Tetracyclic derivatives, their preparation and medical uses

Abstract

The present invention relates to tetracyclic derivatives, their preparation methods, and their pharmaceutical uses. Specifically, the present invention relates to tetracyclic derivatives represented by general formula (I), their preparation methods, and pharmaceutically acceptable salts thereof, as well as their use as therapeutic agents, particularly as K-Ras GTPase inhibitors. The definitions of each substituent in general formula (I) are the same as those in the specification. JPEG2023539188000064.jpg6581

Description

[Technical field]

[0001] [Cross reference] This application claims priority to the following Chinese patent applications: 1) Chinese patent application No. 202010847583.7, filed with the China Patent Office on August 21, 2020, with the title "Tetracyclic derivatives and their manufacturing method and medicinal use" 2) Chinese patent application No. 202011277650.2, filed with the China Patent Office on November 16, 2020, with the title "Tetracyclic derivatives, their manufacturing method and their medicinal use" 3) Chinese patent application No. 202110323813.4, filed with the China Patent Office on March 26, 2021, with the title "Tetracyclic derivatives, their preparation method and their medicinal use" 4) Chinese patent application No. 202110543513.7, filed with the China Patent Office on May 19, 2021, with the title "Tetracyclic derivatives, their preparation method and their medicinal use" 5) Chinese patent application No. 202110816014.0, filed with the China Patent Office on July 20, 2021, with the title "Tetracyclic derivatives, their manufacturing method and their medicinal use." The contents of each of the above priority applications are incorporated herein by reference in their entirety. The present invention relates to tetracyclic derivatives, processes for their preparation and pharmaceutical compositions containing said derivatives, as well as their use as therapeutic agents, in particular as K-Ras GTPase inhibitors. [Background technology]

[0002] RAS represents a series of related monomeric globular proteins (21 kDa molecular weight) that have 189 amino acids, are associated with the plasma membrane, and bind GDP or GTP. Under normal developmental and physiological conditions, RAS is activated by receiving growth factors and various other extracellular signals and is responsible for regulating functions such as cell growth, survival, migration, and differentiation. RAS acts as a molecular switch, and the on / off state of the RAS protein is determined by binding to nucleotides, such that the active signaling conformation binds GTP and the inactive conformation binds GDP. When RAS contains bound GDP, it is in a dormant or quiescent or off state and is "inactive". When cells are exposed to and respond to certain growth-promoting stimuli, RAS is induced to convert its bound GDP to GTP. Once GTP is bound, RAS is "on" and can interact with and activate other proteins (its "downstream targets"). The RAS protein itself has a very low intrinsic ability to turn itself off by hydrolyzing GTP back to GDP. To turn off RAS, exogenous proteins called GTPase-activating proteins (GAPs) are required that interact with RAS and greatly promote the conversion of GTP to GDP. Mutations in any RAS that affect its ability to interact with GAPs and convert GTP to GDP result in prolonged activation of the protein, resulting in prolonged signals that tell cells to continue growing and dividing. Thus, overactive RAS signaling can ultimately lead to cancer, as these signals cause cells to grow and divide.

[0003] Structurally, RAS proteins contain a G domain, which is responsible for the enzymatic activity of RAS, which is the binding and hydrolysis of guanine nucleotides (GTPase reaction), and also a C-terminal extension region called the CAAX box. The C-terminal extension region may be translated to target the modified protein to the membrane. The G domain is approximately 21-25 kDa in size and contains a phosphate-binding loop (P-loop). The P-loop represents the pocket of the protein that binds nucleotides and is a rigid part of the domain with conserved amino acid residues (glycine 12, threonine 26 and lysine 16) required for nucleotide binding and hydrolysis. The G domain further contains the so-called switch I region (residues 30-40) and switch II region (residues 60-76). Both the I and II regions are dynamic parts of the protein, which are often referred to as a "spring-loaded" mechanism due to their ability to convert between a resting and a loaded state. The main interactions are hydrogen bonds formed by threonine-35 with glycine-60 and the γ-phosphate of GTP, which maintain the switch I and switch II domains in their active conformations, respectively. After GTP is hydrolyzed and the phosphate is released, the two domains relax to the inactive GDP conformation.

[0004] Among the members of the RAS family, KRAS (85%) is the most commonly mutated oncogenic gene, with NRAS (12%) and HRAS (3%) being the least common. KRAS mutations are commonly found in pancreatic (95%), colorectal (45%) and lung (25%) cancers, the three most lethal cancers in the United States. KRAS mutations are also found in other cancer types, including multiple myeloma, uterine cancer, cholangiocarcinoma, gastric cancer, bladder cancer, diffuse large B-cell lymphoma, rhabdomyosarcoma, squamous cell carcinoma of the skin, cervical cancer, and testicular germ cell carcinoma, but are rarely found in breast, ovarian and brain cancers (<2%). In non-small cell lung cancer (NSCLC), KRAS G12C is the most common mutation, accounting for nearly half of all KRAS mutations, followed by G12V and G12D. In non-small cell lung cancer, increased mutation frequency of specific alleles is often due to smoking-induced representative mutations (G:C to T:A substitutions), resulting in KRAS G12C (GGT to TGT) and G12V (GGT to GTT) mutations.

[0005] Large-scale genomic studies have demonstrated that lung cancer KRAS mutations, including G12C, are mutually exclusive with other known driver oncogenic mutations in NSCLC (including EGFR, ALK, ROS1, RET, and BRAF), demonstrating the uniqueness of KRAS mutations in lung cancer. Meanwhile, KRAS mutations often occur simultaneously with certain co-mutations, such as STK11, KEAP1, and TP53, which cooperate with mutated RAS to transform cells into highly malignant and aggressive tumor cells.

[0006] These three RAS oncogenes constitute the most frequently mutated gene family in human cancer.Unfortunately, despite more than 30 years of research efforts, there is still no clinically effective anti-RAS therapy, and targeting this gene using small molecules is challenging.Therefore, there is an urgent need in the art for small molecules that target RAS (e.g., K-RAS, H-RAS and / or N-RAS) and use them to treat various diseases, such as cancer.

[0007] Currently, the clinical development competition for KRAS inhibitors is fierce both at home and abroad. Among them, MRTX-849, a KRASase inhibitor developed by Mirati Therapeutics Inc., has entered phase II clinical trials for the prevention and treatment of diseases such as advanced solid tumors, metastatic colorectal cancer, and metastatic non-small cell lung cancer. Other KRAS inhibitors, including AMG-510 (Amgen, phase 3), are also under development. Early clinical studies have shown that KRAS inhibitors can control and alleviate disease progression in patients with non-small cell lung cancer and reduce tumor size in patients with advanced lung cancer and colorectal cancer. Currently, a series of patent applications related to KRAS inhibitors have been published, including WO2020047192, WO2019099524, and WO2018217651. Although research and application of KRAS inhibitors have progressed to a certain extent, existing KRAS inhibitors are still unsatisfactory in terms of efficacy and safety, and there is still a lot of room for improvement, so research and development of new KRAS inhibitors is necessary. Summary of the Invention

[0008] The present inventors have unexpectedly found in the course of research that a tetracyclic derivative represented by the following general formula (I), or a stereoisomer, tautomer, or a pharma- ceutically acceptable salt thereof can be used as an effective KRAS inhibitor and has good efficacy and safety.

[0009] Therefore, in a first aspect, the present invention provides a tetracyclic derivative represented by the following general formula (I), or a stereoisomer, tautomer, or a pharma- ceutically acceptable salt thereof:

[0010] [ka]

[0011] During the ceremony,

[0012] [ka]

[0013] L is selected from the group consisting of a chemical bond and a C1-C6 alkylene group; wherein said alkylene group is optionally further substituted with one or more substituents selected from the group consisting of an alkyl group, a halogen, and a hydroxy group; preferably, L is selected from the group consisting of a chemical bond, -CH2-, -CH2CH2-, or -CH(CH3)-; more preferably, L is a chemical bond; X and Y each independently represent N or CR c is selected from the group consisting of Z is O and NR 6 is selected from the group consisting of

[0014] Ring A is selected from the group consisting of 5-8 membered monocyclic heterocyclyl groups and 5-10 membered bridged heterocyclyl groups; provided that said monocyclic heterocyclyl group or bridged heterocyclyl group contains one or more N, O or S(O) r Including, Ring B is a 4- to 12-membered heterocycle containing two nitrogen atoms; Ring C is selected from the group consisting of aryl groups, heteroaryl groups, and fused rings;

[0015] R a is selected from the group consisting of a hydrogen atom and a fluorine atom;

[0016] [ka]

[0017] R c is selected from the group consisting of a hydrogen atom, a halogen, an alkyl group, and an alkoxy group; provided that said alkyl or alkoxy group is optionally further substituted with one or more substituents selected from the group consisting of a halogen, a hydroxy group, a cyano group, an alkyl group, and an alkoxy group; R c is preferably a halogen, more preferably fluorine or chlorine;

[0018] R 1is selected from the group consisting of a hydrogen atom, a halogen, an alkyl group, and an alkoxy group; provided that said alkyl or alkoxy group is optionally further substituted with one or more substituents selected from the group consisting of a halogen, a hydroxy group, a cyano group, an alkyl group, and an alkoxy group; R 1 is preferably a hydrogen atom,

[0019] R 2 are the same or different and each independently represent a hydrogen atom, an alkyl group, a halogen atom, a nitro group, a cyano group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, ═O, -OR 7 , -C(O)R 7 , -C(O)OR 7 , -NHC(O)R 7 , -NHC(O)OR 7 , -NR 8 R 9 , -C(O)NR 8 R 9 , -CH2NHC(O)OR 7 , -CH2NR 8 R 9 and -S(O) r R 7 wherein the alkyl, cycloalkyl, heterocyclyl, aryl or heteroaryl group is selected from the group consisting of an alkyl group, a halogen, a nitro group, a cyano group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, an oxo group (=O), -OR 7 , -C(O)R 7 , -C(O)OR 7 , -NHC(O)R 7 , -NHC(O)OR 7 , -NR 8 R 9 , -C(O)NR 8 R 9 , -CH2NHC(O)OR 7 , -CH2NR 8 R 9 and -S(O) r R 7 and optionally further substituted with one or more substituents selected from the group consisting of:

[0020] R 3 is selected from the group consisting of an alkyl group, an aryl group, and a heteroaryl group; provided that the alkyl group, the aryl group, or the heteroaryl group is selected from the group consisting of one or more R A Optionally further substituted with R 3 is preferably a heteroaryl group,

[0021] R A are the same or different and each independently represent an alkyl group, a halogen atom, a nitro group, a cyano group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, ═O, -OR 7 , -C(O)R 7 , -C(O)OR 7 , -NHC(O)R 7 , -NHC(O)OR 7 , -NR 8 R 9 , -C(O)NR 8 R 9 , -CH2NHC(O)OR 7 , -CHNR 8 R 9 and -S(O) r R 7 wherein the alkyl, cycloalkyl, heterocyclyl, aryl or heteroaryl group is selected from the group consisting of an alkyl group, a halogen, a nitro group, a cyano group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, =O, -OR 7 , -C(O)R 7 , -C(O)OR 7 , -NHC(O)R 7 , -NHC(O)OR 7 , -NR 8 R 9 , -C(O)NR 8 R 9 , -CH2NHC(O)OR 7 , -CHNR 8 R 9 and -S(O) r R 7 with the proviso that at least one R A -S(O)r R 7 and preferably, R 3 There are two R A Heteroaryl groups further substituted with one R A is an alkyl group, and another R A -S(O) r R 7 is selected from the group consisting of

[0022] R 4 are the same or different and each independently represent a hydrogen atom, a hydroxyl group, a halogen atom, a nitro group, a cyano group, an alkyl group, an alkoxy group, a haloalkyl group, a haloalkoxy group, a deuterated alkyl group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, =O, -C(O)R 7 , -C(O)OR 7 , -OC(O)R 7 , -NR 8 R 9 , -C(O)NR 8 R 9 , -SO2NR 8 R 9 or -NR 8 C(O)R 9 Preferably, R 4 is a hydrogen atom, a methyl group, a deuterated methyl group or =O,

[0023] R 5 are the same or different and each independently selected from the group consisting of a hydrogen atom, a halogen, a hydroxyl group, an alkyl group, and an alkoxy group, and are preferably a hydrogen atom or an alkyl group; R 6 is a hydrogen atom, an alkyl group, -C(O)R 13 and -S(O)2R 13 is selected from the group consisting of R 7is selected from the group consisting of a hydrogen atom, an alkyl group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group; provided that the alkyl group, the cycloalkyl group, the heterocyclyl group, the aryl group, or the heteroaryl group is selected from the group consisting of a hydroxy group, a halogen, a nitro group, a cyano group, an alkyl group, an alkoxy group, a haloalkyl group, a haloalkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, =O, -C(O)R 10 , -C(O)OR 10 , -OC(O)R 10 , -NR 11 R 12 , -C(O)NR 11 R 12 , -SO2NR 11 R 12 and -NR 11 C(O)R 12 and optionally further substituted with one or more substituents selected from the group consisting of:

[0024] R 8 and R 9 are each independently selected from the group consisting of a hydrogen atom, a hydroxy group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group; provided that the alkyl group, the alkoxy group, the cycloalkyl group, the heterocyclyl group, the aryl group, and the heteroaryl group are each independently selected from the group consisting of a hydroxy group, a halogen, a nitro group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, =O, -C(O)R 10 , -C(O)OR 10 , -OC(O)R 10 , -NR 11 R 12 , -C(O)NR 11 R 12 , -SO2NR 11 R 12 and -NR 11 C(O)R 12 optionally further substituted with one or more substituents selected from the group consisting of:

[0025] Or, R 8 and R9 together with the atom to which they are attached form a 4-8 membered heterocyclyl group; provided that said 4-8 membered heterocyclyl group is selected from the group consisting of one or more N, O or S(O) r and the 4-8 membered heterocyclyl group is selected from the group consisting of a hydroxy group, a halogen atom, a nitro group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, ═O, —C(O)R 10 , -C(O)OR 10 , -OC(O)R 10 , -NR 11 R 12 , -C(O)NR 11 R 12 , -SO2NR 11 R 12 and -NR 11 C(O)R 12 and optionally further substituted with one or more substituents selected from the group consisting of:

[0026] R 10 , R 11 and R 12 are each independently selected from the group consisting of a hydrogen atom, an alkyl group, an amino group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group; wherein the alkyl group, the cycloalkyl group, the heterocyclyl group, the aryl group, or the heteroaryl group is optionally further substituted with one or more substituents selected from the group consisting of a hydroxy group, a halogen, a nitro group, an amino group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, a carboxy group, and a carboxylate group; R 13 is an alkyl group, preferably a methyl group;

[0027] n is 0, 1, 2 or 3; p is 0, 1 or 2; q is 0, 1 or 2; r is 0, 1 or 2.

