF), 1,4-dioxane (dioxane), dimethyl sulfoxide (DMSO), and any combination thereof.

[0412] In some embodiments, this step is carried out in the presence of a suitable base, which comprises an organic base or an inorganic base; the organic base may be selected from the group consisting of N,N-diisopropylethylamine (DIPEA), triethylamine (TEA), potassium tert-butoxide (t-BuOK) and pyridine (Py); the inorganic base may be selected from the group consisting of potassium phosphate (KPO), sodium hydride (NaH), potassium carbonate (KCO), sodium carbonate (NaCO), sodium bicarbonate (NaHCO), cesium carbonate (CsCO) and NaOH, preferably NaCO or NaHCO.

[0413] Step 7 The compound of formula (IV) can be obtained by a condensation reaction between a compound of formula (IV)-IM16 and a compound of formula (IV)-SM4.

[0414] In some embodiments, this step is carried out in the presence of a suitable condensing reagent, which may be selected from the group consisting of HATU, HBTU, EDCI, DCC and HOBT, preferably HBTU.

[0415] In some embodiments, this step is carried out at a suitable temperature, which may be 20°C, 25°C, 40°C, 50°C, 60°C, 100°C, preferably 25°C.

[0416] In some embodiments, this step is carried out in a suitable organic solvent, which may be selected from the group consisting of methanol, tetrahydrofuran, dichloromethane, N,N-dimethylformamide, N-methylpyrrolidone, dimethylsulfoxide, n-heptane, n-hexane, ethyl acetate, preferably N,N-dimethylformamide.

[0417] In some embodiments, this step is carried out in the presence of a suitable base, which comprises an organic base or an inorganic base, where the organic base may be selected from the group consisting of DIPEA, TEA, t-BuOK and Py, and the inorganic base may be selected from the group consisting of K3PO4, NaH, K2CO3, Na2CO3, Cs2CO3 and NaOH, preferably DIPEA.

[0418] The compound of formula (V)-1 of the present invention can be prepared by the reaction of a starting material [ka] It may also be synthesized and prepared by a route similar to that of formula (III) using

[0419] Beneficial effects of the invention The present invention provides camptothecin compounds represented by Formulas (I) to (IV), as well as pharmaceutical compositions, preparation methods and applications thereof. Compounds of this type have good antitumor activity, have the potential to combat drug resistance, and can be used to treat diseases associated with abnormal cell proliferation, including, but not limited to, advanced solid tumors.

[0420] A specific model for carrying out the invention The present invention will be further illustrated by the description of specific embodiments, which are not intended to limit the present invention. Those skilled in the art can make various modifications or improvements based on the teachings of the present invention without departing from the basic concept and scope of the present invention.

[0421] The abbreviations used in the present invention have the following meanings.

[0422] [Table 1]

[0423] The structures of the compounds described in the following examples were determined by nuclear magnetic resonance ( 1 The results were confirmed by 1 H NMR or mass spectrometry (MS).

[0424] Nuclear magnetic resonance method ( 1 The instrument for 1 H NMR was a Bruker 400 MHz nuclear magnetic resonance spectrometer; hexadeuteriodimethylsulfoxide (DMSO-d6); the internal standard was tetramethylsilane (TMS).

[0425] The abbreviations used in the nuclear magnetic resonance (NMR) spectra of the examples are shown below.

[0426] s: singlet, d: doublet, t: triplet, q: quartet, m: multiplet, br: broadband, J: coupling constant, Hz: hertz, DMSO-d6: deuterated dimethyl sulfoxide. δ values ​​are expressed in ppm.

[0427] The equipment for mass spectrometer (MS) was an Agilent (ESI) mass spectrometer, Agilent 6120B.

[0428] Example 1: (R)—N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypent-3-ynamide and (S)—N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypent-3-ynamide [ka]

[0429] The compound 2-hydroxypent-3-ynoic acid (4.29 mg, 37.63 μmol) was dissolved in DMF (1 mL), and HATU (21.46 mg, 56.44 μmol), SM1-1 (10.00 mg, 22.94 μmol), and DIPEA (7.29 mg, 56.44 μmol) were added, followed by reaction for 2 hours at 25° C. The reaction solution was concentrated under reduced pressure, and the concentrate was directly purified by preparative high-performance liquid chromatography (conditions as follows) to obtain the title compounds 1-1-A (3.24 mg) and 1-1-B (3.98 mg).

[0430] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 2]

[0431] Retention time: 1-1-A: 10.8 minutes; 1-1-B: 11.1 minutes.

[0432] The structural property data of 1-1-A was as follows: 1HNMR (400 MHz, DMSO-d6) δ 8.58 (d, J = 8.4Hz, 1H), 7.78 (d, J = 11.2 Hz, 1H), 7.31 (s, 1H), 6.54 (s, 1H), 6.17 (d, J = 6.0Hz, 1H), 5.58-5.47 (m, 1H), 5.42 (s, 2H), 5.23 (s, 2H), 4.73-4.64 (m, 1H), 3.25-3.06 (m, 2H), 2.39 (s, 3H), 2.27-2.05 (m, 2H), 1.96-1.77 (m, 5H), 0.87 (t,J = 7.2Hz, 3H). ESI-MS (m / z): 532.2 [M+H] + . 1-1-B's structural characteristics are as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.62 (d, J = 8.8Hz, 1H), 7.78 (d, J = 10.8 Hz, 1H), 7.31 (s, 1H), 6.54 (s, 1H), 6.19 (d, J =6.0 Hz, 1H), 5.58-5.47 (m, 1H), 5.42 (s, 2H), 5.21 (d, J = 4.8 Hz, 2H), 4.73-4.65 (m, 1H), 3.27- 3.06 (m, 2H), 2.39 (s, 3H), 2.27-2.05 (m, 2H),1.93-1.80 (m, 2H), 1.80 (d, J = 2.0 Hz, 3H), 0.87 (t, J = 7.2Hz, 3H). ESI-MS (m / z): 532.2 [M+H] + .

[0433] Example 2: (S)—N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4]:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxy-3-enamide and (R)—N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4]:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxy-3-enamide [ka]

[0434] Compound ethylene glycolic acid (9.61 mg, 94.07 μmol) was dissolved in DMF (2 mL), and HATU (44.70 mg, 117.58 μmol), compound SM1-1 (25.00 mg, 0.047 mmol), and DIPEA (24.30 mg, 188.13 μmol) were added, followed by reaction for 2 hours at 25° C. The reaction solution was concentrated under reduced pressure, and the concentrate was directly purified by preparative high-performance liquid chromatography to give the title compounds 1-7-A (4.00 mg) and 1-7-B (1.38 mg).

[0435] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 3]

[0436] Retention time: 1-7-A: 8.7 minutes; 1-7-B: 9.1 minutes.

[0437] The structural property data of 1-7-A was as follows: 1HNMR (400 MHz, DMSO-d6) δ 8.51 (s, J = 8.8Hz, 1H), 7.76 (s, J = 10.8 Hz, 1H), 7.29 (s, 1H), 6.52 (s, 1H), 6.15 -6.04 (m,1H), 5.58-5.49 (m, 1H), 5.42 (s, 2H), 5.39 (s, 1H), 5.24-5.01 (m, 3H), 4.54 (s,J = 4.9 Hz, 1H), 3.24-3.05 (m, 2H), 2.37 (s, 3H), 2.16 (s, 2H), 1.91-1.79 (m,2H), 0.87 (t, J = 7.2 Hz, 3H). ESI-MS (m / z): 520.1 [M+H] + . 1-7-B's structural characteristics are as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.48 (s, 1H), 7.78(s, 1H), 7.30 (s, 1H), 6.12-5.92 (m, 1H), 5.54-5.47 (m, 1H), 5.42 (s, 2H), 5.37(dt, J = 1.7 Hz, 1H), 5.18 (s, 2H), 5.16-5.14 (m, 1H), 4.5-4.52 (m, 1H), 3.22-3.08 (m, 2H), 2.38 (s, 3H), 2.25-2.16 (m, 1H), 2.16-2.06 (m, 1H),1.93-1.79 (m, 2H), 0.87 (t, J = 7.2 Hz, 3H). ESI-MS (m / z): 520.1 [M+H] + .

[0438] Example 3: N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide and N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide [ka]

[0439] Step 1: Synthesis of 1-chloro-3-bromo-2-methyl-5-nitrobenzene Compound 2-1-01 (5.00 g, 29.14 mmol) was dissolved in n-heptane (25 mL) at 25 ° C., concentrated sulfuric acid (25 mL) was added, and the mixture was heated to 50 ° C., and NBS (6.22 g, 34.97 mmol) was added in batches at 50 ° C., and the mixture was reacted at 50 ° C. for 2 hours. The reaction was monitored by thin layer chromatography (ethyl acetate: petroleum ether = 1:10). The reaction solution was cooled to room temperature and added dropwise to ice water. Extraction was performed with toluene. The organic phases were combined, washed with sodium sulfite solution, water, and saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by preparative high performance liquid chromatography. The fraction was freeze-dried to obtain 4.88 g of the title compound.

[0440] Chromatography column: C18 ODS 45mm x 450mm x 8.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 4]

[0441] Step 2: Synthesis of 3-chloro-5-bromo-4-methylaniline At 25 ° C., compound 2-1-02 (4.88 g, 19.48 mmol) was dissolved in ethyl acetate (100 mL), and platinum on carbon (2.00 g, 19.48 mmol, 5% content) was added. The mixture was subjected to hydrogen substitution and reacted under the protection of a hydrogen balloon at 60 ° C. for 4 hours. The reaction was monitored by LCMS. The reaction solution was filtered, and the filtrate was concentrated to obtain 3.68 g of the crude title compound, which was used directly in the next reaction without further purification.

[0442] Step 3: Synthesis of N-(3-chloro-5-bromo-4-methylphenyl)acetamide Compound 2-1-03 (3.63 g, 14.82 mmol) was dissolved in ethyl acetate (70 mL) at 20 ° C., and triethylamine (4.50 g, 44.45 mmol) and acetic anhydride (2.27 g, 22.23 mmol) were added. The reaction was carried out at 20 ° C. for 20 hours, and the reaction was monitored by LCMS. Water was added to the reaction solution, and the mixture was extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. This was slurried in a mixed solvent of ethyl acetate:petroleum ether = 1:5 to obtain 2.86 g of the title compound.

[0443] Step 4: Synthesis of (Z)-4-(5-acetamido-3-chloro-2-methylphenyl)but-3-enoic acid At 20 ° C., compound 2-1-04 (1.80 g, 6.86 mmol) was dissolved in THF (20 mL) and water (5 mL), and vinylacetic acid (708.31 mg, 8.23 ​​mmol), DIPEA (1.95 g, 15.08 mmol), and tris(o-methylphenyl)phosphorus (62.60 mg, 0.20 mmol) were added. The reaction system was purged with nitrogen and then heated to 70 ° C. for 5 hours. The reaction was monitored by LCMS. The pH of the reaction solution was adjusted to 8 with 1N sodium hydroxide solution, and ethyl acetate was added for extraction. The aqueous phase was adjusted to pH 3 with 1N hydrochloric acid and extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give 0.82 g of the title compound, which was used directly in the next reaction.

[0444] Step 5: Synthesis of 4-(5-acetamido-3-chloro-2-methylphenyl)butanoic acid At 20 ° C., compound 2-1-05 (2.60 g, 9.71 mmol) was dissolved in THF (50 mL), Pd / C (0.52 g, content 10%) was added, the system was subjected to hydrogen substitution, and then reacted under the protection of a hydrogen balloon at 40 ° C. for 2 hours, and the reaction was monitored by LCMS. The reaction solution was filtered, and the filtrate was concentrated under reduced pressure to obtain 2.43 g of the title compound, which was used directly in the next reaction without further purification.

[0445] Step 6: Synthesis of N-(3-chloro-4-methyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide Compound 2-1-06 (2.43 g, 9.01 mmol) was dissolved in trifluoroacetic acid (10 mL), cooled to 5 ° C., and trifluoroacetic anhydride (3.78 g, 18.02 mmol, 2.50 mL) was added dropwise. The reaction was allowed to proceed at 5 ° C. for 4 hours, and the reaction was monitored by LCMS. The reaction solution was added to water, adjusted to pH 9 with 10 N sodium hydroxide, extracted with ethyl acetate, and the organic phases were combined, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified using a flash silica gel column (ethyl acetate:petroleum ether = 0 to 20%) to obtain 1.53 g of the title compound.

[0446] Step 7: Synthesis of (Z)-N-(3-chloro-7-(hydroxyimino)-4-methyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide At 5 ° C., potassium tert-butoxide (1.50 g, 13.37 mmol) was dissolved in THF (16 mL) and tert-butanol (4 mL), and compound 2-1-07 (1.53 g, 6.08 mmol) in THF solution (16 mL) was added dropwise, followed by amyl nitrite (1.14 g, 9.73 mmol) after 10 minutes. The reaction was carried out at 5 ° C. for 1 hour, and the reaction was monitored by LCMS. The reaction solution was adjusted to pH = 5 with 1 N hydrochloric acid, extracted with ethyl acetate, and the combined organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The concentrate was slurried with methyl tert-butyl ether to give 1.20 g of the title compound.

[0447] Step 8: N-(7-amino-3-chloro-4-methyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide At 20°C, compound 2-1-08 (0.50 g, 1.78 mmol) was dissolved in methanol (8 mL) and 2N hydrochloric acid (8 mL), Pd / C (0.15 g, 10% content) was added, the system was subjected to hydrogen substitution, and the reaction was carried out under the protection of a hydrogen balloon at 5°C for 2 hours, and the reaction was monitored by LCMS. The reaction solution was filtered, and the filtrate was concentrated under reduced pressure to obtain 0.52 g of the hydrochloride salt of the title compound, which was used directly in the next reaction without further purification.

[0448] Step 9: Synthesis of N,N'-(3-chloro-4-methyl-8-oxo-5,6,7,8-tetrahydronaphthalene-1,7-diyl)diacetamide Compound 2-1-09 (0.52 g, 1.70 mmol) was dissolved in pyridine (5 mL) at 20° C., and acetic anhydride (2 mL) was added. The reaction was allowed to proceed for 2 hours at 20° C. The reaction was monitored by LCMS. The reaction solution was added to water, extracted with ethyl acetate, and the organic phase was washed with water, combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The concentrate was purified using a flash silica gel column (ethyl acetate:petroleum ether = 0 to 30%) to obtain 0.22 g of the title compound.

[0449] Step 10: Synthesis of N-(8-amino-6-chloro-5-methyl-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)acetamide At 20°C, compound 2-1-10 (0.45 g, 1.46 mmol) was dissolved in methanol (16 mL), 2N hydrochloric acid (16 mL) was added, and the mixture was heated to 60°C and reacted for 2 hours. The reaction was monitored by LCMS. The reaction solution was cooled, saturated sodium bicarbonate solution was added to adjust the pH to 8, and extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain 0.23 g of the title compound, which was used directly in the next step without further purification.

[0450] Step 11: Synthesis of N-((9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzopyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)acetamide Compound 2-1-11 (0.23 g, 0.78 mmol) was dissolved in toluene (10 mL), (S)-4-ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (0.23 g, 0.87 mmol), and p-toluenesulfonic acid (26.73 mg, 0.16 mmol) were added, and the mixture was heated to 140 °C and reacted for 5 hours. The reaction was monitored by LCMS. The reaction solution was concentrated, and the crude product was purified by flash silica gel column (methanol:dichloromethane = 0 to 10%) to give 0.15 g of the title compound.

[0451] Step 12: Synthesis of (9S)-1-amino-5-chloro-9-ethyl-9-hydroxy-4-methyl-1,2,3,9,12,15-hexahydro-10H,13H-benzenepyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione Compound 2-1-12 (40.00 mg, 0.08 mmol) was added to concentrated hydrochloric acid (1 mL), heated to 100° C., and reacted for 5 hours. The reaction was monitored by LCMS. The reaction solution was filtered, the filtrate was purified by preparative high-performance liquid chromatography, and the fraction was lyophilized to obtain 12.00 mg of the trifluoroacetate salt of the title compound 2-23.

[0452] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% trifluoroacetic acid) [Table 5]

[0453] The structural property data were as follows: ESI-MS (m / z): 452.1 [M+H] + .

[0454] Step 13: Synthesis of 2-((tert-butyldiphenylsilyl)oxy)-N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)acetamide and 2-((tert-butyldiphenylsilyl)oxy)-N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)acetamide At 25°C, the trifluoroacetate salt of compound 2-23 (40.00 mg, 81.91 μmol) was dissolved in N,N-dimethylformamide (1 mL), and 2-((tert-butyldiphenylsilyl)oxy)acetic acid (30.91 mg, 98.29 μmol), HATU (62.25 mg, 163.81 μmol), and N,N-diisopropylethylamine (42.34 mg, 327.63 μmol) were added sequentially. The reaction was carried out at 25°C for 0.5 hours, and the reaction was monitored by LCMS. Upon completion of the reaction, water was added to the reaction solution, which was then extracted with dichloromethane / methanol (volume / volume=10 / 1). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified and separated by preparative thin-layer chromatography (dichloromethane:methanol=20:1) to obtain two isomers. According to their Rf values, the two isomers were named 2-1-13-A (15.00 mg, Rf value 0.3) and 2-1-13-B (12.00 mg, Rf value 0.35).

[0455] Step 14: Synthesis of N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide and N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide At 25°C, 2-1-13-A (15.00 mg) and 2-1-13-B (12.00 mg) were dissolved in tetrahydrofuran (1 mL) in two reaction flasks, and a mixed solution (50 µL) of tetrabutylammonium fluoride (1 M in tetrahydrofuran) / glacial acetic acid (volume / volume = 13 / 1) was added dropwise. The reaction was carried out at 25°C for 0.5 hours, and the reaction was monitored by LCMS. Upon completion of the reaction, the reaction solution was purified by preparative high-performance liquid chromatography, and the fractions were lyophilized to give the title compounds 2-1-A (6.94 mg) and 2-1-B (4.00 mg).

[0456] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 6]

[0457] The structural property data of 2-1-A was as follows: 1HNMR (400 MHz, DMSO-d6) δ 8.43 (d, J = 8.8Hz, 1H), 8.16 (s, 1H), 7.31 (s, 1H), 6.55 (s, 1H), 5.65-5.36 (m, 4H), 5.21 (q,J = 19.0 Hz, 2H), 3.95 (d, J = 5.7 Hz, 2H), 3.26-3.11 (m, 2H), 2.53 (s, 3H), 2.30-2.08 (m, 2H), 1.94-1.79 (m, 2H), 0.87 (t, J = 7.3 Hz, 3H). ESI-MS (m / z): 510.1[M+H] + . 2-1-B's structural characteristics are as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.45 (d, J = 8.9Hz, 1H), 8.15 (s, 1H), 7.31 (s, 1H), 6.54 (s, 1H), 5.64-5.35 (m, 4H), 5.19 (q,J = 19.0 Hz, 2H), 3.97 (d, J = 5.2 Hz, 2H), 3.27-3.10 (m, 2H), 2.51 (s, 3H), 2.27-2.10 (m, 2H), 1.93-1.80 (m, 2H), 0.88 (t, J = 7.3 Hz, 3H). ESI-MS (m / z):510.1[M+H] + .

[0458] Example 4: (2S)—N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropylamine and (2S)—N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropylamine [ka]

[0459] Step 1: (2S)-2-((tert-butyldiphenylsilyl)oxy)-N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)propyl amine Synthesis of (2S)-2-((tert-butyldiphenylsilyl)oxy)-N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)propylamine At 25°C, the hydrochloride salt of 2-23 (30.00 mg, 61.43 μmol) was dissolved in N,N-dimethylformamide (1 mL), and (S)-2-((tert-butyldiphenylsilyl)oxy)propionic acid (24.21 mg, 73.72 μmol), HATU (35.01 mg, 92.14 μmol), and N,N-diisopropylethylamine (23.82 mg, 184.29 μmol) were added sequentially. The reaction was carried out at 25°C for 1 hour, and the reaction was monitored by LCMS. Upon completion of the reaction, water was added to the reaction solution, which was then extracted with dichloromethane / methanol (volume / volume=10 / 1). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified and separated by preparative thin-layer chromatography (dichloromethane:methanol=15:1) to obtain two isomers. According to their Rf values, the two isomers were named 2-7-01-A (6.00 mg, Rf value 0.35) and 2-7-01-B (6.00 mg, Rf value 0.40).

[0460] Step 2: Synthesis of (2S)-N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropylamine and (2S)-N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropylamine At 25 °C, 2-7-01-A (6.00 mg, 7.87 μmol) and 2-7-01-B (6.00 mg, 7.87 μmol) were dissolved in anhydrous THF (1 mL) in two reaction flasks, and a mixture of tetrabutylammonium fluoride (1 M in tetrahydrofuran) and glacial acetic acid (volume / volume = 13 / 1) (50 μL) was added dropwise. The reaction was allowed to proceed at 25 °C for 0.5 hours, and the reaction was monitored by LCMS. Upon completion of the reaction, the reaction solution was purified by preparative high-performance liquid chromatography, and the fractions were lyophilized to give the title compounds 2-7-A (2.50 mg) and 2-7-B (3.00 mg).

