Heterocyclic derivatives and their preparation methods and uses

Abstract

The present invention relates to heterocyclic derivatives, a method for preparing the same, and their use in medicine. Specifically, the present invention relates to a heterocyclic derivative represented by the general formula (I), a pharmaceutically acceptable salt thereof, and their use as therapeutic agents, particularly as KRas G12D inhibitors, wherein the definitions of the substituents in the general formula (I) are the same as those defined in the specification.

Description

Technical Field

[0001] The present invention relates to a heterocyclic derivative, a method for preparing the same, a pharmaceutical composition containing the derivative, and the use thereof as a therapeutic agent, particularly as a K-Ras GTPase inhibitor. Background Art

[0002] RAS represents a group of closely related monomeric globular proteins (21 kDa molecular weight), which 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 by receiving growth factors and various other extracellular signals and is responsible for regulating functions such as cell growth, survival, migration, and differentiation. RAS acts as a molecular switch, and the on / off state of the RAS protein is determined by nucleotide binding. The active signal transduction conformation binds GTP, and the inactive conformation binds GDP. When RAS contains bound GDP, it is in a dormant or quiescent or off state and is "inactivated". When a cell is exposed to certain growth-promoting stimuli in response, RAS is induced to convert the bound GDP to GTP. As GTP is bound, RAS is "on" and is able to interact with other proteins and activate other proteins (its "downstream targets"). The RAS protein itself has a very low inherent ability to hydrolyze GTP back to GDP and thereby turn itself off. Converting RAS to the off state requires an exogenous protein called a GTPase-activating protein (GAPs), which interacts with RAS and can 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 the generation of a prolonged signal to the cell, which tells it to continue growing and dividing. Therefore, these signals cause the cell to grow and divide, and overactivated RAS signal transduction may ultimately lead to cancer.

[0003] Structurally, the RAS protein contains a G domain responsible for the enzymatic activity of RAS - guanine nucleotide binding and hydrolysis (the GTPase reaction). It also includes a C - terminal extension region containing a so - called CAAX box, which can be post - translationally modified and targets 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 nucleotide - binding pocket in the protein and is the rigid part of the domain with conserved amino acid residues (glycine 12, threonine 26, and lysine 16) that are essential for nucleotide binding and hydrolysis. 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 dynamic parts of the protein and are often represented as a "spring - loaded" mechanism due to their ability to switch between a resting and a loaded state. The main interaction is the hydrogen bond formed by threonine - 35 and glycine - 60 with the γ - phosphate of GTP, which keeps the switch I region and switch II region in their active conformations respectively. After hydrolyzing GTP and releasing phosphate, both relax into an inactive GDP conformation.

[0004] Among RAS family members, oncogenic mutations are most common in KRAS (85%), while they are less common in NRAS (12%) and HRAS (3%). KRAS mutations are prevalent in three major fatal cancer types in the United States: pancreatic cancer (95%), colorectal cancer (45%), and lung cancer (25%). KRAS mutations have also been found in other cancer types including multiple myeloma, uterine cancer, cholangiocarcinoma, gastric cancer, bladder cancer, diffuse large B - cell lymphoma, rhabdomyosarcoma, cutaneous squamous cell carcinoma, cervical cancer, testicular germ cell carcinoma, etc., while they are rarely found in breast cancer, ovarian cancer, and brain cancer (<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 NSCLC, the increase in the frequency of specific allelic mutations mostly comes from classical smoking - induced canonical mutations (G:C to T:A substitutions), resulting in KRAS G12C (GGT to TGT) and G12V (GGT to GTT) mutations.

[0005] Large - scale genomics 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. At the same time, KRAS mutations often occur concomitantly with certain co - mutations, such as STK11, KEAP1, and TP53, which cooperate with mutant RAS to transform cells into highly malignant and invasive tumor cells.

[0006] 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 effective anti-RAS therapy clinically, and targeting this gene with small molecules is challenging. Therefore, there is an urgent need in the art for small molecules that target RAS (e.g., K-RAS, H-RAS, and / or N-RAS) and utilize them to treat various diseases, such as cancer.

