WO2022199170A1

WO2022199170A1 - Tetracyclic derivative, and preparation method therefor and medical use thereof

Info

Publication number: WO2022199170A1
Application number: PCT/CN2021/141061
Authority: WO
Inventors: 陈友喜; 龚亮; 向清; 毛文涛; 赵雯雯; 赵伟峰; 程超英; 叶成; 钱文建; 陈磊
Original assignee: 浙江海正药业股份有限公司; 上海昂睿医药技术有限公司
Priority date: 2021-03-26
Filing date: 2021-12-24
Publication date: 2022-09-29
Also published as: CN116113416A; CN115124533A

Abstract

The present invention relates to a tetracyclic derivative, and a preparation method therefor and the medical use thereof. Specifically, the present invention relates to a tetracyclic derivative, as represented by general formula (I), a preparation method therefor, a pharmaceutically acceptable salt thereof, and the use of the tetracyclic derivative and the pharmaceutically acceptable salt as therapeutic agents, especially as KRas G12D inhibitors. The definition of each substituent in general formula (I) are the same as that in the description.

Description

Tetracyclic derivatives, their preparation methods and their medical uses

technical field

The present invention relates to a tetracyclic derivative, a preparation method thereof, a pharmaceutical composition containing the derivative and its use as a therapeutic agent, especially as a KRas G12D inhibitor.

Background technique

RAS represents a group of closely related monomeric globular proteins (21 kDa molecular weight) that have 189 amino acids and are associated with the plasma membrane and bind GDP or GTP. Under normal developmental or physiological conditions, RAS is activated upon receipt of 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 RAS proteins is determined by nucleotide binding, with the active signaling conformation bound to GTP and the inactive conformation bound to GDP. When RAS contains bound GDP, it is in a dormant or quiescent or off state and is "inactive". When cells respond to exposure to certain growth-promoting stimuli, RAS is induced to convert bound GDP to GTP. With GTP bound, RAS is "on" and is able to interact with and activate other proteins (its "downstream targets"). The RAS protein itself has a very low intrinsic ability to hydrolyze GTP back to GDP and thereby turn itself into the off state. Switching RAS off requires exogenous proteins called GTPases activating proteins (GAPs), which interact with RAS and greatly facilitate the conversion of GTP to GDP. Any mutation in RAS that affects its ability to interact with GAP or convert GTP back to GDP will result in prolonged activation of the protein and thus an extended signal to the cell that tells it to continue growing and split. So these signals allow cells to grow and divide, and overactive RAS signaling may ultimately lead to cancer.

Structurally, RAS proteins contain a G domain responsible for the enzymatic activity of RAS, guanine nucleotide binding and hydrolysis (GTPase reaction). It also includes a C-terminal extension called the CAAX box, which can be post-translationally modified and target the 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 in the protein that binds nucleotides, and this is the rigid part of the domain with conserved amino acid residues necessary for nucleotide binding and hydrolysis (glycine 12, threo amino acid 26 and lysine 16). The G domain also contains the so-called switch I region (residues 30-40) and switch II region (residues 60-76), both of which are the dynamic part of the protein as it switches between rest and load states capability is often denoted as a "spring loaded" mechanism. The main interaction is the hydrogen bond formed by threonine-35 and glycine-60 with the γ-phosphate of GTP, which allows switch I and switch II regions, respectively, to maintain their active conformations. After hydrolysis of GTP and release of phosphate, both relax to the inactive GDP conformation.

Among RAS family members, oncogenic mutations were most frequently found in KRAS (85%), whereas NRAS (12%) and HRAS (3%) were less common. KRAS mutations are prevalent in the three most lethal cancer types in the United States: pancreatic cancer (95%), colorectal cancer (45%), and lung cancer (25%). KRAS mutations are also found in other cancer types including carcinoma, diffuse large B-cell lymphoma, rhabdomyosarcoma, cutaneous squamous cell carcinoma, cervical cancer, testicular germ cell carcinoma, etc. Rarely found (<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, the increased frequency of allele-specific mutations is mostly due to classic smoking-induced mutations (G:C to T:A substitutions), resulting in KRAS G12C (GGT to TGT) and G12V (GGT) to GTT) mutation.

Large genomic studies have shown that KRAS mutations in lung cancer, including G12C, are mutually exclusive with other known driver oncogenic mutations in NSCLC, including EGFR, ALK, ROS1, RET, and BRAF, indicating the uniqueness of KRAS mutations in lung cancer. Meanwhile, KRAS mutations frequently co-occur 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.

Three RAS oncogenes constitute the most frequently mutated gene family in human cancers. Disappointingly, despite more than three decades of research efforts, there is still no clinically effective anti-RAS therapy, and targeting this gene with small molecules is challenging. Therefore, there is an urgent need in the art for small molecules for targeting RAS (eg, K-RAS, H-RAS and/or N-RAS) and using them to treat various diseases, such as cancer.

At present, there is fierce competition for clinical development of KRas G12D inhibitors at home and abroad. Among them, MRTX-1133, a KRas G12D inhibitor developed by Mirati Therapeutics Inc, has entered the preclinical stage for the treatment of colorectal tumors, non-small cell lung cancer and pancreatic cancer. . At present, there are a few patent applications for KRas G12D inhibitors published, including WO2021041671 of Mirati Therapeutics Inc. Although some progress has been made in the research and application of KRas G12D inhibitors, the room for improvement is still huge, and it is still necessary to continue the research and development of new KRas G12D inhibitors.

SUMMARY OF THE INVENTION

The object of the present invention is to provide a tetracyclic derivative represented by the general formula (I), or a stereoisomer, a tautomer or a pharmaceutically acceptable salt thereof:

in:

Ring A is selected from aryl, heteroaryl or fused rings; preferably phenyl, naphthyl, pyridyl, benzothiazolyl or benzopyrazolyl;

X and Y are each independently selected from N or CR ^a ;

R ^a is selected from hydrogen atom, halogen, alkyl or alkoxy; wherein said alkyl or alkoxy is optionally further selected by one or more groups selected from halogen, hydroxyl, cyano, alkyl or alkoxy Substituents are substituted; R ^a is preferably halogen, more preferably fluorine or chlorine;

R ¹ are the same or different, each independently selected from hydrogen atom, halogen, hydroxyl, alkyl or alkoxy, preferably alkyl;

Alternatively, the two R1 together with the atoms to which they are attached form ^a cycloalkyl or heterocyclyl;

R ² is selected from a hydrogen atom, an alkyl group or a deuterated alkyl group, preferably an alkyl group or a deuterated alkyl group, more preferably a methyl group or a deuterated methyl group;

R ³ is selected from aryl or heteroaryl; wherein said aryl or heteroaryl is optionally further substituted by one or more ^RA ; R ³ is preferably heteroaryl;

R ^A is selected from alkyl, halogen, nitro, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, =O, ^-OR5 , -C(O) ^R5 , -C(O) OR ⁵ , -NHC(O)R ⁵ , -NHC(O)OR ⁵ , -NR ⁶ R ⁷ , -C(O)NR ⁶ R ⁷ , -CH ₂ NHC(O)OR ⁵ , -CH ₂ NR ⁶ R ⁷ or -S(O) _r R ⁵ is substituted by the substituents; wherein the alkyl, cycloalkyl, heterocyclyl, aryl or heteroaryl is optionally further selected from one or more alkyl groups , halogen, nitro, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, =O, -OR ⁵ , -C(O)R ⁵ , -C(O)OR ⁵ , -NHC ( O)R ⁵ , -NHC(O)OR ⁵ , -NR ⁶ R ⁷ , -C(O)NR ⁶ R ⁷ , -CH ₂ NHC(O)OR ⁵ , -CH ₂ NR ⁶ R ⁶ or -S( O) the substituent of _r R ⁵ is substituted;

R ⁴ is the same or different, each independently selected from hydrogen atom, alkyl group, alkynyl group, halogen, nitro group, cyano group, cycloalkyl group, heterocyclic group, aryl group, heteroaryl group, =O, -OR ⁵ , -C(O)R ⁵ , -C(O)OR ⁵ , -NHC(O)R ⁵ , -NHC(O)OR ⁵ , -NR ⁶ R ⁷ , -C(O)NR ⁶ R ⁷ , -CH ₂ NHC(O)OR ⁵ , -CH ₂ NR ⁶ R ⁷ or -S(O) _r R ⁵ substituents; wherein said alkyl, cycloalkyl, heterocyclic, aryl or heteroaryl group is optionally further selected from one or more groups selected from alkyl, halogen, nitro, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, =O, ^-OR5 , -C(O)R ⁵ , -C(O)OR ⁵ , -NHC(O)R ⁵ , -NHC(O)OR ⁵ , -NR ⁶ R ⁷ , -C(O)NR ⁶ R ⁷ , -CH ₂ NHC(O)OR ^5. Substituents of -CH ₂ NR ⁶ R ⁷ or -S(O) _r R ⁵ are substituted;

R ⁵ is selected from a hydrogen atom, an alkyl group, a cycloalkyl group, a heterocyclyl group, an aryl group or a heteroaryl group, wherein said alkyl group, cycloalkyl group, heterocyclyl group, aryl group or heteroaryl group is optionally further One or more selected from hydroxy, halogen, nitro, cyano, alkyl, alkoxy, haloalkyl, haloalkoxy, cycloalkyl, heterocyclyl, aryl, heteroaryl, =O, -C (O)R ⁸ , -C(O)OR ⁸ , -OC(O)R ⁸ , -NR ⁹ R ¹⁰ , -C(O)NR ⁹ R ¹⁰ , -SO ₂ NR ⁹ R ¹⁰ or -NR ⁹ C (O) Substituents of R ¹⁰ are substituted;

R ⁶ and R ⁷ are each independently selected from a hydrogen atom, a hydroxyl group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein the alkyl group, alkoxy group, A cycloalkyl, heterocyclyl, aryl or heteroaryl group is optionally further selected from one or more groups selected from hydroxy, halogen, nitro, cyano, alkyl, alkoxy, cycloalkyl, heterocyclyl, aryl radical, heteroaryl, =O, -C(O)R ⁸ , -C(O)OR ⁸ , -OC(O)R ⁸ , -NR ⁹ R ¹⁰ , -C(O)NR ⁹ R ¹⁰ , - SO ₂ NR ⁹ R ¹⁰ or the substituent of -NR ⁹ C(O)R ¹⁰ ;

Alternatively, R ⁶ and R ⁷ together with the atoms to which they are attached form a 4- to 8-membered heterocyclic group, wherein the 4- to 8-membered heterocyclic group contains one or more N, O or S(O) _r , and the The 4-8 membered heterocyclic group is optionally further selected from one or more groups selected from hydroxyl, halogen, nitro, cyano, alkyl, alkoxy, cycloalkyl, heterocyclic, aryl, heteroaryl , =O, -C(O)R ⁸ , -C(O)OR ⁸ , -OC(O)R ⁸ , -NR ⁹ R ¹⁰ , -C(O)NR ⁹ R ¹⁰ , -SO ₂ NR ⁹ R ¹⁰ or -NR ⁹ C(O)R ¹⁰ is substituted by the substituent;

R ⁸ , R ⁹ and R ¹⁰ are each independently selected from a hydrogen atom, an alkyl group, an amino group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein said alkyl group, cycloalkyl group, heterocyclic group , aryl or heteroaryl are optionally further selected from one or more groups selected from hydroxy, halogen, nitro, amino, cyano, alkyl, alkoxy, cycloalkyl, heterocyclyl, aryl, heteroaryl , substituted by a substituent of a carboxyl group or a carboxylate group;

m is selected from 0, 1, 2, 3 or 4;

n is selected from 0, 1, 2, 3 or 4;

r is selected from 0, 1 or 2.