[0461] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 7]

[0462] The structural property data of 2-7-A was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.38 (d, J = 8.8Hz, 1H), 8.15 (s, 1H), 7.31 (s, 1H), 6.55 (s, 1H), 5.56 - 5.47 (m, 2H), 5.42(s, 2H), 5.26 - 5.11 (m, 2H), 4.17 - 4.06 (m, 1H), 3.27 - 3.10 (m, 2H), 2.52(s, 3H), 2.27 - 2.08 (m, 2H), 1.86 (tt, J = 14.1, 7.3 Hz, 2H), 1.30 (d, J = 6.7Hz, 3H), 0.87 (t, J = 7.3 Hz, 3H). ESI-MS (m / z): 524.2 [M+H] + . The structural property data of 2-7-B was as follows: 1HNMR (400 MHz, DMSO-d6) δ 8.48 (d, J = 9.1Hz, 1H), 8.12 (s, 1H), 7.30 (s, 1H), 6.54 (s, 1H), 5.67 (d, J = 4.8 Hz, 1H),5.55 (dd, J = 14.6, 7.3 Hz, 1H), 5.43 (s, 2H), 5.24 (d, J = 19.0 Hz, 1H), 5.03(d, J = 19.0 Hz, 1H), 4.21-4.08 (m, 1H), 3.28 -3.08 (m, 2H), 2.51 (s, 3H), 2.16(d, J = 6.2 Hz, 2H), 1.94-1.82 (m, 2H), 1.42 (d, J = 6.8 Hz, 3H), 0.88 (t, J =7.3 Hz, 3H). ESI-MS (m / z): 524.2 [M+H] + .

[0463] Example 5: (2S)—N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropyl-2-hydroxyacetamide and (2S)—N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropyl-2-hydroxyacetamide and (2R)-N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropyl-2-hydroxyacetamide and (2R)-N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropyl-2-hydroxyacetamide [ka]

[0464] Step 1: (2S)-2-((tert-butyldiphenylsilyl)oxy)-N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropylacetate amide and (2S)-2-((tert-butyldiphenylsilyl)oxy)-N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropylacetamide and (2R)-2-((tert-butyldiphenylsilyl)oxy)-N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropylacetamide and Synthesis of (2R)-2-((tert-butyldiphenylsilyl)oxy)-N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropylacetamide At 25°C, the hydrochloride salt of 2-23 (30.00 mg, 61.43 μmol) was dissolved in N,N-dimethylformamide (1 mL), and 2-((tert-butyldiphenylsilyl)oxy)-2-cyclopropylacetic acid (26.13 mg, 73.72 μmol), HATU (35.01 mg, 92.14 μmol), and N,N-diisopropylethylamine (23.82 mg, 184.29 μmol) were added sequentially. The reaction was carried out at 25°C for 1 hour, and the reaction was monitored by LCMS. Upon completion of the reaction, water was added to the reaction solution, which was then extracted with dichloromethane / methanol (volume / volume=10 / 1). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified and separated by preparative thin-layer chromatography (dichloromethane:methanol=15:1) to obtain two isomers. According to their Rf values, the two isomers were named 2-12-01-A (8.00 mg, Rf value 0.35) and 2-12-01-B (10.00 mg, Rf value 0.40).

[0465] Step 2: (2S)-N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropyl-2-hydroxyacetamide and (2S)-N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropyl-2-hydroxyacetamide and synthesis of (2R)-N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropyl-2-hydroxyacetamide and (2R)-N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropyl-2-hydroxyacetamide At 25°C, 2-12-01-A (8.00 mg, 10.15 μmol) and 2-12-01-B (10.00 mg, 12.68 μmol) were dissolved in anhydrous THF (1 mL) in two reaction flasks, and a mixed solution (50 μL) of tetrabutylammonium fluoride (1 M in tetrahydrofuran) / glacial acetic acid (volume / volume = 13 / 1) was added dropwise. The reaction was carried out at 25°C for 0.5 hours, and the reaction was monitored by LCMS. Upon completion of the reaction, the reaction solution was purified by preparative high-performance liquid chromatography. 2-12-01-A was used as the raw material for the reaction, and two isomeric products were obtained after separation. The corresponding fractions were freeze-dried to obtain compounds 2-12-A (0.77 mg) and 2-12-B (1.03 mg). At the same time, 2-12-01-B was used as the raw material for the reaction, and two isomeric products were obtained after separation. The corresponding fractions were freeze-dried to obtain compounds 2-12-C (2.50 mg) and 2-12-D (1.00 mg). The purification conditions for 2-12-A / 2-12-B were as follows: Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 8]

[0466] Peak retention time: 2-12-A: 10.0 to 11.0 minutes, 2-12-B: 11.0 to 12.5 minutes The purification conditions for 2-12-C / 2-12-D were as follows: Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 9]

[0467] Peak retention time: 2-12-C: 10.6 to 11.4 minutes, 2-12-D: 11.4 to 12.5 minutes The structural property data of 2-12-A was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.38 (d, J = 8.9Hz, 1H), 8.15 (s, 1H), 7.30 (s, 1H), 6.54 (s, 1H), 5.63-5.53 (m, 1H), 5.51 (d,J = 5.1 Hz, 1H), 5.42 (s, 2H), 5.28 (d, J = 19.2 Hz, 1H), 5.16 (d, J = 19.1 Hz,1H), 3.60 (t, J = 5.6 Hz, 1H), 3.28-3.11 (m, 2H), 2.52 (s, 3H), 2.22-2.10 (m,2H), 1.86 (tt, J = 14.1, 7.3 Hz, 2H), 1.23 (d, J = 4.9 Hz, 1H), 0.87 (t, J =7.2 Hz, 3H), 0.56-0.36 (m, 4H). ESI-MS (m / z): 550.2 [M+H] + . The structural property data of 2-12-B was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.36 (d, J = 8.7Hz, 1H), 8.16 (s, 1H), 7.31 (s, 1H), 6.55 (s, 1H), 5.57-5.48 (m, 1H), 5.42 (s,2H), 5.40 (d, J = 5.4 Hz, 1H), 5.26 (d, J = 19.3 Hz, 1H), 5.18 (d, J = 19.0 Hz,1H), 3.65-3.60 (m, 1H), 3.26-3.12 (m, 2H), 2.52 (s, 3H), 2.26-2.09 (m, 2H),1.86 (tt, J = 14.1, 7.2 Hz, 2H), 1.19-1.08 (m, 1H), 0.87 (t, J = 7.3 Hz, 3H),0.51-0.27 (m, 4H). ESI-MS (m / z): 550.2 [M+H] + . The structural property data of 2-12-C was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.42 (d, J = 9.0Hz, 1H), 8.15 (s, 1H), 7.31 (s, 1H), 6.54 (s, 1H), 5.56 (dd, J = 14.8, 6.8 Hz,1H), 5.52 (d, J = 5.2 Hz, 1H), 5.42 (s, 2H), 5.29 (d, J = 19.2 Hz, 1H), 5.16(d, J = 19.1 Hz, 1H), 3.62-3.58 (m, 1H), 3.27 -3.08 (m, 2H), 2.51 (s, 3H),2.27-2.08 (m, 2H), 1.87 (tt, J = 14.0, 7.2 Hz, 2H), 1.25 (dd, J = 13.2, 6.8 Hz,1H), 0.87 (t, J = 7.3 Hz, 3H), 0.64-0.28 (m, 4H). ESI-MS (m / z): 550.1 [M+H] + . The structural property data of 2-12-D was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.38 (d, J = 8.7Hz, 1H), 8.16 (s, 1H), 7.31 (s, 1H), 6.54 (s, 1H), 5.59-5.49 (m, 1H), 5.43 (d,J = 5.3 Hz, 1H), 5.43 (s, 2H), 5.26 (d, J = 19.1 Hz, 1H), 5.17 (d, J = 19.0 Hz,1H), 3.64 (t, J = 5.8 Hz, 1H), 3.17 (dd, J = 15.6, 8.3 Hz, 2H), 2.27-2.07 (m,2H), 1.94-1.79 (m, 2H), 1.19-1.06 (m, 1H), 0.87 (t, J = 7.3 Hz, 3H), 0.49-0.28(m, 4H). ESI-MS (m / z): 550.1 [M+H] + .

[0468] Example 6: N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxy-2-methylpropylamine and N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxy-2-methylpropylamine [ka]

[0469] The hydrochloride salt of 2-23 (30.00 mg, 61.43 μmol) was dissolved in N,N-dimethylformamide (1 mL) at 25 °C, and 2-((tert-butyldimethylsilyl)oxy)-2-methylpropionic acid (16.10 mg, 73.72 μmol), HATU (35.01 mg, 92.14 μmol), and N,N-diisopropylethylamine (23.82 mg, 184.29 μmol) were added sequentially. The reaction was allowed to proceed at 25 °C for 1 h. The reaction was monitored by LCMS. Upon completion, the reaction solution was concentrated, and the crude product was purified and separated by preparative high-performance liquid chromatography to obtain two isomers. The fractions were lyophilized, and the two isomers were designated 2-17-A (2.65 mg) and 2-17-B (2.69 mg).

[0470] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 10]

[0471] Peak retention times: 2-17-A: 9.5 to 10.2 minutes, and 2-17-B: 10.4 to 10.6 minutes.

[0472] The structural property data of 2-17-A was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.36 (d, J = 9.1Hz, 1H), 8.13 (s, 1H), 7.30 (s, 1H), 6.54 (s, 1H), 5.55-5.45 (m, 2H), 5.42 (s,2H), 5.28 (d, J = 19.0 Hz, 1H), 5.05 (d, J = 19.0 Hz, 1H), 3.27-3.11 (m, 2H),2.51 (s, 1H), 2.24-2.10 (m, 2H), 1.86 (tt, J = 14.0, 7.2 Hz, 2H), 1.46 (s, 3H),1.35 (s, 3H), 0.87 (t, J = 7.3 Hz, 3H). MS m / z (ESI): 538.2 [M+H] + . The structural property data of 2-17-B was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.40 (d, J = 9.2Hz, 1H), 8.13 (s, 1H), 7.30 (s, 1H), 6.53 (s, 1H), 5.60-5.46 (m, 2H), 5.42 (s,2H), 5.29 (d, J = 19.0 Hz, 1H), 5.02 (d, J = 19.0 Hz, 1H), 3.28 - 3.08 (m, 2H),2.50 (s, 3H), 2.22-2.10 (m, 2H), 1.87 (tt, J = 14.2, 7.2 Hz, 2H), 1.47 (s, 3H),1.35 (s, 3H), 0.88 (t, J = 7.3 Hz, 3H). MS m / z (ESI): 538.2 [M+H] + .

[0473] Example 7: N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-1-hydroxycyclopropane-1-carboxamide and N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-1-hydroxycyclopropane-1-carboxamide [ka]

[0474] Step 1: 1-((tert-butyldiphenylsilyl)oxy)-N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)cyclopropane-1-carboxamide Synthesis of 1-((tert-butyldiphenylsilyl)oxy)-N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)cyclopropane-1-carboxamide At 25°C, the hydrochloride salt of 2-23 (30.00 mg, 61.43 μmol) was dissolved in N,N-dimethylformamide (1 mL), and 1-((tert-butyldiphenylsilyl)oxy)cyclopropane-1-carboxylic acid (25.10 mg, 73.72 μmol), HATU (35.01 mg, 92.14 μmol), and N,N-diisopropylethylamine (23.82 mg, 184.29 μmol) were added sequentially. The reaction was carried out at 25°C for 1 hour, and the reaction was monitored by LCMS. When the reaction was completed, water was added to the reaction solution, and the mixture was extracted with dichloromethane / methanol (volume / volume=10 / 1). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified and separated by preparative thin-layer chromatography (dichloromethane:methanol=15:1) to obtain two isomers. According to their Rf values, the two isomers were named 2-20-01-A (4.00 mg, Rf value 0.30) and 2-20-01-B (4.00 mg, Rf value 0.35).

[0475] Step 2: Synthesis of N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-1-hydroxycyclopropane-1-carboxamide and N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-1-hydroxycyclopropane-1-carboxamide At 25 °C, 2-20-01-A (4.00 mg, 5.17 μmol) and 2-20-01-B (4.00 mg, 5.17 μmol) were dissolved in anhydrous THF (1 mL) in two reaction flasks, and a mixture of tetrabutylammonium fluoride (1 M in tetrahydrofuran) and glacial acetic acid (volume / volume = 13 / 1) (50 μL) was added dropwise. The reaction was allowed to proceed at 25 °C for 0.5 h, and the reaction was monitored by LCMS. Upon completion of the reaction, the reaction solution was purified by preparative high-performance liquid chromatography, and the fractions were lyophilized to give the title compounds 2-20-A (0.71 mg) and 2-20-B (1.05 mg).

[0476] The purification conditions for 2-20-A were as follows: Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 11]

[0477] The purification conditions for 2-20-B were as follows: Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 12]

[0478] The structural property data of 2-20-A was as follows: 1HNMR (400 MHz, DMSO-d6) δ 8.58 (d, J = 9.0Hz, 1H), 8.15 (s, 1H), 7.31 (s, 1H), 6.55 (s, 1H), 6.30 (s, 1H), 5.55 (dd, J =13.2, 8.2 Hz, 1H), 5.43 (s, 2H), 5.26 (d, J = 19.0 Hz, 1H), 5.10 (d, J = 19.0Hz, 1H), 3.29-3.09 (m, 2H), 2.52 (s, 3H), 2.31-2.15 (m, 2H), 1.93-1.80 (m, 2H),1.25-1.14 (m, 2H), 0.98-0.90 (m, 2H), 0.87 (t, J = 7.3 Hz, 3H). ESI-MS (m / z):536.2 [M+H] + . 2-20-B's structural characteristics are as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.63 (d, J = 9.0Hz, 1H), 8.15 (s, 1H), 7.31 (s, 1H), 6.54 (s, 1H), 6.35 (s, 1H), 5.55 (dd, J =13.5, 8.6 Hz, 1H), 5.43 (s, 2H), 5.29 (d, J = 19.1 Hz, 1H), 5.08 (d, J = 19.1Hz, 1H), 3.28-3.10 (m, 2H), 2.51 (s, 3H), 2.30- 2.14 (m, 2H), 1.93-1.81 (m,2H), 1.26-1.14 (m, 2H), 1.02-0.90 (m, 2H), 0.89 (d, J = 10.9 Hz, 3H). ESI-MS (m / z):536.2 [M+H] + .

[0479] Example 8: (1S,9S)-1-Amino-4-chloro-9-ethyl-5-fluoro-9-hydroxy-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione and (1R,9S)-1-Amino-4-chloro-9-ethyl-5-fluoro-9-hydroxy-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione [ka]

[0480] Step 1: Synthesis of 3-bromo-4-chloro-5-fluoroaniline Compound 3-1-01 (2.00 g, 10.53 mmol) was dissolved in N,N-dimethylformamide (30 mL), and then N-chlorosuccinimide (1.69 g, 12.63 mmol) was slowly added. After the addition, the reaction was carried out at room temperature for 16 hours and monitored by LCMS. The reaction solution was concentrated under reduced pressure to give the crude product, which was purified by flash silica gel column (ethyl acetate:petroleum ether = 0-25%) to give 0.95 g of the title compound.

[0481] The structural property data were as follows: 1 HNMR (400 MHz, DMSO-d6) δ 6.77 (dd, J = 2.5,1.4 Hz, 1H), 6.51 (dd, J = 11.7, 2.5 Hz, 1H), 5.84 (s, 2H).

[0482] Step 2: Synthesis of N-(3-bromo-4-chloro-5-fluorophenyl)acetamide Compound 3-1-02 (0.95 g, 4.23 mmol) was dissolved in ethyl acetate (20 mL) and acetic anhydride (648.13 mg, 6.35 mmol) was added under nitrogen protection. After the addition, the temperature was raised to 50 °C and the reaction was carried out for 15 hours and monitored by LCMS. The reaction solution was quenched with methanol (5 mL) and directly evaporated to dryness under reduced pressure to obtain the crude product, which was purified by flash silica gel column (ethyl acetate: petroleum ether = 0-40%) to obtain 1.01 g of the title compound.

[0483] The structural property data were as follows: ESI-MS (m / z): 265.9[M+H] + .

[0484] Step 3: Synthesis of (E)-4-(5-acetylamino-2-chloro-3-fluorophenyl)-3-butenoic acid Compound 3-1-03 and 3-butenoic acid (387.65 mg, 4.50 mmol) were dissolved in a mixed solvent of 1,4-dioxane (24 mL) and water (8 mL), and then N,N-diisopropylethylamine (1.45 g, 11.26 mmol), tris(o-methylphenyl)phosphine (114.21 mg, 375.24 μmol), and palladium acetate (42.12 mg, 187.62 μmol) were added. After the addition, the reaction system was subjected to nitrogen purge three times and heated to 100 °C. The reaction was carried out under a nitrogen atmosphere for 16 hours and monitored by LCMS. After the reaction solution was cooled to room temperature, 1N aqueous sodium hydroxide solution (60 mL) and ethyl acetate (50 mL) were added, and the mixture was shaken to form layers. After separating the lower aqueous phase, the pH was adjusted to about 3 with 4 mol / L aqueous hydrochloric acid, and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated to dryness under reduced pressure to give 1.00 g of the crude product of the title compound.

[0485] The structural property data were as follows: ESI-MS (m / z): 272.0 [M + H] + .

[0486] Step 4: Synthesis of 4-(5-acetylamino-2-chloro-3-fluorophenyl)butanoic acid The crude product of compound 3-1-04 (1.00 g, 3.68 mmol) was dissolved in tetrahydrofuran (15 mL), and then 10% palladium on carbon (0.10 g) was added. After the addition, the reaction system was subjected to hydrogen balloon replacement three times, and the reaction was carried out under a hydrogen atmosphere for 4 hours and monitored by LCMS. The reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain 1.00 g of the crude product of the title compound.

[0487] The structural property data were as follows: ESI-MS (m / z): 274.0 [M + H] + .

[0488] Step 5: Synthesis of N-(4-chloro-3-fluoro-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide The crude product of compound 3-1-05 (1.00 g, 3.65 mmol) was dissolved in trifluoroacetic acid (5 mL) and cooled to 5 ° C., and then trifluoroacetic anhydride (3.84 g, 18.27 mmol, 2.54 mL) was slowly added. After the addition, the reaction was carried out at 5 ° C. for 2 hours and monitored by LCMS. The reaction solution was slowly poured into water and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and then filtered. The filtrate was evaporated to dryness under reduced pressure to obtain the crude product, which was purified by flash silica gel column to obtain 0.43 g of the title compound.

[0489] The structural property data were as follows: ESI-MS (m / z): 256.1 [M+H] + .

[0490] Step 6: Synthesis of N-(4-chloro-3-fluoro-7-(hydroxyimino)-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide Tetrahydrofuran (16 mL) and tert-butanol (4 mL) were added to a reaction flask, cooled to 5 ° C. in an ice bath, potassium tert-butoxide (415.18 mg, 3.70 mmol) was added, and then compound 3-1-06 (0.43 mg, 1.68 mmol) was dissolved in tetrahydrofuran (1 mL) and slowly added thereto. After 10 minutes, isoamyl nitrite (315.24 mg, 2.69 mmol) was added. After the addition, the reaction was carried out at 5 ° C. for 1 hour, and the reaction was monitored by LCMS. The reaction solution was quenched with saturated aqueous ammonium chloride solution, extracted with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and then filtered. The filtrate was concentrated under reduced pressure to obtain 455.00 mg of the crude product of the title compound.

[0491] The structural property data were as follows: ESI-MS (m / z): 285.0 [M+H] + .

[0492] Step 7: Synthesis of N-(7-amino-4-chloro-3-fluoro-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide The crude product of compound 3-1-07 (0.40 g, 1.41 mmol) was dissolved in methanol (10 mL), and then 3 mol / L aqueous hydrochloric acid (1 mL) and 10% palladium on carbon (40.00 mg) were added. After the addition, the reaction system was subjected to hydrogen balloon purge three times. The reaction was carried out under a hydrogen atmosphere at room temperature for 1 hour and monitored by LCMS. The reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain 0.43 g of the crude hydrochloride salt of the title compound.

[0493] The structural property data were as follows: ESI-MS (m / z): 271.0 [M + H] + .

[0494] Step 8: Synthesis of (9H-fluoren-9-yl)methyl (8-acetamido-5-chloro-6-fluoro-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate The crude hydrochloride salt of compound 3-1-08 (0.43 g, 1.19 mmol) was dissolved in 1,4-dioxane (15 mL), followed by the addition of sodium bicarbonate (400.35 mg, 4.77 mmol), water (5 mL), and 9-fluorenylmethyl-N-succinimidyl carbonate (481.81 mg, 1.43 mmol). After the addition, the reaction was stirred at room temperature for 2 hours and monitored by LCMS. The reaction solution was poured into water and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give the crude product. The crude product was purified using a C18 reverse-phase column (acetonitrile:0.05% formic acid in water = 20% to 100%) to give 301.00 mg of the title compound.

[0495] The structural property data were as follows: ESI-MS (m / z): 493.2 [M+H] + .

[0496] Step 9: Synthesis of (9H-fluoren-9-yl)methyl (8-amino-5-chloro-6-fluoro-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate Compound 3-1-09 (300.00 mg, 608.61 μmol) was dissolved in dioxane (5 mL) and 12 mol / L concentrated hydrochloric acid (1 mL) was added. After the addition, the temperature was raised to 60 °C, and the reaction was carried out for 2 hours and monitored by LCMS. The reaction solution was poured into water and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was purified using a flash silica gel column (ethyl acetate:petroleum ether = 0-50%) to obtain 198.00 mg of the title compound.

[0497] The structural property data were as follows: ESI-MS (m / z): 451.1 [M+H] + .

[0498] Step 10: Synthesis of (9H-fluoren-9-yl)methyl ((9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)carbamate (S)-4-Ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (138.72 mg, 526.96 μmol) and compound 3-1-10 (198.00 mg, 439.13 μmol) were added to toluene (10 mL), and then p-toluenesulfonic acid (75.53 mg, 439.13 μmol) was added. After the addition, the temperature was raised to 140 ° C. and the reaction was carried out for 4 hours. The reaction solution was directly evaporated to dryness under reduced pressure at 140 ° C. to obtain the crude product. The crude product was purified by flash silica gel column (methanol:dichloromethane = 0 to 5%) to obtain 256.00 mg of the title compound.