[0028] In a preferred embodiment of the present invention, the compound represented by general formula (I), or a stereoisomer, tautomer, or pharma- ceutically acceptable salt thereof is a compound represented by the following general formula (II), or a stereoisomer, tautomer, or pharma- ceutically acceptable salt thereof:

[0029] [ka]

[0030] During the ceremony, G is O, C=O and CR d R e is selected from the group consisting of W is NR f , O and CR d R e is selected from the group consisting of The condition is that if G is O, then W is CR. d R e and W is NR f then G is C=O; R d and R e are the same or different and each independently selected from the group consisting of a hydrogen atom, a halogen, an alkyl group, and an alkoxy group, and are preferably a hydrogen atom; R f is selected from the group consisting of a hydrogen atom, an alkyl group, and a deuterated alkyl group, is preferably an alkyl group or a deuterated alkyl group, and is more preferably a methyl group or a deuterated methyl group; R 5 is a hydrogen atom or an alkyl group; provided that the alkyl group is preferably a methyl group; Ring C, R 2 , R 3 , R c The definitions of E, L and n are as described in general formula (I).

[0031] In a preferred embodiment of the present invention, the compound represented by general formula (II), or a stereoisomer, tautomer, or pharma- ceutically acceptable salt thereof, is a compound represented by the following general formula (III) or (IV), or a stereoisomer, tautomer, or pharma-ceutically acceptable salt thereof:

[0032] [ka]

[0033] In the formula, ring C, R 2 , R 3 , R 5 , R c The definitions of E, L, G, W and n are as described in general formula (II).

[0034] In a preferred embodiment of the present invention, the compound represented by general formula (I) or (II), or a stereoisomer, tautomer, or pharma- ceutically acceptable salt thereof, is a compound represented by the following general formula (V) or (VI), or a stereoisomer, tautomer, or pharma-ceutically acceptable salt thereof:

[0035] [ka]

[0036] During the ceremony, R g represents a hydrogen atom, an alkyl group, or -SR 7 is preferably a methyl group or -S-CH3; R h is a hydrogen atom or an alkyl group, preferably a methyl group or an isopropyl group; R 4 is selected from the group consisting of alkyl groups and deuterated alkyl groups, and is preferably a methyl group or a deuterated methyl group; R 7 is an alkyl group, preferably a methyl group; Ring C, R 2 , R 5 , Rc The definitions of E and n are as described in general formula (II).

[0037] In a preferred embodiment of the present invention, the compound represented by the general formula (I), (II), (III), (IV), (V) or (VI), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof is

[0038] [ka]

[0039] In a preferred embodiment of the present invention, the compound represented by the general formula (I), (II), (III), (IV), (V) or (VI), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof is

[0040] [ka]

[0041] In a preferred embodiment of the present invention, the compound represented by the general formula (I), (II), (III), (IV), (V) or (VI), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof is R c is a halogen, preferably fluorine or chlorine.

[0042] In a preferred embodiment of the present invention, the compound represented by the general formula (I), (II), (III), (IV), (V) or (VI), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof is R 2 is a hydrogen atom, a halogen atom, a hydroxyl group, an alkyl group, an alkoxy group, a cycloalkyl group, or -NR 8 R 9 wherein the alkyl, alkoxy or cycloalkyl group is selected from the group consisting of halogen, hydroxyl, alkyl, alkoxy and -NR 8 R 9and optionally further substituted with one or more substituents selected from the group consisting of: 2 is a fluorine, chlorine, bromine, a hydroxyl group, an amino group, a methyl group, an ethyl group, a trifluoromethyl group, a cyclopropyl group, and

[0043] [ka]

[0044] and even more preferably, R 2 is a hydroxy group or fluorine; R 8 and R 9 is as defined in general formula (I).

[0045] In a preferred embodiment of the present invention, the compound represented by the general formula (I), (II), (III) or (IV), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof is R 3 teeth,

[0046] [ka]

[0047] During the ceremony, R j is a hydrogen atom, halogen, nitro group, cyano group, hydroxyl group, amino group, alkyl group, alkoxy group, -SR 7 At least one R is selected from the group consisting of haloalkyl groups and haloalkoxy groups. j Ha-SR 7 R j is preferably an alkyl group and -SR 7 and more preferably a methyl group, an ethyl group, or an isopropyl group. R 7 is an alkyl group, preferably a methyl group; k is 0, 1, 2, 3, 4 or 5.

[0048] In a preferred embodiment of the present invention, the compound represented by the general formula (I), (II), (III) or (IV), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof is R 3 teeth,

[0049] [ka]

[0050] During the ceremony, R j is a hydrogen atom, halogen, nitro group, cyano group, hydroxyl group, amino group, alkyl group, alkoxy group, -SR 7 At least one Rj is selected from the group consisting of haloalkyl groups and haloalkoxy groups; 7 ;R j is preferably an alkyl group or -SR 7 and more preferably a methyl group, an ethyl group, or an isopropyl group. The conditions are: One R j Ha-SR 7 and Another R j is an alkyl group; with the proviso that the alkyl group is preferably a methyl group, an ethyl group or an isopropyl group, more preferably an isopropyl group; R 7 is an alkyl group; preferably, R 7 is a methyl group, k is 2.

[0051] In a preferred embodiment of the present invention, the compound represented by the general formula (I), (II), (III) or (IV), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof is

[0052] [ka]

[0053] In a preferred embodiment of the present invention, the compound represented by the general formula (I), or a stereoisomer, a tautomer, or a pharma- ceutically acceptable salt thereof, is 4 is selected from the group consisting of alkyl groups and deuterated alkyl groups, and is preferably a methyl group or a deuterated methyl group.

[0054] In a preferred embodiment of the present invention, in the compound represented by general formula (II), (III) or (IV), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof, G is O and W is CH2.

[0055] In a preferred embodiment of the present invention, in the compound represented by general formula (II), (III) or (IV), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof, G is CH2 and W is O.

[0056] In a preferred embodiment of the present invention, in the compound represented by general formula (II), (III) or (IV), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof, G is C=O and W is NCH3.

[0057] In a preferred embodiment of the present invention, the compound represented by the general formula (I), (II), (III), (IV), (V) or (VI), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof is R 5 is a hydrogen atom or a methyl group.

[0058] In a preferred embodiment of the present invention, the compound represented by the general formula (I), (II), (III) or (IV), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof is L is selected from the group consisting of a chemical bond, -CH2-, -CH2CH2-, and -CH(CH3)-; more preferably, L is a chemical bond.

[0059] In a preferred embodiment of the present invention, the compound represented by the general formula (I), (II), (III) or (IV), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof is L is a chemical bond and R 3 is a heteroaryl group, More preferably, L is a chemical bond and R 3 is a heteroaryl group substituted with a methylthio group (-S-CH3), Particularly preferably, L is a chemical bond and R 3 is a pyridyl group substituted with a methylthio group, Particularly preferably, L is a chemical bond and R 3 teeth

[0060] [ka]

[0061] It is. Alternatively, in a preferred embodiment of the present invention, the compound represented by the general formula (V) or (VI), or a stereoisomer, tautomer, or pharma- ceutically acceptable salt thereof, is g is -S-CH3.

[0062] In a preferred embodiment of the present invention, the compound represented by the general formula (I), (II), (III) or (IV), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof is Ring C is a bicyclic heteroaryl or naphthyl group, the bicyclic heteroaryl or naphthyl group being optionally substituted with a hydroxy or amino group; More preferably,

[0063] [ka]

[0064] Representative compounds of the present invention include, but are not limited to, the compounds set forth in the table below and their stereoisomers, tautomers, or pharma- ceutically acceptable salts:

[0065] [ka]

[0066] [ka]

[0067] Note: If there is a mismatch between a structure and the name given to that structure, the structure takes precedence.

[0068] The present invention also provides a method for producing a method for manufacturing a semiconductor device comprising the steps of:

[0069] [ka]

[0070] reacting a compound of general formula (IA) with a compound of general formula (IB) under basic conditions, optionally with further removal of a protecting group, to obtain a compound of general formula (I), 1 is a leaving group, preferably chlorine; ring A, ring B, ring C, R 1 ~R 5 The definitions of X, Y, Z, E, L, n, p and q are as described in general formula (I), or a stereoisomer, tautomer or a pharma- ceutically acceptable salt thereof.

[0071] Furthermore, the present invention relates to compounds of the general formula (IA):

[0072] [ka]

[0073] (In the formula, ring A, ring B, ring C, R 1 ~R 5The definitions of X, Y, Z, L, n, p and q are as described in general formula (I).

[0074] Representative compounds of general formula (IA) include, but are not limited to, the compounds set forth in the table below, and their stereoisomers, tautomers, or pharma- ceutically acceptable salts.

[0075] [ka]

[0076] In another aspect, the present invention provides a pharmaceutical composition comprising an effective amount of a compound represented by general formula (I), (II), (III), (IV), (V) or (VI), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof, and optionally a pharma-ceutically acceptable carrier, excipient or combination thereof.

[0077] In another aspect, the present invention provides a method for inhibiting K-Ras GTPase, the method comprising administering to a subject (including patients and healthy subjects) a pharmaceutical composition comprising an effective amount of a compound represented by general formula (I), (II), (III), (IV), (V) or (VI), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof, and optionally a pharma- ceutically acceptable carrier, excipient or combination thereof, wherein the K-Ras GTPase is preferably KRAS G12Case.

[0078] The present invention also provides the use of a compound represented by general formula (I), (II), (III), (IV), (V) or (VI), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof, or a pharmaceutical composition thereof, in the manufacture of a medicament for treating a disease mediated by a KRAS mutation. Wherein, the disease mediated by the KRAS mutation is cancer, and the cancer is selected from the group consisting of pancreatic cancer, colorectal cancer, lung cancer, multiple myeloma, uterine cancer, bile duct cancer, gastric cancer, bladder cancer, diffuse large B-cell lymphoma, rhabdomyosarcoma, cutaneous squamous cell carcinoma, cervical cancer, and testicular germ cell carcinoma, and is preferably pancreatic cancer, colorectal cancer, and lung cancer. The lung cancer is preferably non-small cell lung cancer. The KRAS mutation is preferably KRAS G12C mutation.

[0079] In another aspect, the present invention provides the use of a compound of general formula (I), (II), (III), (IV), (V) or (VI), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof, or a pharmaceutical composition thereof, in the manufacture of a K-Ras GTPase inhibitor, which is preferably a KRAS G12C inhibitor.

[0080] Another aspect of the present invention relates to a method for preventing and / or treating a disease mediated by a KRAS mutation, comprising administering to a patient a therapeutically effective amount of a compound of general formula (I), (II), (III), (IV), (V) or (VI), or in the form of a tautomer, meso-isomer, racemate, enantiomeric isomer, diastereomeric isomer or mixture thereof, or a pharma- ceutically acceptable salt thereof, or a pharmaceutical composition comprising the same, wherein the KRAS mutation is preferably a KRAS G12C mutation.

[0081] The present invention also provides the use of a compound represented by general formula (I), (II), (III), (IV), (V) or (VI), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof, or a pharmaceutical composition thereof, in the manufacture of a medicament for treating cancer. The cancer is selected from the group consisting of pancreatic cancer, colorectal cancer, lung cancer, multiple myeloma, uterine cancer, bile duct cancer, gastric cancer, bladder cancer, diffuse large B-cell lymphoma, rhabdomyosarcoma, cutaneous squamous cell carcinoma, cervical cancer, and testicular germ cell carcinoma, and is preferably pancreatic cancer, colorectal cancer, and lung cancer. Here, the lung cancer is preferably non-small cell lung cancer.

[0082] The pharmaceutical formulations of the present invention can be administered topically, orally, transdermally, rectally, vaginally, parenterally, intranasally, intrapulmonary, intraocularly, intravenously, intramuscularly, intraarterially, intrathecally, intravesically, intradermally, intraperitoneally, subcutaneously, subcorneally, or by inhalation. Pharmaceutical compositions containing the active ingredient may be in a form suitable for oral administration, such as tablets, sugar tablets, troches, water or oil suspensions, dispersible powders, dispersible granules, emulsions, hard capsules, soft capsules, or syrups or elixir. Tablets may contain the active ingredient and non-toxic pharma- ceutical acceptable excipients for suitable mixing for the manufacture of tablets.

[0083] The preparation of the present invention is preferably in the form of a unit dose. The preparation can be prepared by any method known in the pharmaceutical art. The amount of active ingredient that can be combined with a carrier material to produce a single dosage form can vary depending on the host treated and the particular mode of administration. The amount of active ingredient that can be combined with a carrier material to produce a single dosage form generally refers to the amount of compound that can produce a therapeutic effect.