[0007] Currently, the clinical development of KRas G12D inhibitors is highly competitive both at home and abroad. Among them, the KRas G12D inhibitor MRTX-1133 developed by Mirati Therapeutics Inc has entered the preclinical stage for the treatment of diseases such as colorectal tumors, non-small cell lung cancer, and pancreatic cancer. Currently, there are few publicly disclosed patent applications for KRas G12D inhibitors, including WO2021041671 of Mirati Therapeutics Inc. Although certain progress has been made in the research and application of KRas G12D inhibitors, there is still a huge room for improvement, and it is still necessary to continue researching and developing new KRas G12D inhibitors. Summary of the Invention

[0008] The object of the present invention is to provide a heterocyclic derivative represented by the general formula (I), or its stereoisomer, tautomer, or pharmaceutically acceptable salt:

[0009]

[0010] Wherein:

[0011] Ring B is selected from aryl, heteroaryl, or fused ring;

[0012] Q is selected from N or CR a ;

[0013] Y is selected from a bond, -O-, or -NR b ;

[0014] R a is selected from a hydrogen atom, halogen, alkyl, alkoxy, or cyano; wherein the alkyl or alkoxy is optionally further substituted by one or more substituents selected from halogen, hydroxy, cyano, alkyl, or alkoxy;

[0015] R b is selected from a hydrogen atom or alkyl;

[0016] R c is selected from a hydrogen atom, halogen, cyano, alkyl, or alkoxy; wherein the alkyl or alkoxy is optionally further substituted by one or more substituents selected from halogen, hydroxy, cyano, alkyl, or alkoxy; R cPreferably a halogen, more preferably fluorine or chlorine;

[0017] R 1 Selected from -L-cycloalkyl, -L-(6- to 9-membered) monocyclic heterocyclic group, -L-bicyclic heterocyclic group, -L-tricyclic heterocyclic group, -L-aryl, -L-heteroaryl or -L-fused ring; wherein the cycloalkyl, (6- to 9-membered) monocyclic heterocyclic group, bicyclic heterocyclic group, -tricyclic heterocyclic group, aryl, heteroaryl or fused ring is optionally further substituted by one or more substituents selected from alkyl, halogen, haloalkyl, hydroxyalkyl, benzyl, cyano, cycloalkyl, heterocyclic group, 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 , -CH2NHC(O)OR 5 , -CH2NR 6 R 7 or -S(O) r R 5 ;

[0018] L is independently selected from a bond or -C1-C6 alkylene, wherein the alkylene is optionally further substituted by one or more R D ;

[0019] R D are independently selected from a hydrogen atom, halogen, hydroxy or hydroxymethyl;

[0020] Alternatively, two R D connected to the same carbon atom, together with the carbon atom to which they are connected, form a cycloalkyl; preferably cyclopropyl;

[0021] R 2 are the same or different and are independently selected from a hydrogen atom, halogen, hydroxy, alkyl or alkoxy, preferably a hydrogen atom or alkyl;

[0022] Two R 3 together with the atom to which they are connected form a cycloalkyl or heterocyclic group;

[0023] Provided that when Q is selected from N, is not selected from

[0024] R 4are the same or different and are each independently selected from hydrogen, alkyl, halogen, nitro, cyano, alkenyl, alkynyl, cycloalkyl, heterocyclic, 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 、-CH2NHC(O)OR 5 、-CH2NR 6 R 7 or -S(O) r R 5 wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclic, aryl or heteroaryl is optionally further substituted by one or more selected from alkyl, halogen, nitro, cyano, cycloalkyl, heterocyclic, 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 、-CH2NHC(O)OR 5 、-CH2NR 6 R 7 or -S(O) r R 5 substituted by a substituent;