[0499] The structural property data were as follows: ESI-MS (m / z): 678.1 [M+H] + .

[0500] Step 11: Synthesis of (1S,9S)-1-amino-4-chloro-9-ethyl-5-fluoro-9-hydroxy-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione and (1R,9S)-1-amino-4-chloro-9-ethyl-5-fluoro-9-hydroxy-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione Compound 3-1-11 (201.18 mg, 296.67 μmol) was dissolved in N,N-dimethylformamide (4 mL), and then diethylamine (108.49 mg, 1.48 mmol) was added. After the addition, the reaction was carried out at room temperature for 0.5 h and monitored by LCMS. The reaction solution was distilled under reduced pressure to remove ethylenediamine, and the pH was adjusted to 2-3 with 1 mol / L aqueous hydrochloric acid. The reaction solution was directly purified by preparative high-performance liquid chromatography to give the title compounds 3-1-A (44.00 mg) and 3-1-B (43.00 mg).

[0501] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 13]

[0502] 3-1-A (6 min LCMS, earlier peak with retention time 1.276 min) The structural property data were as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.00 (d, J = 10.3Hz, 1H), 7.33 (s, 1H), 6.54 (s, 1H), 5.62 (d, J = 19.3 Hz, 1H), 5.44 (s, 2H),5.38 (d, J = 19.3 Hz, 1H), 4.43-4.38 (m, 1H), 3.28-3.10 (m, 2H), 2.22-2.12 (m,1H), 2.12-2.02 (m, 1H), 1.93-1.80 (m, 2H), 0.87 (t, J = 7.3 Hz, 3H). ESI-MS (m / z): 456.1 [M+H] + . 3-1-B (6 min LCMS, later peak with retention time 1.300 min) These structural property data were as follows: 1HNMR (400 MHz, DMSO-d6) δ7.98 (d, J = 10.3Hz, 1H), 7.32 (s, 1H), 5.61 (d, J = 19.4 Hz, 1H), 5.44 (s, 2H), 5.32 (d, J =19.4 Hz, 1H), 4.44-4.36 (m, 1H), 3.33-3.25 (m, 1H), 3.22-3.11 (m, 1H), 2.23 -2.13 (m, 1H), 2.11- 2.03 (m, 1H), 1.96 -1.82 (m, 2H), 0.89 (t, J = 7.3 Hz, 3H). ESI-MS (m / z): 456.1 [M+H] + .

[0503] 6 min LCMS conditions: Chromatography column: Waters SunFire C18 OBD 4.6 mm x 50 mm x 5.0 μm Mobile phase A: 0.05% acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 14]

[0504] Example 9: N-((1S,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide and N-((1R,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide [ka]

[0505] Step 1: Synthesis of (S)-10-benzyl-23-(2-(methylsulfonyl)pyrimidin-5-yl)-6,9,12,15,18-pentoxo-3-oxo-5,8,11,14,17-pentazooctadecane-22-yne-carboxylic acid Compound 3-4-01 (30.00 mg, 70.00 μmol) was dissolved in N,N-dimethylformamide (1 mL), and 2,5-dioxopyrrolidin-1-yl 6-(2-(methylsulfonyl)pyrimidin-5-yl)hex-5-ynyl-amide (28.00 mg, 77.00 μmol) was added. The mixture was allowed to react at room temperature for 1 hour, and the reaction was monitored by LCMS. The reaction solution was directly purified by preparative high-performance liquid chromatography, and the fraction was lyophilized to give the title compound 3-4-03 (20.00 mg).

[0506] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 15]

[0507] The structural property data were as follows: ESI-MS (m / z): 691.0 [M+18]+.

[0508] Step 2: N-((S)-10-benzyl-1-((1S,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)amino)-1,6,9,12,15-pentaoxo-3-oxa-5,8,11,14-tetraazahexadecan-16-yl)-6-(2-(methylsulfonyl)pyrimidin-5-yl)hex-5-ynyl-amino Synthesis of N-((S)-10-benzyl-1-((1R,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)amino)-1,6,9,12,15-pentoxo-3-oxa-5,8,11,14-tetraazahexadecan-16-yl)-6-(2-(methylsulfonyl)pyrimidin-5-yl)hex-5-ynyl-amide Compound 3-1-A (36.00 mg, 79.70 μmol) and compound 3-4-03 (64.43 mg, 95.64 μmol) were dissolved in N,N-dimethylformamide (2 mL) in a single configuration, and then 4-(4,6-dimethoxytriazin-2-yl)-4-methylmorpholine hydrochloride (46.98 mg, 159.40 μmol) and triethylamine (24.19 mg, 239.10 μmol) were added. After the addition, the reaction was carried out at room temperature for 1 hour and monitored by LCMS. The reaction solution was directly purified by high-performance liquid chromatography to obtain the title compound 3-4-04-A (51.00 mg) in a single configuration.

[0509] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 16]

[0510] The structural property data were as follows: ESI-MS (m / z): 1111.0 [M+H] + .

[0511] Compound 3-1-B (36.00 mg, 79.70 μmol) and compound 3-4-03 (64.43 mg, 95.64 μmol) were dissolved in N,N-dimethylformamide (2 mL) in a single configuration, and then 4-(4,6-dimethoxytriazin-2-yl)-4-methylmorpholine hydrochloride (46.98 mg, 159.40 μmol) and triethylamine (24.19 mg, 239.10 μmol) were added. After the addition, the reaction was carried out at room temperature for 1 hour and monitored by LCMS. The reaction solution was directly purified by high-performance liquid chromatography to obtain the title compound 3-4-04-B (52.00 mg) in a single configuration.

[0512] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 17]

[0513] The structural property data were as follows: ESI-MS (m / z): 1111.0 [M+H] + .

[0514] Step 3: Synthesis of N-((1S,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide and N-((1R,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide Compound 3-4-04-A (40.00 mg, 35.99 μmol) was weighed and dissolved in a mixed solvent of dichloromethane (2 mL) and methanol (1 mL). 4 mol / L ethyl acetate hydrochloride (1 mL) was then added. After the addition, the reaction was carried out at room temperature for 0.5 hours and monitored by LCMS. The reaction solution was directly concentrated to dryness under reduced pressure to obtain the crude product. The crude product was purified by high-performance liquid chromatography to obtain the title compound 3-4-A (4.75 mg) in a single configuration.

[0515] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 18]

[0516] The structural property data were as follows: 1HNMR (400 MHz, DMSO-d6) δ 8.50 (d, J = 8.9Hz, 1H), 8.05 (d, J = 10.3 Hz, 1H), 7.33 (s, 1H), 6.55 (s, 1H), 5.67 -5.60 (m,1H), 5.49 (t, J = 5.8 Hz, 1H), 5.43 (s, 2H), 5.21 (s, 2H), 3.96 (d, J = 5.8 Hz,2H), 3.32-3.22 (m, 2H), 2.28-2.15 (m, 2H), 1.93-1.80 (m, 2H), 0.87 (t, J = 7.3Hz, 3H). ESI-MS (m / z): 514.0 [M+H] + .

[0517] Compound 3-4-04-B (40.00 mg, 35.99 μmol) was weighed and dissolved in a mixed solvent of dichloromethane (2 mL) and methanol (1 mL). 4 mol / L ethyl acetate hydrochloride (1 mL) was then added. After the addition, the reaction was carried out at room temperature for 0.5 hours and monitored by LCMS. The reaction solution was directly concentrated to dryness under reduced pressure to obtain the crude product. The crude product was purified by high-performance liquid chromatography to obtain the title compound 3-4-B (8.24 mg) in a single configuration.

[0518] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 19]

[0519] The structural property data were as follows: 1HNMR (400 MHz, DMSO-d6) δ 8.52 (d, J = 9.0Hz, 1H), 8.05 (d, J = 10.3 Hz, 1H), 7.34 (s, 1H), 6.55 (s, 1H), 5.68 -5.58 (m,1H), 5.53 (t, J = 5.8 Hz, 1H), 5.43 (d, J = 2.9 Hz, 2H), 5.20 (d, J = 7.3 Hz,2H), 3.97 (d, J = 5.7 Hz, 2H), 3.31-3.21 (m, 2H), 2.26-2.15 (m, 2H), 1.92-1.82(m, 2H), 0.87 (t, J = 7.3 Hz, 3H). ESI-MS (m / z): 514.0 [M+H] + .

[0520] Example 10: (S)—N-(2-(4-ethyl-8-fluoro-4-hydroxy-9-methyl-3,14-dioxo-3,4,12,14-tetrahydro-1H-pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-11-yl)ethyl)-2-hydroxy-N-isopropylacetamide [ka]

[0521] Step 1: Synthesis of 1-(4-fluoro-3-methylphenyl)-3-(isopropylamino)propan-1-one At 20°C, compound 4-12-01 (500.00 mg, 3.29 mmol), formaldehyde solution (2.5 mL, 37%), and isopropylamine (388.46 mg, 6.57 mmol) were dissolved in isopropanol (5 mL), and concentrated hydrochloric acid (2.5 mL) was added dropwise at 0°C. The reaction solution was stirred at 100°C for 16 hours and monitored by LCMS. The reaction solution was concentrated under reduced pressure to obtain a crude product, which was purified by preparative high-performance liquid chromatography. The fractions were lyophilized to obtain 200.00 mg of the title compound.

[0522] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 20]

[0523] The structural property data were as follows: ESI-MS (m / z): 224.1 [M+H] + .

[0524] Step 2: Synthesis of 1-(4-fluoro-5-methyl-2-nitrophenyl)-3-(isopropylamino)propan-1-one At 0 ° C., compound 4-12-02 (100.00 mg, 0.49 mmol) was added to concentrated sulfuric acid (0.5 mL), and then potassium nitrate (54.34 mg, 0.54 mmol) was added, and the reaction mixture was reacted at 0 ° C. for 1 hour and monitored by LCMS. The reaction solution was poured into ice water and purified by reverse phase column (acetonitrile: 0.05% formic acid water = 0 to 30%) to obtain 90.00 mg of the title compound.

[0525] The structural property data were as follows: ESI-MS (m / z): 269.0 [M+H] + .

[0526] Step 3: Synthesis of 1-(2-amino-4-fluoro-5-methylphenyl)-3-(isopropylamino)propan-1-one At 25°C, compound 4-12-03 (200.00 mg, 0.75 mmol) was added to methanol (20.0 mL), and then 10% palladium on carbon (10.00 mg) was added. The reaction solution was subjected to hydrogen substitution and reacted under a hydrogen atmosphere at 20°C for 16 hours. The reaction was monitored by LCMS. The reaction solution was filtered and concentrated under reduced pressure to give 183.00 mg of the title compound.

[0527] The structural property data were as follows: ESI-MS (m / z): 239.1 [M+H] + .

[0528] Step 4: Synthesis of (S)-4-ethyl-8-fluoro-4-hydroxy-11-(2-(isopropylamino)ethyl)-9-methyl-1,12-dihydro-14H-pyrano[3',4':6,7]indolizino[1,2-b]quinoline-3,14(4H)-dione Compound 4-12-04 (50.00 mg, 0.21 mmol) and (S)-4-ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (55.23 mg, 0.21 mmol) were added to toluene (3 mL) at 25 ° C., followed by p-toluenesulfonic acid (3.61 mg, 0.02 mmol). The reaction solution was reacted at 130 ° C. for 4 hours and monitored by LCMS. The reaction solution was concentrated under reduced pressure, and the crude product was purified by preparative high-performance liquid chromatography. The fractions were lyophilized to give 2.00 mg of the trifluoroacetate salt of the title compound.

[0529] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% trifluoroacetic acid) [Table 21]

[0530] The structural property data were as follows: 1HNMR (400 MHz, DMSO-d6) δ 8.58 (s, 2H), 8.22(d, J = 8.1 Hz, 1H), 7.97 (d, J = 10.7 Hz, 1H), 7.35 (s, 1H), 6.59 (s, 1H),5.47 (s, 2H), 5.41 (s, 2H), 3.58-3.45 (m, 3H), 3.31-3.23 (m, 2H), 2.56 (s, 3H),1.98-1.80 (m, 2H), 1.26 (d, J = 6.3 Hz, 6H), 0.89 (t, J = 7.3 Hz, 3H). ESI-MS (m / z): 466.2 [M+H] +

[0531] Step 5: Synthesis of (S)-2-((tert-butyldiphenylsilyl)oxy)-N-(2-(4-ethyl-8-fluoro-4-hydroxy-9-methyl-3,14-dioxo-3,4,12,14-tetrahydro-1H-pyrano[3',4':6,7]indolizino[1,2-b]quinolin-11-yl)ethyl)-N-isopropylacetamide Compound 4-12-05 (22.00 mg, 47.26 μmol) and 2-((tert-butyldiphenylsilyl)oxy)acetic acid (16.35 mg, 51.99 μmol) were dissolved in N,N-dimethylformamide (1 mL), and then HATU (21.55 mg, 56.71 μmol) and N,N-diisopropylethylamine (18.32 mg, 141.78 μmol) were added. After the addition, the reaction was carried out at room temperature for 0.5 h and monitored by LCMS. The reaction solution was directly purified using a C18 reverse-phase column (acetonitrile:0.05% formic acid aqueous solution = 30% to 100%) to give 18.00 mg of the title compound.

[0532] The structural property data were as follows: ESI-MS (m / z): 762.3 [M+H] + .

[0533] Step 6: Synthesis of (S)-N-(2-(4-ethyl-8-fluoro-4-hydroxy-9-methyl-3,14-dioxo-3,4,12,14-tetrahydro-1H-pyrano[3',4':6,7]indolizino[1,2-b]quinolin-11-yl)ethyl)-2-hydroxy-N-isopropylacetamide Compound 4-12-06 (18.00 mg, 23.62 μmol) was dissolved in N,N-dimethylformamide (1 mL), and then potassium fluoride (6.86 mg, 118.12 μmol) was added. The mixture was then heated to 50 ° C. and reacted for 1 hour. The reaction was monitored by LCMS. The reaction solution was directly purified by high-performance liquid chromatography to obtain 1.53 mg of the title compound.

[0534] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 22]

[0535] The structural property data were as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.53 (d, J = 8.2Hz, 1H), 7.91 (d, J = 10.8 Hz, 1H), 7.32 (s, 1H), 6.55 (s, 1H), 5.44 (d, J =13.8 Hz, 4H), 4.72 (t, J = 5.5 Hz, 1H), 4.21 (d, J = 5.5 Hz, 2H), 3.99-3.90 (m,1H), 3.54-3.38 (m, 4H), 2.54 (s, 3H), 1.92-1.83 (m, 2H), 1.17 (dd, J = 6.6, 3.1Hz, 6H), 0.87 (t, J = 7.3 Hz, 3H). ESI-MS (m / z): 524.2 [M+H] + .

[0536] Example 11: (S)—N-((4-ethyl-8-fluoro-4-hydroxy-9-methyl-3,14-dioxo-3,4,12,14-tetrahydro-1H-pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-11-yl)methyl)-1-hydroxycyclopropane-carboxamide [ka]

[0537] The raw materials (S)-11-(aminomethyl)-4-ethyl-8-fluoro-4-hydroxy-9-methyl-1H-pyrano[3',4':6,7]indolizino[1,2-b]quinoline-3,14(4H,12H)-dione (4-10-01, 30.00 mg, 67.00 μmol) and 1-hydroxycyclopropanecarboxylic acid (7.56 mg, 0.074 mmol) were dissolved in DMF (1 mL), and HBTU (34.30 mg, 0.14 mmol) and diisopropylethylamine (26.09 mg, 0.20 mmol) were added with stirring. The mixture was allowed to react at room temperature for 4 hours. Water and ethyl acetate were added, stirred, and the layers were separated by settling. The organic phase was washed with saturated brine and concentrated under reduced pressure. The concentrate was purified by preparative thin-layer chromatography (dichloromethane:methanol=20:1) and then by preparative high-performance liquid chromatography to give 1.20 mg of a solid.

[0538] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 23]

[0539] The structural property data were as follows: 1HNMR (400 MHz, DMSO-d6) δ 8.96 (t, J = 6.0Hz, 1H), 8.51 (d, J = 8.0 Hz, 1H), 7.90 (d, J = 10.8 Hz, 1H), 7.31 (s, 1H),6.53 (s, 1H), 6.30 (s, 2H), 6.30 (s, 1H), 5.52 (s, 2H), 5.44 (s, 2H), 4.84 (d,J = 6.0 Hz, 2H),2.51(s,3H), 1.91-1.81 (m, 2H), 1.01 (dd, J = 7.2, 4.1 Hz, 2H),0.87 (t, J = 7.3 Hz, 3H), 0.83 (t, J = 3.6 Hz, 2H). ESI-MS (m / z): 494.1[M+1]+.

[0540] Example 12: (1S,9S)-1-Amino-9-ethyl-5-fluoro-9-hydroxy-1,4-dimethyl-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione and (1R,9S)-1-Amino-9-ethyl-5-fluoro-9-hydroxy-1,4-dimethyl-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione [ka]

[0541] Step 1: Synthesis of (E)-4-(5-acetylamino-3-fluoro-2-methylphenyl)-2-methyl-3-butenoic acid N-(3-bromo-5-fluoro-4-methylphenyl)acetamide (2.00 g, 8.13 mmol) and 2-methyl-3-butenoic acid (976.44 mg, 9.75 mmol) were weighed and dissolved in a mixed solvent of 1,4-dioxane (15 mL) and water (5 mL). Tris(o-methylphenyl)phosphine (247.37 mg, 812.76 μmol), palladium acetate (91.24 mg, 406.38 μmol), and N,N-diisopropylethylamine (2.31 g, 17.88 mmol) were then added. After the addition, the reaction system was subjected to nitrogen purge three times, the temperature was raised to 80 °C, and the reaction was carried out under a nitrogen atmosphere for 3 hours and monitored by LCMS. After the reaction solution was cooled to room temperature, 1 mol / L aqueous sodium hydroxide solution (60 mL) and ethyl acetate (50 mL) were added, and the mixture was shaken to separate the layers. After separating the lower aqueous layer, the pH was adjusted to about 3 with 4 mol / L aqueous hydrochloric acid, and then extraction was carried out with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give 1.90 g of the title compound.

[0542] The structural property data were as follows: ESI-MS (m / z): 266.1 [M+H] + .

[0543] Step 2: Synthesis of 4-(5-acetylamino-3-fluoro-2-methylphenyl)-2-methylbutanoic acid (E)-4-(5-acetylamino-3-fluoro-2-methylphenyl)-2-methyl-3-butenoic acid (1.90 g, 7.16 mmol) was dissolved in methanol (40 mL), and 10% palladium on carbon (0.15 g) was added under the protection of nitrogen. The reaction system was then subjected to hydrogen balloon replacement three times, and the reaction was carried out under hydrogen atmosphere for 2 hours and monitored by LCMS. The reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain 1.51 g of the title compound.

[0544] The structural property data were as follows: ESI-MS (m / z): 268.1 [M+H] + .

[0545] Step 3: Synthesis of N-(3-fluoro-4,7-dimethyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide 4-(5-Acetylamino-3-fluoro-2-methylphenyl)-2-methylbutanoic acid (1.50 g, 5.61 mmol) was weighed and dissolved in trifluoroacetic acid (20 mL), cooled to 5 °C, and then trifluoroacetic anhydride (2.36 g, 11.22 mmol) was added dropwise. After the addition, the reaction was carried out at 5 °C for 2 h and monitored by LCMS. The reaction solution was slowly poured into saturated aqueous sodium bicarbonate solution and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was purified using a flash silica gel column (ethyl acetate:petroleum ether = 0 to 30%) to obtain 1.05 g of the title compound.

[0546] The structural property data were as follows: ESI-MS (m / z): 250.1 [M+H] + .

[0547] Step 4: Synthesis of N-(7-bromo-3-fluoro-4,7-dimethyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide N-(3-fluoro-4,7-dimethyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (0.55 g, 2.21 mmol) was weighed and dissolved in acetic acid (8 mL), and then bromine (387.85 mg, 2.43 mmol) was added. After the addition, the temperature was raised to 50 °C, and the reaction was carried out for 15 hours and monitored by LCMS. The reaction solution was directly evaporated to dryness under reduced pressure to obtain the crude product, which was purified by flash silica gel column (ethyl acetate:petroleum ether = 0-30%) to obtain 461.00 mg of the title compound.

[0548] The structural property data were as follows: ESI-MS (m / z): 328.0 [M+H] + .

[0549] Step 5: Synthesis of N-(7-azido-3-fluoro-4,7-dimethyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide N-(7-bromo-3-fluoro-4,7-dimethyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (460.00 mg, 1.40 mmol) was weighed and dissolved in N,N-dimethylformamide (10 mL). Sodium azide (273.37 mg, 4.21 mmol) was then added. After the addition, the reaction was carried out at room temperature for 1 hour and monitored by LCMS. The reaction solution was slowly poured into water and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated to dryness under reduced pressure to give the crude product, which was purified using a flash silica gel column (ethyl acetate:petroleum ether = 0-50%) to give 347.00 mg of the title compound.

[0550] The structural property data were as follows: ESI-MS (m / z): 291.1 [M + H] + .

[0551] Step 6: Synthesis of N-(7-amino-3-fluoro-4,7-dimethyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide N-(7-azido-3-fluoro-4,7-dimethyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (347.00 mg, 1.20 mmol) was weighed and dissolved in tetrahydrofuran (10 mL). Under nitrogen protection, 10% palladium on carbon (30.00 mg) was added. The reaction system was then subjected to three hydrogen balloon replacements. The reaction was carried out under a hydrogen atmosphere for 2 hours and monitored by LCMS. The reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain the crude product. This was purified using a C18 reverse-phase column (acetonitrile: 0.05% formic acid aqueous solution = 0% to 30%) to obtain 205.00 mg of the title compound.