[0084] Dosage forms for topical or transdermal administration of a compound of the present invention include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches and inhalants. The active compound may be mixed under sterile conditions with a pharma- ceutically acceptable carrier, and with any preservatives, buffers, or propellants that may be required.

[0085] When the compounds according to the present invention are administered to humans and animals in the form of pharmaceuticals, the compounds may be provided alone or in the form of pharmaceutical compositions, which may contain, for example, 0.1% to 99.5% (more preferably, 0.5% to 90%) of the active ingredient in combination with a pharma- ceutical acceptable carrier.

[0086] Examples of pharma- ceutically acceptable carriers include, but are not limited to, (1) sugars such as lactose, glucose, and sucrose; (2) starches such as corn starch and potato starch; (3) cellulose and its derivatives such as sodium carboxymethylcellulose, ethylcellulose, and cellulose acetate; (4) powdered trout gum; (5) malt; (6) gelatin; (7) talc; (8) excipients such as cocoa butter and suppository wax; (9) oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; (10) diols such as propylene glycol; (11) polyols such as glycerin, sorbitol, mannitol, and polyethylene glycol; (12) esters such as ethyl oleate and ethyl laurate; (13) agar; (14) buffers such as magnesium hydroxide and aluminum hydroxide; (15) alginic acid; and (16) pyrogen-free water. water), (17) isotonic saline, (18) Ringer's solution, (19) ethanol, (20) phosphate buffer, (21) targeting ligands linked to the nanoparticles such as cyclodextrins, e.g., Accurins™, and (22) other non-toxic compatible substances used in pharmaceutical formulations such as polymer-based compositions.

[0087] Examples of pharma- ceutically acceptable antioxidants include, but are not limited to, (1) water-soluble antioxidants such as ascorbic acid, cysteine ​​hydrochloride, sodium bisulfate, sodium metabisulfite, sodium sulfite, and analogs thereof; (2) oil-soluble antioxidants such as ascorbic acid palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, α-tocopherol, and analogs thereof; and (3) metal chelators such as citric acid, ethylenediaminetetraacetic acid (EDTA), sorbitol, tartaric acid, phosphoric acid, and analogs thereof. Solid dosage forms (e.g., capsules, lozenges, dragees, powders, granules, and the like) may include one or more pharma- ceutically acceptable carriers, such as sodium citrate and dicalcium phosphate, and / or (1) fillers or extenders, such as starch, lactose, sucrose, glucose, mannitol, and / or silicic acid; (2) binders, such as carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose, and / or gum arabic; (3) humectants, such as glycerin; (4) agar, calcium carbonate, oyster, or the like; Disintegrating agents such as mozzarella or tapioca starch, alginic acid, certain silicates and sodium carbonate, (5) dissolution retarders such as paraffin, (6) absorption accelerators such as quaternary ammonium compounds, (7) wetting agents such as cetyl alcohol and monoglyceride stearate, (8) absorbents such as kaolin and bentonite, (9) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate and mixtures thereof, and (10) colorants. Liquid dosage forms can include pharma- ceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs.Liquid dosage forms may contain, in addition to the active ingredient, an inert diluent commonly used in the art, such as water or other solvents; solubilizers and emulsifiers, such as ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butanediol, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil, and sesame oil), glycerin, tetrahydrofuran methanol, polyethylene glycols, and fatty acid esters of sorbitan, and mixtures thereof.

[0088] Suspensions may contain, in addition to the active compounds, suspending aids such as ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum hydroxide, bentonite, agar-agar and trant gum, and mixtures thereof.

[0089] The ointments, pastes, creams and gels may contain, in addition to the active compound, excipients such as animal and vegetable fats, oils, waxes, paraffin, starch, trant gum, cellulose derivatives, polyethylene glycols, polysiloxanes, bentonite, silicic acid, talc and zinc oxide, or mixtures thereof.

[0090] Powders and sprays can contain, in addition to the active compound, excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicates and polyamide powder, or mixtures thereof. The sprays can contain other customary propellants, such as chlorofluorohydrocarbons, and volatile unsubstituted hydrocarbons, such as butane and propane. [Brief description of the drawings]

[0091] [Figure 1] FIG. 1 is a graph showing changes in tumor volume in BALB / c nude mice bearing NCI-H358 cell xenograft tumors treated with Compound 17 according to the present invention in Test Example 6. [Diagram 2]FIG. 2 is a graph showing changes in body weight of BALB / c nude mice bearing NCI-H358 cell xenograft tumors treated with Compound 17 according to the present invention in Test Example 6. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0092] Unless otherwise specified to the contrary, several terms used in the present invention in the specification and claims are defined as follows.

[0093] By "chemical bond" it is meant that the depicted substituent is not present and that both ends of the substituent are directly joined to form a chemical bond.

[0094] An "alkyl group" is a C1-C 20 The alkyl group is preferably a C1 to C6 alkyl group. 10 and more preferably C1-C6 alkyl groups or C1-C4 alkyl groups. Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, t-butyl, s-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, and 2,3-dimethylbutyl. The alkyl groups may be substituted or unsubstituted.

[0095] The term "alkenyl group" refers to an alkyl group as defined above, which is composed of at least two carbon atoms and at least one carbon-carbon double bond. Representative examples include, but are not limited to, ethenyl, 1-propenyl, 2-propenyl, 1-, 2- or 3-butenyl groups. Preferably, the alkenyl group is a C2-C 10 More preferably, it is an alkenyl group having from C2 to C6, and most preferably, it is an alkenyl group having from C2 to C4. The alkenyl group may be optionally substituted or unsubstituted.

[0096] The term "alkynyl group" refers to an aliphatic hydrocarbon group containing one carbon-carbon triple bond, and may be either linear or branched. The alkynyl group is preferably a C2-C 10 Alkynyl groups are preferably C2-C6 alkynyl groups, more preferably C2-C4 alkynyl groups. Examples of alkynyl groups include, but are not limited to, ethynyl, 1-propynyl, 2-propynyl, 1-, 2- or 3-butynyl groups. The alkynyl group may be substituted or unsubstituted.

[0097] The term "alkylene group" refers to a divalent alkyl group. The alkylene group is preferably a C1 to C 10 The alkylene group is preferably a C1-C6 alkylene group, and more preferably a C1-C4 alkylene group. Examples of the alkylene group include, but are not limited to, a methylene group, an ethylene group, -CH(CH3)2, and a propylidene group. The alkylene group may be substituted or unsubstituted.

[0098] The term "cycloalkyl group" refers to a saturated or partially saturated monocyclic, fused polycyclic, bridged, or spiro-cyclic carbon ring. The cycloalkyl group is preferably a C3-C 12It is a cycloalkyl group of the formula (I), more preferably a cycloalkyl group of C3 to C8, and most preferably a cycloalkyl group of C3 to C6. Examples of monocyclic cycloalkyl groups include, but are not limited to, a cyclopropyl group, a cyclobutyl group, a cyclopentyl group, a cyclopentenyl group, a cyclohexyl group, a cyclohexenyl group, a cyclohexadienyl group, a cycloheptyl group, a cycloheptatrienyl group, and a cyclooctyl group. It is preferably a cyclopropyl group or a cyclohexenyl group. The cycloalkyl group may be any substituted or unsubstituted one.

[0099] The terms "heterocyclyl group", "heterocycle" or "heterocyclic" may be used interchangeably herein and refer to a non-aromatic heterocyclyl group in which one or more of the ring atoms is a heteroatom such as oxygen, nitrogen or sulfur atom, including monocyclic, fused polycyclic, bridged and spirocyclic rings. Preferably, the ring has a 5-7 membered monocyclic ring or a 7-10 membered bicyclic or tricyclic ring, and may contain 1, 2 or 3 atoms selected from the group consisting of nitrogen, oxygen and / or sulfur. Illustrative examples of "heterocyclyl groups" include, but are not limited to, morpholinyl, oxetanyl, thiomorpholinyl, tetrahydropyranyl, 1,1-dioxothiomorpholinyl, piperidinyl, 2-oxopiperidinyl, pyrrolidinyl, 2-oxopyrrolidinyl, piperazin-2-one, 8-oxa-3-aza-biscyclo[3.2.1]octyl, and piperazinyl. Heterocyclyl groups may be substituted or unsubstituted.

[0100] "Aryl group" refers to a carbocyclic aromatic system containing one or two rings, which may be linked in a fused manner. The term "aryl group" includes monocyclic or bicyclic aryl groups, such as phenyl, naphthyl, and tetrahydronaphthyl aromatic groups. Preferably, the aryl group is a C6-C 10 More preferably, the aryl groups are phenyl and naphthyl groups. The aryl groups may be substituted or unsubstituted.

[0101] The term "heteroaryl group" refers to an aromatic 5- to 6-membered monocyclic ring or 8- to 10-membered bicyclic ring, which may contain 1 to 4 atoms selected from the group consisting of nitrogen, oxygen and / or sulfur. The "heteroaryl group" is preferably a bicyclic heteroaryl group, illustrative examples of which include, but are not limited to, for example, a furanyl group, a pyridyl group, a 2-oxo-1,2-dihydropyridyl group, a pyridazinyl group, a pyrimidinyl group, a pyrazinyl group, a thienyl group, an isoxazolyl group, an oxazolyl group, an oxadiazolyl group, an imidazolyl group, a pyrrolyl group, a pyrazolyl group, a triazolyl group, a tetrazolyl group, a thiazolyl group, an isothiazolyl group, a 1,2,3-thiadiazolyl group, a benzodioxolyl group, a benzothienyl group, a benzimidazolyl group, an indolyl group, an isoindolyl group, a 1,3-dioxo-isoindolyl group, a quinolinyl group, an indazolyl group, a benzoisothiazolyl group, a benzoxazolyl group, a benzisoxazolyl group,

[0102] [ka]

[0103] The heteroaryl group may be substituted or unsubstituted.

[0104] "Fused ring" refers to an aromatic system in which two or more ring structures share a pair of atoms in common, one or more of which may contain one or more double bonds, but at least one of the rings does not have fully conjugated pi-electrons. As used herein, the ring atoms may include nitrogen, oxygen, or S(O). r (r is 0, 1 or 2), and the other ring atoms are carbon. The fused ring is preferably a bicyclic or tricyclic fused ring, and the bicyclic fused ring is preferably a fused ring of an aryl group or a heteroaryl group with a monocyclic heterocyclyl group or a monocyclic cycloalkyl group. The "fused ring" is preferably 7 to 14 members, more preferably 8 to 10 members, and examples include, but are not limited to, the following:

[0105] [ka]

[0106] [ka]

[0107] [ka]

[0108] The term "alkoxy group" refers to a group of (alkyl group -O-). Alkyl group is as defined herein. Preferred are C1-C6 and C1-C4 alkoxy groups. Examples include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, and t-butoxy groups.

[0109] "Haloalkyl group" refers to an alkyl group that is optionally further substituted with one or more halogens, where alkyl group is as defined herein.

[0110] The term "deuterated alkyl group" refers to an alkyl group optionally further substituted with one or more deuterium atoms, where alkyl group is as defined herein. The "deuterated alkyl group" is preferably a deuterated methyl group, including a mono-deuterated methyl group, a di-deuterated methyl group, and a tri-deuterated methyl group, and more preferably a tri-deuterated methyl group.

[0111] "Hydroxyalkyl group" refers to an alkyl group which is optionally further substituted with one or more hydroxy groups, where alkyl is as defined herein.

[0112] "Haloalkoxy" refers to an (alkyl-O-) group in which the alkyl group is optionally further substituted with one or more halogens, where alkoxy is as defined herein.

[0113] "Hydroxy" refers to an -OH group. "Halogen" refers to fluorine, chlorine, bromine and iodine. An "amino group" refers to -NH2. A "cyano group" refers to -CN. A "nitro group" refers to --NO.sub.2. The term "benzyl" refers to the group -CH2-phenyl. A "carboxy group" refers to -C(O)OH.

[0114] The term "carboxylate group" refers to a -C(O)O-alkyl group or a -C(O)O-cycloalkyl group, where the alkyl group and the cycloalkyl group are defined above.

[0115] "DMSO" refers to dimethyl sulfoxide. "BOC" refers to a tert-butoxycarbonyl group. "Ts" refers to p-toluenesulfonyl. "T3P" refers to propyl phosphate anhydride. "DPPA" refers to diphenylphosphoryl azide. "DEA" refers to diethylamine. "TFA" refers to trifluoroacetic acid. "CaCl2" refers to calcium chloride. "MgCl2" refers to magnesium chloride. "KCl" refers to potassium chloride. "NaCl" refers to sodium chloride. "Glucose" refers to glucose. "HEPES" refers to N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid. "EGTA" refers to ethylene glycol bis(2-aminoethyl ether) tetraacetic acid.

[0116] "Substituted" means that one or more hydrogen atoms, preferably up to 5, more preferably 1 to 3 hydrogen atoms in the atomic group, are each independently replaced with the corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and the possibility or impossibility of the substitution can be determined (by experiment or theory) by the skilled artisan without undue effort. For example, an amino group or a hydroxy group having a free hydrogen may be unstable when combined with a carbon atom having an unsaturated (e.g., olefinic) bond.