Provided that, excluding compounds:

A compound represented by general formula (I), or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, which is a compound represented by general formula (II) and/or (III) or Its stereoisomers, tautomers or pharmaceutically acceptable salts thereof:

Wherein: the definitions of ring A, R ¹ to R ⁴ , X, Y, m and n are as described in claim 1 .

A compound represented by general formula (I), (II) and/or (III) or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, wherein:

X is selected from N;

Y is selected from CR ^a ;

X is selected from CR ^a ;

Y is selected from N;

R ^a is selected from hydrogen atom, halogen, alkyl or alkoxy; wherein said alkyl or alkoxy is optionally further selected by one or more groups selected from halogen, hydroxyl, cyano, alkyl or alkoxy Substituents are substituted; R ^a is preferably halogen, more preferably fluorine or chlorine.

A compound represented by general formula (I), (II) and/or (III) or its stereoisomer, tautomer or its pharmaceutically acceptable salt, wherein R ² is selected from alkyl or deuterium Alkyl, preferably methyl or deuterated methyl.

A compound represented by general formula (I), (II) and/or (III) or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, wherein ^R is selected from:

in:

R ^j is selected from hydrogen atom, halogen, nitro, cyano, hydroxyl, amino, alkyl, alkoxy, haloalkyl or haloalkoxy, preferably alkyl, more preferably methyl, ethyl or isopropyl ;

k is selected from 0, 1, 2 or 3.

A compound represented by general formula (I), (II) and/or (III) or its stereoisomer, tautomer or its pharmaceutically acceptable salt, wherein: ^R is selected from:

^R4 is selected from hydrogen atom, halogen, hydroxyl, amino, alkyl, alkoxy, alkynyl or cycloalkyl, wherein said alkyl, alkoxy, alkynyl or cycloalkyl is optionally further substituted by one or Substituted with a plurality of substituents selected from halogen, hydroxy, amino, alkyl or alkoxy.

R ⁴ is selected from fluorine, chlorine, bromine, hydroxy, amino, methyl, ethyl, trifluoromethyl, ethynyl or cyclopropyl, preferably hydroxy, fluorine or ethynyl.

Selected from:

Typical compounds of the present invention include, but are not limited to:

or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof.

NOTE: If there is a discrepancy between the drawn structure and the given name of that structure, the drawn structure will be given greater weight.

The present invention provides a method for preparing a compound of general formula (I) or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, the method comprising:

The compound of general formula (IA) and the compound of general formula (IB) are subjected to a coupling reaction under the action of a palladium catalyst, and the protecting group is further removed to obtain the compound of general formula (I);

in:

X ¹ is selected from halogen or -Sn(R ¹⁴ ) ₃ ; wherein the halogen is preferably chlorine;

R ¹⁴ is selected from alkyl, preferably methyl;

M is selected from -B(OH) ₂ , -BF ₃ K,

or halogen;

PG is a protecting group, preferably tert-butoxycarbonyl;

The definitions of ring A, R ¹ to R ⁴ , X, Y, m and n are as described in the general formula (I).

Further, the present invention provides a method for preparing a compound of general formula (I) or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, the method comprising:

The compound of general formula (IC) is reacted with the compound of general formula (ID), and the protecting group is further removed to obtain the compound of general formula (I);

in:

X ² is halogen, preferably iodine;

R ² is alkyl or deuterated alkyl, preferably methyl or deuterated methyl;

PG is a protecting group, preferably tert-butoxycarbonyl;

Rings A, R ¹ , R ³ , R ⁴ , X, Y, m and n are as defined in general formula (I).

Further, the present invention provides a compound of the general formula (IA) or a stereoisomer, a tautomer and a pharmaceutically acceptable salt thereof,

in:

R ¹⁴ is selected from alkyl, preferably methyl;

PG is selected from protecting groups, preferably tert-butoxycarbonyl;

The definitions of R ¹ to R ³ , X, Y and n are as described in the general formula (I).

Typical compounds of formula (IA) include, but are not limited to:

or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof.

Further, the present invention provides a compound of the general formula (IC) or a stereoisomer, a tautomer and a pharmaceutically acceptable salt thereof,

in:

PG is selected from protecting groups, preferably tert-butoxycarbonyl;

Typical compounds of formula (IC) include, but are not limited to:

or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof.

In another aspect, the present invention provides a pharmaceutical composition containing an effective dose of the compound of general formula (I), (II) or (III) or its stereoisomers, interconversions Isomers or pharmaceutically acceptable salts thereof, and pharmaceutically acceptable carriers, excipients or combinations thereof.

In another aspect, the present invention provides a method for inhibiting KRas G12D enzyme, wherein the method comprises administering to a patient a pharmaceutical composition containing an effective dose of general formula (I), (II) Or (III) or the compound or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof, and pharmaceutically acceptable carriers, excipients or combinations thereof.

The present invention also provides a compound of general formula (I), (II) or (III) or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof in Use in the preparation of a medicine for treating diseases mediated by KRas G12D mutation, wherein said disease mediated by KRas G12D mutation is selected from cancer, wherein said cancer is selected from cardiac myxoma, lung cancer, gastric cancer, colorectal tumor, Rectal cancer, pancreatic cancer, prostate cancer, bladder cancer, hepatocellular carcinoma, cholangiocarcinoma, cholangiocarcinoma, chondrosarcoma, multiple myeloma, uterine cancer, cervical cancer, seminoma, malignant melanoma, skin squamous cell carcinoma , adrenal neuroblastoma, myeloid leukemia, acute lymphoblastic leukemia or glioblastoma; wherein the lung cancer is selected from non-small cell lung cancer or small cell lung cancer.

In another aspect, the present invention provides a compound of general formula (I), (II) or (III) or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, or a medicament thereof Use of the composition in the preparation of a KRas G12D inhibitor.

Another aspect of the present invention relates to a method of preventing and/or treating a disease mediated by KRas G12D mutation, comprising administering to a patient a therapeutically effective amount of a compound of general formula (I), (II) or (III) or a tautomer, meso, racemate, enantiomer, diastereomer or mixture thereof or a pharmaceutically acceptable salt thereof or a pharmaceutical composition comprising the same.

The present invention also provides a compound of general formula (I), (II) or (III) or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof in Use in the preparation of a medicament for the treatment of cancer, wherein the cancer is selected from cardiac myxoma, lung cancer, gastric cancer, colorectal cancer, rectal cancer, pancreatic cancer, prostate cancer, bladder cancer, hepatocellular carcinoma, cholangiocarcinoma, cholangiocarcinoma, cartilage Sarcoma, multiple myeloma, uterine cancer, cervical cancer, seminoma, malignant melanoma, cutaneous squamous cell carcinoma, adrenal neuroblastoma, myeloid leukemia, acute lymphoblastic leukemia, or glioblastoma; of which The lung cancer is selected from non-small cell lung cancer or small cell lung cancer.

The pharmaceutical formulations of the present invention can be topical, oral, transdermal, rectal, vaginal, parenteral, intranasal, intrapulmonary, intraocular, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intradermal , intraperitoneal, subcutaneous, substratum corneum, or by inhalation. Pharmaceutical compositions containing the active ingredient may be in a form suitable for oral administration, such as tablets, dragees, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups or Elixirs. Tablets contain the active ingredient in admixture with nontoxic pharmaceutically acceptable excipients suitable for the manufacture of tablets.

The formulations of the present invention are suitably presented in unit dosage form, and such formulations may be prepared by any method well known in the art of pharmacy. The amount of active ingredient that can be combined with carrier materials to produce a single dosage form can vary depending upon the host treated and the particular mode of administration. The amount of active ingredient which, in combination with a carrier material, can produce a single dosage form generally refers to that amount of compound which produces a therapeutic effect.

Dosage forms for topical or transdermal administration of the compounds of this invention may include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches and inhalants. The active compound can be mixed under sterile conditions with a pharmaceutically acceptable carrier, and it can be mixed with any preservatives, buffers or propellants that may be required.

When the compounds of the present invention are administered in pharmaceutical form to humans and animals, the compounds may be provided alone or in a pharmaceutical composition containing in combination with a pharmaceutically acceptable carrier active ingredient, for example, 0.1% to 99.5% (more preferably, 0.5% to 90%) of the active ingredient.

Examples of pharmaceutically 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 derivatives thereof such as carboxylate (4) powdered gum tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients such as cocoa butter and Suppository waxes; (9) oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols such as propylene glycol; (11) polyols such as glycerol, 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) seaweed (16) pyrogen-free water; (17) isotonic saline; (18) Ringer's solution; (19) ethanol; (20) phosphate buffer solution; (21) cyclodextrin, e.g. linked Targeting ligands in nanoparticles, such as Accurins™; and (22) other non-toxic compatible substances in pharmaceutical formulations, such as polymer-based compositions.