[0025] R 5 Each is independently selected from a hydrogen atom, an alkyl group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein the alkyl group, the cycloalkyl group, the heterocyclic group, the aryl group or the heteroaryl group is optionally further substituted by one or more selected from hydroxyl, halogen, nitro, cyano, alkyl, alkoxy, haloalkyl, haloalkoxy, cycloalkyl, heterocyclic group, 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 、-SO2NR 9 R 10 or -NR 9 C(O)R 10substituted by substituents;

[0026] 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, cycloalkyl group, heterocyclic group, aryl group or heteroaryl group is optionally further substituted by one or more substituents selected from a hydroxyl group, a halogen, a nitro group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, =O, -C(O)R 8 、-C(O)OR 8 、-OC(O)R 8 、-NR 9 R 10 、-C(O)NR 9 R 10 、-SO2NR 9 R 10 or -NR 9 C(O)R 10 substituted by substituents;

[0027] 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 4- to 8-membered heterocyclic group is optionally further substituted by one or more substituents selected from a hydroxyl group, a halogen, a nitro group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, =O, -C(O)R 8 、-C(O)OR 8 、-OC(O)R 8 、-NR 9 R 10 、-C(O)NR 9 R 10 、-SO2NR 9 R 10 or -NR 9 C(O)R 10 substituted by substituents;

[0028] 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 the alkyl group, cycloalkyl group, heterocyclic group, aryl group or heteroaryl group is optionally further substituted by one or more substituents selected from a hydroxyl group, a halogen, a nitro group, an amino group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, a carboxyl group or a carboxylic acid ester group;

[0029] m is selected from 0, 1, 2, 3 or 4;

[0030] n is selected from 0, 1, 2 or 3;

[0031] r is 0, 1 or 2.

[0032] In a preferred embodiment of the present invention, the heterocyclic derivative represented by the general formula (I), or its stereoisomer, tautomer or pharmaceutically acceptable salt, is a compound represented by the general formula (II) or its stereoisomer, tautomer or pharmaceutically acceptable salt:

[0033]

[0034] Wherein: Ring B, R 1 , R 2 , R 4 , R c , Y, m and n are defined as in the general formula (I).

[0035] In a preferred embodiment of the present invention, the heterocyclic derivative represented by the general formula (I) or (II), or its stereoisomer, tautomer or pharmaceutically acceptable salt, wherein:

[0036] R 1 is -L-bicyclic heterocyclic group; wherein the bicyclic heterocyclic group is optionally further substituted by one or more substituents selected from alkyl, halogen, alkoxy or =O; wherein the halogen is preferably fluorine;

[0037] L is selected from a bond or -C1-C3 alkylene, wherein the alkylene is optionally further substituted by one or more R D ;

[0038] R D are each independently selected from a hydrogen atom, halogen, hydroxyl or hydroxymethyl;

[0039] Alternatively, two R D connected to the same carbon atom, together with the carbon atom to which they are connected, form a cycloalkyl group; preferably cyclopropyl.

[0040] In a preferred embodiment of the present invention, the heterocyclic derivative represented by the general formula (I) or (II), or its stereoisomer, tautomer or pharmaceutically acceptable salt, wherein:

[0041] L is selected from a bond, -CH2-, -CH2CH2- or

[0042] In a preferred embodiment of the present invention, the heterocyclic derivative represented by the general formula (I) or (II), or its stereoisomer, tautomer or pharmaceutically acceptable salt, wherein R 1 is selected from:

[0043]

[0044]

[0045] In a preferred embodiment of the present invention, the heterocyclic derivative represented by the general formula (I), or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein two Rs 3 together with the atom to which it is attached form a 4- to 8-membered cycloalkyl or 4- to 8-membered heterocyclic group.

[0046] In a preferred embodiment of the present invention, the heterocyclic derivative represented by the general formula (I) or (II), or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein:

[0047] R 4 are the same or different and each independently selected from a hydrogen atom, an alkyl group, a halogen, an alkoxy group, an alkynyl group, a hydroxyl group, an amino group, a hydroxyalkyl group, a haloalkyl group or a haloalkoxy group;

[0048] preferably a hydrogen atom, a methyl group, fluorine, chlorine, a hydroxyl group, an amino group, a hydroxymethyl group or an ethynyl group.