[0117] As used herein, unless otherwise specified, "substituted" or "substituted" means that the group is an alkyl group, an alkenyl group, an alkynyl group, an alkoxy group, an alkylthio group, an alkylamino group, a halogen atom, a mercapto group, a hydroxy group, a nitro group, a cyano group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, a cycloalkoxy group, a heterocycloalkoxy group, a cycloalkylthio group, a heterocycloalkylthio group, an amino group, a haloalkyl group, a hydroxyalkyl group, a carboxy group, a carboxylate group, =O, -C(O)R 7 , -C(O)OR 7 , -NHC(O)R 7 , -NHC(O)OR 7 , -NR 8 R 9 , -C(O)NR 8 R 9 , -CH2NHC(O)OR 7 , -CHNR 8 R 9 and -S(O) r R 7 This means that it may be substituted with one or more groups selected from the group consisting of:

[0118] Here, R 7is selected from the group consisting of a hydrogen atom, an alkyl group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group, provided that the alkyl group, the cycloalkyl group, the heterocyclyl group, the aryl group, or the heteroaryl group is not selected from the group consisting of a hydroxy group, a halogen atom, a nitro group, a cyano group, an alkyl group, an alkoxy group, a haloalkyl group, a haloalkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, =O, -C(O)R 10 , -C(O)OR 10 , -OC(O)R 10 , -NR 11 R 12 , -C(O)NR 11 R 12 , -SO2NR 11 R 12 and -NR 11 C(O)R 12 and optionally further substituted with one or more substituents selected from the group consisting of:

[0119] R 8 and R 9 are each independently selected from the group consisting of a hydrogen atom, a hydroxy group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group, provided that the alkyl group, the alkoxy group, the cycloalkyl group, the heterocyclyl group, the aryl group, and the heteroaryl group are each independently selected from the group consisting of a hydroxy group, a halogen, a nitro group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, =O, -C(O)R 10 , -C(O)OR 10 , -OC(O)R 10 , -NR 11 R 12 , -C(O)NR 11 R 12 , -SO2NR 11 R 12 and -NR 11 C(O)R 12 and optionally further substituted with one or more substituents selected from the group consisting of:

[0120] Or, R8 and R 9 together with the atom to which they are attached form a 4-8 membered heterocyclyl group, provided that the 4-8 membered heterocyclyl group does not include one or more N, O or S(O) r The 4- to 8-membered heterocyclyl group includes a hydroxy group, a halogen atom, a nitro group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, ═O, —C(O)R 10 , -C(O)OR 10 , -OC(O)R 10 , -NR 11 R 12 , -C(O)NR 11 R 12 , -SO2NR 11 R 12 and -NR 11 C(O)R 12 and optionally further substituted with one or more substituents selected from the group consisting of:

[0121] R 10 , R 11 and R 12 are each independently selected from the group consisting of a hydrogen atom, an alkyl group, an amino group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group, with the proviso that the alkyl group, the cycloalkyl group, the heterocyclyl group, the aryl group, or the heteroaryl group is optionally further substituted with one or more substituents selected from the group consisting of a hydroxy group, a halogen, a nitro group, an amino group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, a carboxy group, and a carboxylate group.

[0122] r is 0, 1 or 2.

[0123] The compounds of the present invention contain asymmetric or chiral centers and therefore can exist in different stereoisomeric forms. All stereoisomeric forms of the compounds of the present invention are contemplated, including, but not limited to, diastereoisomers, enantiomers, atropisomers and geometric (conformational) isomers, as well as mixtures thereof, such as racemic mixtures. All of these forms are within the scope of the present invention.

[0124] Unless otherwise specified, structures depicted in the present invention further include all isomeric (e.g., diastereoisomers, enantioisomers, atropisomers and geometric (conformational) isomeric) forms of the structures, such as the R and S configurations of each asymmetric center, (Z) and (E) double bond isomers, and (Z) and (E) conformational isomers. Thus, individual stereoisomers and enantiomeric mixtures, diastereoisomeric mixtures, and geometric (conformational) isomeric mixtures of the compounds of the present invention are within the scope of the present invention.

[0125] The term "pharmaceutical acceptable salt" refers to a specific salt suitable for medical use while retaining the original biological activity of the above compound. The pharmaceutical acceptable salt of the compound represented by formula (I) may be a metal salt or an amine salt formed with a suitable acid.

[0126] By "pharmaceutical composition" is meant a mixture of one or more compounds described herein or physiologically pharmaceutical acceptable salts or prodrugs thereof with other chemical components, and any other components such as physiologically pharmaceutical acceptable carriers, excipients, etc. The pharmaceutical composition is intended to facilitate administration to a living body and facilitate absorption of the active ingredient, and thus the exertion of biological activity.

[0127] Methods for synthesizing the compounds according to the present invention In order to achieve the objectives of the present invention, the present invention adopts the following technical solutions:

[0128] The method for preparing a compound of the present invention represented by general formula (I), or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof, comprises the steps of reacting a compound of general formula (IA) with a compound of general formula (IB) under basic conditions, and optionally further removing a protecting group to obtain a compound of general formula (I).

[0129] [ka]

[0130] During the ceremony, X 1 is a leaving group, preferably chlorine; Ring A, Ring B, Ring C, R 1 ~R 5 The definitions of X, Y, Z, E, L, n, p and q are as described in general formula (I).

[0131] The present invention will be further described below in conjunction with examples, but these examples are not intended to limit the scope of the present invention. EXAMPLES

[0132] In the Examples, the preparation of representative compounds of formula (I) and related structural identification data are described, and the following Examples are used to illustrate the present invention, but not to limit the present invention. 1 H NMR spectra were recorded on a Bruker instrument (400 MHz) and chemical shifts are expressed in ppm. Tetramethylsilane was used as the internal standard (0.00 ppm). 1 HNMR designation: s = singlet, d = doublet, t = triplet, m = multiplet, br = broad, dd = doublet of doublets, dt = doublet of triplets. Coupling constants, when given, are in Hz.

[0133] Mass spectra were measured on an LC / MS instrument with the ionization mode being ESI or APCI.

[0134] Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plates were used as silica gel plates for thin layer chromatography (TLC). The specifications of the silica gel plates used for thin layer chromatography (TLC) were 0.15mm to 0.2mm, and the specifications of the thin layer chromatography separated and purified products were 0.4mm to 0.5mm.

[0135] For column chromatography, Yantai Yellow Sea silica gel 200-300 mesh silica gel was generally used as the carrier.

[0136] In the following examples, all temperatures are in degrees Celsius, and starting materials and reagents were either commercially available or synthesized according to known methods, provided that commercially available materials and reagents were used as is without further purification, unless otherwise specified, commercial suppliers include, but are not limited to, SAHNGHAI HAOHONG BIOMEDICAL TECHNOLOGY CO., LTD., Shanghai Shaoyuan Co. Ltd., Bide Pharmatech Ltd., Sun Chemical Technology(Shanghai) Co., Ltd., and LinkChem Co., Ltd.

[0137] CD3OD: deuterated methanol. CDCl3: deuterated chloroform. DMSO-d6: deuterated dimethyl sulfoxide. Unless otherwise specified, reaction solutions in the examples refer to aqueous solutions.

[0138] The compounds were purified using column chromatography or thin layer chromatography with the following eluent systems: A: petroleum ether and ethyl acetate, B: dichloromethane and methanol, C: dichloromethane and ethyl acetate, D: dichloromethane and ethanol, E: tetrahydrofuran / petroleum ether, or F: tetrahydrofuran and methanol. The volume ratio of the solvents in the solvent system varies depending on the polarity of the compounds, but may be adjusted by adding a small amount of an acidic reagent such as acetic acid or a basic reagent such as triethylamine. Example 1 - Compound 7

[0139] (2R,4aR)-3-Acryloyl-10-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0140] [ka]

[0141] [ka]

[0142] Step 1 6-Chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)nicotinic acid

[0143] 2-Isopropyl-4-methylpyridin-3-amine 1d (21.46 g, 142.86 mmol) was dissolved in 200 mL of tetrahydrofuran, cooled to -78 ° C., and lithium bis(trimethylsilyl)amide (1.0 M, 238.11 mL) was added dropwise under a nitrogen atmosphere. After stirring at -78 ° C. for 15 minutes, a solution of 2,6-dichloro-5-fluoronicotinic acid 1a (20 g, 95.24 mmol) in tetrahydrofuran (100 mL) was added dropwise. After reacting at -78 ° C. for 1 hour, the reaction was continued at 25 ° C. for 3 hours. After the reaction was completed, the reaction solution was poured into 100 mL of ice water, and methyl-tert-butyl ether (100 mL) was added. The acidity of the aqueous phase was adjusted to pH=4 with 2M dilute hydrochloric acid, the layers were separated, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 6-chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)nicotinic acid 7a (15 g, 46.33 mmol) in a yield of 48.65%.

[0144] MS m / z(ESI):323.8[M+1] +

[0145] Step 2 Ethyl 3-(6-chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)pyridin-3-yl)-2-nitro-3-oxopropionate

[0146] 6-Chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)nicotinic acid 7a (5 g, 15.44 mmol) was dissolved in N,N-dimethylformamide (50 mL), potassium carbonate (6.40 g, 46.33 mmol) and ethyl 2-nitroacetate (6.17 g, 46.33 mmol) were added, and 2-chloro-1-methylpyridinium iodide (7.89 g, 30.89 mmol) was further added, and the mixture was allowed to react at 25° C. for 3 hours. 10 mL of ethyl acetate and 10 mL of saturated saline were added, the layers were separated, the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure, and the obtained residue was isolated by flash silica gel column chromatography (eluent: E system) to obtain ethyl 3-(6-chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)pyridin-3-yl)-2-nitro-3-oxopropionate 7b (2.3 g, 5.24 mmol) in a yield of 33.94%.

[0147] MS m / z(ESI):438.9[M+1] +

[0148] Step 3 7-Chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H)-dione

[0149] Ethyl 3-(6-chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)pyridin-3-yl)-2-nitro-3-oxopropionate 7b (2.3 g, 5.24 mmol) was dissolved in N,N-dimethylformamide (20 mL), cesium carbonate (2.56 g, 7.86 mmol) was added, and the mixture was stirred at 50° C. for 16 hours to react. After the reaction was completed, the mixture was cooled to 25°C, 10mL of ethyl acetate and 10mL of saturated saline were added, the liquids were separated, the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure, and the resulting residue was isolated by flash silica gel column chromatography (eluent: E system-F system) to obtain 7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H)-dione 7c (1.7g, 4.33mmol) in a yield of 82.58%.

[0150] MS m / z(ESI):393.0[M+1] +

[0151] Step 4 4,7-Dichloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one

[0152] 7-Chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridin-2,4(1H,3H)-dione 7c (400 mg, 1.02 mmol) was dissolved in phosphoryl chloride (3 mL) and reacted at 90° C. for 1 hour. The progress of the reaction was monitored by LC-MS. After completion of the reaction, the mixture was concentrated under reduced pressure, and the resulting residue was isolated by flash silica gel column chromatography (eluent: E system) to obtain 4,7-dichloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one 7d (350 mg, 851.14 μmol) in a yield of 83.58%.

[0153] MS m / z(ESI):410.8[M+1] +

[0154] Step 5 (3R,6R)-1-N-tert-butoxycarbonyl-4-(7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-3-carboxylate methyl

[0155] 4,7-Dichloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one 7d (1.3 g, 3.16 mmol) was dissolved in acetonitrile (15 mL), and 1-(t-butyl)-3-methyl(3R,6R)-6-methylpiperazine-1,3-dicarboxylate 1j (1.63 g, 6.32 mmol) was added and reacted at 80° C. for 16 hours. The progress of the reaction was monitored by LC-MS. After completion of the reaction, the mixture was concentrated under reduced pressure, and the resulting residue was isolated by flash silica gel column chromatography (eluent: E system) to obtain methyl (3R,6R)-1-N-tert-butoxycarbonyl-4-(7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-3-carboxylate 7e (1 g, 1.58 mmol) in a yield of 49.97%.

[0156] MS m / z(ESI):633.0[M+1] +

[0157] Step 6 (3R,6R)-1-N-tert-butoxycarbonyl-4-(3-amino-7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-3-carboxylate methyl

[0158] (3R,6R)-1-N-tert-butoxycarbonyl-4-(7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-3-carboxylate methyl 7e (1 g, 1.58 mmol) and Raney nickel (10 mg, 157.96 μmol) were dissolved in tetrahydrofuran (10 mL), hydrogen replacement was performed three times, and the reaction was carried out under a hydrogen gas atmosphere at 25° C. for 2 hours. Filtration and concentration of the filtrate under reduced pressure gave the crude product (3R,6R)-1-N-tert-butoxycarbonyl-4-(3-amino-7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-3-carboxylate methyl 7f (0.83 g, 1.38 mmol), which was used directly in the next reaction. The yield was 87.13%.

[0159] MS m / z(ESI):603.3[M+1] +

[0160] Step 7 (2R,4aR)-10-Chloro-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2-methyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester

[0161] (3R,6R)-1-N-tert-butoxycarbonyl-4-(3-amino-7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-3-carboxylate methyl 7f (0.83 g, 1.38 mmol) and potassium carbonate (570.64 mg, 4.13 mmol) were dissolved in N,N-dimethylformamide (10 mL) and reacted at 50° C. for 1 hour. After the reaction was completed, 10 mL of ethyl acetate and 10 mL of saturated saline were added, the liquids were separated, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the crude product (2R,4aR)-10-chloro-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2-methyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 7g (0.7 g, 1.23 mmol), which was used as it was in the next reaction.

[0162] MS m / z(ESI):571.0[M+1] +

[0163] Step 8 (2R,4aR)-10-Chloro-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester

[0164] (2R,4aR)-10-chloro-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2-methyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 7g (0.7g, 1.23mmol), iodomethane (521.98mg, 3.68mmol) and potassium carbonate (508.27mg, 3.68mmol) were dissolved in N,N-dimethylformamide (10mL) and reacted at 25°C for 16 hours. 10 mL of ethyl acetate and 10 mL of water were added, the layers were separated, the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was isolated by flash silica gel column chromatography (eluent: E system) to give (2R,4aR)-10-chloro-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 7h (0.45 g, 769.14 μmol) in a yield of 62.74%.