Examples of pharmaceutically acceptable antioxidants include, but are not limited to: (1) water-soluble antioxidants such as ascorbic acid, cysteine hydrochloride, sodium bisulfite, sodium metabisulfite, sodium sulfite, and the like; ( 2) Oil-soluble antioxidants such as ascorbyl palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, alpha-tocopherol and the like; and (3) metal chelating agents such as citric acid, ethylenediaminetetraacetic acid (EDTA), sorbitol, tartaric acid, phosphoric acid and the like. Solid dosage forms (eg, capsules, dragees, dragees, powders, granules, and the like) may include one or more pharmaceutically acceptable carriers, such as sodium citrate or dicalcium phosphate, and/or the following Any one of: (1) fillers or bulking agents, such as starch, lactose, sucrose, glucose, mannitol and/or silicic acid; (2) binders, such as carboxymethyl cellulose, alginate, Gelatin, polyvinylpyrrolidone, sucrose and/or acacia; (3) humectants, such as glycerol; (4) disintegrants, such as agar, calcium carbonate, potato 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) humectants, such as cetyl alcohol and glycerol monostearate; (8) absorption agents 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 pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active ingredient, liquid dosage forms may contain inert diluents commonly used in the art, such as water or other solvents; solubilizers and emulsifiers, such as ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzene Methanol, 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 Diols and fatty acid esters of sorbitan, and mixtures thereof.

Suspensions, in addition to the active compounds, may also contain suspending agents such as ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum hydroxide oxide, bentonite, agar-agar and tragacanth and mixtures thereof.

Ointments, pastes, creams and gels can contain, in addition to the active compound, excipients such as animal and vegetable fats, oils, waxes, paraffins, starches, tragacanth, cellulose derivatives, poly Ethylene glycol, polysiloxane, bentonite, silicic acid, talc and zinc oxide or mixtures thereof.

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 of these substances. The sprays can contain other customary propellants, such as chlorofluorohydrocarbons, and volatile unsubstituted hydrocarbons, such as butane and propane.

Detailed description of the invention

Unless stated to the contrary, some terms used in the specification and claims of the present invention are defined as follows:

"Chemical bond" means that the indicated substituent does not exist, and both ends of the substituent are directly connected to form a bond.

"Alkyl" when taken as a group or part of a group is meant to include _C1 - _C20 straight or branched chain aliphatic hydrocarbon groups. Preferably it is a C ₁ -C ₁₀ alkyl group, more preferably a C ₁ -C ₆ alkyl group. Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-di Methylpropyl, 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-dimethyl Butyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl Wait. Alkyl groups can be substituted or unsubstituted.

"Alkenyl" refers to an alkyl group as defined above consisting 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, etc. Alkenyl groups can be optionally substituted or unsubstituted.

"Alkynyl" refers to an aliphatic hydrocarbon group containing a carbon-carbon triple bond, which may be straight or branched. Preferred are C2 _- _C10 alkynyl groups, more preferably C2 _- _C6 alkynyl groups, and most 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, and the like. Alkynyl groups can be substituted or unsubstituted.

"Alkylene" is a divalent alkyl group. It is preferably a C ₁ -C ₁₀ alkylene group, more preferably a C1 -C6 alkylene group, and particularly preferably a C ₁ -C ₄ alkylene group. Examples of alkylene groups include, but are not limited to, methylene, ethylene, -CH( _CH3 ) ₂ -n-propylene, and the like. Alkylene groups can be substituted or unsubstituted.

"Cycloalkyl" refers to saturated or partially saturated monocyclic, fused, bridged and spirocyclic carbocyclic rings. Preferably it is C ₃ -C ₁₂ cycloalkyl, more preferably C ₃ -C ₈ cycloalkyl, most preferably C ₃ -C ₆ cycloalkyl. Examples of monocyclic cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptyl Alkenyl, cyclooctyl, etc., preferably cyclopropyl and cyclohexenyl. Cycloalkyl groups can be optionally substituted or unsubstituted.

"Spirocycloalkyl" refers to a polycyclic group with 5 to 18 members, two or more cyclic structures, and the single rings share one carbon atom (called spiro atom) with each other, and the ring contains one or more aromatic systems with double bonds but none of the rings have fully conjugated pi electrons. Preferably it is 6 to 14 yuan, more preferably 7 to 10 yuan. According to the number of spiro atoms shared between the rings, spirocycloalkyl groups are divided into mono-spiro, double-spiro or poly-spirocycloalkyl groups, preferably mono-spiro and double-spirocycloalkyl groups, preferably 4-membered/5-membered, 4-membered Yuan/6 Yuan, 5 Yuan/5 Yuan or 5 Yuan/6 Yuan. Non-limiting examples of "spirocycloalkyl" include, but are not limited to, spiro[4.5]decyl, spiro[4.4]nonyl, spiro[3.5]nonyl, spiro[2.4]heptyl.

"Fused cycloalkyl" refers to a 5- to 18-membered all-carbon polycyclic group containing two or more cyclic structures sharing a pair of carbon atoms with each other, one or more rings may contain one or more double bonds, But none of the rings have an aromatic system with fully conjugated pi electrons, preferably 6 to 12 membered, more preferably 7 to 10 membered. According to the number of constituent rings, it can be divided into bicyclic, tricyclic, tetracyclic or polycyclic fused cycloalkyl, preferably bicyclic or tricyclic, more preferably 5-membered/5-membered or 5-membered/6-membered bicycloalkyl. Non-limiting examples of "fused cycloalkyl" include, but are not limited to: bicyclo[3.1.0]hexyl, bicyclo[3.2.0]hept-1-enyl, bicyclo[3.2.0]heptyl, Decalinyl or tetrahydrophenanthryl.

"Bridged cycloalkyl" refers to an all-carbon polycyclic group of 5 to 18 members, containing two or more cyclic structures, sharing two carbon atoms that are not directly connected to each other, and one or more rings may contain one or more Aromatic systems in which multiple double bonds, but none of the rings have fully conjugated pi electrons, are preferably 6 to 12 membered, more preferably 7 to 10 membered. Preferably it is 6 to 14 yuan, more preferably 7 to 10 yuan. According to the number of constituent rings, it can be divided into bicyclic, tricyclic, tetracyclic or polycyclic bridged cycloalkyl, preferably bicyclic, tricyclic or tetracyclic, more preferably bicyclic or tricyclic. Non-limiting examples of "bridged cycloalkyl" include, but are not limited to: (1s,4s)-bicyclo[2.2.1]heptyl, bicyclo[3.2.1]octyl, (1s,5s)-dicyclo[3.2.1]octyl Cyclo[3.3.1]nonyl, bicyclo[2.2.2]octyl, (1r,5r)-bicyclo[3.3.2]decyl.

"Heterocyclyl," "heterocycle," or "heterocyclic" are used interchangeably herein to refer to a non-aromatic heterocyclyl in which one or more of the ring-forming atoms is a heteroatom, such as oxygen, Nitrogen, sulfur atoms, etc., including monocyclic, fused, bridged and spiro rings. It preferably has a 5- to 7-membered monocyclic ring or a 7- to 10-membered bi- or tricyclic ring, which may contain 1, 2 or 3 atoms selected from nitrogen, oxygen and/or sulfur. Examples of "heterocyclyl" 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-bicyclo[3.2.1]octyl and piperazinyl . Heterocyclyl groups can be substituted or unsubstituted.

"Spiroheterocyclyl" refers to a polycyclic group with 5 to 18 members, two or more cyclic structures, and single rings share one atom with each other, and the ring contains one or more double bonds, but no An aromatic system with fully conjugated pi electrons in one ring, wherein one or more ring atoms are selected from nitrogen, oxygen, or a heteroatom of S(O) _r (wherein r is selected from 0, 1, or 2), and the remaining ring atoms are carbon. Preferably it is 6 to 14 yuan, more preferably 7 to 10 yuan. Spirocycloalkyl groups are classified into mono-spiroheterocyclyl, bis-spiroheterocyclyl or poly-spiroheterocyclyl according to the number of spiro atoms shared between rings, preferably mono-spiroheterocyclyl and bis-spiroheterocyclyl. More preferably, it is a 4-membered/4-membered, 4-membered/5-membered, 4-membered/6-membered, 5-membered/5-membered or 5-membered/6-membered monospiroheterocyclyl group. Non-limiting examples of "spiroheterocyclyl" include, but are not limited to: 1,7-dioxaspiro[4.5]decyl, 2-oxa-7-azaspiro[4.4]nonyl, 7-oxaspiro[4.4]nonyl Heterospiro[3.5]nonyl and 5-oxaspiro[2.4]heptyl.

"Fused heterocyclic group" refers to an all-carbon polycyclic group containing two or more ring structures sharing a pair of atoms with each other, one or more rings may contain one or more double bonds, but no ring has complete Conjugated pi-electron aromatic systems wherein one or more ring atoms are selected from nitrogen, oxygen or heteroatoms of S(O) _r (wherein r is selected from 0, 1 or 2) and the remaining ring atoms are carbon. Preferably it is 6 to 14 yuan, more preferably 7 to 10 yuan. According to the number of constituent rings, it can be divided into bicyclic, tricyclic, tetracyclic or polycyclic fused heterocyclic groups, preferably bicyclic or tricyclic, more preferably 5-membered/5-membered or 5-membered/6-membered bicyclic fused heterocyclic groups. Non-limiting examples of "fused heterocyclyl" include, but are not limited to: octahydropyrrolo[3,4-c]pyrrolyl, octahydro-1H-isoindolyl, 3-azabicyclo[3.1. 0] Hexyl, octahydrobenzo[b][1,4]dioxine.

"Bridged heterocyclyl" refers to a 5- to 14-membered, 5- to 18-membered polycyclic group containing two or more cyclic structures that share two atoms that are not directly connected to each other, and one or more rings may be Aromatic systems containing one or more double bonds but none of the rings have fully conjugated pi electrons, wherein one or more ring atoms are selected from nitrogen, oxygen or S(O) _r (wherein r is selected from 0, 1 or 2), the remaining ring atoms are carbon. Preferably it is 6 to 14 yuan, more preferably 7 to 10 yuan. According to the number of constituent rings, it can be divided into bicyclic, tricyclic, tetracyclic or polycyclic bridged heterocyclic groups, preferably bicyclic, tricyclic or tetracyclic, more preferably bicyclic or tricyclic. Non-limiting examples of "bridged heterocyclyl" include, but are not limited to: 2-azabicyclo[2.2.1]heptyl, 2-azabicyclo[2.2.2]octyl, and 2-azabicyclo Cyclo[3.3.2]decyl.

"Aryl" refers to a carbocyclic aromatic system containing one or two rings, wherein the rings may be joined together in a fused fashion. The term "aryl" includes monocyclic or bicyclic aryl groups such as phenyl, naphthyl, tetrahydronaphthyl aromatic groups. Preferred aryl groups are _C6 - _C10 aryl groups, more preferred aryl groups are phenyl and naphthyl, and most preferred are naphthyl. Aryl groups can be substituted or unsubstituted.