[0049] In a preferred embodiment of the present invention, the heterocyclic derivative represented by the general formula (I) or (II), or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein:

[0050] ring B is selected from phenyl, naphthyl, pyridyl, quinolinyl, isoquinolinyl, indolyl, indazolyl, benzothiazolyl, tetrahydronaphthyl,

[0051] preferably naphthyl and benzothiazolyl.

[0052] In a preferred embodiment of the present invention, the heterocyclic derivative represented by the general formula (I) or (II), or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein is selected from the following groups:

[0053]

[0054] In a preferred embodiment of the present invention, the heterocyclic derivative represented by the general formula (I), its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein is selected from the following groups:

[0055]

[0056] In a preferred embodiment of the present invention, the heterocyclic derivative represented by the general formula (I) or (II), or its stereoisomer, tautomer or pharmaceutically acceptable salt thereof, wherein R c is selected from halogens, preferably fluorine.

[0057] Exemplary compounds of the present invention include, but are not limited to:

[0058]

[0059] or their stereoisomers, tautomers or pharmaceutically acceptable salts thereof.

[0060] Note: If there is a difference between the drawn structure and the name given for that structure, the drawn structure will be given greater weight. On the other hand, the present invention provides a pharmaceutical composition comprising an effective dose of a compound of general formula (I) or (II) or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof, and a pharmaceutically acceptable carrier, excipient or a combination thereof.

[0061] On the other hand, the present invention provides a method for inhibiting the KRas G12D enzyme, which method comprises administering to a patient a pharmaceutical composition comprising an effective dose of a compound of general formula (I) or (II) or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof, and a pharmaceutically acceptable carrier, excipient or a combination thereof.

[0062] The present invention also provides the use of a compound of general formula (I) or (II) or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof, or a pharmaceutical composition thereof in the preparation of a medicament for the treatment of a disease mediated by KRas G12D mutation, wherein the disease mediated by KRas G12D mutation is selected from cancers, wherein the cancers are selected from cardiac myxoma, lung cancer, gastric cancer, colorectal cancer, rectal cancer, pancreatic cancer, prostate cancer, bladder cancer, hepatocellular carcinoma, cholangiocarcinoma, chondrosarcoma, multiple myeloma, uterine cancer, cervical cancer, seminoma, malignant melanoma, cutaneous squamous cell carcinoma, adrenocortical neuroblastoma, myeloid leukemia, acute lymphoblastic leukemia or glioblastoma, preferably pancreatic cancer, colorectal cancer, rectal cancer and lung cancer; wherein the lung cancer is selected from non-small cell lung cancer or small cell lung cancer.

[0063] On the other hand, the present invention provides the use of a compound of general formula (I) or (II) or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof, or a pharmaceutical composition thereof in the preparation of a KRas G12D inhibitor.

[0064] Another aspect of the present invention relates to a method for preventing and / or treating a disease mediated by KRas G12D mutation, which comprises administering to a patient a therapeutically effective dose of a compound of general formula (I) or (II) or its tautomer, meso form, racemate, enantiomer, diastereomer or a mixture thereof or a pharmaceutically acceptable salt thereof or a pharmaceutical composition comprising the same.

[0065] The present invention also provides the use of a compound of formula (I) or (II), or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof in the preparation of a medicament for treating 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, chondrosarcoma, multiple myeloma, uterine cancer, cervical cancer, seminoma, malignant melanoma, cutaneous squamous cell carcinoma, adrenocortical neuroblastoma, myeloid leukemia, acute lymphoblastic leukemia or glioblastoma, preferably pancreatic cancer, colorectal cancer, rectal cancer and lung cancer; wherein the lung cancer is preferably non-small cell lung cancer.