[0165] MS m / z(ESI):585.0[M+1] +

[0166] Step 9 (2R,4aR)-10-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester

[0167] (2R,4aR)-10-chloro-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 7h (0.12 g, 205.10 μmol), (2-((tert-butoxycarbonyl (N-amino)-7-fluorobenzo[d]thiazol-4-yl)boronic acid 7i (192.05 mg, 615.31 μmol), tetrakis(triphenylphosphine)palladium (23.70 mg, 20.51 μmol) and potassium phosphate (217.68 mg, 1.03 mmol) were dissolved in a mixed solvent of 0.2 mL of water and 1 mL of 1,4-dioxane, and the mixture was purged with nitrogen three times and reacted at 100° C. for 16 hours under a nitrogen atmosphere. The progress of the reaction was monitored by LC-MS. After completion of the reaction, the mixture was concentrated under reduced pressure, and the resulting residue was isolated by flash silica gel column chromatography (eluent: E system) to obtain (2R,4aR)-10-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 7j (0.1 g, 122.41 μmol) in a yield of 59.68%.

[0168] MS m / z(ESI):817.4[M+1] +

[0169] Step 10 (2R,4aR)-10-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0170] (2R,4aR)-10-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 7j (0.1 g, 122.41 μmol) was dissolved in dichloromethane (2 mL), trifluoroacetic acid (300 mg, 2.63 mmol) was added, and the mixture was allowed to react at 20 ° C. for 16 hours. Concentration under reduced pressure gave the crude product (2R,4aR)-10-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 7k (100 mg, 136.85 μmol), which was used directly in the next reaction.

[0171] MS m / z(ESI):617.5[M+1] +

[0172] Step 11 (2R,4aR)-3-Acryloyl-10-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0173] Acrylic acid (18.33 mg, 254.41 μmol), (2R, 4aR)-10-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 7k (110 mg, 150.54 μmol) and N,N-diisopropylethylamine (194.56 mg, 1.51 mmol) were dissolved in acetonitrile (1 mL), propylphosphoric anhydride (191.59 mg, 301.08 μmol, purity 50%) was added, and the mixture was reacted at 25 ° C. for 16 hours. After the reaction was completed, 20 mL of water was added, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phases were combined, dried over anhydrous magnesium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was isolated by preparative liquid chromatography (separation column: Boston Prime C18, 150 x 30 mm ID, 5 μm; mobile phase A: water (0.05% NH3H2O ​​+ 10 mM NH4HCO3), mobile phase B: acetonitrile; flow rate: 25 mL / min), (2R,4aR)-3-acryloyl-10-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 7 (30 mg) was obtained.

[0174] MS m / z(ESI):671.1[M+1] + Example 2 - Compound 8 and Compound 9

[0175] (2R,4aR,8R)-3-Acryloyl-10-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 8 (2R,4aR,8S)-3-Acryloyl-10-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 9

[0176] [ka]

[0177] [ka]

[0178] (2R,4aR)-3-acryloyl-10-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 7 (30 mg) was subjected to preparative SFC chiral separation (column type: (s,s)WHELK-O1, 250×30 mm ID, 5 μm; mobile phase: A for CO2 and B for After purification with EtOH (0.1% NH3H2O); column pressure: 100 bar; flow rate: 70 mL / min; detection wavelength: 220 nm; column temperature: 40 °C), monocoordinate compound 8 (shorter retention time) and monocoordinate compound 9 (longer retention time) were obtained.

[0179] Monoconformal compound 8 (shorter retention time): MS m / z(ESI):671.1[M+1] + 2.05 mg; retention time 3.446 min; chiral purity 100% ee. 11H NMR (400 MHz, DMSO-d6) δ 8.46 (d, J = 4.9 Hz, 1H), 8.02 - 7.83 (m, 3H), 7.25 (d, J = 4.9 Hz, 1H), 7.10 - 6.80 (m, 3H), 6.24 - 6.08 (m, 1H), 5.82 - 5.69 (m, 1H), 5.17 - 4.37 (m, 2H), 4.08 - 3.92 (m, 1H), 3.75 (br dd, J = 3.9, 13.8 Hz, 1H), 3.34 (s, 3H), 3.02 - 2.71 (m, 3H), 1.83 (s, 3H), 1.62 - 1.45 (m, 3H), 1.11 (br d, J = 6.6 Hz, 3H), 0.99 (br d, J = 6.5 Hz, 3H).

[0180] Compound 9 in a single conformation (the longer retention time): MS m / z (ESI): 671.1 [M+1] + 9.35 mg; retention time 4.235 min; chiral purity 100% ee. 1 1H NMR (400 MHz, DMSO-d6) δ 8.47 (d, J = 4.8 Hz, 1H), 8.00 - 7.87 (m, 3H), 7.25 (d, J = 4.9 Hz, 1H), 7.09 - 6.83 (m, 3H), 6.23 - 6.09 (m, 1H), 5.76 - 5.69 (m, 1H), 5.10 - 4.42 (m, 2H), 4.13 - 3.96 (m, 1H), 3.75 (dd, J = 4.0, 14.0 Hz, 1H), 3.37 (s, 3H), 3.32 - 3.22 (m, 2H), 2.99 - 2.77 (m, 1H), 2.00 (s, 3H), 1.60 - 1.51 (m, 3H), 1.05 (br d, J = 6.6 Hz, 3H), 0.92 (br d, J = 6.6 Hz, 3H). Example 3 - Compound 10

[0181] (2R,4aR)-3-Acryloyl-10-(6-amino-3-chloropyridin-2-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0182] [ka]

[0183] [ka]

[0184] Step 1 (2R,4aR)-11-Fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-10-(trimethylstannyl)-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester

[0185] (2R,4aR)-10-chloro-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 7h (0.1g, 170.92μmol), hexamethylditin (139.99mg, 427.30μmol) and tetrakis(triphenylphosphine)palladium (19.75mg, 17.09μmol) were dissolved in 1,4-dioxane (1mL), nitrogen substitution was performed three times, and the mixture was reacted at 110 ° C. under a nitrogen atmosphere for 16 hours. Concentration under reduced pressure gave a residue, which was isolated by flash silica gel column chromatography (eluent: E system) to give (2R,4aR)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-10-(trimethylstannyl)-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 10a (95 mg, 133.16 μmol) in a yield of 77.91%.

[0186] MS m / z(ESI):714.8[M+1] +

[0187] Step 2 (2R,4aR)-10-(6-amino-3-chloropyridin-2-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester

[0188] (2R,4aR)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-10-(trimethylstannyl)-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 10a (40 mg, 56. 07 μmol), 6-bromo-5-chloropyridin-2-amine 10b (13.96 mg, 67.28 μmol), cuprous iodide (1.07 mg, 5.61 μmol) and tetrakis(triphenylphosphine)palladium (3.24 mg, 2.80 μmol) were dissolved in 1,4-dioxane (0.5 mL), substituted with nitrogen three times, and the reaction was carried out under a nitrogen atmosphere at 100° C. for 16 hours. Concentration under reduced pressure gave a residue, which was isolated by flash silica gel column chromatography (eluent: E system) to give (2R,4aR)-10-(6-amino-3-chloropyridin-2-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 10c (15 mg, 22.15 μmol) in a yield of 39.51%.

[0189] MS m / z(ESI):677.3[M+1] +

[0190] Step 3 (2R,4aR)-10-(6-amino-3-chloropyridin-2-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0191] (2R,4aR)-10-(6-amino-3-chloropyridin-2-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 10c (30 mg, 44.30 μmol) was dissolved in dichloromethane (3 mL), trifluoroacetic acid (1 g, 8.77 mmol) was added, and the mixture was allowed to react at 20 ° C. for 16 hours. Concentration under reduced pressure gave the crude product (2R,4aR)-10-(6-amino-3-chloropyridin-2-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 10d (30 mg, 51.99 μmol), which was used directly in the next reaction.

[0192] MS m / z(ESI):577.0[M+1] +

[0193] Step 4 (2R,4aR)-3-Acryloyl-10-(6-amino-3-chloropyridin-2-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0194] Acrylic acid (4.41 mg, 61.14 μmol), (2R, 4aR)-10-(6-amino-3-chloropyridin-2-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 10d (25 mg, 36.18 μmol) and N,N-diisopropylethylamine (46.75 mg, 361.76 μmol) were dissolved in acetonitrile (5 mL), propylphosphoric anhydride (46.04 mg, 72.35 μmol, purity 50%) was added, and the mixture was reacted at 25 ° C. for 16 hours. After the reaction was completed, 20 mL of water was added, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phases were combined, dried over anhydrous magnesium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was isolated by preparative liquid chromatography (separation column: Boston Prime C18, 150 x 30 mm ID, 5 μm; mobile phase A: water (0.05% NH3H2O ​​+ 10 mM NH4HCO3), mobile phase B: acetonitrile; flow rate: 25 mL / min), (2R,4aR)-3-acryloyl-10-(6-amino-3-chloropyridin-2-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 10 (13 mg, 20.60 μmol) was obtained in 56.94% yield.

[0195] MS m / z(ESI):631.4[M+1] + 1H NMR(400MHz, DMSO-d6) δ8.46(dd, J = 2.3,4.8 Hz,1H),8.09-7.99(m,1H),7.49(d, J=8.8 Hz,1H),7.26(dd, J =4.9, 12.3 Hz,1H),7.10-6.80(m,1H),6.52(d, J =8.9 Hz,1H),6.33(br s, 2H),6.22-6.11(m,1H),5.83-5.70(m,1H),5.10-4.29(m,2H),4.09-3.92(m,1H),3.74(dd, J =4.0, 14.1 Hz,1H),3.3-3.33(m,3H),2.97-2.72(m,2H),2.46-2.36(m,1H),2.04-1.75(m,3H),1.61-1.49(m,3H),1.14-0.83(m,6H). Example 4 - Compound 11

[0196] (2R,4aR)-3-Acryloyl-11-fluoro-10-(3-hydroxynaphthalen-1-yl)-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0197] [ka]

[0198] [ka]

[0199] Step 1 (2R,4aR)-11-Fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-10-(3-methoxynaphthalen-1-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester

[0200] (2R,4aR)-10-chloro-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 7h (100 mg, 170.92 μmol), (3-Methoxy Cinaphthalene-1-yl)boronic acid 11a (103.58 mg, 512.76 μmol), tetrakis(triphenylphosphine)palladium (19.75 mg, 17.09 μmol) and potassium phosphate (181.40 mg, 854.60 μmol) were dissolved in a mixed solvent of 0.3 mL of water and 1.5 mL of 1,4-dioxane, and the mixture was purged with nitrogen three times and reacted under a nitrogen atmosphere at 100° C. for 16 hours. The progress of the reaction was monitored by LC-MS. Concentration under reduced pressure gave a residue, which was isolated by flash silica gel column chromatography (eluent: E system) to give (2R,4aR)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-10-(3-methoxynaphthalen-1-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 11b (0.1 g, 141.48 μmol) in a yield of 82.78%.

[0201] MS m / z(ESI):707.7[M+1] +

[0202] Step 2 (2R,4aR)-11-Fluoro-10-(3-hydroxynaphthalen-1-yl)-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0203] (2R,4aR)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-10-(3-methoxynaphthalen-1-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 11b (70 mg, 99.04 μmol) was dissolved in 5 mL of dichloromethane, boron tribromide (1.40 g, 5.59 mmol) was added, and the mixture was reacted at 20 ° C. for 16 hours. The reaction was quenched by adding 10 mL of methanol, concentrated under reduced pressure, diluted with 20 mL of water, washed with ethyl acetate (20 mL x 2), collected the aqueous phase, and lyophilized the aqueous phase to give the crude product (2R,4aR)-11-fluoro-10-(3-hydroxynaphthalen-1-yl)-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 11c (100 mg, 168.73 μmol).

[0204] MS m / z(ESI):593.4[M+1] +

[0205] Step 3 4-((2R,4aR)-3-Acryloyl-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-2,3,4,4a,5,6,7,8-octahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridin-10-yl)naphthalen-2-yl Butyl acrylate

[0206] (2R,4aR)-11-fluoro-10-(3-hydroxynaphthalen-1-yl)-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 11c (100 mg, 168.73 μmol) and triethylamine (85.37 mg, 843.65 μmol) were dissolved in dichloromethane (5 mL), and acryloyl chloride (15.27 mg, 168.73 μmol) was added dropwise at 10° C. and reacted at 10 to 20° C. for 16 hours. The progress of the reaction was monitored by LC-MS. After the reaction was completed, 10 mL of water was added, and the mixture was extracted with dichloromethane (10 mL × 2). The organic phases were combined, dried over anhydrous magnesium sulfate, filtered, and concentrated under reduced pressure to obtain the crude product 4-((2R,4aR)-3-acryloyl-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-2,3,4,4a,5,6,7,8-octahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridin-10-yl)naphthalene-2-group butyl acrylate 11d (150 mg, 214.05 μmol), which was used as it is in the next reaction.

[0207] MS m / z(ESI):701.2[M+1] +

[0208] Step 4 (2R,4aR)-3-Acryloyl-11-fluoro-10-(3-hydroxynaphthalen-1-yl)-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0209] 4-((2R,4aR)-3-acryloyl-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-2,3,4,4a,5,6,7,8-octahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridin-10-yl)naphthalen-2-yl butyl acrylate 11d (150 mg, 214.05 μmol) was dissolved in 1 mL of tetrahydrofuran, and an aqueous solution of lithium hydroxide monohydrate (26.95 mg, 642.16 μmol) (1 mL) was added dropwise and reacted at 15° C. for 16 hours. The progress of the reaction was monitored by LC-MS. After the reaction was completed, the pH was adjusted to 7 by adding 1M dilute hydrochloric acid dropwise, and the mixture was extracted with ethyl acetate (10 mL x 2). The organic phases were combined, dried over anhydrous magnesium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was isolated by preparative liquid chromatography (separation column: Boston Prime C18, 150 x 30 mm ID, 5 μm; mobile phase A: water (0.05% NH3H2O ​​+ 10 mM NH4HCO3), mobile phase B: acetonitrile; flow rate: 25 mL / min), (2R,4aR)-3-acryloyl-11-fluoro-10-(3-hydroxynaphthalen-1-yl)-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 11 (26 mg, 40.20 μmol) was obtained in 18.78% yield.