"Heteroaryl" refers to an aromatic 5 to 6 membered monocyclic or 8 to 10 membered bicyclic ring, which may contain 1 to 4 atoms selected from nitrogen, oxygen and/or sulfur. Preferably bicyclic heteroaryl, examples of "heteroaryl" include but are not limited to furyl, pyridyl, 2-oxo-1,2-dihydropyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, thienyl, isoxazolyl, oxazolyl, oxadiazolyl, imidazolyl, pyrrolyl, pyrazolyl, triazolyl, tetrazolyl, thiazolyl, isothiazolyl, 1,2,3-thiazolyl oxadiazolyl, benzodioxolyl, benzothienyl, benzimidazolyl, indolyl, isoindolyl, 1,3-dioxo-isoindolyl, quinolinyl , Indazolyl, Benzisothiazolyl, Benzoxazolyl, Benzisoxazolyl,

Heteroaryl groups can be substituted or unsubstituted.

"Fused ring" refers to a polycyclic group in which two or more ring structures share a pair of atoms with each other, one or more rings may contain one or more double bonds, but at least one ring is not completely conjugated A pi-electron aromatic system, wherein the ring atoms are selected from 0, one or more heteroatoms selected from nitrogen, oxygen or S(O) _r (wherein r is selected from 0, 1 or 2), and the remaining ring atoms are carbon . The condensed ring preferably includes a bicyclic or tricyclic condensed ring, wherein the bicyclic condensed ring is preferably a condensed ring of an aryl group or a heteroaryl group and a monocyclic heterocyclic group or a monocyclic cycloalkyl group. Preferably it is 7 to 14 yuan, more preferably 8 to 10 yuan. Examples of "fused rings" include, but are not limited to:

"Alkoxy" refers to a group (alkyl-O-). Wherein, alkyl is as defined herein. Ci _- _C6 alkoxy groups are preferred. Examples include, but are not limited to: methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, and the like.

"Haloalkyl" means a group in which an alkyl group is optionally further substituted with one or more halogens, wherein alkyl is as defined herein.

"Deuterated alkyl" refers to a group in which an alkyl group is optionally further substituted with one or more deuterium atoms, wherein alkyl is as defined herein. "Deuterated alkyl" is preferably deuterated methyl, including: mono-deuterated methyl, di-deuterated methyl and tri-deuterated methyl, preferably tri-deuterated methyl.

"Hydroxyalkyl" refers to a group in which an alkyl group is optionally further substituted with one or more hydroxy groups, wherein alkyl is as defined herein.

"Haloalkoxy" refers to a group in which the alkyl group of (alkyl-O-) is optionally further substituted with one or more halogens, wherein alkoxy is as defined herein.

"Hydroxy" refers to the -OH group.

"Halogen" refers to fluorine, chlorine, bromine and iodine.

"Amino" refers to _-NH2 .

"Cyano" refers to -CN.

"Nitro" refers to _-NO2 .

"Benzyl" refers to _-CH2 -phenyl.

"Carboxyl" refers to -C(O)OH.

"Carboxylate" means -C(O)O-alkyl or -C(O)O-cycloalkyl, wherein alkyl and cycloalkyl are as defined above.

"DMSO" refers to dimethyl sulfoxide.

"BOC" refers to t-butoxycarbonyl.

"Ts" refers to p-toluenesulfonyl.

"T3P" refers to propylphosphoric 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.

"Substituted" means that one or more hydrogen atoms in a group, preferably up to 5, more preferably 1 to 3 hydrogen atoms, independently of one another, are substituted by the corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and the person skilled in the art can determine (either experimentally or theoretically) possible or impossible substitutions without undue effort. For example, amino or hydroxyl groups with free hydrogens may be unstable when combined with carbon atoms with unsaturated (eg, olefinic) bonds.

"Substituted" or "substituted" mentioned in this specification, unless otherwise specified, means that the group may be substituted by one or more groups selected from the following groups: alkyl, alkenyl, alkynyl, alkoxy , alkylthio, alkylamino, halogen, mercapto, hydroxyl, nitro, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkane Thio, heterocycloalkylthio, amino, haloalkyl, hydroxyalkyl, carboxyl, carboxylate, =O, -C(O)R ⁵ , -C(O)OR ⁵ , -NHC(O)R ⁵ , -NHC(O)OR ⁵ , -NR ⁶ R ⁷ , -C(O)NR ⁶ R ⁷ , -CH ₂ NHC(O)OR ⁵ , -CH ₂ NR ⁶ R ⁷ or -S(O) _r Replaced by the substituent of R ⁵ ;

r is 0, 1 or 2.

The compounds of the present invention may contain asymmetric centers or chiral centers and therefore exist in different stereoisomeric forms. All stereoisomeric forms of the compounds of the present invention are contemplated, including but not limited to diastereomers, enantiomers, and atropisomers and geometric (conformational) isomers and Their mixtures, such as racemic mixtures, are within the scope of the present invention.

Unless otherwise indicated, the structures depicted herein also include all isomeric (eg, diastereomeric, enantiomeric, and atropisomeric and geometric (conformational) isomeric forms of such structures; for example, , the R and S configuration of each asymmetric center, the (Z) and (E) double bond isomers, and the (Z) and (E) conformational isomers. Thus the individual stereoisomers of the compounds of the present invention as well as Enantiomeric mixtures, diastereomeric mixtures and geometric (conformational) isomer mixtures are all within the scope of the present invention.

"Pharmaceutically acceptable salts" refers to certain salts of the above-mentioned compounds that retain their original biological activity and are suitable for medicinal use. The pharmaceutically acceptable salt of the compound represented by the formula (I) may be a metal salt or an amine salt with a suitable acid.

"Pharmaceutical composition" means a mixture containing one or more of the compounds described herein, or a physiologically pharmaceutically acceptable salt or prodrug thereof, with other chemical components, and other components such as physiologically pharmaceutically acceptable carriers and excipients Form. The purpose of the pharmaceutical composition is to facilitate the administration to the organism, facilitate the absorption of the active ingredient and then exert the biological activity.

Synthetic method of the compound of the present invention

In order to accomplish the purpose of the present invention, the present invention adopts following technical scheme:

The preparation method of the compound described in the general formula (I) of the present invention or its stereoisomer, tautomer or its pharmaceutically acceptable salt comprises the following steps:

in:

R ¹⁴ is selected from alkyl, preferably methyl;

M is selected from -B(OH) ₂ , -BF ₃ K,

or halogen;

PG is a protecting group, preferably tert-butoxycarbonyl;

Further, the preparation method of the compound described in the general formula (I) of the present invention or its stereoisomer, tautomer or its pharmaceutically acceptable salt comprises the following steps:

in:

X ² is halogen, preferably iodine;

R ² is alkyl or deuterated alkyl, preferably methyl or deuterated methyl;

PG is a protecting group, preferably tert-butoxycarbonyl;

Detailed ways

The following examples are used to further describe the present invention, but these examples do not limit the scope of the present invention.

Example

The examples give the preparation and related structural identification data of the representative compounds represented by formula (I). It must be noted that the following examples are used to illustrate the present invention rather than limit it. The ¹ H NMR spectrum was measured with a Bruker instrument (400 MHz), and the chemical shifts were expressed in ppm. A tetramethylsilane internal standard (0.00 ppm) was used. ¹ HNMR representation: s=singlet, d=doublet, t=triplet, m=multiplet, br=broadened, dd=doublet of doublet, dt=doublet of triplet. When coupling constants are provided, they are in Hz.

Mass spectrum is obtained by LC/MS instrument, and the ionization mode can be ESI or APCI.

The thin layer chromatography silica gel plate uses Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plate, the size of the silica gel plate used for thin layer chromatography (TLC) is 0.15mm ~ 0.2mm, and the size of the TLC separation and purification products is 0.4mm ~0.5mm.

Column chromatography generally uses Yantai Huanghai silica gel 200-300 mesh silica gel as the carrier.

In the following examples, all temperatures are in degrees Celsius unless otherwise indicated, and various starting materials and reagents were obtained from commercially available or synthesized according to known methods without further purification. For direct use, unless otherwise specified, commercially available manufacturers include but are not limited to Shanghai Haohong Biomedical Technology Co., Ltd., Shanghai Shaoyuan Reagent Co., Ltd., Shanghai Bide Pharmaceutical Technology Co., Ltd., Saen Chemical Technology (Shanghai) Co., Ltd. and Shanghai Lingkai Pharmaceutical Technology Co., Ltd., etc.

_CD3OD : Deuterated methanol.

CDCl ₃ : deuterated chloroform.

DMSO-d ₆ : deuterated dimethyl sulfoxide.

There is no special description in the examples, and the solution in the reaction refers to an aqueous solution.

The compound was purified using column chromatography and thin layer chromatography eluent system, wherein the system was selected from: A: petroleum ether and ethyl acetate system; B: dichloromethane and methanol system; C: dichloromethane and ethyl acetate system, D: dichloromethane and ethanol system, E: tetrahydrofuran and petroleum ether system, F: tetrahydrofuran and methanol system, the volume ratio of the solvent varies according to the polarity of the compound, and a small amount of acid can also be added Or alkaline reagents, such as acetic acid or triethylamine, etc.

Example 1

(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

(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

first step

6-chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)nicotinic acid

6-Chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)nicotinic acid

2-Isopropyl-4-methylpyridin-3-amine 1d (21.46g, 142.86mmol) was dissolved in 200mL of tetrahydrofuran, cooled to -78°C, and bis(trimethylsilyl)amino was added dropwise under nitrogen protection Lithium (1.0 M, 238.11 mL) was stirred at -78°C for 15 minutes, and a solution of 2,6-dichloro-5-fluoronicotinic acid 1a (20 g, 95.24 mmol) in tetrahydrofuran (100 mL) was added dropwise. After 1 hour of reaction at -78°C, 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 pH of the aqueous phase was adjusted to pH=4 with 2M dilute hydrochloric acid, the layers were separated, the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 6-chloro-5-fluoro-2-((2-isopropyl- 4-Methylpyridin-3-yl)amino)nicotinic acid 1b (15 g, 46.33 mmol), yield: 48.65%.