[0552] The structural property data were as follows: ESI-MS (m / z): 265.1 [M+H] + .

[0553] Step 7: Synthesis of (9H-fluoren-9-yl)methyl (8-acetylamino-6-fluoro-2,5-dimethyl-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate N-(7-amino-3-fluoro-4,7-dimethyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (200.00 mg, 756.73 μmol) was weighed and dissolved in a mixed solvent of 1,4-dioxane (6 mL) and water (3 mL), then sodium bicarbonate (254.28 mg, 3.03 mmol) and 9-fluorenylmethyl-N-succinimidocarbonate (650.55 mg, 1.14 mmol) were added, after the addition, the reaction was stirred at room temperature for 2 hours and monitored by LCMS. The reaction solution was slowly poured into water and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated to dryness under reduced pressure to obtain a crude product, which was purified using a C18 reverse-phase column (acetonitrile: 0.05% formic acid aqueous solution = 20% to 80%) to obtain 301.00 mg of the title compound.

[0554] The structural property data were as follows: ESI-MS (m / z): 487.0 [M+H] + .

[0555] Step 8: Synthesis of (9H-fluoren-9-yl)methyl (8-amino-6-fluoro-2,5-dimethyl-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate (9H-Fluoren-9-yl)methyl (8-acetylamino-6-fluoro-2,5-dimethyl-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl) carbamate (101.00 mg, 207.59 μmol) was dissolved in 1,4-dioxane (5 mL), and then 3 mol / L aqueous hydrochloric acid (5 mL) was added. The temperature was raised to 50 ° C., and the reaction was carried out for 15 hours and monitored by LCMS. The reaction solution was slowly poured into saturated aqueous sodium bicarbonate solution, and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product, which was purified by flash silica gel column (methanol:dichloromethane = 0% to 5%) to obtain 71.00 mg of the title compound.

[0556] The structural property data were as follows: ESI-MS (m / z): 445.2 [M+H] + .

[0557] Step 9: Synthesis of (9H-fluoren-9-yl)methyl ((9S)-9-ethyl-5-fluoro-9-hydroxy-1,4-dimethyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)carbamate (9H-Fluoren-9-yl)methyl (8-amino-6-fluoro-2,5-dimethyl-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate (35.00 mg, 132.96 μmol) and (S)-4-ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolazine-3,6,10(4H)-trione (49.25 mg, 110.80 μmol) were added to toluene (3 mL), followed by p-toluenesulfonic acid (19.08 mg, 110.80 μmol), and the temperature was raised to 140 ° C. and the reaction was carried out for 4 hours. The liquid reaction solution was directly evaporated to dryness under reduced pressure at 140 ° C. to obtain the crude product. The crude product was purified using a C18 reverse phase column (acetonitrile:0.05% aqueous formic acid solution=20% to 80%) to give 21.00 mg of the title compound.

[0558] The structural property data were as follows: ESI-MS (m / z): 672.2 [M+H] + .

[0559] Step 10: Synthesis of (1S,9S)-1-amino-9-ethyl-5-fluoro-9-hydroxy-1,4-dimethyl-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione and (1R,9S)-1-amino-9-ethyl-5-fluoro-9-hydroxy-1,4-dimethyl-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione (9H-Fluoren-9-yl)methyl ((9S)-9-ethyl-5-fluoro-9-hydroxy-1,4-dimethyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)carbamate (21.00 mg, 31.26 μmol) was dissolved in N,N-dimethylformamide (1 mL), then diethylamine (0.2 mL) was added and the reaction was carried out at room temperature for 0.5 h and monitored by LCMS. The reaction solution was distilled under reduced pressure to remove ethylenediamine, and then the pH was adjusted to 2-3 with 1 mol / L aqueous hydrochloric acid. The reaction solution was directly purified by preparative high-performance liquid chromatography to obtain two isomers, 5-13-A (1.30 mg) and 5-13-B (1.68 mg).

[0560] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 24]

[0561] The structural characterization data for 5-13-A (6 min LCMS, earlier peak with retention time 1.373 min) was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 7.88 (d, J = 10.6Hz, 1H), 7.36 (s, 1H), 6.58 (s, 1H), 5.60 (d, J = 3.5 Hz, 2H), 5.46 (d, J = 2.5Hz, 2H), 3.25-3.17 (m, 2H), 2.41 (s, 3H), 2.38-2.28 (m, 2H), 1.91-1.84 (m, 2H),1.79 (s, 3H)), 0.88 (t, J = 7.3 Hz, 3H). ESI-MS (m / z): 450.2 [M+H] + . The structural characterization data for 5-13-B (6 min LCMS, later peak with retention time 1.523 min) was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 7.76 (d, J = 10.8Hz, 1H), 7.30 (s, 1H), 6.52 (s, 1H), 5.73 (d, J = 19.8 Hz, 1H), 5.50 -5.40 (m,3H), 3.26-3.17 (m, 1H), 3.08-2.96 (m, 1H), 2.38 (s, 3H), 2.19-2.11 (m, 1H),2.04 (td, J = 13.0, 5.1 Hz, 1H), 1.91-1.79 (m, 2H), 1.34 (s, 3H), 0.87 (t, J =7.3 Hz, 3H). ESI-MS (m / z): 450.2 [M+H] + .

[0562] 6 min LCMS conditions: Chromatography column: Waters SunFire C18 OBD 4.6 mm x 50 mm x 5.0 μm Mobile phase A: 0.05% acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 25]

[0563] Example 13: (1S,9S)-1-(aminomethyl)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione and (1R,9S)-1-(aminomethyl)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione [ka]

[0564] Step 1: Synthesis of N-(7-((dimethylamino)methylene)-3-fluoro-4-methyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide Compound 5-13-04 (1.00 g, 4.25 mmol) was dissolved in N,N-dimethylformamide dimethyl acetal (10 mL) and then heated to 120 ° C. for 3 hours. The reaction was monitored by LCMS. After the reaction solution was cooled to room temperature, it was directly evaporated to dryness under reduced pressure to obtain the crude product, which was purified by flash silica gel column (ethyl acetate:petroleum ether = 20% to 100%) to obtain 891.00 mg of the title compound.

[0565] The structural property data were as follows: ESI-MS (m / z): 291.1 [M + H] + .

[0566] Step 2: Synthesis of N-(7-(aminomethylene)-3-fluoro-4-methyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide Compound 5-7-01 (0.89 g, 3.07 mmol) was dissolved in ethanol (25 mL), and then ammonium acetate (2.36 g, 30.65 mmol) was added. After the addition, the reaction was carried out at room temperature for 16 hours and monitored by LCMS. The reaction solution was evaporated to dryness under reduced pressure, and then dichloromethane (30 mL) and water (20 mL) were added, stirred, and allowed to stand. The organic phase was separated, dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated to dryness under reduced pressure to give 785.00 mg of the title compound.

[0567] The structural property data were as follows: ESI-MS (m / z): 263.1 [M+H] + .

[0568] Step 3: Synthesis of N-(7-(aminomethyl)-3-fluoro-4-methyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide Compound 5-7-02 (0.80 g, 3.05 mmol) was dissolved in ethanol (200 mL), and then 10% palladium on carbon (0.40 mg) and concentrated hydrochloric acid (0.2 mL) were added. The reaction system was subjected to hydrogen balloon replacement three times. The reaction was then carried out under a hydrogen atmosphere at room temperature for 3 hours and monitored by LCMS. The reaction solution was directly filtered, and the filtrate was evaporated to dryness under reduced pressure to give 905.00 mg of the hydrochloride salt of the title compound.

[0569] The structural property data were as follows: ESI-MS (m / z): 265.1 [M+H] + .

[0570] Step 4: Synthesis of (9H-fluoren-9-yl)methyl ((8-acetamido-6-fluoro-5-methyl-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)methyl)carbamate The hydrochloride salt of compound 5-7-03 (0.90 g, 2.99 mmol) was dissolved in 1,4-dioxane (20 mL), followed by the addition of sodium bicarbonate (1.01 g, 11.97 mmol), water (10 mL), and 9-fluorenylmethyl-N-succinimidyl carbonate (1.21 g, 3.59 mmol). After the addition, the reaction was stirred at room temperature for 1 hour and monitored by LCMS. The reaction solution was poured into water and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was dried under reduced pressure to give the crude product. The crude product was purified using a flash silica gel column (ethyl acetate:petroleum ether = 0-50%) to give 1.30 g of the title compound.

[0571] The structural property data were as follows: ESI-MS (m / z): 487.1 [M+H] + .

[0572] Step 5: Synthesis of (9H-fluoren-9-yl)methyl ((8-amino-6-fluoro-5-methyl-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)methyl)carbamate Compound 5-7-04 (0.80 g, 1.64 mmol) was dissolved in 1,4-dioxane (20 mL) and 3 mol / L aqueous hydrochloric acid (20 mL) was added under nitrogen protection. After the addition, the temperature was raised to 60 °C, and the reaction was carried out for 15 hours and monitored by LCMS. The reaction solution was slowly poured into water and then extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was purified using a flash silica gel column (ethyl acetate:petroleum ether = 0-40%) to obtain 561.00 mg of the title compound.

[0573] The structural property data were as follows: ESI-MS (m / z): 445.1 [M+H] + .

[0574] Step 6: Synthesis of (9H-fluoren-9-yl)methyl (((9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)methyl)carbamate (S)-4-Ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (597.00 mg, 2.27 mmol) and compound 5-7-05 (840.00 mg, 1.89 mmol) were added to toluene (60 mL), followed by p-toluenesulfonic acid (325.00 mg, 1.89 mmol). After the addition, the temperature was raised to 140 °C and the reaction was carried out for 4 hours. The reaction solution was then directly evaporated to dryness under reduced pressure at 140 °C to obtain the crude product. The crude product was purified using a flash silica gel column (methanol:dichloromethane = 0-5%) to obtain 563.00 mg of the title compound.

[0575] The structural property data were as follows: ESI-MS (m / z): 672.2 [M+H] + .

[0576] Step 7: Synthesis of (1S,9S)-1-(aminomethyl)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione and (1R,9S)-1-(aminomethyl)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione Compound 5-7-06 (454.00 mg, 675.89 μmol) was dissolved in N,N-dimethylformamide (5 mL), and then diethylamine (1 mL) was added and reacted at room temperature for 0.5 hours. The reaction was monitored by LCMS. After evaporating ethylenediamine from the reaction solution under reduced pressure, the pH was adjusted to 2-3 with formic acid. The reaction solution was directly purified by preparative high-performance liquid chromatography, and the fractions were lyophilized to give the title compounds 5-7-A (32.00 mg) and 5-7-B (56.00 mg).

[0577] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 26]

[0578] The structural characterization data of 5-7-A (6 min LCMS, earlier peak with retention time 1.488 min) was as follows: 1HNMR (400 MHz, DMSO-d6) δ 7.88 (d, J = 10.6Hz, 1H), 7.36 (s, 1H), 6.58 (s, 1H), 5.60 (d, J = 3.5 Hz, 2H), 5.46 (d, J = 2.5Hz, 2H), 3.25-3.17 (m, 2H), 2.41 (s, 3H), 2.38-2.28 (m, 2H), 1.91-1.84 (m, 2H),1.79 (s, 3H)), 0.88 (t, J = 7.3 Hz, 3H). ESI-MS (m / z): 450.2 [M+H] + . The structural characterization data for 5-7-B (6 min LCMS, later peak with retention time 1.596 min) was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 7.76 (d, J = 10.8Hz, 1H), 7.30 (s, 1H), 6.52 (s, 1H), 5.73 (d, J = 19.8 Hz, 1H), 5.50 -5.40 (m,3H), 3.26-3.17 (m, 1H), 3.08-2.96 (m, 1H), 2.38 (s, 3H), 2.19-2.11 (m, 1H),2.04 (td, J = 13.0, 5.1 Hz, 1H), 1.91- 1.79 (m, 2H), 1.34 (s, 3H), 0.87 (t, J =7.3 Hz, 3H). ESI-MS (m / z): 450.2 [M+H] + .

[0579] 6 min LCMS conditions: Chromatography column: Waters SunFire C18 OBD 4.6 mm x 50 mm x 5.0 μm Mobile phase A: 0.05% acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 27]

[0580] Example 14: N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-1,4-dimethyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-1-hydroxycyclopropane-1-carboxamide or N-((1R,9S)-9-ethyl-5-fluoro-9-hydroxy-1,4-dimethyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-1-hydroxycyclopropane-1-carboxamide [ka]

[0581] Step 1: 1-((tert-butyldiphenylsilyl)oxy)-N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-1,4-dimethyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)cyclopropane-1-carboxamide Synthesis of 1-((tert-butyldiphenylsilyl)oxy)-N-((1R,9S)-9-ethyl-5-fluoro-9-hydroxy-1,4-dimethyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)cyclopropane-1-carboxamide Compound 5-13-A (10.00 mg, 22.25 μmol) in a single configuration and 1-((tert-butyldiphenylsilyl)oxy)cyclopropane-1-carboxylic acid (11.36 mg, 33.37 μmol) were dissolved in N,N-dimethylformamide (1 mL), followed by the addition of HATU (12.68 mg, 33.37 μmol) and N,N-diisopropylethylamine (8.63 mg, 66.74 μmol). After the addition, the reaction was carried out at room temperature for 0.5 h and monitored by LCMS. The reaction solution was directly purified using a C18 reverse-phase column (acetonitrile:0.05% formic acid aqueous solution = 30% to 100%) to give the title compound 5-16-01-A (7 mg) in a single configuration.

[0582] The structural property data were as follows: ESI-MS (m / z): 772.3 [M+H] + .

[0583] Step 2: Synthesis of N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-1,4-dimethyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-1-hydroxycyclopropane-1-carboxamide or N-((1R,9S)-9-ethyl-5-fluoro-9-hydroxy-1,4-dimethyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-1-hydroxycyclopropane-1-carboxamide Compound 5-16-01-A (7.00 mg, 9.07 μmol) was dissolved in N,N-dimethylformamide (1 mL), and then potassium fluoride (2.63 mg, 45.34 μmol) was added. The temperature was then raised to 50 °C, and the reaction was carried out for 1 hour and monitored by LCMS. The reaction solution was directly purified by high-performance liquid chromatography to give the title compound 5-16-A as a single stereoisomer (1.73 mg).

[0584] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 28]

[0585] The structural property data were as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.26 (s, 1H), 7.78(d, J = 10.8 Hz, 1H), 7.30 (s, 1H), 6.56 (s, 1H), 6.52 (s, 1H), 5.52 (d, J =19.3 Hz, 1H), 5.43 (d, J = 4.4 Hz, 2H), 4.94 (d, J = 19.2 Hz, 1H), 3.30-3.24(m,1H), 3.11-3.00 (m, 1H), 2.95-2.84 (m, 1H), 2.39 (s, 3H), 1.98-1.80 (m, 3H),1.62 (s, 3H), 0.87 (t, J = 7.9 Hz, 3H). ESI-MS (m / z): 534.2 [M+H] + .

[0586] Example 15: N-((10S)-10-benzyl-1-(((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4';6,7]indolizino[1,2-b]quinolin-1-yl)amino)-1,6,9,12,15-pentaoxo-3-oxa-5,8,11,14-tetraazahexadecan-16-yl)-6-(2-(methylsulfonyl)pyrimidin-5-yl)hexadecane amide or N-((10S)-10-benzyl-1-(((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4';6,7]indolizino[1,2-b]quinolin-1-yl)amino)-1,6,9,12,15-pentoxo-3-oxa-5,8,11,14-tetraazahexadecan-16-yl)-6-(2-(methylsulfonyl)pyrimidin-5-yl)hexadecanamide [ka]

[0587] Step 1: Isolation and purification of (9S)-1-amino-5-chloro-9-ethyl-9-hydroxy-4-methyl-1,2,3,9,12,15-hexahydro-10H,13H-benzopyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione Compound 2-23 (16.00 mg) was purified by preparative high-performance liquid chromatography, and the two diastereoisomers were separated under the following purification conditions to obtain 5.10 mg of trifluoroacetate salt of 2-23-A (retention time 9.85 min) and 7.12 mg of trifluoroacetate salt of 2-23-B (retention time 10.62 min).

[0588] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% trifluoroacetic acid) Table 29

[0589] Structural characteristics: 2-23-A: 1 HNMR (400 MHz, DMSO-d6) δ 8.42 (s, 3H), 8.27 (s, 1H), 7.36 (s, 1H), 6.59 (s, 1H), 5.78-5.63 (m, 1H), 5.50- 5.36 (m, 3H),5.10-5.06 (m, 1H), 3.20-3.04 (m, 2H), 2.56 (s, 3H), 2.26-2.13 (m, 2H), 1.93-1.79 (m, 2H), 0.88 (t, J = 7.2 Hz, 3H). ESI-MS (m / z):452.1[M+H] + . 2-23-B: 1 HNMR (400 MHz, DMSO-d6) δ 8.42 (s, 3H), 8.27 (s, 1H), 7.36 (s, 1H), 6.58 (s, 1H), 5.78-5.63 (m, 1H), 5.50- 5.36 (m, 3H),5.10-5.06 (m, 1H), 3.20-3.04 (m, 2H), 2.55 (s, 3H), 2.26-2.13 (m, 2H), 1.93-1.79 (m, 2H), 0.88 (t, J = 7.2 Hz, 3H). ESI-MS (m / z):452.0[M+H] + .

[0590] Step 2: N-((10S)-10-benzyl-1-(((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4';6,7]indolizino[1,2-b]quinolin-1-yl)amino)-1,6,9,12,15-pentaoxo-3-oxa-5,8,11,14-tetraazahexadecan-16-yl)-6-(2-(methylsulfonyl)pyrimidin-5-yl)hexadecanami Synthesis of N-((10S)-10-benzyl-1-(((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4';6,7]indolizino[1,2-b]quinolin-1-yl)amino)-1,6,9,12,15-pentoxo-3-oxa-5,8,11,14-tetraazahexadecan-16-yl)-6-(2-(methylsulfonyl)pyrimidin-5-yl)hexadecanamide At 25 ° C., the trifluoroacetic acid salt of 2-23-A (34.71 mg, 61.43 μmol) was dissolved in N,N-dimethylformamide (1 mL), and 3-4-03 (49.66 mg, 73.72 μmol), HATU (35.01 mg, 92.14 μmol) and N,N-diisopropylethylamine (23.82 mg, 184.29 μmol) were added sequentially. The reaction was carried out at 25 ° C. for 0.5 hours, and the reaction was monitored by LCMS. Upon completion of the reaction, the reaction solution was purified by preparative high performance liquid chromatography (conditions were as follows). The fraction was lyophilized to obtain the title compound DL-15 (11.04 mg) with a retention time of 7.5 minutes.

[0591] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 30]

[0592] The structural property data were as follows: DL-15: ESI-MS (m / z): 1107.3 [M+H] + .

[0593] Example 16: N-((9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropyl-2-hydroxyacetamide [ka]

[0594] Step 1: Synthesis of N-((9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-cyclopropyl-2-hydroxyacetamide The formate salt of the single stereoisomer compound 3-1-A (50 mg, 109.68 μmol) and 2-cyclopropyl-2-hydroxyacetic acid (25.47 mg, 219.36 μmol) were dissolved in N,N-dimethylformamide (2 mL), and then HATU (7.57 mg, 219.36 μmol) and N,N-diisopropylethylamine (42.53 mg, 329.04 μmol) were added. After the addition, the reaction was carried out at room temperature for 0.5 h and monitored by LCMS. The reaction solution was directly purified by preparative high-performance liquid chromatography to give two isomers of the title compound (3-12-A: 12.96 mg, 3-12-B: 13.56 mg).

[0595] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 31]

[0596] The structural characterization data for 3-12-A (6 min LCMS, earlier peak) was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.44 (d, J = 9.0Hz, 1H), 8.05 (d, J = 10.2 Hz, 1H), 7.33 (s, 1H), 6.54 (s, 1H), 5.62 (q, J =6.7 Hz, 1H), 5.52 (d, J = 5.1 Hz, 1H), 5.42 (s, 2H), 5.24 (q, J = 19.2 Hz, 2H),3.61 (dd, J = 6.2, 5.1 Hz, 1H), 3.32 - 3.21 (m, 2H), 2.19 (q, J = 6.5 Hz, 2H),1.92 - 1.80 (m, 2H), 1.26 - 1.20 (m, 1H), 0.87 (t, J = 7.3 Hz, 3H), 0.57 - 0.34(m, 4H). ESI-MS (m / z): 554.0 [M+H] + . The structural characterization data for 3-12-B (6 min LCMS, slower peak) was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.43 (d, J = 8.7Hz, 1H), 8.06 (d, J = 10.3 Hz, 1H), 7.33 (s, 1H), 5.57 (q, J = 6.7 Hz, 1H),5.43 (s, 2H), 5.30 - 5.17 (m, 2H), 3.64 (d, J = 6.2 Hz, 1H), 3.29 (q, J = 6.7Hz, 2H), 2.28 - 2.13 (m, 2H), 1.93 - 1.78 (m, 2H), 1.18 - 1.08 (m, 1H), 0.87(t, J = 7.3 Hz, 3H), 0.50 - 0.29 (m, 4H). ESI-MS (m / z): 554.0 [M+H] +.

[0597] The formate salt of the other single stereoisomer, compound 3-1-B (50 mg, 109.68 μmol) and 2-cyclopropyl-2-hydroxyacetic acid (25.47 mg, 219.36 μmol) were dissolved in N,N-dimethylformamide (2 mL), and then HATU (7.57 mg, 219.36 μmol) and N,N-diisopropylethylamine (42.53 mg, 329.04 μmol) were added. After the addition, the reaction was carried out at room temperature for 0.5 hours and monitored by LCMS. The reaction solution was directly purified by preparative high-performance liquid chromatography to give two isomers of the title compound (3-12-C: 20.19 mg, 3-12-D: 18.33 mg).