[0210] MS m / z(ESI):647.2[M+1] + 1H NMR(400MHz, DMSO-d6) δ 10.02(s,1H),8.42-8.37(m,1H),8.14-8.04(m,1H),7.75(d, J =8.3 Hz,1H),7.45-7.33(m,2H),7.27-7.12(m,3H),7.11-6.83(m,2H),6.22-6.12(m,1H),5.83-5.71(m,1H),5.05(br d, J = 13.8 Hz,1H),4.81(br s, 1H),4.63(br d, J = 13.3 Hz,1H),4.46(s,1H),4.07-3.95(m,1H),3.77(dd, J =4.1, 14.2 Hz,1H),3.54-3.41(m,2H),3.29-3.22(m,1H),2.95-2.79(m,1H),2.07-1.81(m,3H),1.64-1.52(m,3H),1.14-0.84(m,6H). Example 5 - Compound 16

[0211] (2R,4aR)-3-Acryloyl-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0212] [ka]

[0213] [ka]

[0214] [ka]

[0215] Step 1 3-Isocyanato-2-isopropyl-4-(methylthio)pyridine

[0216] 2-Isopropyl-4-(methylthio)pyridine-3-amine 16a (3.38g, 18.45mmol, prepared by the inventors themselves according to patent WO2020239077) was added to tetrahydrofuran (20mL), cooled to zero degrees Celsius, triethylamine (1.86g, 18.45mmol) was added, triphosgene (5.48g, 18.45mmol) was added slowly in batches, reacted at zero degrees Celsius for 0.5 hours, filtered, and 3-isocyanate-2-isopropyl-4-(methylthio)pyridine 16b (3.83g) was obtained in 100% yield, which was directly used for the next reaction without purification.

[0217] Step 2 N-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-nitroacetamide

[0218] Nitromethane (1.12 g, 18.45 mmol) was added to tetrahydrofuran (20 mL), cooled to zero degrees Celsius, potassium tert-butoxide (4.41 g, 36.9 mmol) was added, and the mixture was reacted at zero degrees Celsius for 0.5 hours. 3-isocyanate-2-isopropyl-4-(methylthio)pyridine 16b (3.83 g, 18.45 mmol) was slowly added dropwise. After the reaction was completed, ethyl acetate (30 mL) and water (30 mL) were added for extraction. The organic phase was washed with saturated saline (30 mL x 3), dried over anhydrous sodium sulfate, and the obtained residue was purified by silica gel column chromatography (eluent: A system) to obtain N-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-nitroacetamide 16c (2.2 g) in a yield of 44.95%.

[0219] MS m / z(ESI):270.1[M+1] +

[0220] Step 3 N-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-nitro-3-oxo-3-(2,5,6-trichloropyridin-3-yl)propionamide

[0221] N-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-nitroacetamide 16c (820 mg, 3.05 mmol), 2,5,6-trichloronicotinic acid 4a (686.25 mg, 3.05 mmol), tetramethylfluorourea hexafluorophosphate (1.2 g, 4.57 mmol) and N,N-diisopropylethylamine (923 mg, 7.1 mmol) were added to acetonitrile (20 mL) and reacted at room temperature for 3 hours. After the reaction was completed, ethyl acetate (30 mL) and water (30 mL) were added for extraction, the organic phase was washed with saturated saline (30 mL x 3), dried over anhydrous sodium sulfate, and the obtained residue was purified by silica gel column chromatography (eluent: A system) to obtain N-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-nitro-3-oxo-3-(2,5,6-trichloropyridin-3-yl)propionamide 16e (1.2 g) in a yield of 82.56%.

[0222] MS m / z(ESI):477.1[M+1] +

[0223] Step 4 6,7-Dichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H)-dione

[0224] N-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-nitro-3-oxo-3-(2,5,6-trichloropyridin-3-yl)propionamide 16e (1.2 g, 2.52 mol) was added to N,N-dimethylformamide (20 mL), cesium carbonate (1.6 g, 5.04 mmol) was added, and the mixture was heated to 85° C. and reacted overnight. After the reaction was completed, ethyl acetate (30 mL) and water (30 mL) were added for extraction, the organic phase was washed with saturated saline (30 mL x 3), dried over anhydrous sodium sulfate, and the obtained residue was purified by silica gel column chromatography (eluent: A system) to obtain 6,7-dichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H)-dione 16f (980 mg) in a yield of 87.64%.

[0225] MS m / z(ESI):441.1[M+1] +

[0226] Step 5 4,6,7-Trichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one

[0227] 6,7-Dichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridin-2,4(1H,3H)-dione 16f (610mg, 1.38mol) was added to phosphoryl chloride (10mL), heated to 110℃, and reacted for 3 hours. After the reaction was completed, the reaction solution was poured into ice water, the PH was adjusted to basic, and dichloromethane (50mL) was added for extraction, dried over anhydrous sodium sulfate, and the resulting residue was purified by silica gel column chromatography (eluent: A system) to obtain 4,6,7-trichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one 16g (460mg) in a yield of 72.24%.

[0228] MS m / z(ESI):459.1[M+1] +

[0229] Step 6 1-(t-Butyl)-3-methyl(3R,6R)-4-(6,7-dichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylate

[0230] 4,6,7-Trichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one 16g (460 mg, 1 mmol) and 1-(t-butyl)-3-methyl(3R,6R)-6-methylpiperazine-1,3-dicarboxylate 1j (309.42 mg, 1.15 mmol) were added to acetonitrile (20 mL) and refluxed overnight under an argon atmosphere. After completion of the reaction, ethyl acetate (20 mL) and water (20 mL) were added for extraction, the organic phase was washed with saturated saline (20 mL x 3), dried over anhydrous sodium sulfate, and the obtained residue was purified by silica gel column chromatography (eluent: A system) to obtain 1-(t-butyl)-3-methyl(3R,6R)-4-(6,7-dichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylate 16i (544 mg) in a yield of 80%.

[0231] MS m / z(ESI):681.1[M+1] +

[0232] Step 7 1-(t-Butyl)-3-methyl(3R,6R)-4-(6,7-dichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-amino-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylate

[0233] 1-(t-Butyl)-3-methyl(3R,6R)-4-(6,7-dichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylic acid 16i (544 mg, 800 μmol) was added to acetonitrile (20 mL), cooled to zero degrees Celsius, N,N-diisopropylethylamine (520.0 mg, 4.0 mmol) and trichlorosilane (379.26 mg, 2.5 mmol) were added, and the mixture was reacted at room temperature for 2 hours. After the reaction was completed, Ethyl acetate (20 mL) and water (20 mL) were added for extraction, the organic phase was washed with saturated saline (20 mL × 3), dried over anhydrous sodium sulfate, and the obtained residue was purified by silica gel column chromatography (eluent: A system) to obtain 1-(t-butyl)-3-methyl(3R,6R)-4-(6,7-dichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-amino-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylate 16j (420 mg) in a yield of 81.37%.

[0234] MS m / z(ESI):651.1[M+1] +

[0235] Step 8 (2R,4aR)-10,11-dichloro-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-methyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester

[0236] 1-(t-butyl)-3-methyl(3R,6R)-4-(6,7-dichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-amino-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylate 16j (420 mg, 651.22 μmol) was added to N,N-dimethylformamide (10 mL), potassium carbonate (179.73 mg, 1.31 mmol) and the mixture was reacted at room temperature for 3 hours. After the reaction was completed, ethyl acetate (20 mL) and water (20 mL) were added for extraction, the organic phase was washed with saturated saline (20 mL × 3), dried over anhydrous sodium sulfate, and the obtained residue was purified by silica gel column chromatography (eluent: A system) to obtain (2R, 4aR)-10,11-dichloro-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-methyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 16k (330 mg) in a yield of 82.04%.

[0237] MS m / z(ESI):619.1[M+1] +

[0238] Step 9 (2R,4aR)-10,11-dichloro-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester

[0239] (2R,4aR)-10,11-dichloro-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-methyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 16k (330 mg, 533.98 μmol) and potassium carbonate (147.72 mg, 1.06 mmol) were added to acetone (10 mL), iodomethane (149.46 mg, 1.06 mmol) was added dropwise, and the mixture was heated to reflux for 2 hours. After the reaction was completed, ethyl acetate (20 mL) and water (20 mL) were added for extraction, the organic phase was washed with saturated saline (20 mL × 3), dried over anhydrous sodium sulfate, and the obtained residue was purified by silica gel column chromatography (eluent: A system) to obtain (2R, 4aR)-10,11-dichloro-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 16l (300 mg) in a yield of 87.34%.

[0240] MS m / z(ESI):633.1[M+1] +

[0241] Step 10 (2R,4aR)-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester

[0242] (2R,4aR)-10,11-dichloro-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 16l (300 mg, 474.68 μmol), potassium (2- Fluoro-6-hydroxyphenyl)trifluoroborate (155.22 mg, 712.02 μmol), [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (52.84 mg, 71.21 μmol) and potassium acetate (139.16 mg, 1.42 mmol) were added to 13 mL of a mixed solvent (1,4-dioxane:water=10:3), heated to 100° C., and reacted for 5 hours. After the reaction was completed, the reaction solution was cooled to room temperature, filtered, the filtrate was collected, and ethyl acetate (20 mL) and water (10 mL) were added for extraction. The organic phase was washed with saturated saline (10 mL × 3), dried over anhydrous sodium sulfate, and the resulting residue was purified by silica gel column chromatography (eluent: A system) to obtain (2R, 4aR)-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 16m (150 mg) in a yield of 43.67%.

[0243] MS m / z(ESI):709.1[M+1] +

[0244] Step 11 (2R,4aR)-11-Chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0245] (2R,4aR)-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 16m (150 mg, 207.75 μmol) was added to dichloromethane (5 mL), cooled to zero degrees Celsius, and hydrochloric acid dioxane solution (4 M, 2 mL) was added dropwise and reacted at room temperature for 2 hours. After the reaction was completed, poured into ice water, added dichloromethane (50 mL) and extracted, the aqueous phase was adjusted to weak base with saturated aqueous sodium carbonate solution, added 10 mL of ethyl acetate and water (10 mL) and extracted, the organic phase was washed with saturated saline (10 mL × 3), dried over anhydrous sodium sulfate, and the obtained residue was purified by silica gel column chromatography (eluent: A system) to obtain (2R, 4aR)-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 16n (80 mg) in a yield of 64.19%.

[0246] MS m / z(ESI):609.1[M+1] +

[0247] Step 12 (2R,4aR)-3-Acryloyl-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0248] (2R,4aR)-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 16n (80 mg, 131.34 μmol) was added to dichloromethane (5 mL), cooled to zero degrees Celsius, triethylamine (26.53 mg, 262.68 μmol) was added dropwise, and a dichloromethane solution of acryloyl chloride (12.92 mg, 142.71 μmol) was slowly added dropwise to continue the reaction at zero degrees Celsius. After the reaction was completed, the mixture was extracted with dichloromethane (10 mL) and water (10 mL) at room temperature. The organic phase was washed with saturated saline (10 mL x 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting residue was separated and purified to obtain the product (2R,4aR)-3-acryloyl-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 16 (25 mg) in a yield of 27.08%.

[0249] MS m / z(ESI):663.1[M+1] + 1H NMR(400mHz, CDCl3) δ8.61(d, J =5.3 Hz,1H),8.29(s,1H),7.74(s,1H),7.24(d, J = 2.0 Hz,1H),7.11(d, J =5.4 Hz,1H),7.08 -6.99(m,1H),6.73 -6.70(m,1H),6.69(t, J = 1.3 Hz,1H),6.36(dd, J = 16.9, 2.0 Hz,1H),5.81(dd, J = 10.6, 1.9 Hz,1H),5.07(s,1H),4.77(d, J = 14.0 Hz,1H),3.82(dd, J = 14.1,4.4 Hz,1H),3.64(d, J =4.0 Hz,1H),3.49(s,3H),3.22(d, J = 12.2 Hz,1H),3.00(dd, J = 12.0,3.6 Hz,1H),2.53 - 2.46(m,1H), 2.45(s,3H), 1.68(s,3H), 1.20(d, J =6.7 Hz,3H), 0.95(d, J =6.7 Hz,3H).

[0250] [ka]

[0251] Step 1 6-Chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H)-dione

[0252] 6,7-Dichloro-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H)-dione 16f (500 mg, 1.13 mmol) was added to a mixture of dioxane (20 mL) and water (5 mL), and (2-fluoro-6-methoxyphenyl)boronic acid 16o (386 mg, 2.26 mmol), sodium carbonate (220 mg, 2.26 mmol) and tetrakistriphenylphosphine palladium (12.7 mg, 0.01 mmol) were added, and the mixture was heated to 100° C. overnight under an argon atmosphere. After the reaction was completed, the mixture was filtered, the filtrate was collected, and extracted with ethyl acetate (20 mL) and water (20 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluent: A system) to obtain the product 6-chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H)-dione 16p (530 mg, 1 mmol) in a yield of 72.24%.

[0253] MS m / z(ESI):531.1[M+1] +

[0254] Step 2 4,6-Dichloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one

[0255] 6-Chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H)-dione 16p (530 mg, 1 mmol) was added to phosphoryl chloride (10 mL) and heated to 110° C. for 3 h. The reaction mixture was poured into ice water, the system was adjusted to basicity with saturated sodium carbonate solution, and dichloromethane (50 mL) was added for extraction, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluent: A system) to obtain the product 4,6-dichloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one 16q (439.2 mg, 800 μmol) in 80% yield.