MS m/z(ESI): 323.8[M+1] ⁺

second step

ethyl

3-(6-chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)pyridin-3-yl)-2-nitro-3-oxopropanoate

3-(6-Chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)pyridin-3-yl)-2-nitro-3-oxopropane Ethyl acetate

6-Chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)nicotinic acid 1b (5 g, 15.44 mmol) was dissolved in N,N-dimethylmethane To the amide (50 mL), potassium carbonate (6.40 g, 46.33 mmol) and ethyl 2-nitroacetate (6.17 g, 46.33 mmol) were added, followed by 2-chloro-1-picoline iodide (7.89 g, 30.89 g) mmol), and reacted at 25°C for 3 hours. 10 mL of ethyl acetate and 10 mL of saturated brine were added to separate the layers. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was separated and purified by flash silica gel column chromatography (eluent: E system). yielded 3-(6-chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)pyridin-3-yl)-2-nitro-3-oxo Ethyl propionate 1c (2.3 g, 5.24 mmol), yield: 33.94%.

MS m/z(ESI): 438.9[M+1] ⁺

third step

7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H)-dione

7-Chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H)-di ketone

3-(6-Chloro-5-fluoro-2-((2-isopropyl-4-methylpyridin-3-yl)amino)pyridin-3-yl)-2-nitro-3-oxo Ethyl propionate 1c (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 reaction was stirred at 50° C. for 16 hours. After the reaction was completed, it was cooled to 25°C, 10 mL of ethyl acetate and 10 mL of saturated brine were added, and the layers were separated. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was subjected to flash silica gel column chromatography (washed). Removal agent: E system-F system) separation and purification to obtain 7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8- Naphthyridine-2,4(1H,3H)-dione 1e (1.7 g, 4.33 mmol), yield: 82.58%.

MS m/z(ESI): 393.0[M+1] ⁺

the fourth step

4,7-dichloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one

7-Chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H)- The diketone 1e (400 mg, 1.02 mmol) was dissolved in phosphorus oxychloride (3 mL) and reacted at 90° C. for 1 hour. The progress of the reaction was monitored by LC-MS. After the reaction was completed, concentrated under reduced pressure, and the obtained residue was separated and purified 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 1f (350 mg, 851.14 μmol), yield: 83.58%.

MS m/z(ESI): 410.8[M+1] ⁺

the fifth step

1-(tert-butyl)3-methyl

(3R,6R)-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-1,3-dicarboxylate

(3R,6R)-1-N-tert-butoxycarbonyl-4-(7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro Methyl-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-3-carboxylate

4,7-Dichloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one 1f (1.3 g, 3.16 mmol) was dissolved in acetonitrile (15 mL), and (3R,6R)-1-N-tert-butoxycarbonyl-6-methylpiperazine-3-carboxylic acid methyl ester 1j (1.63 g, 6.32 mmol), reacted at 80°C for 16 hours. The progress of the reaction was monitored by LC-MS. After the reaction was completed, concentrated under reduced pressure, and the obtained residue was separated and purified by flash silica gel column chromatography (eluent: E system) to obtain (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-naphthalene Methyl pyridin-4-yl)-6-methylpiperazine-3-carboxylate 1 g (1 g, 1.58 mmol), yield: 49.97%.

MS m/z(ESI): 633.0[M+1] ⁺

Step 6

1-(tert-butyl)3-methyl

(3R,6R)-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-1,3-dicarboxylate

(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-carboxylic acid methyl ester

(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-carboxylic acid methyl ester 1g (1g, 1.58mmol) and Raney nickel (10 mg, 157.96 μmol) was dissolved in tetrahydrofuran (10 mL), replaced with hydrogen 3 times, and reacted at 25° C. for 2 hours under the protection of hydrogen. Filtration, the filtrate was concentrated under reduced pressure to give crude (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-carboxylic acid methyl ester 1h (0.83 g, 1.38 mmol), used directly in the next reaction. Yield: 87.13%.

MS m/z(ESI): 603.3[M+1] ⁺

Step 7

tert-butyl

(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

(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-Carboxylic acid tert-butyl ester

(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-carboxylic acid methyl ester 1h (0.83g, 1.38mmol) and potassium carbonate ( 570.64 mg, 4.13 mmol) was dissolved in N,N-dimethylformamide (10 mL) and reacted at 50°C for 1 hour. After the reaction, 10 mL of ethyl acetate and 10 mL of saturated brine were added, and the layers were separated. 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 ester 1k (0.7 g, 1.23 mmol), used directly in the next reaction.

MS m/z(ESI): 571.0[M+1] ⁺

Step 8

tert-butyl

(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

(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

(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-Carboxylic acid tert-butyl ester 1k (0.7 g, 1.23 mmol), iodomethane (521.98 mg, 3.68 mmol) and potassium carbonate (508.27 mg, 3.68 mmol) were dissolved in N,N-dimethylformamide (10 mL) in the reaction at 25°C for 16 hours. 10 mL of ethyl acetate and 10 mL of water were added, and the layers were separated. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was separated and purified by flash silica gel column chromatography (eluent: E system) to obtain (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 1 m (0.45 g, 769.14 μmol), yield: 62.74%.

MS m/z(ESI): 585.0[M+1] ⁺

Step 9

tert-butyl

(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]p yrazino [2,3-c][1,8]naphthyridine-3-carboxylate

(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-pyrazine tert-Butyl [1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate

(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 1m (0.12g, 205.10μmol), (2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)boronic acid 1p (192.05mg, 615.31μmol), tetrakis(triphenylphosphine)palladium (23.70mg, 20.51μmol) and potassium phosphate (217.68mg, 1.03mmol) were dissolved in a mixed solvent of 0.2mL of water and 1mL of 1,4-dioxane During the reaction, nitrogen was replaced three times, and the reaction was carried out at 100° C. for 16 hours under a nitrogen atmosphere. The progress of the reaction was monitored by LC-MS. After the reaction was completed, concentrated under reduced pressure, and the obtained residue was separated and purified 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 ester In (0.1 g, 122.41 μmol), yield: 59.68%.

MS m/z(ESI): 817.4[M+1] ⁺

Step 10

(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-pyridine Azino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester 1n (0.1 g, 122.41 μmol) in dichloro To methane (2 mL), trifluoroacetic acid (300 mg, 2.63 mmol) was added, and the reaction was carried out at 20° C. for 16 hours. Concentration under reduced pressure gave crude (2R,4aR)-10-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-11-fluoro-8-(2-isopropyl-4-methanone) 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 1 (100 mg, 136.85 μmol).

MS m/z(ESI): 617.5[M+1] ⁺

Example 2

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

(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

first step

tert-butyl

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

(2R,4aR)-11-Fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-5,7-dioxo-10-(tri methylstannyl)-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

(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 1m (0.1g, 170.92μmol), hexamethyldistin (139.99mg, 427.30μmol) and tetrakis(triphenylphosphine)palladium (19.75mg, 17.09μmol) were dissolved in In 1,4-dioxane (1 mL), nitrogen was replaced three times, and the reaction was carried out at 110° C. for 16 hours under a nitrogen atmosphere. Concentrated under reduced pressure, the obtained residue was separated and purified by flash silica gel column chromatography (eluent: E system) to obtain (2R,4aR)-11-fluoro-8-(2-isopropyl-4-methyl) Pyridin-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 2a (95 mg, 133.16 μmol ), yield: 77.91%.

MS m/z(ESI): 714.8[M+1] ⁺

second step

tert-butyl

(2R,4aR)-10-(6-amino-3-chloropyridin-2-yl)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-di 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

(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]pyrazine tert-Butyl [2,3-c][1,8]naphthyridine-3-carboxylate

(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 2a (40 mg, 56.07 μmol), 6-bromo-5-chloropyridin-2-amine 2b (13.96 mg, 67.28 μmol), iodide Copper (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), replaced with nitrogen three times, under nitrogen atmosphere, 100 The reaction was carried out at °C for 16 hours. Concentrated under reduced pressure, the obtained residue was separated and purified by flash silica gel column chromatography (eluent: E system) to obtain (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 2c (15 mg, 22.15 μmol), yield: 39.51%.

MS m/z(ESI): 677.3[M+1] ⁺

third step

(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]pyridine Azino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester 2c (30 mg, 44.30 μmol) was dissolved in dichloromethane (3 mL), trifluoroacetic acid (1 g, 8.77 mmol) was added ) for 16 hours at 20°C. Concentration under reduced pressure gave (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 2 (30 mg, 51.99 μmol).

MS m/z(ESI): 577.0[M+1] ⁺

Example 3

(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

(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

first step

tert-butyl

(2R,4aR)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-10-(3-methoxynaphthalen-1-yl)-2,6-dime thyl-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

(2R,4aR)-11-Fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-10-(3-methoxynaphthalen-1-yl)-2,6-di 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

(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 1m (100mg, 170.92μmol), (3-methoxynaphthalen-1-yl)boronic acid 3a (103.58mg, 512.76μmol), tetrakis(triphenylphosphine)palladium ( 19.75mg, 17.09μmol) and potassium phosphate (181.40mg, 854.60μmol) were dissolved in a mixed solvent of 0.3mL of water and 1.5mL of 1,4-dioxane, nitrogen was replaced 3 times, under nitrogen atmosphere, at 100 ℃ The reaction was carried out for 16 hours. The progress of the reaction was monitored by LC-MS. Concentrated under reduced pressure, the obtained residue was separated and purified by flash silica gel column chromatography (eluent: E system) to obtain (2R,4aR)-11-fluoro-8-(2-isopropyl-4-methyl) Pyridin-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 3b ( 0.1 g, 141.48 μmol), yield: 82.78%.

MS m/z(ESI): 707.7[M+1] ⁺

second step

(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 3b (70 mg, 99.04 μmol) was dissolved in 5 mL of dichloromethane, and boron tribromide (1.40 g, 5.59 mmol) was added, The reaction was carried out at 20°C for 16 hours. Add 10 mL of methanol to quench the reaction, concentrate under reduced pressure, add 20 mL of water to dilute the reaction, wash with ethyl acetate (20 mL×2), collect the aqueous phase, and freeze the aqueous phase to obtain (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 3 (100mg, 168.73 μmol).