[0598] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 32]

[0599] The structural characterization data for 3-12-C (6 min LCMS, earlier peak) was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.47 (d, J = 9.0Hz, 1H), 8.06 (d, J = 10.3 Hz, 1H), 7.33 (s, 1H), 6.54 (s, 1H), 5.62 (q, J =6.5 Hz, 1H), 5.53 (d, J = 5.1 Hz, 1H), 5.43 (s, 2H), 5.32 - 5.16 (m, 2H), 3.61(dd, J = 6.3, 5.1 Hz, 1H), 3.32 - 3.22 (m, 2H), 2.19 (q, J = 6.5 Hz, 2H), 1.92- 1.80 (m, 2H), 1.28 - 1.20 (m, 1H), 0.87 (t, J = 7.3 Hz, 3H), 0.54 - 0.35 (m,4H). ESI-MS (m / z): 554.0 [M+H] + . The structural characterization data for 3-12-D (6 min LCMS, slower peak) was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.44 (d, J = 8.8 Hz,1H), 8.05 (d, J = 10.2 Hz, 1H), 7.34 (s, 1H), 6.55 (s, 1H), 5.58 (q, J = 6.7Hz, 1H), 5.45 (d, J = 5.2 Hz, 1H), 5.43 (s, 2H), 5.31 - 5.14 (m, 2H), 3.65 (t,J = 5.7 Hz, 1H), 3.33 - 3.21 (m, 2H), 2.28 - 2.13 (m, 2H), 1.95 - 1.80 (m, 2H),1.16 - 1.09 (m, 1H), 0.88 (t, J = 7.3 Hz, 3H), 0.46 - 0.31 (m, 4H). ESI-MS (m / z): 554.0 [M+H] + .

[0600] Example 17 Preparation of (S)—N-((1S,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropylamine and (S)—N-((1R,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropylamine [ka]

[0601] At 25 ° C., (9S)-1-amino-4-chloro-9-ethyl-5-fluoro-9-hydroxy-2,3,12,15-tetrahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13(1H,9H)-dione (80.0 mg, 175.5 μmol) and L-lactic acid (31.6 mg, 351.0 μmol) were dissolved in DMF (3 mL), and then HATU (121.1 mg, 351.0 μmol) and DIPEA (68.0 mg, 526.5 μmol) were added. The mixture was reacted at room temperature for 2 hours. The reaction solution was directly purified by preparative high-performance liquid chromatography to give compound 3-7-A (6.1 mg, 12% yield) and compound 3-7-B (9.6 mg, 20% yield).

[0602] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 33]

[0603] The structural characterization data of compound 3-7-A (6 min LC-MS, earlier peak with retention time 2.49 min) was as follows: MS m / z (ESI): 528.2 [M+H] + 1HNMR (400 MHz, DMSO-d6) δ 8.55 (d, J = 9.2Hz, 1H), 8.06 (d, J = 10.4 Hz, 1H), 7.33 (s, 1H), 6.55 (s, 1H), 6.67 (d, J =4.8 Hz, 1H), 5.65 - 5.59 (m, 1H), 5.43 (s, 2H), 5.29 - 5.21 (m, 1H), 5.14 -5.10 (m, 1H), 4.15 - 4.10 (m,, 1H), 3.27 - 3.20 (m, 1H), 2.22 - 2.15 (m, 2H),1.92 - 1.81 (m, 2H), 1.41 (d, J = 6.8 Hz, 3H), 1.30 - 1.23 (m, 1H), 0.89 - 0.85(t, J = 7.2 Hz, 3H). The structural characterization data of compound 3-7-B (6 min LC-MS, later peak with retention time 2.50 min) was as follows: MS m / z (ESI): 528.2 [M+H] + 1 HNMR (400 MHz, DMSO-d6) δ 8.46 (d, J = 8.8Hz, 1H), 8.06 (d, J = 10.4 Hz, 1H), 7.33 (s, 1H), 6.56 (s, 1H), 5.60 - 5.53(ms, 1H), 5.51 (d, J = 5.2 Hz, 1H), 5.43 (s, 2H), 5.27 - 5.14 (m, 2H), 4.16 -4.08 (m, 1H), 3.28 - 3.22 (m, 1H), 2.22 - 2.19 (m, 2H), 1.92 - 1.81 (m, 2H),1.49 - 1.39 (m, 1H), 1.29 (d, J = 6.8 Hz, 3H), 0.87 (t, J = 7.2 Hz, 3H).

[0604] Example 18 Synthesis of N-((1S,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-1,2,3,9,10,12,13,15-octahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl-1-hydroxycyclopropylcarboxamide and N-((1R,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-1,2,3,9,10,12,13,15-octahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl-1-hydroxycyclopropylcarboxamide [ka]

[0605] (9S)-1-amino-4-chloro-9-ethyl-5-fluoro-9-hydroxy-2,3,12,15-tetrahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13(1H,9H)-dione (80 mg, 175.49 μmol) and 1-hydroxycyclopropanecarboxylic acid (35.83 mg, 350.98 μmol) were dissolved in DMF at 25°C. (2 mL), and then HATU (121.14 mg, 350.98 μmol) and DIPEA (68.04 mg, 526.47 μmol) were added. After the addition, the reaction was carried out at room temperature for 0.5 h and monitored by HPLC-MS. The reaction solution was concentrated to dryness and directly purified by preparative high performance liquid chromatography to give the title compounds 3-17-A (5.3 mg) and 3-17-B (3.5 mg).

[0606] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 34]

[0607] The structural property data were as follows: The structural characterization data for 3-17-A (6 min LCMS, earlier peak with retention time 2.657 min) was as follows: ESI-MS (m / z): 540.0 [M+H] + . 1 HNMR (400 MHz, DMSO-d6) δ 8.69 (d, J = 8.8Hz, 1H), 8.06 (d, J = 10.2 Hz, 1H), 7.35 (s, 1H), 6.57 (s, 1H), 6.32 (s, 1H),5.62 (s, 1H), 5.45 (s, 2H), 5.34 - 5.24 (m, 1H), 5.20 - 5.10(m, 1H), 2.26 (s,2H), 2.00 (s, 1H), 1.88 (s, 2H), 1.47 - 1.12 (m, 8H), 1.01 - 0.80(m, 6H). The structural characterization data for 3-17-A (6 min LCMS, later peak with retention time 2.724 min) was as follows: ESI-MS (m / z): 540.0 [M+H] + . 1 HNMR (400 MHz, DMSO-d6) δ 8.69 (d, J = 8.8 Hz,1H), 8.06 (d, J = 10.2 Hz, 1H), 7.35 (s, 1H), 6.57 (s, 1H), 6.32 (s, 1H), 5.62(s, 1H), 5.45 (s, 2H), 5.34 - 5.24 (m, 1H), 5.20 - 5.10(m, 1H), 2.26 (s, 2H),2.00 (s, 1H), 1.88 (s, 2H), 1.47 - 1.12 (m, 8H), 1.01 - 0.80(m, 6H).

[0608] Example 19: Synthesis of N-((1S,9S)-4-cyclopropyl-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-1,2,3,9,10,12,13,15-tetrahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide [ka]

[0609] Step 1: Synthesis of N-(4-chloro-3-fluoro-7-(hydroxyimino)-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide N-(4-chloro-3-fluoro-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (570 mg, 2.23 mmol) and cyclopropylboronic acid (574.56 mg, 6.69 mmol) were dissolved in 1,4-dioxane, followed by dichlorodi-tert-butyl-(4-dimethylaminophenyl)phosphine palladium(II) (480 mg, 677.97 μmol) and cesium carbonate (2.17 g, 6.69 mmol). l) was added, and under nitrogen protection, the reaction was carried out in a microwave at 115°C for 2 hours and monitored by LCMS. The reaction solution was diluted with ethyl acetate and filtered, the filtrate was extracted with ethyl acetate (30 ml x 3), the organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated to dryness under reduced pressure, and the obtained crude product was purified by column chromatography on a silica gel column (PE:EA = 1:4) to give 550 mg of the title compound.

[0610] The structural property data were as follows: ESI-MS (m / z): 262.1 [M+H] + .

[0611] Step 2: Synthesis of N-(4-cyclopropyl-3-fluoro-7-(hydroxyimino)-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide Tetrahydrofuran (30 mL) and tert-butanol (10 mL) were added to the reaction flask and cooled to 5° C. in an ice bath, followed by the addition of potassium tert-butoxide (945 mg, 8.42 mmol), followed by the addition of N-(4-chloro-3-fluoro-7-(hydroxyimino)-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (1.0 g, 3.83 mmol) dissolved in tetrahydrofuran (1 mL). After 10 minutes, isoamyl nitrite (718 mg, 6.12 mmol) was added slowly. After the addition, the reaction was carried out at 5° C. for 1 hour and monitored by LCMS. The reaction solution was quenched with saturated aqueous ammonium chloride solution (50 mL) and extracted with ethyl acetate (40 ml×3). The organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, and then filtered. The filtrate was evaporated to dryness under reduced pressure to give 1.2 g of the crude product of the title compound.

[0612] The structural property data were as follows: ESI-MS (m / z): 291.1 [M + H] + .

[0613] Step 3: Synthesis of N-(7-amino-4-cyclopropyl-3-fluoro-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide hydrochloride The crude product of N-(4-cyclopropyl-3-fluoro-7-(hydroxyimino)-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (1.2 g, 1.41 mmol) was dissolved in methanol (7.5 mL) and tetrahydrofuran (7.5 mL), and then 1 mol / L aqueous hydrochloric acid solution (7.5 mL) and 10% palladium on carbon (450 mg) were added. After the addition, the reaction system was subjected to substitution using a hydrogen balloon three times, and the reaction was carried out under a hydrogen atmosphere at room temperature for 1 hour and monitored by LCMS. The reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain 1.05 g of crude product.

[0614] The structural property data were as follows: ESI-MS (m / z): 277.1 [M + H]+ .

[0615] Step 4: Synthesis of (9H-fluoren-9-yl)methyl (8-acetamido-5-cyclopropyl-6-fluoro-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate The crude product of N-(7-amino-4-cyclopropyl-3-fluoro-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide hydrochloride (1.05 g, 3.80 mmol) was dissolved in 1,4-dioxane (10 mL), and then sodium bicarbonate (1.3 g, 15.20 mmol), water (10 mL) and 9-fluorenylmethyl-N-succinimidyl carbonate (1.54 g, 4.56 mmol) were added. After the addition, the reaction was stirred at room temperature for 2 hours and monitored by LCMS. The reaction solution was poured into water (50 mL), and then extracted with ethyl acetate (40 mL × 3). The organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by C18 reverse-phase column to obtain 2.0 g of the title compound.

[0616] The structural property data were as follows: ESI-MS (m / z): 499.2 [M + H] + .

[0617] Step 5: Synthesis of (9H-fluoren-9-yl)methyl (8-amino-5-cyclopropyl-6-fluoro-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate (9H-fluoren-9-yl) methyl (8-acetamido-5-cyclopropyl-6-fluoro-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl) carbamate (2.0 g, 3.21 mmol, 80%) was dissolved in dioxane (20 mL), then 12 mol / L concentrated hydrochloric acid (5 mL) was added, after the addition, the temperature was raised to 70 ° C, the reaction was carried out for 2 hours, monitored by LCMS, the reaction solution was poured into water (40 mL), then extracted with ethyl acetate (30 mL × 3), the organic phase was combined, washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was dried under reduced pressure to obtain crude product. The crude product was purified by column chromatography silica gel column (PE: EA = 2: 1) to obtain 740 mg of the title compound.

[0618] The structural property data were as follows: ESI-MS (m / z): 457.3 [M+H] + .

[0619] Step 6: Synthesis of (9H-fluoren-9-yl)methyl ((9S)-4-cyclopropyl-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyran[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)carbamate (S)-4-Ethyl-4-hydroxyl-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (442 mg, 1.68 mmol) and (9H-fluoren-9-yl)methyl (8-amino-5-cyclopropyl-6-fluoro-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate (640 mg, 1.40 mmol) were added to toluene (30 mL), followed by p-toluenesulfonic acid (242 mg, 1.40 mmol). After the addition, the temperature was raised to 135 ° C. and the reaction was carried out for 2 hours. The reaction solution was directly evaporated to dryness under reduced pressure at 140 ° C. to obtain the crude product. The crude product was purified by column chromatography on a silica gel column (DCM:MeOH = 33:1) to obtain 1.02 g of the title compound.

[0620] The structural property data were as follows: ESI-MS (m / z): 684.1 [M+H] + .

[0621] Step 7: Synthesis of (1S,9S)-1-amino-4-cyclopropyl-9-ethyl-5-fluoro-9-hydroxy-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione (compound 5-29-1) and (1R,9S)-1-amino-4-cyclopropyl-9-ethyl-5-fluoro-9-hydroxy-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione (9H-Fluoren-9-yl)methyl ((9S)-4-cyclopropyl-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyran[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)carbamate (1.02 g, 1.49 mmol) was dissolved in N,N-dimethylformamide (15 mL). After the addition, the reaction was carried out at room temperature for 0.5 hours and monitored by LCMS. The reaction solution was distilled under reduced pressure to remove ethylenediamine, and then the pH was adjusted to 2-3 with 1 mol / L aqueous hydrochloric acid. The reaction solution was then directly purified by preparative high performance liquid chromatography to obtain two isomers of the title compound (5-22-7-A: 60 mg; 5-22-7-B: 55 mg).

[0622] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 35]

[0623] The structural characterization data for 5-22-7-A (6 min LCMS, earlier peak with retention time 2.28 min) was as follows: ESI-MS (m / z): 462.2 [M+H] + . The structural characterization data for 5-22-7-B (6 min LCMS, later peak with retention time 2.35 min) was as follows: ESI-MS (m / z): 462.2 [M+H] + .

[0624] Step 8: Synthesis of N-((1S,9S)-4-cyclopropyl-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-1,2,3,9,10,12,13,15-tetrahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide and N-((1R,9S)-4-cyclopropyl-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-1,2,3,9,10,12,13,15-tetrahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide At 25°C, single stereoisomer compound 5-28-7-A (40 mg, 86 μmol) and glycolic acid (8 mg, 104 μmol) were dissolved in DMF (2 mL), and then HATU (40 mg, 104 μmol) and DIPEA (36 mg, 258 μmol) were added. After the addition, the reaction was carried out at room temperature for 0.5 h and monitored by LCMS. The reaction solution was directly purified by preparative high-performance liquid chromatography to give compound 5-22-A (12.5 mg).

[0625] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 36]

[0626] The structural property data were as follows: The structural characterization data for 5-22-A (6 min LCMS, earlier peak with retention time 2.540 min) was as follows: ESI-MS (m / z): 520.0 [M+H] + . 1 HNMR (400 MHz, DMSO-d6) δ 8.46 (d, J = 8.9Hz, 1H), 7.74 (d, J = 11.9 Hz, 1H), 7.30 (s, 1H), 6.53 (s, 1H), 5.64 - 5.56 (m,1H), 5.49 (t, J = 5.8 Hz, 1H), 5.42 (s, 2H), 5.19 (s, 2H), 3.96 (d, J = 5.7 Hz,2H), 2.25 - 2.10 (m, 2H), 2.04 - 1.79 (m, 4H), 1.23 (s, 2H), 1.15 - 1.05 (m,2H), 0.87 (t, J = 7.2 Hz, 3H), 0.80 - 0.70 (m, 2H).

[0627] At 25°C, single stereoisomer compound 5-28-7-B (30 mg, 65 μmol) and glycolic acid (6 mg, 78 μmol) were dissolved in DMF (2 mL), and then HATU (40 mg, 104 μmol) and DIPEA (17 mg, 130 μmol) were added. After the addition, the reaction was carried out at room temperature for 0.5 h and monitored by LCMS. The reaction solution was concentrated to dryness and directly purified by preparative high-performance liquid chromatography to give compound 5-22-B (13.83 mg).

[0628] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 37]

[0629] The structural property data were as follows: The structural characterization data for 5-22-B (6 min LCMS, earlier peak with retention time 2.612 min) was as follows: ESI-MS (m / z): 520.0 [M+H] + . 1 HNMR (400 MHz, DMSO-d6) δ 8.49 (d, J = 8.9Hz, 1H), 7.74 (d, J = 11.9 Hz, 1H), 7.31 (s, 1H), 6.53 (s, 1H), 5.60 (s, 1H),5.51 (t, J = 5.9 Hz, 1H), 5.43 (s, 2H), 5.25 - 5.13 (m, 2H), 3.97 (d, J = 5.8Hz, 2H), 2.18 (s, 2H), 2.04 - 1.91 (m, 4H), 1.90 - 1.80 (m, 1H), 1.23 (s, 6H),1.15 - 1.05(m, 2H), 0.87 (t, J = 7.2 Hz, 4H), 0.80 - 0.70 (m, 2H).

[0630] Example 20 Synthesis of (R)-3-(dimethylamino)-N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-1,2,3,9,10,12,13,15-octahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropionamide and (S)-3-(dimethylamino)-N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-1,2,3,9,10,12,13,15-octahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropionamide [ka]

[0631] Step 1: (9H-Fluoro-9-yl)methyl((S)-3-(((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-1,2,3,9,10,12,13,15-octahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)amino)-2-hydroxy-3-oxopropyl)carbamate Synthesis of (9H-fluoro-9-yl)methyl ((R)-3-(((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-1,2,3,9,10,12,13,15-octahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)amino)-2-hydroxy-3-oxopropyl)carbamate A mixture of (1S,9S)-1-amino-9-ethyl-5-fluoro-9-hydroxy-4-methyl-2,3,12,15-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13(1H,9H)-dione mesylate (72 mg, 166 μmol) and 3-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-2-hydroxypropionic acid (65 mg) was added at 25°C. g, 199 μmol) was dissolved in DMF (2 mL), and then HATU (95 mg, 250 μmol) and DIPEA (65 mg, 498 μmol) were added. After the addition, the reaction was carried out at room temperature for 0.5 h and monitored by LCMS. The reaction solution was concentrated to dryness and directly purified by preparative high performance liquid chromatography to give the title compounds (compound 1-10-1-A, 24 mg; and compound 1-10-1-B, 28 mg).

[0632] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 38]

[0633] The structural characterization data of compound 1-10-1-A (6 min LCMS, earlier peak with retention time 3.283 min) was as follows: ESI-MS (m / z): 745.4 [M+H] + . The structural characterization data of compound 1-10-1-B (6 min LCMS, later peak with retention time 3.465 min) was as follows: ESI-MS (m / z): 745.4 [M+H] + .

[0634] Step 2: Synthesis of (R)-3-amino-N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-1,2,3,9,10,12,13,15-octahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropionamide and (S)-3-amino-N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-1,2,3,9,10,12,13,15-octahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropionamide At 25°C, compound 1-10-1-B (28 mg, 37 μmol) was dissolved in DMF (2 mL), and then diethylamine (1 mL) was added. After the addition, the reaction was carried out at room temperature for 1.0 hour, and the reaction solution was concentrated to dryness to obtain 28 mg of crude product (compound 1-10-2-B), which was used directly in the next reaction.

[0635] At 25°C, compound 1-10-1-A (24 mg, 33 μmol) was dissolved in DMF (2 mL), and then diethylamine (1 mL) was added. After the addition, the reaction was carried out at room temperature for 1.0 hour, and the reaction solution was concentrated to dryness to obtain 24 mg of crude product (compound 1-10-2-A), which was used directly in the next reaction.

[0636] The structural property data were as follows: ESI-MS (m / z): 523.2 [M+H] + .

[0637] Step 3: Synthesis of (R)-3-(dimethylamino)-N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-1,2,3,9,10,12,13,15-octahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropionamide and (S)-3-(dimethylamino)-N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-1,2,3,9,10,12,13,15-octahydrobenzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropionamide At 25°C, compound 1-10-2-B (28 mg, 37 μmol, 70%) was dissolved in methanol (2 mL), and then formaldehyde solution (1 mL) was added. After the addition, the reaction was carried out at room temperature for 16.0 hours. Then sodium cyanoborohydride (7.07 mg, 96.45 μmol) was added and the reaction was carried out at room temperature for 1.0 hours. The reaction was monitored by LCMS. The reaction solution was concentrated to dryness and directly purified by preparative high performance liquid chromatography to obtain 1.3 mg of the title compound (compound 1-10B).

[0638] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: methanol; Mobile phase B: water (0.05% formic acid) [Table 39]

[0639] The structural characterization data of compound 1-10-B (6 min LCMS, earlier peak with retention time 1.937 min) was as follows: ESI-MS (m / z): 551.2 [M+H] + . 1HNMR (400 MHz, DMSO-d6) δ 8.48 (d, J = 8.6Hz, 1H), 8.31 (s, 2H), 7.80 (d, J = 11.0 Hz, 1H), 7.31 (s, 1H), 6.55 (s, 1H),5.54 (s, 1H), 5.43 (s, 2H), 5.34 (d, J = 19.2 Hz, 1H), 5.19 (d, J = 19.1 Hz,1H), 4.09 - 4.06 (m, 1H), 3.20 - 3.15 (m, 2H), 2.59 - 2.53 (m, 1H), 2.45 -2.42(m, 1H), 2.42 - 2.38 (s, 3H), 2.23 - 2.19 (d, J = 7.0 Hz, 1H), 2.13 (s, 6H),2.12 - 2.08 (m, 1H), 2.02 - 1.95 (m, 1H), 1.90 - 1.85 (m, 2H), 1.23 (s, 2H),0.88 (d, J = 7.2 Hz, 3H).