[0066] The pharmaceutical preparation of the present invention can be administered locally, orally, transdermally, rectally, vaginally, parenterally, intranasally, intrapulmonary, intraocularly, intravenously, intramuscularly, intraarterially, intrathecally, intracapsularly, intradermally, intraperitoneally, subcutaneously, subcuticularly or by inhalation. The pharmaceutical composition containing the active ingredient can be in a form suitable for oral administration, such as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft gelatin capsules, or syrups or elixirs. Tablets contain the active ingredient and non-toxic pharmaceutically acceptable excipients suitable for mixing for the preparation of tablets.

[0067] The preparation of the present invention is suitable to exist in the form of unit dosage, and the preparation can be prepared by any method well known in the pharmaceutical art. The amount of the active ingredient capable of producing a single dosage form by combination with a carrier substance can vary depending on the host to be treated and the specific mode of administration. The amount of the active ingredient capable of producing a single dosage form by combination with a carrier substance generally refers to the amount of the compound capable of producing a therapeutic effect.

[0068] The dosage forms for local or transdermal administration of the compounds of the present invention may include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches and inhalants. The active compound can be mixed with a pharmaceutically acceptable carrier under sterile conditions, and it can be mixed with any preservatives, buffers or propellants that may be required.

[0069] When the compounds of the present invention are administered to humans and animals in the form of a medicament, the compounds can be provided alone or in the form of a pharmaceutical composition containing the active ingredient in combination with a pharmaceutically acceptable carrier, such as 0.1% to 99.5% (more preferably, 0.5% to 90%) of the active ingredient.

[0070] 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 its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose, and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients, such as cocoa butter and suppository wax; (9) oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; (10) glycols, such as propylene glycol; (11) polyols, such as glycerin, sorbitol, mannitol, and polyethylene glycol; (12) esters, such as ethyl oleate and ethyl laurate; (13) agar; (14) buffering agents, such as magnesium hydroxide and aluminum hydroxide; (15) alginic acid; (16) pyrogen-free water; (17) isotonic saline; (18) Ringer's solution; (19) ethanol; (20) phosphate buffer solution; (21) cyclodextrins, such as targeting ligands attached to nanoparticles, such as AccurinsTM; and (22) other non-toxic compatible substances for pharmaceutical formulations, such as polymer-based compositions.

[0071] Examples of pharmaceutically acceptable antioxidants include, but are not limited to: (1) water-soluble antioxidants such as ascorbic acid, cysteine hydrochloride, sodium bisulfate, sodium metabisulfite, sodium sulfite, and the like; (2) oil-soluble antioxidants such as ascorbyl palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, α-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 (such as capsules, tablets, pills, dragees, powders, granules, and the like) may include one or more pharmaceutically acceptable carriers such as sodium citrate or calcium phosphate, and / or any one of the following: (1) fillers or extenders 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) solution retarders such as paraffin; (6) absorption accelerators such as quaternary ammonium compounds; (7) wetting agents such as cetyl alcohol and glycerol monostearate; (8) absorbents such as kaolin and bentonite; (9) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, and mixtures thereof; and (10) coloring agents. Liquid dosage forms may 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; solubilizing agents and emulsifying agents such as ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butanediol, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil, and sesame oil), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols, and fatty acid esters of sorbitan, and mixtures thereof.

[0072] In addition to the active compound, the suspension may also contain suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol, and sorbitan esters, microcrystalline cellulose, aluminum hydroxide oxide, bentonite, agar, and tragacanth, and mixtures thereof.

[0073] In addition to the active compound, ointments, pastes, creams, and gels may also contain excipients such as animal and vegetable fats, oils, waxes, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycols, polysiloxanes, bentonite, silicic acid, talc, and zinc oxide, or mixtures thereof.

[0074] In addition to the active compound, the powders and sprays may also contain excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicate and polyamide powder or mixtures of these substances. The sprays may contain other common propellants such as chlorofluorocarbons, as well as volatile unsubstituted hydrocarbons such as butane and propane.