[0256] MS m / z(ESI):549.1[M+1] +

[0257] Step 3 1-(t-Butyl) 3-methyl (3R,6R)-4-(6-chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylate

[0258] 4,6-Dichloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one 16q (267 mg, 487.8 μmol) and 1-(t-butyl) 3-methyl(3R,6R)-6-methylpiperazine-1,3-dicarboxylate 1j (196.8 mg, 731.7 μmol) were added to acetonitrile (20 mL) and refluxed overnight under an argon atmosphere. After the reaction was completed, ethyl acetate (20 mL) and water (20 mL) were added for extraction, and the organic phase was washed with saturated saline (20 mL x 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluent: A system) to obtain the product 1-(t-butyl) 3-methyl (3R, 6R)-4-(6-chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylic acid ester 16r (300 mg, 390.24 μmol) in a yield of 80%.

[0259] MS m / z(ESI):771.1[M+1] +

[0260] Step 4 1-(t-Butyl) 3-methyl(3R,6R)-4-(3-amino-6-chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylate

[0261] 1-(t-butyl)3-methyl(3R,6R)-4-(6-chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-3-nitro-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylic acid ester 16r (300 mg, 390.24 μmol) was added to a mixed solvent of N,N-dimethylformamide (10 mL) and acetonitrile (10 mL), cooled to 0 ° C., diisopropylethylamine (176 mg, 1.75 mmol) and trichlorosilane (188.92 mg, 1.38 mmol) were added, and the mixture was reacted at room temperature for 2 hours. After the reaction was completed, ethyl acetate (20 mL) and water (20 mL) were added for extraction, and the organic phase was washed with saturated saline (20 mL x 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluent: A system) to obtain the product 1-(t-butyl) 3-methyl (3R, 6R)-4-(3-amino-6-chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylic acid ester 16s (230 mg, 320 μmol) in a yield of 82.04%.

[0262] MS m / z(ESI):741.1[M+1]+

[0263] Step 5 (2R,4aR)-11-chloro-10-(2-fluoro-6-methoxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-methyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester

[0264] 1-(t-butyl) 3-methyl (3R, 6R)-4-(3-amino-6-chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylate 16s (230 mg, 320 μmol) was added to N,N-dimethylformamide (10 mL), potassium carbonate (88.32 mg, 640 μmol) was added, and the mixture was allowed to react at room temperature for 3 hours. After the reaction was completed, ethyl acetate (20 mL) and water (20 mL) were added for extraction, and the organic phase was washed with saturated saline (20 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluent: A system) to obtain the product (2R, 4aR)-11-chloro-10-(2-fluoro-6-methoxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-methyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 16t (182 mg, 256.48 μmol) in a yield of 80.04%.

[0265] MS m / z(ESI):709.1[M+1] +

[0266] Step 6 (2R,4aR)-11-chloro-10-(2-fluoro-6-methoxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester

[0267] (2R,4aR)-11-chloro-10-(2-fluoro-6-methoxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2-methyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 16t (182 mg, 256.48 μmol) and potassium carbonate (70.89 mg, 512.9 μmol) were added to acetone (5 mL), iodomethane (72.46 mg, 512.96 μmol) was added dropwise, and the mixture was heated to reflux for 2 hours. After the reaction was completed, ethyl acetate (20 mL) and water (20 mL) were added for extraction, and the organic phase was washed with saturated saline (20 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluent: B system) to obtain the product (2R, 4aR)-11-chloro-10-(2-fluoro-6-methoxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 16u (150 mg, 207.75 μmol) in a yield of 81%.

[0268] MS m / z(ESI):723.1[M+1] +

[0269] Step 7 (2R,4aR)-11-Chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0270] (2R,4aR)-11-chloro-10-(2-fluoro-6-methoxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl 16u (150 mg, 207.75 μmol) was added to dichloromethane (5 mL), cooled to 0 ° C, and boron tribromide (1 M, 13.34 mL) was added dropwise, returned to room temperature and reacted overnight. After the reaction was completed, the mixture was poured into ice water (50 mL), extracted with dichloromethane (50 mL), and the aqueous phase was adjusted to a weak base with saturated aqueous sodium carbonate solution, and extracted with ethyl acetate (10 mL) and water (10 mL). The organic phases were combined, washed with saturated aqueous sodium chloride (10 mL × 3), and dried over anhydrous sodium sulfate to obtain the crude product (2R, 4aR)-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 16n (80 mg, 131.34 μmol) in a yield of 64.19%.

[0271] MS m / z(ESI):609.1[M+1] +

[0272] Step 8 (2R,4aR)-3-Acryloyl-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

[0273] (2R,4aR)-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 16n (80 mg, 131.34 μmol) was added to dichloromethane (5 mL), cooled to 0 °C, triethylamine (26.53 mg, 262.68 μmol) was added dropwise, and a dichloromethane solution of acryloyl chloride (12.92 mg, 142.71 μmol) was slowly added dropwise to continue the reaction at 0 °C. After the reaction was completed, the mixture was extracted with dichloromethane (10 mL) and water (10 mL) at room temperature. The organic phase was washed with saturated saline (10 mL x 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting residue was isolated by preparative liquid chromatography (separation column: AKZONOBEL Kromasil; 250 x 21.2 mm). ID; 5 μm; mobile phase A: 0.05% TFA + HO, mobile phase B: acetonitrile; flow rate: 20 mL / min), the product (2R,4aR)-3-acryloyl-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 16 (25 mg, 37.76 μmol) was obtained in a yield of 27.08%.

[0274] MS m / z(ESI):663.1[M+1] + 1H NMR(400mHz, CDCl3) δ8.61(d, J =5.3 Hz,1H),8.29(s,1H),7.74(s,1H),7.24(d, J = 2.0 Hz,1H),7.11(d, J =5.4 Hz,1H),7.08 -6.99(m,1H),6.73 -6.70(m,1H),6.69(t, J = 1.3 Hz,1H),6.36(dd, J = 16.9, 2.0 Hz,1H),5.81(dd, J = 10.6, 1.9 Hz,1H),5.07(s,1H),4.77(d, J = 14.0 Hz,1H),3.82(dd, J = 14.1,4.4 Hz,1H),3.64(d, J =4.0 Hz,1H),3.49(s,3H),3.22(d, J = 12.2 Hz,1H),3.00(dd, J = 12.0,3.6 Hz,1H),2.53 - 2.46(m,1H), 2.45(s,3H), 1.68(s,3H), 1.20(d, J =6.7 Hz,3H), 0.95(d, J =6.7 Hz,3H). Example 6 - Compound 17 and Compound 18

[0275] [ka]

[0276] (2R,4aR)-3-acryloyl-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-(methylthio)pyridin-3-yl)-2,6-dimethyl-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 16 (45 mg) was subjected to preparative SFC chiral separation (column type: ChiralPak AD, 250 × 30 mm ID, 10 μm, 250 × 30 mm ID, 10 μm; mobile phase: A for CO2 and B for 100 μM). After purification with 100 mL of ethyl acetate (ethanol; column pressure: 100 bar; flow rate: 80 mL / min; detection wavelength: 220 nm; column temperature: 38 °C), monodentate compound 17 (shorter retention time) and monodentate compound 18 (longer retention time) were obtained.

[0277] Monoconformal compound 17 (shorter retention time): MS m / z(ESI):663.2[M+1] + 20 mg; retention time 1.109 min; chiral purity 100% ee.

[0278] Monodentate compound 18 (longer retention time): MS m / z(ESI):663.3[M+1] + 10mg; retention time 2.525min; chiral purity 99.28%ee.

[0279] Biological evaluation Test Example 1. Measurement of the covalent binding ability of the compound according to the present invention to KRAS G12C protein

[0280] The following method was used to determine the ability of the compounds according to the invention to covalently bind to recombinant human KRAS G12C protein under in vitro conditions.

[0281] The experimental procedure was briefly described as follows: Recombinant human KRAS G12C protein (aa1-169) was prepared using reaction buffer (20 mM HEPES, 150 mM NaCl, 1 mM MgCl2, 1 mM DTT) and stocked at a concentration of 4 μM. Test compounds were prepared as 10 mM stock solutions by dissolving in DMSO, and then diluted using reaction buffer to stock. First, 1.5 μL of test compound diluted with reaction buffer (final concentration of reaction system was 3 μM) was added to the well, then 23.5 μL of reaction buffer was added and mixed well, then 25 μL of 4 μM recombinant human KRAS G12C protein was added, and incubated at room temperature for 5 minutes, after which 5 μL of acetic acid was added to stop the reaction, and the sample was transferred to a vial. The ratio of covalent binding of test compounds to KRAS G12C protein was detected using an Agilent 1290 / 6530 instrument. However, the samples were analyzed using a liquid phase chromatography column (XBridge Protein BEH C4, 300 Å, 3.5 μm, 2.1 mm × 50 mm), and during the detection process, mobile phase A was 0.1% formic acid aqueous solution, and mobile phase B was acetonitrile. The mobile phase elution program was 0-0.5 min, mobile phase A was 95%, 2.5 min, mobile phase A was changed to 30% and maintained for 0.5 min, and at 3.1 min, mobile phase A was changed to 95% and maintained for 1.9 min, and the flow rate was 0.5 mL / min. Finally, the data was analyzed using MassHunter Workstation Software Bioconfirm Version B.08.00 software, and the covalent binding rate (Binding Rate) to KRAS G12C protein under the condition of incubation for 5 min at a concentration of 3 μM of the test compound was obtained, and is shown in Table 1.

[0282] [Table 1]

[0283] Conclusion: The compounds according to the present invention have good covalent binding rate with KRAS G12C protein.

[0284] Test Example 2. Inhibitory activity of the compounds according to the present invention on NCI-H358 cell proliferation The following method was used to measure the effect of the compounds of the present invention on NCI-H358 cell proliferation. NCI-H358 cells (containing KRAS G12C mutation) were purchased from Cell Resource Center, Shanghai Institutes for Life Sciences, Chinese Academy of Sciences, and cultured in RPMI 1640 medium containing 10% fetal bovine serum, 100 U penicillin, 100 μg / mL streptomycin, and 1 mM sodium pyruvate. Cell viability was measured using CellTiter-Glo. (R) Measured by Luminescent Cell Viability Assay kit (Promega, Cat. No. G7573).

[0285] The experimental method was performed according to the procedure in the kit's manual and was briefly described as follows. The test compounds were first dissolved in DMSO to prepare a 10 mM stock solution, then diluted with culture medium to prepare test samples so that the final compound concentration was in the range of 1000 nM to 0.015 nM. Logarithmic growth phase cells were seeded in a 96-well cell culture plate at a density of 800 cells per well, and cultured overnight in a 37°C, 5% CO2 incubator, after which the test compounds were added and cultured for 120 hours. After the culture was completed, a 50 μL volume of CellTiter-Glo detection solution was added to each well, shaken for 5 minutes, and allowed to stand for 10 minutes, after which the luminescence value of each well of the sample was read in the Luminescence mode of the microplate reader. The percent inhibition rate of the compound at each concentration point was calculated by comparing with the value of the control group (0.3% DMSO), and then a nonlinear regression analysis was performed on the compound concentration logarithm-inhibition rate using GraphPad Prism5 software to obtain the IC of cell proliferation inhibition by the compound. 50 The values ​​were obtained and are shown in Table 2.

[0286] [Table 2]

[0287] Conclusion: The compounds according to the present invention have good proliferation inhibitory effects on NCI-H358 (human non-small cell lung cancer) cells.

[0288] Test Example 3. Inhibitory activity of p-ERK1 / 2 in NCI-H358 cells by the compounds according to the present invention The following method was used to measure the inhibitory activity of the compounds of the present invention against p-ERK1 / 2 in NCI-H358 cells. In this method, Cisbio's Advanced phospho-ERK1 / 2 (Thr202 / tyr204) kit (catalog number 64AERPEH) was used, and the detailed experimental procedure can be referred to the kit's instruction manual. NCI-H358 cells (containing KRAS G12C mutation) were purchased from Cell Resource Center, Shanghai Institutes of Life Sciences, Chinese Academy of Sciences.

[0289] The experimental procedure was briefly described as follows. NCI-H358 cells were cultured in RPMI 1640 complete medium containing 10% fetal bovine serum, 100U penicillin, 100μg / mL streptomycin and 1mM sodium pyruvate. NCI-H358 cells were seeded at 30000 cells per well in a 96-well plate, and cultured overnight at 37℃ in a 5% CO2 incubator using complete medium as the culture medium. The test compounds were dissolved in DMSO to prepare 10mM stock solutions, which were then diluted with RPMI 1640 basal medium, and 90μL of RPMI 1640 basal medium containing the corresponding concentration of the test compound was added to each well, so that the final concentration of the test compound in the reaction system was in the range of 1000nM to 0.015nM, and the cells were placed in a cell incubator and cultured for 3 hours and 40 minutes. Then, 10 μL of hEGF (purchased from Roche, catalog number 11376454001) prepared in RPMI 1640 basal medium was added so that the final concentration was 5 nM, and the incubator was turned on and the cells were cultured for 20 minutes. The cell supernatant was discarded, the cells were washed with ice-bathed PBS, and then 45 μL of 1 × cell phospho / total protein lysis buffer (Advanced phospho-ERK1 / 2 kit component) was added to each well to decompose the cells. The 96-well plate was decomposed on ice for 30 minutes, and the decomposition solution was detected according to the instructions of the Advanced phospho-ERK1 / 2 (Thr202 / tyr204) kit. Finally, the fluorescence intensity of each well was measured at emission wavelengths of 620 nM and 665 nM at an excitation wavelength of 304 nM in the TF-FRET mode of the microplate reader, and the 665 / 620 fluorescence intensity ratio of each well was calculated. The percent inhibition rate of the test compound at each concentration was calculated by comparing with the fluorescence intensity ratio of the control group (0.1% DMSO). Nonlinear regression analysis was performed on the logarithm of the test compound concentration versus inhibition rate using GraphPad Prism5 software to obtain the IC value of the compound. 50 The values ​​were obtained and are shown in Table 3.