[0640] At 25°C, compound 1-10-2-A (24 mg, 33 μmol, 70%) was dissolved in methanol (2 mL), and then formaldehyde solution (1 mL) was added. After the addition, the reaction was carried out at room temperature for 16.0 hours. Then sodium cyanoborohydride (6.06 mg, 96.45 μmol) was added and the reaction was carried out at room temperature for 1.0 hours. The reaction was monitored by LCMS. The reaction solution was concentrated to dryness and directly purified by preparative high performance liquid chromatography to obtain 4.44 mg of the title compound (compound 1-10-A).

[0641] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: methanol; Mobile phase B: water (0.05% formic acid) [Table 40]

[0642] The structural characterization data of compound 1-10-A (6 min LCMS, earlier peak with retention time 1.920 min) was as follows: ESI-MS (m / z): 551.2 [M+H] + . 1 HNMR (400 MHz, DMSO-d6) δ 8.58 (d, J = 9.0Hz, 1H), 8.28 (s, 1H), 7.84 (d, J = 10.9 Hz, 1H), 7.37 (s, 1H), 6.61 (s, 1H),5.66 - 5.59 (m, 1H), 5.49 (s, 2H), 5.35 (d, J = 19.1 Hz, 1H), 5.17 (d, J = 18.9Hz, 1H), 4.23 - 4.16 (m, 1H), 3.23 (d, J = 7.8 Hz, 2H), 2.75 - 2.67 (m, 2H),2.45 (s, 3H), 2.30 (s, 6H), 2.27 - 2.17 (m, 2H), 2.14 - 1.99 (m, 1H), 1.98 -1.87 (m, 2H), 1.30 (s, 2H), 0.93 (t, J = 7.3 Hz, 3H).

[0643] Example 21: (S)-14-(2-(cyclopropylamino)ethyl)-7-ethyl-7-hydroxy-7H-[1,3]dioxolano[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(10H,13H)-dione (Compound 4-14) [ka]

[0644] Step 1: Synthesis of 2-nitro-4,5-methylenedioxyacetophenone Compound 4-14-1 (10.0 g, 60.92 mmol) was dissolved in nitromethane (100 mL), and then concentrated nitric acid (26 mL) was slowly added dropwise with stirring. The reaction was allowed to proceed at room temperature for 2 hours. TLC monitoring of the reaction indicated that a small amount of starting material remained, and the product was clear. The reaction solution was neutralized by slowly adding saturated aqueous sodium bicarbonate solution dropwise, and extracted three times with dichloromethane. The combined organic phases were washed three times with saturated brine, then dried and concentrated to give the crude product, which was purified using a silica gel column (eluent: 0-20% ethyl acetate / petroleum ether) to give 9.8 g of the title compound.

[0645] Step 2: Synthesis of 6-amino-3,4-methylenedioxyacetophenone Compound 4-14-2 (2.0 g, 9.56 mmol) was dissolved in ethyl acetate (20 mL), and then 10% palladium on carbon (0.2 g) was added. The mixture was subjected to hydrogen substitution and reacted for 4 hours with stirring under hydrogen protection. After filtration, the filtrate was concentrated under reduced pressure to give 1.7 g of the crude product of the title compound.

[0646] Step 3: Synthesis of 6-acetamido-3,4-methylenedioxyacetophenone Compound 4-14-3 (1.7 g, 9.49 mmol) was dissolved in acetic anhydride (17 mL) and stirred for 1 hour. After evaporating the solvent under reduced pressure, water was added and stirred. The solid was filtered, washed with water, and evaporated to dryness in vacuo to give 2.08 g of the crude product of the title compound.

[0647] Step 4: Synthesis of (E)-N-(6-(3-(dimethylamino)acryloyl)benzo[d][1,3]dioxol-5-yl)acetamide Compound 4-14-4 (1.88 g, 8.50 mmol) was dissolved in DMF-DMA (30 mL), heated to 120° C., and reacted for 2 hours. The solvent was evaporated under reduced pressure to give 2.33 g of the crude product of the title compound.

[0648] The structural property data were as follows: ESI-MS (m / z): 277.2[M+1]+.

[0649] Step 5: Synthesis of (E)-N-(6-(3-(cyclopropylamino)acryloyl)benzo[d][1,3]dioxol-5-yl)acetamide Compound 4-14-5 (200 mg, 0.72 mmol) was dissolved in ethanol (5 mL), and then cyclopropylamine (413.3 mg, 7.24 mmol) was added dropwise, heated to 50° C., and reacted for 16 hours. The solvent was evaporated under reduced pressure to give 208 mg of the crude product of the title compound.

[0650] The structural property data were as follows: ESI-MS (m / z): 289.2[M+1]+.

[0651] Step 6: Synthesis of N-(6-(3-(cyclopropylamino)propionyl)benzo[d][1,3]dioxol-5-yl)acetamide Compound 4-14-6 (208 mg, 0.72 mmol) was dissolved in glacial acetic acid (4 mL), and then sodium borohydride (13.65 mg, 0.36 mmol) was added while stirring in an ice-water bath. The mixture was then warmed to room temperature and reacted with stirring for 3 hours. The solvent was evaporated under reduced pressure to give 209 mg of the crude product of the title compound.

[0652] The structural property data were as follows: ESI-MS (m / z): 291.1[M+1]+.

[0653] Step 7: Synthesis of (9H-fluoren-9-yl)methyl (3-(6-acetamidobenzo[d][1,3]dioxol-5-yl)-3-oxopropyl)(cyclopropyl)carbamate Compound 4-14-7 (200 mg, 0.69 mmol) was dissolved in 1,4-dioxane (20 mL) and water (20 mL). 9-Fluorenylmethyl-N-succinimidocarbonate (395 mg, 0.68 mmol) and sodium bicarbonate (231.5 mg, 2.76 mmol) were added with stirring, and the mixture was allowed to react at room temperature for 2 hours. Water and ethyl acetate were added with stirring, and the mixture was allowed to stand for liquid separation. The organic phase was washed with saturated brine, dried, concentrated, and purified using a silica gel column (eluent: 30% ethyl acetate / petroleum ether) to give the title compound (350 mg).

[0654] Step 8: Synthesis of (9H-fluoren-9-yl)methyl (3-(6-aminobenzo[d][1,3]dioxol-5-yl)-3-oxopropyl)(cyclopropyl)carbamate Compound 4-14-8 (350 mg, 0.68 mmol) was dissolved in 1,4-dioxane (10 mL), and then 3N aqueous hydrochloric acid solution (10 mL) was added dropwise. The mixture was heated to 60° C. and reacted for 16 hours with stirring. Water and ethyl acetate were added, and the mixture was stirred and allowed to stand for liquid separation. The organic phase was washed with water, dried, concentrated, and purified using a silica gel column (eluent: 33% ethyl acetate / petroleum ether) to give 218 mg of the title compound.

[0655] Step 9: Synthesis of (S)-(9H-fluoren-9-yl)methylcyclopropyl(2-(7-ethyl-7-hydroxy-8,11-dioxo-8,10,11,13-tetrahydro-7H-[1,3]dioxolano[4,5-g]pyrano[3,4]:6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)carbamate Compound 4-14-9 (40 mg, 0.085 mmol) and (S)-4-ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (24.62 mg, 0.094 mmol) were dissolved in toluene (1 mL), and then p-toluenesulfonic acid (2.93 mg, 0.017 mmol) was added, heated to 120 ° C, and reacted for 4 hours. After concentration under reduced pressure, 59 mg of the title compound was obtained as a crude product.

[0656] The structural property data were as follows: ESI-MS (m / z): 698.1[M+1]+.

[0657] Step 10: Synthesis of (S)-14-(2-(cyclopropylamino)ethyl)-7-ethyl-7-hydroxy-7H-[1,3]dioxolano[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(10H,13H)-dione

[0658] Compound 4-14-10 (59 mg, 0.085 mmol) was dissolved in DMF (1 mL), and then diethylamine (0.5 mL) was added dropwise and allowed to react for 1 hour with stirring. The reaction solution was distilled under reduced pressure to remove diethylamine, and acidified by adding 3N hydrochloric acid dropwise. The mixture was purified by HPLC (purification conditions were as follows) and lyophilized to obtain 12.66 mg of the trifluoroacetate salt of the title compound.

[0659] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% trifluoroacetic acid) [Table 41]

[0660] The structural property data were as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.73 (s, 2H), 7.67(s, 1H), 7.57 (s, 1H), 7.26 (s, 1H), 6.54 (s, 1H), 6.33 (s, 2H), 5.44 (s, 2H),5.34 (s, 2H), 3.40 (s, 4H), 2.82 (s, 1H), 1.91 - 1.81 (m, 2H), 0.87 (t, J = 7.2Hz, 5H), 0.79 (d, J = 7.4 Hz, 2H). ESI-MS (m / z): 476.1[M+1]+.

[0661] Example 22: (S)-7-ethyl-7-hydroxy-14-(2-((2-methoxyethyl)amino)ethyl)-7H-[1,3]dioxolano[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(10H,13H)-dione (Compound 4-15) [ka]

[0662] Step 1: Synthesis of (E)-N-(6-(3-((2-methoxyethyl)amino)acryloyl)benzo[d][1,3]dioxol-5-yl)acetamide Compound 4-14-5 (200 mg, 0.72 mmol) was dissolved in ethanol (5 mL), and then 2-methoxyethylamine (543.7 mg, 7.24 mmol) was added dropwise, heated to 50 °C, and reacted for 16 hours. The reaction was monitored by LCMS, which showed that a small amount of starting material remained, and the product was clear. The solvent was evaporated under reduced pressure to give 221 mg of the title compound, which was used directly in the next reaction.

[0663] The structural property data were as follows: ESI-MS (m / z): 307.1 [M+1]+

[0664] Step 2: Synthesis of N-(6-(3-((2-methoxyethyl)amino)propionyl)benzo[d][1,3]dioxol-5-yl)acetamide Compound 4-15-1 (200 mg, 0.65 mmol) was dissolved in glacial acetic acid (4 mL), and then sodium borohydride (12.35 mg, 0.33 mmol) was added while stirring in an ice-water bath. The mixture was warmed to room temperature and reacted with stirring for 3 hours. The reaction was monitored by LCMS, which showed that the starting material had disappeared and the product was clear. The solvent was evaporated under reduced pressure to give 200 mg of the title compound, which was used directly in the next reaction.

[0665] ESI-MS (m / z): 309.1 [M+1]+

[0666] Step 3: Synthesis of (9H-fluoren-9-yl)methyl (3-(6-acetamidobenzo[d][1,3]dioxol-5-yl)-3-oxopropyl)(2-methoxyethyl)carbamate The crude product of compound 4-15-2 (200 mg, 0.65 mmol) was dissolved in 1,4-dioxane (20 mL) and water (20 mL). Then, 9-fluorenylmethyl-N-succinyliminocarbonate (372 mg, 0.65 mmol) and sodium bicarbonate (231.5 mg, 2.76 mmol) were added with stirring and the mixture was allowed to react at room temperature for 2 hours. The reaction was monitored by TLC, which showed that the starting material had disappeared and the product was clearly visible. Water and ethyl acetate were added, the mixture was stirred, and the mixture was allowed to stand for liquid separation. The organic phase was washed with saturated brine, dried, concentrated, and purified using a silica gel column (eluent: 50% ethyl acetate / petroleum ether) to give 180 mg of the title compound.

[0667] Step 4: Synthesis of (9H-fluoren-9-yl)methyl (3-(6-aminobenzo[d][1,3]dioxol-5-yl)-3-oxopropyl)(2-methoxyethyl)carbamate Compound 4-15-3 (180 mg, 0.68 mmol) was dissolved in 1,4-dioxane (5 mL), 3N aqueous hydrochloric acid solution (5 mL) was added dropwise, and the mixture was heated to 60° C. and reacted for 16 hours with stirring. Water and ethyl acetate were added, and the mixture was stirred and allowed to stand for liquid separation. The organic phase was washed with water, dried, concentrated, and purified using a silica gel column (eluent: 45% ethyl acetate / petroleum ether) to give 132 mg of the title compound.

[0668] Step 5: Synthesis of (S)-(9H-fluoren-9-yl)methyl (2-(7-ethyl-7-hydroxy-8,11-dioxo-8,10,11,13-tetrahydro-7H-[1,3]dioxolano[4,5-g]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)(2-methoxyethyl)carbamate Compound 4-15-4 (130 mg, 0.266 mmol) and rac-(4S)-4-ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10-trione (70.05 mg, 0.266 mmol) were dissolved in toluene (4 mL), and then p-toluenesulfonic acid (9.16 mg, 0.053 mmol) was added. The mixture was heated to 120 °C and reacted for 4 hours. LCMS monitoring of the reaction showed that the starting material had disappeared and the product was clear. After concentration under reduced pressure, 190 mg of the title compound was obtained as a crude product.

[0669] The structural property data were as follows: ESI-MS (m / z): 716.1 [M+1]+

[0670] Step 6: Synthesis of (S)-7-ethyl-7-hydroxy-14-(2-((2-methoxyethyl)amino)ethyl)-7H-[1,3]dioxolano[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(10H,13H)-dione Compound 4-15-5 (190 mg, 0.265 mmol) was dissolved in DMF (3 mL), and then diethylamine (2 mL) was added dropwise and reacted with stirring for 1 hour. The reaction was monitored by LCMS, which showed that the starting material had disappeared and the product was clear. The reaction solution was distilled under reduced pressure to remove diethylamine, acidified by adding 3N hydrochloric acid dropwise, purified, and lyophilized to give 99.28 mg of the title compound.

[0671] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 42]

[0672] The structural property data were as follows: 1 HNMR (400 MHz, DMSO-d6): δ 8.68 (s, 2H), 7.68(s, 1H), 7.56 (s, 1H), 7.26 (s, 1H), 6.52 (s, 1H), 6.32 (s, 2H), 5.44 (s, 2H),5.31 (s, 2H), 3.65 - 3.58 (m, 2H), 3.42 (d, J = 10.2 Hz, 2H), 3.36 (s, 3H),3.22 (d, J = 4.0 Hz, 4H), 1.94 - 1.80 (m, 2H), 0.87 (t, J = 7.3 Hz, 3H). ESI-MS (m / z): 494.2 [M+1]+

[0673] Example 23: N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methoxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide and N-((1R,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methoxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide [ka]

[0674] Step 1: Synthesis of N-(3-bromo-5-fluoro-4-methoxyphenyl)acetamide 3-Bromo-5-fluoro-4-methoxy-aniline (1.7 g, 7.73 mmol) was dissolved in tetrahydrofuran (30 mL), and then triethylamine (2.35 g, 23.18 mmol) and acetic anhydride (1.18 g, 11.59 mmol) were added. After the addition, the temperature was raised to 50° C., and the reaction was stirred for 4 hours and monitored by LCMS. After the reaction solution was cooled to room temperature, it was diluted with ethyl acetate (50 mL), and then washed once with water (30 mL) and saturated brine (30 mL), respectively. The organic phase was separated, dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product, which was slurried (using petroleum ether: ethyl acetate = 5:1) and purified to obtain 1.1 g of the title compound.

[0675] The structural property data were as follows: ESI-MS (m / z): 262.0 [M+H] + .

[0676] Step 2: Synthesis of (E)-4-(5-acetylamino-3-fluoro-2-methoxyphenyl)-3-butenoic acid N-(3-bromo-5-fluoro-4-methoxyphenyl)acetamide (1.1 g, 4.20 mmol) and 3-butenoic acid (397.47 mg, 4.62 mmol) were dissolved in 1,4-dioxahexane (20 mL) and water (5 mL), and then triethylamine (1.27 g, 12.59 mmol), tris(o-methylphenyl)phosphine (127.75 mg, 419.73 μmol) and palladium acetate (47.12 mg, 209.89 μmol) were added. After the addition, the reaction system was subjected to nitrogen substitution three times, heated to 100 ° C., and reacted for 4 hours under a nitrogen atmosphere. The reaction was monitored by LCMS. The reaction solution was cooled to room temperature, and then 1 mol / L aqueous sodium hydroxide solution (50 mL) and ethyl acetate (50 mL) were added and shaken for liquid separation. The lower aqueous phase was separated, the pH adjusted to about 3 with 4 mol / L aqueous hydrochloric acid, and then extracted with ethyl acetate (40 mL × 2). The organic phases were combined, washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated to dryness under reduced pressure to give 1.1 g of the crude product of the title compound.

[0677] The structural property data were as follows: ESI-MS (m / z): 268.1 [M+H] + .

[0678] Step 3: Synthesis of 4-(5-acetylamino-3-fluoro-2-methoxyphenyl)butanoic acid The crude product of (E)-4-(5-acetylamino-3-fluoro-2-methoxyphenyl)-3-butenoic acid (1.1 g, 4.12 mmol) was dissolved in methanol (20 mL), and then 10% palladium on carbon (100 mg) was added. After the addition, the reaction system was subjected to hydrogen substitution using a hydrogen balloon three times, and the reaction was carried out under a hydrogen atmosphere for 4 hours and monitored by LCMS. The reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain 1.05 g of the crude product of the title compound.

[0679] The structural property data were as follows: ESI-MS (m / z): 270.1 [M+H] + .

[0680] Step 4: Synthesis of N-(3-fluoro-4-methoxy-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide The crude product of 4-(5-acetylamino-3-fluoro-2-methoxyphenyl)butanoic acid (1.1 g, 4.09 mmol) was dissolved in trifluoroacetic acid (10 mL), cooled to 5 ° C., and then trifluoroacetic anhydride (4.29 g, 20.43 mmol) was slowly added. After the addition, the reaction system was allowed to warm to room temperature naturally and reacted for 2 hours. The reaction was monitored by LCMS. The reaction solution was slowly poured into water (60 mL), and then extracted with ethyl acetate (40 mL × 3). The organic phases were combined, washed with saturated aqueous sodium bicarbonate solution until neutral, then washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. This was purified by flash silica gel column (ethyl acetate: petroleum ether = 0 ~ 40%) to obtain 503 mg of the title compound.

[0681] The structural property data were as follows: ESI-MS (m / z): 252.1 [M + H] + .

[0682] Step 5: Synthesis of N-(3-fluoro-7-(hydroxyimino)-4-methoxy-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide Tetrahydrofuran (15 mL) and tert-butanol (4 mL) were added to a reaction flask, which was cooled to 5° C. in an ice bath. Potassium tert-butoxide (491.26 mg, 4.38 mmol) was added, and then N-(3-fluoro-4-methoxy-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (500 mg, 1.99 mmol) was dissolved in tetrahydrofuran (5 mL) and slowly added dropwise thereto. , followed by the addition of isoamyl nitrite (373.01 mg, 3.18 mmol) after 10 minutes. After the addition, the reaction was carried out at 5° C. for 1 hour and monitored by LCMS. The reaction solution was quenched with saturated aqueous ammonium chloride solution (50 mL) and extracted with ethyl acetate (40×2). The organic phases were combined, washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, and then filtered. The filtrate was evaporated to dryness under reduced pressure to give 550 mg of crude product of the title compound.

[0683] The structural property data were as follows: ESI-MS (m / z): 281.1 [M+H] + .

[0684] Step 6: Synthesis of N-(7-amino-3-fluoro-4-methoxy-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide The crude product of N-(3-fluoro-7-(hydroxyimino)-4-methoxy-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (520 mg, 1.86 mmol) was dissolved in a mixed solution of methanol (10 mL) and tetrahydrofuran (10 mL), and then 1 mol / L aqueous hydrochloric acid solution (3.71 mL) and 10% palladium on carbon (50 mg) were added. After the addition, the reaction system was subjected to hydrogen substitution using a hydrogen balloon three times, and the reaction was carried out under a hydrogen atmosphere at room temperature for 1 hour. The reaction was monitored by LCMS, the reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain 551 mg of crude hydrochloride salt of the title compound.

[0685] The structural property data were as follows: ESI-MS (m / z): 267.1 [M+H] + .

[0686] Step 7: Synthesis of (9H-fluoren-9-yl)methyl (8-acetamido-6-fluoro-5-methoxy-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate The crude hydrochloride salt of N-(7-amino-3-fluoro-4-methoxy-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (550 mg, 1.64 mmol) was dissolved in 1,4-dioxane (15 mL), and then sodium bicarbonate (549.45 mg, 6.54 mmol), water (5 mL) and 9-fluorenylmethyl-N-succinimidocarbonate (1.12 g, 1.96 mmol) were added. After the addition, the reaction was stirred at room temperature for 2 hours and monitored by LCMS. The reaction solution was poured into water (50 mL) and then extracted with ethyl acetate (40 mL × 2). The organic phases were combined, washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, and then filtered. The filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by C18 (acetonitrile / 0.05% aqueous formic acid, 20% acetonitrile to 100% acetonitrile) reverse phase column to give 410 mg of the title compound.

[0687] The structural property data were as follows: ESI-MS (m / z): 489.1 [M+H] + .

[0688] Step 8: Synthesis of (9H-fluoren-9-yl)methyl (8-amino-6-fluoro-5-methoxy-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate (9H-fluoren-9-yl) methyl (8-acetamido-6-fluoro-5-methoxy-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl) carbamate (410 mg, 839.29 μmol) was dissolved in dioxane (10 mL), and 12 mol / L concentrated hydrochloric acid (2 mL) was added. After the addition, the temperature was raised to 70 ° C., and the reaction was carried out for 2 hours. Monitored by LCMS. The reaction solution was poured into water (30 mL), and then extracted with ethyl acetate (30 mL × 2). The organic phases were combined, washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, and then filtered. The filtrate was dried under reduced pressure to obtain a crude product, which was purified by flash silica gel (ethyl acetate: petroleum ether = 0 ~ 60%) to obtain 351 mg of the title compound.