[0075] Detailed Description of the Invention

[0076] Unless otherwise stated, some of the terms used in the specification and claims of the present invention are defined as follows:

[0077] "Bond" means that the indicated substituent is absent and the two end portions of the substituent are directly connected to form a bond.

[0078] "Alkyl", when regarded as a group or part of a group, refers to a straight-chain or branched-chain aliphatic hydrocarbon group. Preferably C1-C 20 alkyl, more preferably C1-C 10 alkyl, even more preferably C1-C6 alkyl. 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-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, etc. The alkyl can be substituted or unsubstituted.

[0079] "Alkenyl" refers to an alkyl 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, vinyl, 1-propenyl, 2-propenyl, 1-, 2- or 3-butenyl, etc. The alkenyl can be optionally substituted or unsubstituted.

[0080] "Alkynyl" refers to an aliphatic hydrocarbon group containing a carbon-carbon triple bond, which can be straight-chain or branched-chain. Preferred is C2-C 10 alkynyl, more preferably C2-C6 alkynyl, most preferably C2-C4 alkynyl. Examples of alkynyl groups include, but are not limited to, ethynyl, 1-propynyl, 2-propynyl, 1-, 2- or 3-butynyl, etc. The alkynyl can be substituted or unsubstituted.

[0081] "Cycloalkyl" refers to a saturated or partially saturated monocyclic, fused-ring, bridged-ring and spiro carbon ring. Preferably C 3- C 12 cycloalkyl, more preferably C3-C8 cycloalkyl, most preferably C 3-C6 cycloalkyl. Examples of monocyclic cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptatrienyl, cyclooctyl, etc., preferably cyclopropyl and cyclohexenyl. The cycloalkyl may be optionally substituted or unsubstituted.

[0082] "Spirocycloalkyl" refers to a polycyclic group having 5 to 18 members, two or more cyclic structures, and sharing a single carbon atom (referred to as a spiro atom) between the single rings. One or more double bonds are contained within the rings, but none of the rings has an aromatic system with completely conjugated π electrons. Preferably, it is 6 to 14 members, more preferably 7 to 10 members. Spirocycloalkyl is classified into monospiro, dispiro or polyspirocycloalkyl according to the number of spiro atoms shared between the rings, preferably monospiro and dispirocycloalkyl, preferably 4-membered / 5-membered, 4-membered / 6-membered, 5-membered / 5-membered or 5-membered / 6-membered. 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.

[0083] "Fused cycloalkyl" refers to a fully carbon polycyclic group having 5 to 18 members, containing two or more cyclic structures sharing a pair of carbon atoms. One or more of the rings may contain one or more double bonds, but none of the rings has an aromatic system with completely conjugated π electrons. Preferably, it is 6 to 12 members, more preferably 7 to 10 members. 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 bicyclic alkyl. 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, decahydronaphthyl or tetradecahydrophenanthryl.

[0084] "Bridged cycloalkyl" refers to a fully carbon polycyclic group having 5 to 18 members, containing two or more cyclic structures, and sharing two non-adjacent carbon atoms. One or more of the rings may contain one or more double bonds, but none of the rings has an aromatic system with completely conjugated π electrons. Preferably, it is 6 to 12 members, more preferably 7 to 10 members. Preferably, it is 6 to 14 members, more preferably 7 to 10 members. 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)-bicyclo[3.3.1]nonyl, bicyclo[2.2.2]octyl, (1r,5r)-bicyclo[3.3.2]decyl.

[0085] "Heterocyclic group", "heterocycle" or "heterocyclic" are used interchangeably in this application and all refer to non-aromatic heterocyclic groups in which one or more ring-forming atoms are heteroatoms such as oxygen, nitrogen, sulfur atoms, etc., including monocyclic, fused-ring, bridged-ring and spiro-ring. Preferably having a 5- to 7-membered monocyclic or 7- to 10-membered bicyclic or tricyclic ring, which may contain 1, 2 or 3 atoms selected from nitrogen, oxygen and / or sulfur. Examples of "heterocyclic group" 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-azabicyclo[3.2.1]octyl and piperazinyl. The heterocyclic group may be substituted or unsubstituted.