MS m/z(ESI): 593.4[M+1] ⁺

Example 4

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

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

first step

tert-butyl

(2R,4aR)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-10-(5-methyl-1H-indazol-4-yl)-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

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

(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 1m (50mg, 85.46μmol), (5-methyl-1H-indazol-4-yl)boronic acid 4a (45.12mg, 256.38μmol), tetrakis(triphenylphosphine) ) Palladium (9.88 mg, 8.55 μmol) and potassium phosphate (54.42 mg, 256.38 μmol) were dissolved in a mixed solvent of 0.3 mL of water and 1.5 mL of 1,4-dioxane, and the nitrogen was replaced 3 times. The reaction was carried out at 100°C for 16 hours. The progress of the reaction was monitored by LC-MS. After the reaction was completed, concentrated under reduced pressure, the obtained residue was separated and purified by flash silica gel column chromatography (eluent: E system) to obtain (2R,4aR)-11-fluoro-8-(2-isopropyl- 4-Methylpyridin-3-yl)-2,6-dimethyl-10-(5-methyl-1H-indazol-4-yl)-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- tert-Butyl carboxylate 4b (10 mg, 14.69 μmol), yield: 17.19%.

MS m/z(ESI): 681.1[M+1] ⁺

second step

(2R,4aR)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-10-(5-methyl-1H-indone) azol-4-yl)-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 4b (10mg, 14.69μmol) was dissolved in 2mL 1,4-dioxane, and 4M dilute hydrochloric acid was added dropwise (36.72 μL), reacted at 25°C for 2 hours. The progress of the reaction was monitored by LC-MS. After the reaction, concentrated under reduced pressure to obtain crude product (2R,4aR)-11-fluoro-8-(2-isopropyl-4-methylpyridin-3-yl)-2,6-dimethyl-10- (5-Methyl-1H-indazol-4-yl)-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino [2,3-c][1,8]Naphthyridine-5,7-dione 4 (10 mg, 17.22 μmol).

MS m/z(ESI): 581.2[M+1] ⁺

Example 5

(2R,4aR)-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-methylpyridin-3-yl)-2-methyl-6-(methyl-d3)- 2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione

(2R,4aR)-11-Chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl-4-methylpyridin-3-yl)-2-methyl-6 -(Methyl-d3)-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1',2':4,5]pyrazino[2,3-c][ 1,8]Naphthyridine-5,7-dione

first step

2,5-Dichloro-6-(2-fluoro-6-methoxyphenyl)nicotinic acid

2,5,6-Trichloronicotinic acid 5a (20 g, 88.32 mmol) and (2-fluoro-6-methoxyphenyl)boronic acid (30.02 g, 176.64 mmol) were dissolved in 1,4-dioxane (200 mL) and water (40 mL) were added tetrakis(triphenylphosphine) palladium (2.00 g, 1.73 mmol) and sodium carbonate (28.08 g, 264.97 mmol), heated to 100° C. under argon protection, and reacted overnight. After the reaction was completed, cooled, added 100 mL of water, extracted with ethyl acetate (100 mL×2), combined the organic phases, the aqueous phase was adjusted to acidity with 1M dilute hydrochloric acid, extracted with ethyl acetate (100 mL×2), the organic phases were combined, Dry over anhydrous sodium sulfate, filter, and concentrate under reduced pressure to give crude product 2,5-dichloro-6-(2-fluoro-6-methoxyphenyl)nicotinic acid 5b (27 g, 85.41 mmol) in yield : 96.71%.

MS m/z(ESI): 315.8[M+1] ⁺

second step

Methyl 2,5-dichloro-6-(2-fluoro-6-methoxyphenyl)nicotinate

2,5-Dichloro-6-(2-fluoro-6-methoxyphenyl)nicotinic acid 5b (26 g, 82.25 mmol) was dissolved in methanol (200 mL), and thionyl chloride ( 19.57 g, 164.50 mmol, 11.93 mL), heated to 90 °C and reacted for 6 hours. After the reaction, cooled, concentrated under reduced pressure, added 200 mL of water, added dropwise saturated sodium bicarbonate solution to adjust the pH to basic, extracted with ethyl acetate (100 mL×2), combined the organic phases, and added saturated brine (100 mL) to the organic phase. ×3) washed, dried with anhydrous sodium sulfate, filtered and concentrated under reduced pressure, the obtained residue was purified by silica gel column chromatography (eluent: system A) to obtain 2,5-dichloro-6-(2 -Fluoro-6-methoxyphenyl)nicotinic acid methyl ester 5c (17.16 g, 51.98 mmol), yield: 63.20%.

MS m/z(ESI): 329.8[M+1] ⁺

¹ H NMR (400MHz, CDCl ₃ -d) δ 8.28-8.23 (m, 1H), 7.42-7.36 (m, 1H), 6.82-6.77 (m, 2H), 3.99 (s, 3H), 3.78 (s) , 3H).

third step

methyl 5-chloro-6-(2-fluoro-6-methoxyphenyl)-2-((2-isopropyl-4-methylpyridin-3-yl)amino)nicotinate methyl

5-chloro-6-(2-fluoro-6-methoxyphenyl)-2-((2-isopropyl-4-methylpyridin-3-yl)amino)nicotinate

2,5-Dichloro-6-(2-fluoro-6-methoxyphenyl)nicotinic acid methyl ester 5c (16.8 g, 50.89 mmol) and 2-isopropyl-4-methylpyridine-3- Amine 1d (8.41 g, 55.98 mmol) was dissolved in anhydrous 1,4-dioxane (200 mL), cesium carbonate (49.74 g, 152.66 mmol), 2-dicyclohexylphosphorus-2',6'- Diisopropoxy-1,1'-biphenyl Ruphos (4.75 g, 10.18 mmol) and methanesulfonic acid (2-dicyclohexylphosphino-2',6'-diisopropoxy-1,1' -biphenyl)(2-amino-1,1'-biphenyl-2-yl)palladium(II) RuPhos Pd G3 (4.26 g, 5.09 mmol), heated to 100°C under argon protection, and reacted overnight. After the reaction, 250 mL of water was added to the system, extracted with ethyl acetate (200 mL), the organic phase was washed with saturated brine (100 m×2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the residue. Purified by silica gel column chromatography (eluent: A system) to obtain 5-chloro-6-(2-fluoro-6-methoxyphenyl)-2-((2-isopropyl-4-methyl) pyridin-3-yl)amino)nicotinic acid methyl ester 5d (7.4 g, 16.67 mmol), yield: 32.76%.

MS m/z(ESI): 443.9[M+1] ⁺

the fourth step

5-Chloro-6-(2-fluoro-6-methoxyphenyl)-2-((2-isopropyl-4-methylpyridin-3-yl)amino)nicotinic acid

Methyl 5-chloro-6-(2-fluoro-6-methoxyphenyl)-2-((2-isopropyl-4-methylpyridin-3-yl)amino)nicotinate 5d (15 g , 33.79 mmol) was dissolved in a mixed solvent of methanol (50 mL) and water (10 mL), lithium hydroxide monohydrate (7.09 g, 168.96 mmol) was added, heated to 90° C., and reacted overnight. After the reaction, cooled, concentrated under reduced pressure to remove methanol, added 100 mL of water, extracted with 100 mL of ethyl acetate, washed the organic phase with saturated ammonium chloride solution (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain Crude 5-chloro-6-(2-fluoro-6-methoxyphenyl)-2-((2-isopropyl-4-methylpyridin-3-yl)amino)nicotinic acid 5e (11.4 g, 26.52 mmol), yield: 78.48%.

MS m/z(ESI): 429.9[M+1] ⁺

the fifth step

6-chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H) -dione

6-Chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthalene Pyridin-2,4(1H,3H)-dione

5-Chloro-6-(2-fluoro-6-methoxyphenyl)-2-((2-isopropyl-4-methylpyridin-3-yl)amino)nicotinic acid 5e (3 g, 6.98 mmol), ethyl 2-nitroacetate (2.79 g, 20.94 mmol) and potassium carbonate (2.89 g, 20.94 mmol) were dissolved in 30 mL of N,N-dimethylformamide, and 2-chloro-1-methylformamide was added. Pyridine iodide (3.57 g, 13.96 mmol) was reacted at room temperature for 16 hours. After the reaction, 100 mL of water was added, extracted with ethyl acetate (100 mL×2), the organic phases were combined, the organic phases were washed with saturated brine (100 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain The residue was purified by silica gel column chromatography (eluent: system B) to give 6-chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4 -Methylpyridin-3-yl)-3-nitro-1,8-naphthyridine-2,4(1H,3H)-dione 5f (800 mg, 1.60 mmol), yield: 22.9%.

MS m/z(ESI): 499.0[M+1] ⁺

Step 6

4,6-dichloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one

4,6-Dichloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1, 8-Naphthyridin-2(1H)-one

6-Chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1,8- Naphthyridine-2,4(1H,3H)-dione 5f (800 mg, 1.60 mmol) was dissolved in phosphorus oxychloride (18 g, 117.39 mmol), heated to 100° C., and reacted for 3 hours. After the reaction, the reaction solution was poured into 100 mL of ice water, extracted with dichloromethane (100 mL×2), the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was subjected to silica gel column chromatography method (eluent: B system) to obtain the product 4,6-dichloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-methylpyridine) -3-yl)-3-nitro-1,8-naphthyridin-2(1H)-one 5 g (550 mg, 1.1 mmol), yield: 68.75%.

MS m/z(ESI): 517.0[M+1] ⁺

Step 7

(3R,6R)-1-N-tert-butoxycarbonyl-4-(6-chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4- Methylpyridin-3-yl)-3-nitro-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-3-carboxylic acid methyl ester

1-(tert-butyl)3-methyl

(3R,6R)-4-(6-chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-2-oxo-1 ,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-1,3-dicarboxylate

4,6-Dichloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-1 , 8-Naphthyridin-2(1H)-one 5g (550mg, 1.1mmol) and (3R,6R)-1-N-tert-butoxycarbonyl-6-methylpiperazine-3-carboxylic acid methyl ester 1j ( 458.33 mg, 1.77 mmol) was dissolved in acetonitrile (10 mL), under argon protection, and reacted at 90 °C for 16 hours. After the reaction, 100 mL of water was added, extracted with ethyl acetate (100 mL×2), the organic phases were combined, the organic phases were washed with saturated brine (100 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain The residue was purified by silica gel column chromatography (eluent: B system) to give the product (3R,6R)-1-N-tert-butoxycarbonyl-4-(6-chloro-7-(2-fluoro) -6-Methoxyphenyl)-1-(2-isopropyl-4-methylpyridin-3-yl)-3-nitro-2-oxo-1,2-dihydro-1,8 -Naphthyridin-4-yl)-6-methylpiperazine-3-carboxylic acid methyl ester 5h (400 mg, 541.13 μmol), yield: 49.28%.