[0290] [Table 3]

[0291] Conclusion: The compounds according to the present invention have good proliferation inhibitory effects on p-ERK1 / 2 in NCI-H358 cells.

[0292] Test Example 4. Inhibitory activity of the compounds according to the present invention on the hERG potassium ion channel 1, Cell culture 1.1 The cells used in this study were a CHO cell line (provided by Sophion Bioscience, Denmark) transfected with hERG cDNA to stably express hERG channels, and the cell generation was P17. The cells were cultured in a medium containing Ham's F12 medium, 10% (v / v) inactivated fetal bovine serum, 100 μg / mL hygromycin B, and 100 μg / mL geneticin (all purchased from Invitrogen).

[0293] 1.2 CHO hERG cells were grown in Petri dishes containing the above culture medium and incubated in an incubator containing 5% CO2 at 37°C. 24-48 hours before electrophysiological experiments, CHO hERG cells were transferred to circular glass slides placed in Petri dishes and grown in the same culture medium and under the same conditions as above. The density of CHO hERG cells on each circular glass slide was required such that most of the cells were independent and single.

[0294] 2. Experimental Solution The following solutions (recommended by Sophion) were used for electrophysiological recordings:

[0295] [Table 4-1]

[0296] 3. Electrophysiological Recording Process 3.1 Electrophysiological recording system A fully automated QPatch system (Sophion, Denmark) was used for recording whole-cell currents. Cells were voltage clamped at -80 mV. The cell clamp voltage was depolarized to +20 mV to activate the hERG potassium channel and clamped to -50 mV after 2.5 s to override inactivation and generate an outward tail current. The tail current peak was used as a measure of the magnitude of the hERG current.

[0297] 3.2 QPatch Experiment Procedure After reaching a membrane-permeable whole-cell configuration state in the initial stage, the cells were recorded for at least 120 seconds until stabilization was reached. The above voltage pattern was then applied to the cells every 15 seconds throughout the entire process. Only stable cells were allowed to enter the drug treatment process after recording the above parameter thresholds. An external solution containing 0.1% dimethyl sulfoxide (solvent) was applied to the cells to establish a baseline and allow the current to stabilize for 3 minutes. After addition of the compound solution, the cells remained in the test environment until the compound effect reached a plateau or limit for 4 minutes. In experiments testing different concentration gradients of compounds, compounds were added to the clamped cells from low to high concentrations. After the compound test was completed, the cells were washed with external solution until the current returned to a stable state.

[0298] 4. Preparation of Compounds 4.1 10 mM compound stock solutions were serially diluted in extracellular fluid to final μM concentrations. 4.2 The highest test concentration was 30 μM, with six concentrations being 30, 10, 3, 1, 0.3 and 0.1 μM. 4.3 The final concentration of DMSO in the compound solutions of other concentrations was 0.1%, except for the 30 μM compound, which had a final DMSO concentration of 0.3%. All compound solutions were sonicated and shaken for 5-10 min to completely dissolve the compounds.

[0299] 5. Data Analysis The study data was analyzed using Qpatch analysis software provided by Sophion, Excel, and Graphpad Prism.

[0300] 6. Experimental Results The results of inhibition of hERG current by the compounds according to the present invention are shown in Table 4.

[0301] [Table 4-2]

[0302] Conclusion: Drug inhibition of cardiac hERG potassium ion channel is the main reason why drugs cause long QT syndrome. Experimental results show that compound 17 of the present invention has no significant inhibitory effect on cardiac hERG potassium ion channel, and can avoid cardiotoxic side effects at high doses.

[0303] Test Example 5. Pharmacokinetic study of the compound according to the present invention in ICR mice 1. Purpose of the experiment ICR mice were used as test animals, and the compounds of Comparative Example 1 and Example 17 of the present invention were intragastrically administered to the mice. The drug concentrations in plasma at different time points were measured by LC / MS / MS, and the pharmacokinetic properties of the compounds of the present invention in the mouse body were considered.

[0304] 2. Experimental Plan 2.1 Experimental drugs and animals Comparative Example 1: Compound of Example 17 ICR mice, male, weighing 29.2–34.9 g, were purchased from Beijing Weitong Lihua Laboratory Animal Technology Co., Ltd.

[0305] 2.2 Drug Preparation Preparation of formulation for intragastric administration: An appropriate amount of test compound was weighed out, and an appropriate amount of DMSO:PEG200=5%:95% (v / v) was added, followed by shaking with a vortex to prepare a solution with a final concentration of 0.5 mg / mL.

[0306] 2.3 Administration ICR mice, test compound intragastric group (9 mice per group). Intragastric group: After an overnight fast, the compound was administered intragastricly (dosage: 5 mg / kg, administration volume: 10 mL / kg), and food was consumed 4 hours after administration.

[0307] 3.Operation Intragastric group: Approximately 100 μL of blood was collected from the orbital vein before administration and 0.25 hours, 0.5 hours, 1 hour, 2 hours, 4 hours, 8 hours, 12 hours, and 24 hours after administration. Whole blood samples were placed in EDTA-K2 anticoagulant tubes. Plasma was collected by centrifugation (1500 g, 10 min). The upper plasma was stored at -40 to -20°C until analysis. The content of the test compound in mouse plasma after intragastric administration of the compound was measured by LC-MS / MS.

[0308] 4. Pharmacokinetic parameters The pharmacokinetic parameters of the compounds according to the present invention are shown in Table 5.

[0309] [Table 5]

[0310] Conclusion: Compared with Comparative Example 1, compound 17 of the present invention has good pharmacokinetic absorption, and the blood drug concentration, area under the curve and half-life are significantly better than those of Comparative Example 1, and has good pharmacokinetic properties.

[0311] Note: Comparative Example 1 is compound Z27-2 of WO2021083167A1, which is prepared according to Example 27 of WO2021083167A1, and its specific structure is as follows:

[0312] [ka]

[0313] Test Example 6. Pharmacodynamics test of the compound according to the present invention in a subcutaneous transplant model of NCI-H358 cells in BALB / c nude mice 1. Purpose of the experiment This study was conducted to evaluate the antitumor effect and safety of compound 17 of the present invention administered orally, intragastricly, once daily for 14 days in a subcutaneous animal model of NCI-H358 (human non-small cell lung cancer) cell line in BALB / c nude mice.

[0314] 2. Preparation of Test Substances 2.1 Preparation of blank dosage formulation: An appropriate volume of a formulation containing DMA (dimethylacetamide): 30% Solutol HS 15: Saline (physiological saline) = 10:10:80 (v / v / v) was prepared as a blank group administration test liquid.

[0315] 2.2 Preparation of oral formulation of compound 17 An appropriate amount of compound 17 was weighed into a 10 mL centrifuge tube, an appropriate amount of DMA was added, and the tube was vortexed and shaken to completely dissolve the solids. An appropriate volume of 30% Solutol HS 15 was then added, and the tube was vortexed and shaken to mix well. Then, saline was added to adjust the ratio of DMA:30% Solutol HS 15:saline to 10:10:80 (v / v / v), and a dosage formulation with a concentration of 1 mg / mL was prepared.

[0316] 3. Experimental Animals Twelve BALB / c nude mice, female, 6–7 weeks old (age of mice at the time of tumor cell inoculation), purchased from Jiangsu Jisu Yaokang Biotechnology Co., Ltd., license number: SCXK(Su) 2019-0009, animal certification number: 202113149.

[0317] 4.Cell culture NCI-H358 cells were cultured in RPMI 1640 medium containing 10% fetal bovine serum, 1% sodium pyruvate, and 1% glutamine. Exponentially growing NCI-H358 cells were harvested and resuspended in PBS to a suitable concentration for subcutaneous tumor inoculation into nude mice.

[0318] 5. Animal Inoculation and Grouping Approximately 3.7 × 10 6NCI-H358 cells were inoculated subcutaneously, and the average tumor volume was approximately 100–150 mm 3 When the tumors reached this size, they were randomly assigned to two groups of six mice each according to tumor size.

[0319] 6. Administration to animals and observation After tumor inoculation, a subcutaneous tumor model was established. Each treatment group and the vehicle control group were orally and intragastrically administered for 14 days. The animals were weighed daily, and the tumor volumes were measured twice a week.

[0320] Tumor volume (TV), relative tumor proliferation rate (T / C), relative tumor inhibition rate (TGI) and tumor inhibition rate (IR) were calculated according to the following formulas. (1) TV (tumor volume) = 1 / 2 × a × b 2 , where a and b represent the length and width of the tumor, respectively. (2)T / C (relative tumor growth rate, %)=T RTV / C RTV ×100%, but T RTV is the RTV of the treatment group, C RTV is the RTV of the control group. (3) TGI% (tumor growth inhibition rate) = (1-T / C) x 100%, where T and C are the relative tumor volumes at a particular time point in the treatment and control groups, respectively. (4) IR(%)(tumor weight tumor inhibition rate)=(1-TW t / TW c ) × 100%, but TW t is the tumor weight in the treatment group, TW c is the tumor weight in the control group.

[0321] 7.Results FIG. 1 shows the change in tumor volume of nude mouse xenograft tumors of NCI-H358 cells in BALB / c nude mice treated with compound 17 according to the present invention. FIG. 2 shows the change in tumor weight of nude mouse xenograft tumors of NCI-H358 cells in BALB / c nude mice treated with compound 17 according to the present invention.

[0322] [Table 6]

[0323] [Table 7]

[0324] As can be seen from Tables 6-7 and Figures 1-2, at a dose of 10 mg / kg (po, qd), the compound of the present invention (taking compound 17 as an example) has a significant growth inhibitory effect on the establishment of a mouse in vivo tumor model based on NCI-H358 cells within 14 days, no significant body weight change was observed, and it has good safety and tolerability.

Claims

1. A compound represented by the following general formula (III), or a stereoisomer, tautomer, or pharma- ceutically acceptable salt thereof: 【Chemistry 1】 (In the formula, G is C=O; W is NR ｆ and R ｆ is a methyl group or a deuterated methyl group, 【Chemistry 2】 teeth 【Chemistry 3】 and E is 【Chemistry 4】 and L is a chemical bond; R ａ is a hydrogen atom, R ｂ is a hydrogen atom, R ｃ is a halogen, R ３ teeth 【Chemistry 5】 and R ５ is C １ ~C ４ is an alkyl group represented by the formula:

2. R ｆ is a deuterated methyl group, or a stereoisomer, tautomer, or a pharma- ceutically acceptable salt thereof.

3. The compound according to claim 1 or 2, which is a compound represented by the following general formula (V) or (VI), or a stereoisomer, tautomer, or pharma- ceutically acceptable salt thereof: 【Chemistry 6】 (In the formula, R ｇ is a methyl group or -S-CH ３ and R ｈ is an isopropyl group, R ４ is a methyl group or a deuterated methyl group, substructure 【Chemistry 7】 are defined as in claim 1, R ５ , R ｃ The definitions of E and E are as described in claim 1.

4. R ４ is a deuterated methyl group, or a stereoisomer, tautomer, or a pharma- ceutically acceptable salt thereof.

5. R ｃ is fluorine or chlorine, or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof according to any one of claims 1 to 4.

6. 【Chemical 8】 or a stereoisomer, tautomer, or a pharma- ceutically acceptable salt thereof.

7. An effective amount of a compound according to any one of claims 1 to 6, or a stereoisomer, tautomer, or pharma- ceutically acceptable salt thereof; A pharmaceutical composition comprising a pharma- ceutically acceptable carrier, excipient, or combination thereof.

8. Use of a compound according to any one of claims 1 to 6, or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof, or a pharmaceutical composition according to claim 7, in the manufacture of a K-Ras GTPase inhibitor.

9. Use of a compound according to any one of claims 1 to 6, or a stereoisomer, tautomer or a pharma- ceutically acceptable salt thereof, or a pharmaceutical composition according to claim 7, in the manufacture of a medicament for the treatment of a disease mediated by KRAS mutations.

10. The use according to claim 9, wherein the disease mediated by KRAS mutation is cancer.

11. 11. The use of claim 10, wherein the cancer is selected from the group consisting of pancreatic cancer, colorectal cancer, lung cancer, multiple myeloma, uterine cancer, bile duct cancer, gastric cancer, bladder cancer, diffuse large B-cell lymphoma, rhabdomyosarcoma, cutaneous squamous cell carcinoma, cervical cancer, and testicular germ cell carcinoma.

12. The use according to claim 10, wherein the cancer is selected from the group consisting of pancreatic cancer, colorectal cancer and lung cancer.

13. The use according to claim 9, wherein the KRAS mutation is a KRAS G12C mutation.

14. Use of a compound according to any one of claims 1 to 6, or a stereoisomer, tautomer or pharma- ceutically acceptable salt thereof, or a pharmaceutical composition according to claim 7, in the manufacture of a medicament for treating cancer, comprising The cancer is selected from the group consisting of pancreatic cancer, colorectal cancer, lung cancer, multiple myeloma, uterine cancer, bile duct cancer, gastric cancer, bladder cancer, diffuse large B-cell lymphoma, rhabdomyosarcoma, cutaneous squamous cell carcinoma, cervical cancer, and testicular germ cell carcinoma.

15. 15. The use according to claim 14, wherein the cancer is selected from the group consisting of pancreatic cancer, colorectal cancer, and lung cancer.

16. 16. The use according to any one of claims 11, 12, 14 and 15, wherein the lung cancer is non-small cell lung cancer.

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