[0689] The structural property data were as follows: ESI-MS (m / z): 447.1 [M+H] + .

[0690] Step 9: Synthesis of (9H-fluoren-9-yl)methyl ((9S)-9-ethyl-5-fluoro-9-hydroxy-4-methoxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)carbamate (S)-4-Ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (247.64 mg, 940.71 μmol) and (9H-fluoren-9-yl)methyl (8-amino-6-fluoro-5-methoxy-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate (350 mg, 783.93 μmol) were added to toluene (15 mL), followed by p-toluenesulfonic acid (134.84 mg, 783.93 μmol). After the addition, the temperature was raised to 135 °C, and the reaction was carried out for 4 hours. The reaction solution was directly evaporated to dryness under reduced pressure at 135 °C to obtain the crude product. The crude product was purified using a flash silica gel column (methanol:dichloromethane = 0-6%) to obtain 358 mg of the title compound.

[0691] The structural property data were as follows: ESI-MS (m / z): 674.2 [M+H] + .

[0692] Step 10: Synthesis of (9S)-1-amino-9-ethyl-5-fluoro-9-hydroxy-4-methoxy-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione (9H-Fluoren-9-yl)methyl ((9S)-9-ethyl-5-fluoro-9-hydroxy-4-methoxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)carbamate (358 mg, 531.41 μmol) was dissolved in N,N-dimethylformamide (4 mL), and then diethylamine (0.4 mL) was added. After the addition, the reaction was carried out at room temperature for 0.5 hours and monitored by LCMS. The reaction solution was evaporated to dryness under reduced pressure to give the crude product, which was slurried in ethyl acetate for purification to give 220 mg of the title compound.

[0693] The structural property data were as follows: ESI-MS (m / z): 452.1 [M+H] + .

[0694] Step 11: Synthesis of N-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methoxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide and N-((1R,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methoxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide (9S)-1-amino-9-ethyl-5-fluoro-9-hydroxy-4-methoxy-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione (50 mg, 110.76 μmol) and 2-hydroxyacetic acid (16.85 mg, 221.51 μmol) were dissolved in N,N-dimethylformamide ( 2 mL), and then HATU (84.17 mg, 221.51 μmol) and N,N-diisopropylethylamine (42.94 mg, 332.27 μmol) were added. After the addition, the reaction was carried out at room temperature for 0.5 h and monitored by LCMS. The reaction solution was directly purified by preparative high-performance liquid chromatography to give two isomers (5-34-A: 6.22 mg, 5-34-B: 9.81 mg).

[0695] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) [Table 43]

[0696] The structural property data of 5-34-A was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.53 (d, J = 9.0Hz, 1H), 7.89 (d, J = 12.3 Hz, 1H), 7.33 (s, 1H), 5.58 (q, J = 7.5, 7.0 Hz,1H), 5.42 (d, J = 2.1 Hz, 2H), 5.25 - 5.09 (m, 2H), 3.98 (s, 2H), 3.96 (d, J =1.1 Hz, 3H), 3.30 - 3.10 (m, 2H), 2.15 (q, J = 7.4 Hz, 2H), 1.93 - 1.80 (m,2H), 0.87 (t, J = 7.3 Hz, 3H). ESI-MS (m / z): 510.2 [M+H] + . The structural property data for 5-34-B was as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.55 (d, J = 9.0Hz, 1H), 7.90 (d, J = 12.3 Hz, 1H), 7.34 (s, 1H), 5.59 (q, J = 7.4, 6.8 Hz,1H), 5.43 (s, 2H), 5.25 - 5.11 (m, 2H), 3.99 (s, 2H), 3.96 (d, J = 1.1 Hz, 3H),3.30 - 3.13 (m, 2H), 2.15 (q, J = 6.4 Hz, 2H), 1.93 - 1.82 (m, 2H), 0.88 (t, J= 7.3 Hz, 3H). ESI-MS (m / z): 510.2 [M+H] + .

[0697] Example 24: (2R)—N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropylamine and (2R)—N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropylamine [ka]

[0698] Step 1: Synthesis of (2R)-N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropylamine and (2R)-N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropylamine (9S)-1-amino-5-chloro-9-ethyl-9-hydroxy-4-methyl-1,2,3,9,12,15-hexahydro-10H,13H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione (50 mg, 110.64 μmol) and (2R)-2-hydroxypropanoic acid (19.93 mg, 221.29 μmol) were dissolved in N,N-dimethylformamide ( 2 mL), and then HATU (84.09 mg, 221.29 μmol) and N,N-diisopropylethylamine (42.90 mg, 331.93 μmol) were added. After the addition, the reaction was carried out at room temperature for 0.5 h and monitored by LCMS. The reaction solution was directly purified by preparative high-performance liquid chromatography to give two isomers of the title compound (2-27-A: 5.73 mg, 2-27-B: 7.59 mg).

[0699] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% trifluoroacetic acid) [Table 44]

[0700] The structural property data of 2-27-A was as follows: 1HNMR (400 MHz, DMSO-d6) δ 8.45 (d, J = 9.1Hz, 1H), 8.14 (s, 1H), 7.30 (s, 1H), 6.54 (s, 1H), 5.63 (s, 1H), 5.56 (q, J =8.0 Hz, 1H), 5.42 (s, 2H), 5.25 (d, J = 19.0 Hz, 1H), 5.08 (d, J = 19.0 Hz,1H), 4.13 (q, J = 6.7 Hz, 1H), 3.27 - 3.12 (m, 2H), 2.51 (s, 3H), 2.23 - 2.13(m, 2H), 1.92 - 1.80 (m, 2H), 1.41 (d, J = 6.8 Hz, 3H), 0.87 (t, J = 7.3 Hz, 3H). ESI-MS (m / z):524.2[M+H] + . 2-27-B's structural characteristics are as follows: 1 HNMR (400 MHz, DMSO-d6) δ 8.40 (d, J = 8.9Hz, 1H), 8.15 (s, 1H), 7.31 (s, 1H), 6.54 (s, 1H), 5.55 - 5.49 (m, 1H), 5.43(d, J = 2.2 Hz, 2H), 5.18 (q, J = 19.0 Hz, 2H), 4.13 (q, J = 6.6 Hz, 1H), 3.24- 3.12 (m, 2H), 2.51 (s, 3H), 2.22 - 2.10 (m, 2H), 1.92 - 1.82 (m, 2H), 1.30(d, J = 6.7 Hz, 3H), 0.88 (t, J = 7.3 Hz, 3H). ESI-MS (m / z):524.2[M+H] + .

[0701] Example 25 Preparation of (R)—N-((1S,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropylamine and (R)—N-((1R,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxypropylamine [ka]

[0702] At 25°C, 3-1-A (20.0 mg, 43.9 μmol) and D-lactic acid (7.90 mg, 87.8 μmol) were dissolved in DMF (1.0 mL), and then HATU (33.4 mg, 87.8 μmol) and DIPEA (17.0 mg, 131.6 μmol) were added. The mixture was allowed to react at room temperature for 2 hours. The reaction solution was concentrated to remove most of the DMF, and the residue was purified by preparative high-performance liquid chromatography to obtain compound 3-26-A (15.4 mg, 64% yield).

[0703] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) Holding time: 5.3~6.2 minutes [Table 45]

[0704] The structural property data were as follows: MS m / z (ESI): 528.2 [M+H] + 1HNMR (400 MHz, DMSO-d6) δ 8.52 (d, J = 9.2Hz, 1H), 8.05 (d, J = 10.0 Hz, 1H), 7.33 (s, 1H), 6.55 (s, 1H), 5.62 - 5.59 (m,2H), 5.43 (s, 2H), 5.28 - 5.10 (m, 2H), 4.13 - 4.11 (m, 1H), 3.41 - 3.38 (m,1H), 3.28 - 3.22 (m, 1H), 2.20 - 2.18 (m, 2H), 1.92 - 1.80 (m, 2H), 1.40 (d, J= 6.8 Hz, 3H), 0.87 (t, J = 7.2 Hz, 3H).

[0705] At 25°C, 3-1-B (20.0 mg, 43.9 μmol) and D-lactic acid (7.90 mg, 87.8 μmol) were dissolved in DMF (1.0 mL), and then HATU (33.4 mg, 87.8 μmol) and DIPEA (17.0 mg, 131.6 μmol) were added. The mixture was allowed to react at room temperature for 2 hours. The reaction solution was concentrated to remove most of the DMF, and the residue was purified by preparative high-performance liquid chromatography to obtain compound 3-26-B (4.8 mg, 20% yield).

[0706] Chromatography column: SunFire Prep C18 OBD 19mm x 150mm x 5.0μm Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid) Holding time: 7.5~8.5 minutes [Table 46]

[0707] The structural property data were as follows: MS m / z (ESI): 528.2 [M+H] + 1HNMR (400 MHz, DMSO-d6) δ 8.48 (d, J = 8.8Hz, 1H), 8.06 (d, J = 10.4 Hz, 1H), 7.34 (s, 1H), 6.57 (br, 1H), 5.61 - 5.55(m, 1H), 5.48 - 5.39 (m, 2H), 5.27 - 5.16 (m, 2H), 4.15 - 4.10 (m, 1H), 3.32 -3.21 (m, 3H), 2.23 - 2.16 (m, 2H), 1.92 - 1.81 (m, 2H), 1.30 (d, J = 6.4 Hz,3H), 0.87 (t, J = 7.2 Hz, 3H).

[0708] Biological evaluation 1. Tumor cell growth inhibition test 1. Inhibitory effect of compounds on HT29 cell proliferation (1) Cell plating: First, tumor cells HT29 were cultured in the corresponding medium, then digested with trypsin, centrifuged, resuspended, counted, and the cells were adjusted to the appropriate concentration for plating. The sources of tumor cells are listed in Table 1. [Table 47]

[0709] Co-incubation of the compounds of the present invention and tumor cells: After the cells had attached to the wall, the medium was removed and diluted bioactive molecules (compounds of the present invention) were added to the wells of the above plate and incubated for 72 hours.

[0710] In vitro cell viability detection: After incubation, 50 μL of Cell Counting-Lite™ 2.0 Reagent (Vazyme / Novazyme) was added to each well, shaken, thoroughly mixed in the dark, and reacted for 10 minutes. Then, detection was performed using a microplate reader (manufacturer: BMG, model: PHERAStar-FS) for reading. Background RLU was obtained by using cell-free medium (Cell Counting-Lite™), and vehicle RLU was obtained by using cell-containing medium (Cell Counting-Lite™). Cell inhibition rate = 1 - (sample RLU - sample RLU) / (solvent RLU - background RLU) × 100%. The half-maximal inhibitory concentration (IC50) of the compound was calculated according to a four-parameter model fitting curve, and the detection results are shown in Table 2.

[0711] (2) Data results [Table 48]

[0712] The detection results showed that the compounds of the present invention in Table 2 had strong inhibitory effects on the proliferation of HT29 colon cancer cells.

[0713] 2. Inhibitory effect of compounds on A549 cell proliferation (1) Cell plating: First, tumor cells A549 were cultured in the corresponding medium, then digested with trypsin, centrifuged, resuspended, counted, and the cells were adjusted to the appropriate concentration for plating. The sources of tumor cells are listed in Table 3. [Table 49]

[0714] Co-incubation of the compounds of the present invention and tumor cells: After the cells had attached to the wall, the medium was removed and diluted bioactive molecules (compounds of the present invention) were added to the wells of the above plate and incubated for 72 hours.

[0715] In vitro cell activity detection: After incubation, 50 μL of Cell Counting-Rite™ 2.0 Reagent (Vazyme / Novazyme) was added to each well, shaken, thoroughly mixed in the dark, and reacted for 10 minutes. Then, detection was performed using a microplate reader (manufacturer: BMG, model: PHERAStar-FS) for reading. Background RLU was obtained by using cell-free medium (Cell Counting-Rite™), and vehicle RLU was obtained by using cell-containing medium (Cell Counting-Rite™). Cell inhibition rate = 1 - (sample RLU - sample RLU) / (solvent RLU - background RLU) × 100%. The half-maximal inhibitory concentration (IC50) of the compound was calculated according to a four-parameter model fitting curve, and the detection results are shown in Table 4.

[0716] (2) Data results [Table 50]

[0717] The detection results showed that the compounds of the present invention in Table 4 had significant inhibitory effects on the proliferation of A549 human lung cancer cells.

[0718] 3. Inhibitory effect of compounds on NCI-H1806 cell proliferation (1) Cell plating: First, tumor cells HCC1806 were cultured in the corresponding medium, then digested with trypsin, centrifuged, resuspended, counted, and the cells were adjusted to the appropriate concentration for plating. The sources of tumor cells are listed in Table 5. [Table 51]

[0719] Co-incubation of the compounds of the present invention and tumor cells: After the cells had attached to the wall, the medium was removed and diluted bioactive molecules (compounds of the present invention) were added to the wells of the above plate and incubated for 72 hours.

[0720] In vitro cell viability detection: After incubation, 50 μL of Cell Counting-Rite™ 2.0 Reagent (Vazyme / Novazyme) was added to each well, shaken, thoroughly mixed in the dark, and reacted for 10 minutes. Then, detection was performed using a microplate reader (manufacturer: BMG, model: PHERAStar-FS) for reading. Background RLU was obtained by using cell-free medium (Cell Counting-Rite™), and vehicle RLU was obtained by using cell-containing medium (Cell Counting-Rite™). Cell inhibition rate = 1 - (sample RLU - sample RLU) / (solvent RLU - background RLU) × 100%. The half-maximal inhibitory concentration (IC50) of the compound was calculated according to a four-parameter model fitting curve, and the detection results are shown in Table 6.

[0721] (2) Data results [Table 52]

[0722] The detection results showed that the compounds of the present invention in Table 6 had significant inhibitory effects on the proliferation of HCC1806 human breast squamous cell carcinoma cells.

[0723] 4. Inhibitory effect of compounds on SKOV-3 cell proliferation (1) Cell plating: First, tumor cells SKOV-3 were cultured in the corresponding medium, then digested with trypsin, centrifuged, resuspended, counted, and the cells were adjusted to the appropriate concentration for plating. The sources of tumor cells are listed in Table 7. [Table 53]

[0724] Co-incubation of the compounds of the present invention and tumor cells: After the cells had attached to the wall, the medium was removed and diluted bioactive molecules (compounds of the present invention) were added to the wells of the above plate and incubated for 72 hours.

[0725] In vitro cell viability detection: After incubation, 50 μL of Cell Counting-Rite™ 2.0 Reagent (Vazyme / Novazyme) was added to each well, shaken, thoroughly mixed in the dark, and reacted for 10 minutes. Then, detection was performed using a microplate reader (manufacturer: BMG, model: PHERAStar-FS) for reading. Background RLU was obtained by using cell-free medium (Cell Counting-Rite™), and vehicle RLU was obtained by using cell-containing medium (Cell Counting-Rite™). Cell inhibition rate = 1 - (sample RLU - sample RLU) / (solvent RLU - background RLU) × 100%. The half-maximal inhibitory concentration (IC50) of the compound was calculated according to a four-parameter model fitting curve, and the detection results are shown in Table 8.

[0726] (2) Data results [Table 54]

[0727] The detection results showed that the compounds of the present invention in Table 8 had significant inhibitory effects on the proliferation of SKOV-3 human ovarian cancer cells.

[0728] 5. Inhibitory effect of compounds on NCI-H358 cell proliferation (1) Cell plating: First, tumor cells NCI-H358 were cultured in the corresponding medium, then digested with trypsin, centrifuged, resuspended, counted, and the cells were adjusted to the appropriate concentration for plating. The sources of tumor cells are listed in Table 9. [Table 55]

[0729] Co-incubation of the compounds of the present invention and tumor cells: After the cells had attached to the wall, the medium was removed and diluted bioactive molecules (compounds of the present invention) were added to the wells of the above plate and incubated for 72 hours.

[0730] In vitro cell viability detection: After incubation, 50 μL of Cell Counting-Rite™ 2.0 Reagent (Vazyme / Novazyme) was added to each well, shaken, thoroughly mixed in the dark, and reacted for 10 minutes. Then, detection was performed using a microplate reader (manufacturer: BMG, model: PHERAStar-FS) for reading. Background RLU was obtained by using cell-free medium (Cell Counting-Rite™), and vehicle RLU was obtained by using cell-containing medium (Cell Counting-Rite™). Cell inhibition rate = 1 - (sample RLU - sample RLU) / (solvent RLU - background RLU) × 100%. The half-maximal inhibitory concentration (IC50) of the compound was calculated according to a four-parameter model fitting curve, and the detection results are shown in Table 10.

[0731] (2) Data results [Table 56]

[0732] The detection results showed that the compounds of the present invention in Table 10 had significant inhibitory effects on the proliferation of NCI-H358 human non-small cell lung cancer cells.

[0733] 6. Inhibitory effect of compounds on NCI-N87 cell proliferation (1) Cell plating: First, tumor cells NCI-N87 were cultured in the corresponding medium, then digested with trypsin, centrifuged, resuspended, counted, and the cells were adjusted to the appropriate concentration for plating. The sources of tumor cells are listed in Table 11. [Table 57]

[0734] Co-incubation of the compounds of the present invention and tumor cells: After the cells had attached to the wall, the medium was removed and diluted bioactive molecules (compounds of the present invention) were added to the wells of the above plate and incubated for 72 hours.

[0735] In vitro cell viability detection: After incubation, 50 μL of Cell Counting-Rite™ 2.0 Reagent (Vazyme / Novazyme) was added to each well, shaken, thoroughly mixed in the dark, and reacted for 10 minutes. Then, a microplate reader (manufacturer: BMG, model: PHERAStar-FS) was used for reading. The background RLU was obtained by using cell-free medium (Cell Counting-Rite™), and the vehicle RLU was obtained by using cell-containing medium (Cell Counting-Rite™). Cell inhibition rate = 1 - (sample RLU - sample RLU) / (solvent RLU - background RLU) × 100%. The half-maximal inhibitory concentration (IC50) of the compound was calculated according to a four-parameter model fitting curve, and the detection results are shown in Table 12.

[0736] (2) Data results [Table 58]

[0737] The detection results showed that the compounds of the present invention in Table 12 had significant inhibitory effects on the proliferation of NCI-N87 human gastric cancer cells.

[0738] 7. Inhibitory effect of compounds on HeLa cell proliferation (1) Cell plating: First, Hela tumor cells were cultured in the corresponding medium, then digested with trypsin, centrifuged, resuspended, counted, and the cells were adjusted to the appropriate concentration for plating. The sources of tumor cells are listed in Table 13. [Table 59]

[0739] Co-incubation of the compounds of the present invention and tumor cells: After the cells had attached to the wall, the medium was removed and diluted bioactive molecules (compounds of the present invention) were added to the wells of the above plate and incubated for 72 hours.

[0740] In vitro cell viability detection: After incubation, 50 μL of Cell Counting-Rite™ 2.0 Reagent (Vazyme / Novazyme) was added to each well, shaken, mixed thoroughly in the dark, and reacted for 10 minutes. Then, a microplate reader (manufacturer: BMG, model: PHERAStar-FS) was used for reading. The background RLU was obtained by using cell-free medium (Cell Counting-Rite™), and the vehicle RLU was obtained by using cell-containing medium (Cell Counting-Rite™). Cell inhibition rate = 1 - (sample RLU - sample RLU) / (solvent RLU - background RLU) × 100%. The half-maximal inhibitory concentration (IC50) of the compound was calculated according to a four-parameter model fitting curve, and the detection results are shown in Table 14.

[0741] (2) Data results [Table 60]

[0742] The detection results showed that the compounds of the present invention in Table 14 had significant inhibitory effects on the proliferation of Hela human cervical cancer cells.

[0743] 8. Inhibitory effect of compounds on HCC70 cell proliferation (1) Cell plating: First, tumor cells HCC70 were cultured in the corresponding medium, then digested with trypsin, centrifuged, resuspended, counted, and the cells were adjusted to the appropriate concentration for plating. The sources of tumor cells are listed in Table 15. [Table 61]

[0744] Co-incubation of the compounds of the present invention and tumor cells: After the cells had attached to the wall, the medium was removed and diluted bioactive molecules (compounds of the present invention) were added to the wells of the above plate and incubated for 72 hours.

[0745] In vitro cell viability detection: After incubation, 50 μL of Cell Counting-Rite™ 2.0 Reagent (Vazyme / Novazyme) was added to each well, shaken, thoroughly mixed in the dark, and reacted for 10 minutes. Then, a microplate reader (manufacturer: BMG, model: PHERAStar-FS) was used for reading. The background RLU was obtained by using cell-free medium (Cell Counting-Rite™), and the vehicle RLU was obtained by using cell-containing medium (Cell Counting-Rite™). Cell inhibition rate = 1 - (sample RLU - sample RLU) / (solvent RLU - background RLU) × 100%. The half-maximal inhibitory concentration (IC50) of the compound was calculated according to a four-parameter model fitting curve, and the detection results are shown in Table 16.

[0746] (2) Data results [Table 62]

[0747] The detection results showed that the compounds of the present invention in Table 16 had significant inhibitory effects on the proliferation of HCC70 human breast cancer cells.

[0748] 9. Inhibitory effect of compounds on MDA-MB-231 cell proliferation (1) Cell plating: First, tumor cells MDA-MB-231 were cultured in the corresponding medium, then digested with trypsin, centrifuged, resuspended, counted, and the cells were adjusted to the appropriate concentration for plating. The sources of tumor cells are listed in Table 17. [Table 63]

[0749] Co-incubation of the compounds of the present invention and tumor cells: After the cells had attached to the wall, the medium was removed and diluted bioactive molecules (compounds of the present invention) were added to the wells of the above plate and incubated for 72 hours.