MS m/z(ESI): 739.1[M+1] ⁺

Step 8

tert-butyl

(2R,4aR)-11-chloro-10-(2-fluoro-6-methoxyphenyl)-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

(2R,4aR)-11-Chloro-10-(2-fluoro-6-methoxyphenyl)-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

(3R,6R)-1-N-tert-butoxycarbonyl-4-(6-chloro-7-(2-fluoro-6-methoxyphenyl)-1-(2-isopropyl-4 -Methylpyridin-3-yl)-3-nitro-2-oxo-1,2-dihydro-1,8-naphthyridin-4-yl)-6-methylpiperazine-3-carboxylic acid methyl The ester 5h (40 mg, 54.11 μmol) was dissolved in methanol (5 mL), 10% palladium on carbon (16.33 mg, 153.45 μmol) was added, and after hydrogen replacement for 3 times, the reaction was carried out at room temperature for 2 hours under the protection of hydrogen. After the reaction, the reaction solution was filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product (2R,4aR)-11-chloro-10-(2-fluoro-6-methoxyphenyl)-8-(2-iso Propyl-4-methylpyridin-3-yl)-2-methyl-5,7-dioxo-1,2,4,4a,5,6,7,8-octahydro-3H-pyrazine Io[1',2':4,5]pyrazino[2,3-c][1,8]naphthyridine-3-carboxylate tert-butyl ester 5i (36.5 mg).

MS m/z(ESI): 677.3[M+1] ⁺

Step 9

tert-butyl

(2R,4aR)-11-chloro-10-(2-fluoro-6-methoxyphenyl)-8-(2-isopropyl-4-methylpyridin-3-yl)-2-methyl-6-(methyl-d3)- 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

(2R,4aR)-11-Chloro-10-(2-fluoro-6-methoxyphenyl)-8-(2-isopropyl-4-methylpyridin-3-yl)-2-methyl -6-(Methyl-d3)-5,7-dione-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

(2R,4aR)-11-chloro-10-(2-fluoro-6-methoxyphenyl)-8-(2-isopropyl-4-methylpyridin-3-yl)-2-methyl yl-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 5i (1.43g, 2.11mmol) was dissolved in 10mL N,N-dimethylformamide, deuterated iodomethane (918.36mg) was added dropwise , 6.34 mmol, 394.15 μL), reacted overnight at room temperature. 10 mL of water was added to the reaction solution, extracted with ethyl acetate (10 mL×3), the organic phases were combined, washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was used 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) yl-4-methylpyridin-3-yl)-2-methyl-6-(methyl-d3)-5,7-dione-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 5j (1 g, 1.44 mmol ), yield: 68.21%.

MS m/z(ESI): 694.3[M+1] ⁺

Step 10

(2R,4aR)-11-chloro-10-(2-fluoro-6-methoxyphenyl)-8-(2-isopropyl-4-methylpyridin-3-yl)-2-methyl Base-6-(methyl-d3)-5,7-dione-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 5j (1.33g, 1.92mmol) was dissolved in dichloromethane (15mL) and cooled to 0°C , boron tribromide (9.60 g, 38.32 mmol, 3.69 mL) was added dropwise, and the reaction was carried out at room temperature overnight. The reaction solution was lowered to 0°C, saturated sodium bicarbonate solution was added dropwise to quench the reaction, saturated sodium bicarbonate solution was added to adjust the pH to 7-8, extracted with ethyl acetate (50 mL×3), the organic phases were combined, and saturated brine was added. Washed, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and dried to give (2R,4aR)-11-chloro-10-(2-fluoro-6-hydroxyphenyl)-8-(2-isopropyl) -4-Methylpyridin-3-yl)-2-methyl-6-(methyl-d3)-2,3,4,4a,6,8-hexahydro-1H-pyrazino[1', 2':4,5]pyrazino[2,3-c][1,8]naphthyridine-5,7-dione 5 (1.11 g, 1.91 mmol).

MS m/z(ESI): 580.3[M+1] ⁺

Example 6

(2R,4aR)-11-chloro-10-(7,8-difluoro-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 (2R ,4aR)-11-chloro-10-(7,8-difluoro-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

The reaction conditions of the first step and the second step of Example 3 were repeated, except that in the first step, (2R,4aR)-10,11-dichloro-8-(2-isopropyl-4-methyl) was used. 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 6a and (7,8-difluoro-3-(methoxymethyl) Oxy)naphthalene-1-yl)boronic acid 6b was used as the starting material to obtain (2R,4aR)-11-chloro-10-(7,8-difluoro-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 6.

Example 7

(2R,4aR)-11-chloro-10-(8-ethynyl-7-fluoro-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

(2R,4aR)-11-Chloro-10-(8-ethynyl-7-fluoro-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

The reaction conditions of the first step and the second step of Example 3 were repeated, except that in the first step, (2R,4aR)-10,11-dichloro-8-(2-isopropyl-4-methyl) was used. 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 6a and (8-ethynyl-7-fluoro-3-(methoxy) (2R,4aR)-11-chloro-10-(8-ethynyl-7-fluoro-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 7.

Biological evaluation

Test Example 1. Determination of the Inhibitory Activity of the Compounds of the Invention on p-ERK1/2 in AGS Cells

The following method was used to determine the inhibitory activity of the compounds of the present invention on p-ERK1/2 in AGS cells. This method uses the Advanced phospho-ERK1/2 (Thr202/tyr204) kit (Cat. No. 64AERPEH) from Cisbio. For detailed experimental operations, please refer to the kit instructions. AGS cells (containing KRAS G12D mutation) were purchased from the Cell Resource Center, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.

The experimental procedure is briefly described as follows: AGS cells were cultured in F12K complete medium containing 10% fetal bovine serum, 100 U penicillin and 100 μg/mL streptomycin. AGS cells were plated in 96-well plates at 40,000 cells per well, and the medium was complete medium, and cultured overnight in a 37° C., 5% CO2 incubator. The test compound was dissolved in DMSO to prepare a 10mM stock solution, then diluted with F12K complete medium, and 100uL of F12K complete medium containing the corresponding concentration of the test compound was added to each well. The final concentration range of the test compound in the reaction system After culturing in a cell incubator for 3 hours, discard the cell supernatant, wash the cells with ice-bathed PBS, and then add 50 μl of 1×cell phospho/total protein lysis buffer (Advanced phospho-ERK1 /2 kit components) for lysis, the 96-well plate was placed on ice for half an hour, and then the lysate was detected according to the instructions of the Advanced phospho-ERK1/2 (Thr202/tyr204) kit. Finally, under the excitation wavelength of 304nM, the fluorescence intensity of each well at 620nM and 665nM was measured on a microplate reader in TF-FRET mode, and the fluorescence intensity ratio of 665/620 in each well was calculated. By comparing the fluorescence intensity ratio with the control group (0.1% DMSO), the percentage inhibition rate of the test compound at each concentration was calculated, and the numerical-inhibition rate was subjected to nonlinear regression analysis with the test compound concentration by GraphPad Prism 5 software. , to obtain the _IC50 value of the compound.

The preferred compound of the present invention has obvious inhibitory effect on the activity of p-ERK1/2 in AGS cells, the preferred compound has an IC ₅₀ <500nM, and the more preferred compound has an IC ₅₀ <200nM.

Test Example 2. The compound of the present invention inhibits the proliferation of AsPC-1 cells

The following method was used to determine the effect of the compounds of the present invention on the proliferation of AsPC-1 cells. AsPC-1 cells (containing KRAS G12D mutation) were purchased from the Cell Resource Center, Shanghai Institutes for Biological 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 middle. cell viability through

Assays were performed with the Luminescent Cell Viability Assay Kit (Promega, Cat. No. G7573).

The experimental method was operated in accordance with the steps in the kit instructions, which are briefly described as follows: the test compound was first dissolved in DMSO to prepare a 10 mM stock solution, and then diluted with culture medium to prepare a test sample. The final concentration of the compound was in the range of 1000nM-0.015nM . Cells in logarithmic growth phase were seeded into 96-well cell culture plates at a density of 800 cells per well, cultured overnight in a 37°C, 5% _CO2 incubator, followed by the addition of test compounds for 120 hours. After the incubation, 50uL of CellTiter-Glo detection solution was added to each well, shaken for 5 minutes, and then allowed to stand for 10 minutes. Then, the luminescence value of each well of the sample was read on a microplate reader using Luminescence mode. By comparing with the value of the control group (0.3% DMSO), the percentage inhibition rate of the compound at each concentration point was calculated, and then a nonlinear regression analysis was performed in the GraphPad Prism 5 software with the compound concentration logarithm-inhibition rate to obtain the compound inhibiting cell proliferation. the _IC50 value.

The preferred compound of the present invention has obvious inhibitory effect on the proliferation of AsPC-1 cells, the preferred compound has an IC ₅₀ <500nM, and the more preferred compound has an IC ₅₀ <200nM.

Claims

A compound represented by the general formula (I) or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof:

in:

Ring A is selected from aryl, heteroaryl or fused rings; preferably phenyl, naphthyl, pyridyl, benzothiazolyl or benzopyrazolyl;

X and Y are each independently selected from N or CR a ;

R a is selected from hydrogen atom, halogen, alkyl or alkoxy; wherein said alkyl or alkoxy is optionally further selected by one or more groups selected from halogen, hydroxyl, cyano, alkyl or alkoxy Substituents are substituted; R a is preferably halogen, more preferably fluorine or chlorine;

R 1 are the same or different, each independently selected from hydrogen atom, halogen, hydroxyl, alkyl or alkoxy, preferably alkyl;

Alternatively, the two R1 together with the atoms to which they are attached form a cycloalkyl or heterocyclyl;

R 2 is selected from a hydrogen atom, an alkyl group or a deuterated alkyl group, preferably an alkyl group or a deuterated alkyl group, more preferably a methyl group or a deuterated methyl group;

R 3 is selected from aryl or heteroaryl; wherein said aryl or heteroaryl is optionally further substituted by one or more RA ; R 3 is preferably heteroaryl;

R A is selected from alkyl, halogen, nitro, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, =O, -OR5 , -C(O) R5 , -C(O) OR 5 , -NHC(O)R 5 , -NHC(O)OR 5 , -NR 6 R 7 , -C(O)NR 6 R 7 , -CH 2 NHC(O)OR 5 , -CH 2 NR 6 R 7 or -S(O) r R 5 is substituted by the substituents; wherein the alkyl, cycloalkyl, heterocyclyl, aryl or heteroaryl is optionally further selected from one or more alkyl groups , halogen, nitro, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, =O, -OR 5 , -C(O)R 5 , -C(O)OR 5 , -NHC ( O)R 5 , -NHC(O)OR 5 , -NR 6 R 7 , -C(O)NR 6 R 7 , -CH 2 NHC(O)OR 5 , -CH 2 NR 6 R 6 or -S( O) the substituent of r R 5 is substituted;

R 4 is the same or different, each independently selected from hydrogen atom, alkyl group, alkynyl group, halogen, nitro group, cyano group, cycloalkyl group, heterocyclic group, aryl group, heteroaryl group, =O, -OR 5 , -C(O)R 5 , -C(O)OR 5 , -NHC(O)R 5 , -NHC(O)OR 5 , -NR 6 R 7 , -C(O)NR 6 R 7 , -CH 2 NHC(O)OR 5 , -CH 2 NR 6 R 7 or -S(O) r R 5 substituents; wherein said alkyl, cycloalkyl, heterocyclic, aryl or heteroaryl group is optionally further selected from one or more groups selected from alkyl, halogen, nitro, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, =O, -OR5 , -C(O)R 5 , -C(O)OR 5 , -NHC(O)R 5 , -NHC(O)OR 5 , -NR 6 R 7 , -C(O)NR 6 R 7 , -CH 2 NHC(O)OR 5. Substituents of -CH 2 NR 6 R 7 or -S(O) r R 5 are substituted;

R 5 is selected from a hydrogen atom, an alkyl group, a cycloalkyl group, a heterocyclyl group, an aryl group or a heteroaryl group, wherein said alkyl group, cycloalkyl group, heterocyclyl group, aryl group or heteroaryl group is optionally further One or more selected from hydroxy, halogen, nitro, cyano, alkyl, alkoxy, haloalkyl, haloalkoxy, cycloalkyl, heterocyclyl, aryl, heteroaryl, =O, -C (O)R 8 , -C(O)OR 8 , -OC(O)R 8 , -NR 9 R 10 , -C(O)NR 9 R 10 , -SO 2 NR 9 R 10 or -NR 9 C (O) Substituents of R 10 are substituted;

R 6 and R 7 are each independently selected from a hydrogen atom, a hydroxyl group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein the alkyl group, alkoxy group, A cycloalkyl, heterocyclyl, aryl or heteroaryl group is optionally further selected from one or more groups selected from hydroxy, halogen, nitro, cyano, alkyl, alkoxy, cycloalkyl, heterocyclyl, aryl radical, heteroaryl, =O, -C(O)R 8 , -C(O)OR 8 , -OC(O)R 8 , -NR 9 R 10 , -C(O)NR 9 R 10 , - SO 2 NR 9 R 10 or the substituent of -NR 9 C(O)R 10 ;

Alternatively, R 6 and R 7 together with the atoms to which they are attached form a 4- to 8-membered heterocyclic group, wherein the 4- to 8-membered heterocyclic group contains one or more N, O or S(O) r , and the The 4-8 membered heterocyclic group is optionally further selected from one or more groups selected from hydroxyl, halogen, nitro, cyano, alkyl, alkoxy, cycloalkyl, heterocyclic, aryl, heteroaryl , =O, -C(O)R 8 , -C(O)OR 8 , -OC(O)R 8 , -NR 9 R 10 , -C(O)NR 9 R 10 , -SO 2 NR 9 R 10 or -NR 9 C(O)R 10 is substituted by the substituent;

R 8 , R 9 and R 10 are each independently selected from a hydrogen atom, an alkyl group, an amino group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein said alkyl group, cycloalkyl group, heterocyclic group , Aryl or Heteroaryl optionally further selected by one or more groups selected from hydroxy, halogen, nitro, amino, cyano, alkyl, alkoxy, cycloalkyl, heterocyclyl, aryl, heteroaryl , substituted by a substituent of a carboxyl group or a carboxylate group;

m is selected from 0, 1, 2, 3 or 4;

n is selected from 0, 1, 2, 3 or 4;

r is selected from 0, 1 or 2.

Provided that, excluding compounds:
The compound according to claim 1 or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, which is a compound represented by general formula (II) and/or (III) or a stereoisomer thereof isomers, tautomers or their pharmaceutically acceptable salts:

Wherein: the definitions of ring A, R 1 to R 4 , X, Y, m and n are as described in claim 1 .
The compound according to claim 1 or 2 or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein:

X is selected from N;

Y is selected from CR a ;

R a is selected from hydrogen atom, halogen, alkyl or alkoxy; wherein said alkyl or alkoxy is optionally further selected by one or more groups selected from halogen, hydroxyl, cyano, alkyl or alkoxy Substituents are substituted; R a is preferably halogen, more preferably fluorine or chlorine;
The compound according to claim 1 or 2 or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein:

X is selected from CR a ;

Y is selected from N;

R a is selected from hydrogen atom, halogen, alkyl or alkoxy; wherein said alkyl or alkoxy is optionally further selected by one or more groups selected from halogen, hydroxyl, cyano, alkyl or alkoxy Substituents are substituted; R a is preferably halogen, more preferably fluorine or chlorine.
The compound according to any one of claims 1 to 4 or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein R 2 is selected from alkyl or deuterated alkyl, preferably methyl or deuterated methyl.
The compound according to claim 1 or 2, or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein R is selected from:

in:

R j is selected from hydrogen atom, halogen, nitro, cyano, hydroxyl, amino, alkyl, alkoxy, haloalkyl or haloalkoxy, preferably alkyl, more preferably methyl, ethyl or isopropyl ;

k is selected from 0, 1, 2 or 3.
The compound of claim 6 or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein:

R 3 is selected from
The compound according to any one of claims 1 to 4 or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, wherein:

R4 is selected from hydrogen atom, halogen, hydroxyl, amino, alkyl, alkoxy, alkynyl or cycloalkyl, wherein said alkyl, alkoxy, alkynyl or cycloalkyl is optionally further substituted by one or Substituted with a plurality of substituents selected from halogen, hydroxy, amino, alkyl or alkoxy.
The compound according to claim 8 or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein R is selected from fluoro, chloro, bromo, hydroxy, amino, methyl, ethyl, tris Fluoromethyl, ethynyl or cyclopropyl, preferably hydroxy, fluorine or ethynyl.
The compound according to any one of claims 1 to 4 or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, wherein
Selected from:
The compound according to claim 1 or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein said compound is:
A method for preparing the compound of general formula (I) according to claim 1 or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, the method comprising:

The compound of general formula (IA) and the compound of general formula (IB) are subjected to a coupling reaction under the action of a palladium catalyst, and the protecting group is further removed to obtain the compound of general formula (I);

in:

X 1 is selected from halogen or -Sn(R 14 ) 3 ; wherein the halogen is preferably chlorine;

R 14 is selected from alkyl, preferably methyl;

M is selected from -B(OH) 2 , -BF 3 K,
or halogen;

PG is a protecting group, preferably tert-butoxycarbonyl;

Ring A, R 1 to R 4 , X, Y, m and n are as defined in claim 1 .
A method for preparing the compound of general formula (I) according to claim 1 or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, the method comprising:

The compound of general formula (IC) is reacted with the compound of general formula (ID), and the protecting group is further removed to obtain the compound of general formula (I);

in:

X 2 is halogen, preferably iodine;

R 2 is alkyl or deuterated alkyl, preferably methyl or deuterated methyl;

PG is a protecting group, preferably tert-butoxycarbonyl;

Rings A, R 1 , R 3 , R 4 , X, Y, m and n are as defined in claim 1 .
A compound of general formula (IA) or a stereoisomer, a tautomer and a pharmaceutically acceptable salt thereof,

in:

X 1 is selected from halogen or -Sn(R 14 ) 3 ; wherein the halogen is preferably chlorine;

R 14 is selected from alkyl, preferably methyl;

PG is selected from protecting groups, preferably tert-butoxycarbonyl;

The definitions of R 1 to R 3 , X, Y and n are as described in claim 1 .
The compound of claim 14 or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein the compound is:
A compound of the general formula (IC) or a stereoisomer, a tautomer and a pharmaceutically acceptable salt thereof,

in:

PG is selected from protecting groups, preferably tert-butoxycarbonyl;

Rings A, R 1 , R 3 , R 4 , X, Y, m and n are as defined in claim 1 .
The compound of claim 16 or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein the compound is:
A pharmaceutical composition comprising an effective dose of the compound according to any one of claims 1 to 11 or a stereoisomer, tautomer or a pharmaceutically acceptable salt thereof, and pharmaceutically acceptable carriers, excipients or their combinations.
The compound according to any one of claims 1 to 11 or a stereoisomer, a tautomer or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 18 is used in the preparation of KRas G12D inhibition use in medicaments.
The compound according to any one of claims 1 to 11 or a stereoisomer, a tautomer or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 18 is prepared for treatment by KRas Use in medicine for diseases mediated by G12D mutation, wherein said disease mediated by KRas G12D mutation is selected from cancer, wherein said cancer is selected from cardiac myxoma, lung cancer, gastric cancer, colorectal tumor, rectal cancer, pancreas Cancer, prostate cancer, bladder cancer, hepatocellular carcinoma, cholangiocarcinoma, cholangiocarcinoma, chondrosarcoma, multiple myeloma, uterine cancer, cervical cancer, seminoma, malignant melanoma, cutaneous squamous cell carcinoma, adrenal neurogenesis cell tumor, myeloid leukemia, acute lymphoblastic leukemia or glioblastoma, preferably pancreatic cancer, colorectal tumor, rectal cancer and lung cancer.
The compound according to any one of claims 1 to 11 or a stereoisomer, a tautomer or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 18 is used in the preparation of the treatment of cancer. Use in medicine, wherein the cancer is selected from cardiac myxoma, lung cancer, gastric cancer, colorectal tumor, rectal cancer, pancreatic cancer, prostate cancer, bladder cancer, hepatocellular carcinoma, cholangiocarcinoma, cholangiocarcinoma, chondrosarcoma, multiple Myeloma, uterine cancer, cervical cancer, seminoma, malignant melanoma, cutaneous squamous cell carcinoma, adrenal neuroblastoma, myeloid leukemia, acute lymphoblastic leukemia or glioblastoma, preferably pancreatic cancer, Colorectal tumors, rectal cancer and lung cancer.
The use according to claim 20 or 21, wherein the lung cancer is selected from non-small cell lung cancer or small cell lung cancer.