[0750] In vitro cell viability detection: After incubation, 50 μL of Cell Counting-Rite™ 2.0 Reagent (Vazyme / Novazyme) was added to each well, shaken, mixed thoroughly in the dark, and reacted for 10 minutes. Then, a microplate reader (manufacturer: BMG, model: PHERAStar-FS) was used for reading. The background RLU was obtained by using cell-free medium (Cell Counting-Rite™), and the vehicle RLU was obtained by using cell-containing medium (Cell Counting-Rite™). Cell inhibition rate = 1 - (sample RLU - sample RLU) / (solvent RLU - background RLU) × 100%. The half-maximal inhibitory concentration (IC50) of the compound was calculated according to a four-parameter model fitting curve, and the detection results are shown in Table 18.

[0751] (2) Data results [Table 64]

[0752] The detection results showed that the compounds of the present invention in Table 18 had significant inhibitory effects on the proliferation of MDA-MB-231 human breast cancer cells.

[0753] 10. Inhibitory effects of compounds on Jeko-1 cell proliferation (1) Cell plating: First, tumor cells Jeko-1 were cultured in the corresponding medium, then digested with trypsin, centrifuged, resuspended, counted, and the cells were adjusted to the appropriate concentration for plating. The sources of tumor cells are listed in Table 19. [Table 65]

[0754] Co-incubation of the compounds of the present invention and tumor cells: After the cells had attached to the wall, the medium was removed and diluted bioactive molecules (compounds of the present invention) were added to the wells of the above plate and incubated for 72 hours.

[0755] In vitro cell viability detection: After incubation, 50 μL of Cell Counting-Rite™ 2.0 Reagent (Vazyme / Novazyme) was added to each well, shaken, mixed thoroughly in the dark, and reacted for 10 minutes. Then, a microplate reader (manufacturer: BMG, model: PHERAStar-FS) was used for reading. The background RLU was obtained by using cell-free medium (Cell Counting-Rite™), and the vehicle RLU was obtained by using cell-containing medium (Cell Counting-Rite™). Cell inhibition rate = 1 - (sample RLU - sample RLU) / (solvent RLU - background RLU) × 100%. The half-maximal inhibitory concentration (IC50) of the compound was calculated according to a four-parameter model fitting curve, and the detection results are shown in Table 20.

[0756] (2) Data results [Table 66]

[0757] The detection results showed that the compounds of the present invention in Table 20 had significant inhibitory effects on the proliferation of Jeko-1 human mantle cell lymphoma cells.

[0758] 11. Inhibitory effect of compounds on MDA-MB-453 cell proliferation (1) Cell plating: First, tumor cells MDA-MB-453 were cultured in the corresponding medium, then digested with trypsin, centrifuged, resuspended, counted, and the cells were adjusted to the appropriate concentration for plating. The sources of tumor cells are listed in Table 21. [Table 67]

[0759] Co-incubation of the compounds of the present invention and tumor cells: After the cells had attached to the wall, the medium was removed and diluted bioactive molecules (compounds of the present invention) were added to the wells of the above plate and incubated for 72 hours.

[0760] In vitro cell viability detection: After incubation, 50 μL of Cell Counting-Rite™ 2.0 Reagent (Vazyme / Novazyme) was added to each well, shaken, thoroughly mixed in the dark, and reacted for 10 minutes. Then, a microplate reader (manufacturer: BMG, model: PHERAStar-FS) was used for reading. The background RLU was obtained by using cell-free medium (Cell Counting-Rite™), and the vehicle RLU was obtained by using cell-containing medium (Cell Counting-Rite™). Cell inhibition rate = 1 - (sample RLU - sample RLU) / (solvent RLU - background RLU) × 100%. The half-maximal inhibitory concentration (IC50) of the compound was calculated according to a four-parameter model fitting curve, and the detection results are shown in Table 22.

[0761] (2) Data results [Table 68]

[0762] The detection results showed that the compounds of the present invention in Table 22 had significant inhibitory effects on the proliferation of MDA-MB-453 human breast cancer cells.

[0763] The structures of comparative compound 1 and comparative compound 2 are as follows: [ka]

[0764] 2. Antibody conjugation test The preparation and conjugation of the ADC DL-15 sample was as follows: 1.036 mL of hIgG antibody (anti-chicken lysozyme antibody, 19.3 mg / mL) was diluted with 0.1 M edetate disodium solution (pH 7.6), then adjusted to pH 7.6 using 1 M NaHPO solution. 10 mM TCEP (tris(2-carboxyethyl)phosphine) solution (pH 7.6) in an amount 2.4 times the amount of the substance was added, mixed thoroughly, and allowed to stand at room temperature for 90 minutes. Compound DL-15 dissolved in dimethyl sulfoxide in an amount 5 times the amount of the substance was added to the above solution system, mixed thoroughly, and allowed to stand at room temperature for 2 hours. The buffer was exchanged with 10 mM histidine buffer solution, pH 6.0, using a NAP-5 gel column (Cytiva). Sucrose and Tween-20 were then added and mixed thoroughly to obtain the antibody-drug conjugate ADC DL-15 (1.77 mL, 8.60 mg / mL). [ka]

[0765] As shown in Tables 23 and 24, the molecular weight of ADC DL-15 was determined by LC-MS, and the calculated drug-to-antibody ratio, or DAR value, was 4.11.

[0766] [Table 69] [Table 70]

[0767] Chromatographic determination conditions: Liquid chromatography column: Thermo MAbPac RP 3.0*100mm; Mobile phase A: 0.1%FA / H2O; Mobile phase B: 0.1%FA / ACN; Flow rate: 0.25 mL / min; sample chamber temperature: 8 °C; column temperature: 60 °C; injection volume: 2 μL; [Table 71]

[0768] Mass spectrometry conditions: Mass spectrometer model: AB Sciex Triple TOF 5600+; GS1 35; GS2 35; CUR 30; TEM 350; ISVF 5500; DP 250; CE 10; accumulation time 0.5 s; m / z 600–4000; total time bins 40.

[0769] The preparation and conjugation of the ADC 3-4-04-A sample was as follows: 0.518 mL of hIgG antibody (anti-chicken lysozyme antibody, 19.3 mg / mL) was diluted with 0.1 M edetate disodium solution (pH 7.6), and then the pH was adjusted to 7.6 using 1 M NaHPO solution. 10 mM TCEP (tris(2-carboxyethyl)phosphine) solution (pH 7.6) in an amount 5.5 times the amount of the substance was added, mixed thoroughly, and allowed to stand at room temperature for 90 minutes. Compound 3-4-04-A dissolved in dimethyl sulfoxide in an amount 10 times the amount of the substance was added to the above solution system, mixed thoroughly, and allowed to stand at room temperature for 2 hours. The buffer was exchanged with a 10 mM histidine buffer solution at pH 6.0 using a NAP-5 gel column (Cytiva), and then sucrose and Tween-20 were added and mixed thoroughly to obtain the antibody-drug conjugate ADC 3-4-04-A (1.50 mL, 5.60 mg / mL). [ka]

[0770] The molecular weight of ADC 3-4-04-A was determined by LC-MS, and the calculated drug-to-antibody ratio, or DAR, value was 7.47, as shown in Tables 25 and 26. The chromatographic determination conditions were the same as those for ADC DL-15.

[0771] [Table 72] [Table 73]

[0772] The preparation and conjugation of the ADC 3-4-04-B sample was as follows: 0.518 mL of hIgG antibody (anti-chicken lysozyme antibody, 19.3 mg / mL) was diluted with 0.1 M edetate disodium solution (pH 7.6), and then the pH was adjusted to 7.6 using 1 M NaHPO solution. 10 mM TCEP (tris(2-carboxyethyl)phosphine) solution (pH 7.6) in an amount 5.5 times the amount of the substance was added, mixed thoroughly, and allowed to stand at room temperature for 90 minutes. Compound 3-4-04-B dissolved in dimethyl sulfoxide in an amount 10 times the amount of the substance was added to the above solution system, mixed thoroughly, and allowed to stand at room temperature for 2 hours. The buffer was exchanged with a 10 mM histidine buffer solution at pH 6.0 using a NAP-5 gel column (Cytiva), and then sucrose and Tween-20 were added and mixed thoroughly to obtain the antibody-drug conjugate ADC 3-4-04-B (1.50 mL, 5.60 mg / mL). [ka]

[0773] The molecular weight of ADC 3-4-04-B was determined by LC-MS, and the calculated drug-to-antibody ratio, or DAR, value was 8.04, as shown in Tables 27 and 28. The chromatographic determination conditions were the same as those for ADC DL-15.

[0774] [Table 74] [Table 75]

[0775] The above experiments demonstrated that the cytotoxic drug-linker compounds of the present invention could be successfully conjugated with antibodies to obtain antibody drug conjugates.

[0776] Although specific implementations of the present invention have been described in detail, those skilled in the art will understand that, according to all the teachings disclosed, various modifications and substitutions can be made to these details, and all of these modifications fall within the scope of protection of the present invention. The full scope of the present invention is set forth in the appended claims and their equivalents.

Claims

1. Structure of formula (III): 【Chemistry 1】 (In the above formula (III), A″ is 【Chemistry 2】 is selected from one of R ｘ’’ is hydrogen, C １～６ Alkyl, C １～６ Haloalkyl, C １～６ Alkoxyalkyl, C ３～６ selected from the group consisting of cycloalkyl and 3- to 6-membered heterocyclyl; R ｙ’’ and R ｚ’’ is hydrogen, C １～６ Alkyl, C １～６ Haloalkyl, C １～６ Alkoxyalkyl, C １～６ Alkylaminoalkyl, C ３～６ cycloalkyl, 3- to 6-membered heterocyclyl, 3- to 6-membered heterocyclylalkyl, 4- to 6-membered heterocyclyl, C ２～６ Alkenyl, C ２～６ independently selected from the group consisting of alkynyl, aryl, and heteroaryl, or R ｙ’’ and R ｚ’’ bonds with adjacent carbon atoms to form a 3- to 6-membered ring) or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof,

2. Structure of Formula (III)-1: 【Transformation 3】 (In the formula, R ｘ’’ is hydrogen, C １～６ Alkyl, C １～６ Haloalkyl, C １～６ Alkoxyalkyl, C ３～６ selected from the group consisting of cycloalkyl and 3- to 6-membered heterocyclyl; R ｙ’’ and R ｚ’’ is hydrogen, C １～６ Alkyl, C １～６ Haloalkyl, C １～６ Alkoxyalkyl, C １～６ Alkylaminoalkyl, C ３～６ cycloalkyl, 3- to 6-membered heterocyclyl, 3- to 6-membered heterocyclylalkyl, 4- to 6-membered heterocyclyl, C ２～６ Alkenyl, C ２～６ independently selected from the group consisting of alkynyl, aryl, and heteroaryl, or R ｙ’’ and R ｚ’’ bonds with adjacent carbon atoms to form a 3- to 6-membered ring) 10. The compound of claim 1, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, having the formula:

3. R ｙ’’ and R ｚ’’ is hydrogen, C １～６ Alkyl, C １～６ Haloalkyl, C １～６ Alkoxyalkyl, C ３～６ Cycloalkyl, 4- to 6-membered heterocyclyl, C ２～６ Alkenyl, C ２～６ independently selected from the group consisting of alkynyl, aryl, and heteroaryl, or R ｙ’’ and R ｚ’’ are bonded to adjacent carbon atoms to form a 3- to 6-membered ring, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof.

4. R ｙ’’ is hydrogen, and R ｚ’’ is hydrogen, C １～６ Alkyl, C ３～６ selected from the group consisting of cycloalkyl and vinyl, or R ｙ’’ and R ｚ’’ are linked to adjacent carbon atoms to form a 3- to 6-membered carbocyclic ring, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof.

5. R ｘ’’ is hydrogen; R ｙ’’ and R ｚ’’ is hydrogen or R ｙ’’ and R ｚ’’ are bonded to adjacent carbon atoms to form a 3- to 6-membered ring, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof.

6. R ｙ’’ and R ｚ’’ are bonded to adjacent carbon atoms to form a 3- to 6-membered ring, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof.

7. The following structure: 【Chemistry 4】 【change】 10. The compound of claim 1, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, having the formula:

8. The following structure: M-L-E-D Formula (VI) (In the formula, M is a linker moiety of an antibody or antigen-binding fragment thereof; L is a linker connecting linker moieties M and E; E is a structural fragment connecting L and D; D is a structural fragment of a cytotoxic drug, said cytotoxic drug being selected from the compounds of any one of claims 1 to 7. or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, wherein:

9. M has the structure shown below: 【Transformation 5】 9. The compound represented by formula (VI) of claim 8, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, selected from the group consisting of:

10. M has the structure shown below: 【Transformation 6】 9. The compound represented by formula (VI) of claim 8, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, selected from the group consisting of:

11. L is C １～６ alkylene, —N(R′)—, carbonyl, —O—, Val, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn, Val-Cit, Val-Ala, Val-Lys, Val-Lys(Ac), Phe-Lys, Phe-Lys(Ac), D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn, Ala-Ala-Ala, Val-Lys-Ala, Gly-Gly-Gly, Gly-Gly-Phe-Gly (SEQ ID NO: 1), Gly-Gly-Gly-Gly-Gly (SEQ ID NO: 2), 【Transformation 7】 (Wherein R' is hydrogen, C １～６ Alkyl or -(CH ２ CH ２ O) ｒ -containing alkyl; r is an integer selected from 1 to 10; and s is an integer selected from 1 to 10. or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof.

12. L is, 【Transformation 8】 or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, wherein the compound represented by formula (VI) of claim 8 is a compound having a structure selected from the group consisting of:

13. L has the following structure: 【Chemistry 9】 9. A compound represented by formula (VI) according to claim 8, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite or prodrug thereof, selected from:

14. E is a single bond, —NH—CH ２ -, 【Chemistry 10】 9. The compound represented by formula (VI) of claim 8, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, selected from the group consisting of:

15. E is -NH-CH ２ - or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite or prodrug thereof.

16. The structure shown below: 【Chemistry 11】 【change】 9. The compound of claim 8, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, having the formula:

17. 8. A pharmaceutical composition comprising a compound of any one of claims 1 to 7, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, and one or more pharmaceutically acceptable carriers.

18. 10. A pharmaceutical composition comprising the compound of claim 8, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, and one or more pharmaceutically acceptable carriers.

19. 17. A pharmaceutical composition comprising the compound of claim 16, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, and one or more pharmaceutically acceptable carriers.

20. 8. A compound according to any one of claims 1 to 7, or at least one of a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitroxide, isotopically labeled product, metabolite, or prodrug thereof, for use as a first therapeutic agent. A kit product comprising:

21. 10. The compound of claim 8, or at least one of its pharmaceutically acceptable salts, esters, stereoisomers, polymorphs, solvates, nitroxides, isotopically labeled products, metabolites, or prodrugs, used as a first therapeutic agent. A kit product comprising:

22. The pharmaceutical composition according to claim 17, which is used as a first therapeutic agent. A kit product comprising:

23. The pharmaceutical composition according to claim 18, which is used as a first therapeutic agent. A kit product comprising:

24. 20. The pharmaceutical composition of claim 19, which is used as a first therapeutic agent. A kit product comprising:

25. 10. A compound according to any one of claims 1 to 7, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitroxide, isotopically labeled product, metabolite or prodrug thereof, for use in the treatment of a disease associated with abnormal cell proliferation.

26. A compound according to any one of claims 1 to 7, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitroxide, isotopically labeled product, metabolite or prodrug thereof, for use in the treatment of a disease associated with abnormal cell proliferation; The compound, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, wherein the disease associated with abnormal cell proliferation is selected from the group consisting of brain tumor, lung cancer, squamous cell carcinoma, bladder cancer, gastric cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colorectal cancer, liver cancer, kidney cancer, esophageal adenocarcinoma, esophageal squamous cell carcinoma, prostate cancer, cancer of the female reproductive system, carcinoma in situ, lymphoma, neurofibroma, thyroid cancer, bone cancer, skin cancer, brain cancer, colon cancer, testicular cancer, gastrointestinal stromal tumor, prostate tumor, mast cell tumor, multiple myeloma, melanoma, glioma, or sarcoma.

27. 10. The compound of claim 8, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitroxide, isotopically labeled product, metabolite, or prodrug thereof, for use in the treatment of a disease associated with abnormal cell proliferation.

28. 10. A compound of claim 8, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitroxide, isotopically labeled product, metabolite, or prodrug thereof, for use in the treatment of a disease associated with abnormal cell proliferation; The compound, or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotopically labeled product, metabolite, or prodrug thereof, wherein the disease associated with abnormal cell proliferation is selected from the group consisting of brain tumor, lung cancer, squamous cell carcinoma, bladder cancer, gastric cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colorectal cancer, liver cancer, kidney cancer, esophageal adenocarcinoma, esophageal squamous cell carcinoma, prostate cancer, cancer of the female reproductive system, carcinoma in situ, lymphoma, neurofibroma, thyroid cancer, bone cancer, skin cancer, brain cancer, colon cancer, testicular cancer, gastrointestinal stromal tumor, prostate tumor, mast cell tumor, multiple myeloma, melanoma, glioma, or sarcoma.

29. 18. The pharmaceutical composition of claim 17 for use in the treatment of a disease associated with abnormal cell proliferation.

30. 18. The pharmaceutical composition of claim 17 for use in the treatment of a disease associated with abnormal cell proliferation, The pharmaceutical composition, wherein the disease associated with abnormal cell proliferation is selected from the group consisting of brain tumor, lung cancer, squamous cell carcinoma, bladder cancer, gastric cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colorectal cancer, liver cancer, kidney cancer, esophageal adenocarcinoma, esophageal squamous cell carcinoma, prostate cancer, cancer of the female reproductive system, carcinoma in situ, lymphoma, neurofibroma, thyroid cancer, bone cancer, skin cancer, brain cancer, colon cancer, testicular cancer, gastrointestinal stromal tumor, prostate tumor, mast cell tumor, multiple myeloma, melanoma, glioma, or sarcoma.

31. 19. The pharmaceutical composition of claim 18 for use in the treatment of a disease associated with abnormal cell proliferation.

32. 19. The pharmaceutical composition of claim 18 for use in the treatment of a disease associated with abnormal cell proliferation, The pharmaceutical composition, wherein the disease associated with abnormal cell proliferation is selected from the group consisting of brain tumor, lung cancer, squamous cell carcinoma, bladder cancer, gastric cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colorectal cancer, liver cancer, kidney cancer, esophageal adenocarcinoma, esophageal squamous cell carcinoma, prostate cancer, cancer of the female reproductive system, carcinoma in situ, lymphoma, neurofibroma, thyroid cancer, bone cancer, skin cancer, brain cancer, colon cancer, testicular cancer, gastrointestinal stromal tumor, prostate tumor, mast cell tumor, multiple myeloma, melanoma, glioma, or sarcoma.

33. 21. The kit of claim 20 for use in treating a disease associated with abnormal cell proliferation.

34. 21. A kit according to claim 20 for use in treating a disease associated with abnormal cell proliferation, comprising: the disease associated with abnormal cell proliferation is selected from the group consisting of brain cancer, lung cancer, squamous cell carcinoma, bladder cancer, gastric cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colorectal cancer, liver cancer, kidney cancer, esophageal adenocarcinoma, esophageal squamous cell carcinoma, prostate cancer, cancer of the female reproductive system, carcinoma in situ, lymphoma, neurofibroma, thyroid cancer, bone cancer, skin cancer, brain cancer, colon cancer, testicular cancer, gastrointestinal stromal tumor, prostate tumor, mast cell tumor, multiple myeloma, melanoma, glioma, or sarcoma.

35. 22. The kit of claim 21 for use in treating a disease associated with abnormal cell proliferation.

36. 22. A kit according to claim 21 for use in treating a disease associated with abnormal cell proliferation, comprising: the disease associated with abnormal cell proliferation is selected from the group consisting of brain cancer, lung cancer, squamous cell carcinoma, bladder cancer, gastric cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colorectal cancer, liver cancer, kidney cancer, esophageal adenocarcinoma, esophageal squamous cell carcinoma, prostate cancer, cancer of the female reproductive system, carcinoma in situ, lymphoma, neurofibroma, thyroid cancer, bone cancer, skin cancer, brain cancer, colon cancer, testicular cancer, gastrointestinal stromal tumor, prostate tumor, mast cell tumor, multiple myeloma, melanoma, glioma, or sarcoma.

37. 23. The kit of claim 22 for use in treating a disease associated with abnormal cell proliferation.

38. 23. A kit according to claim 22 for use in treating a disease associated with abnormal cell proliferation, comprising: the disease associated with abnormal cell proliferation is selected from the group consisting of brain cancer, lung cancer, squamous cell carcinoma, bladder cancer, gastric cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colorectal cancer, liver cancer, kidney cancer, esophageal adenocarcinoma, esophageal squamous cell carcinoma, prostate cancer, cancer of the female reproductive system, carcinoma in situ, lymphoma, neurofibroma, thyroid cancer, bone cancer, skin cancer, brain cancer, colon cancer, testicular cancer, gastrointestinal stromal tumor, prostate tumor, mast cell tumor, multiple myeloma, melanoma, glioma, or sarcoma.

39. 24. The kit of claim 23 for use in treating a disease associated with abnormal cell proliferation.

40. 24. A kit according to claim 23 for use in treating a disease associated with abnormal cell proliferation, comprising: the disease associated with abnormal cell proliferation is selected from the group consisting of brain cancer, lung cancer, squamous cell carcinoma, bladder cancer, gastric cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colorectal cancer, liver cancer, kidney cancer, esophageal adenocarcinoma, esophageal squamous cell carcinoma, prostate cancer, cancer of the female reproductive system, carcinoma in situ, lymphoma, neurofibroma, thyroid cancer, bone cancer, skin cancer, brain cancer, colon cancer, testicular cancer, gastrointestinal stromal tumor, prostate tumor, mast cell tumor, multiple myeloma, melanoma, glioma, or sarcoma.