[0086] "Spiroheterocyclic group" refers to a polycyclic group having 5 to 18 members, two or more ring structures, and a single atom shared between monocyclic rings, with one or more double bonds in the ring but no ring having a completely conjugated π-electron aromatic system, and one or more ring atoms selected from nitrogen, oxygen or S(O) r (where r is selected from 0, 1 or 2) heteroatoms, and the remaining ring atoms are carbon. Preferably 6 to 14 members, more preferably 7 to 10 members. Spirolalkyl groups are classified into monospiroheterocyclic groups, bisspiroheterocyclic groups or multispiroheterocyclic groups according to the number of spiro atoms shared between rings, preferably monospiroheterocyclic groups and bisspiroheterocyclic groups. More preferably 4-membered / 4-membered, 4-membered / 5-membered, 4-membered / 6-membered, 5-membered / 5-membered or 5-membered / 6-membered monospiroheterocyclic groups. Non-limiting examples of "spiroheterocyclic group" include but are not limited to: 1,7-dioxaspiro[4.5]decyl, 2-oxa-7-azaspiro[4.4]nonyl, 7-oxaspiro[3.5]nonyl and 5-oxaspiro[2.4]heptyl.

[0087] "Fused heterocyclic group" refers to a fully carbon polycyclic group containing two or more ring structures sharing a pair of atoms with each other, and one or more rings may contain one or more double bonds, but no ring has a completely conjugated π-electron aromatic system, and one or more ring atoms are selected from nitrogen, oxygen or S(O) r (where r is selected from 0, 1 or 2) heteroatoms, and the remaining ring atoms are carbon. Preferably 6 to 14 members, more preferably 7 to 10 members. 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 heterocyclic group" include but are not limited to: octahydropyrrolo[3,4-c]pyrrolyl, octahydro-1H-indolizinyl, 3-azabicyclo[3.1.0]hexyl, octahydrobenzo[b][1,4]dioxine or

[0088] "Bridged heterocyclic group" refers to a polycyclic group having 5 to 14 members, 5 to 18 members, containing two or more cyclic structures, sharing two non-adjacent atoms with each other, where one or more rings may contain one or more double bonds, but none of the rings has a completely conjugated π-electron aromatic system, and one or more of the ring atoms are selected from nitrogen, oxygen or S(O) r (where r is selected from 0, 1 or 2) heteroatoms, and the remaining ring atoms are carbon. Preferably it is 6 to 14 members, more preferably 7 to 10 members. 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 heterocyclic group" include but are not limited to: 2-azabicyclo[2.2.1]heptyl, 2-azabicyclo[2.2.2]octyl and 2-azabicyclo[3.3.2]decyl.

[0089] "Aryl" refers to a carbocyclic aromatic system containing one or two rings, where the rings may be linked together in a fused manner. The term "aryl" includes monocyclic or bicyclic aryls, such as phenyl, naphthyl, aromatic groups of tetrahydronaphthyl. Preferably the aryl is C6-C 10 aryl, more preferably the aryl is phenyl and naphthyl, most preferably naphthyl. The aryl may be substituted or unsubstituted.

[0090] "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 it is a 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-thiadiazolyl, benzodioxolyl, benzothienyl, benzimidazolyl, indolyl, isoindolyl, 1,3-dioxo-isoindolyl, quinolinyl, indazolyl, benzisothiazolyl, benzoxazolyl, benzisoxazolyl, benzothiazolyl.

[0091]

[0092] The heteroaryl may be substituted or unsubstituted.

[0093] "Fused ring" refers to a polycyclic group having two or more cyclic structures sharing a pair of atoms with each other, where one or more rings may contain one or more double bonds, but at least one ring does not have a completely conjugated π-electron aromatic system, and the ring atoms are selected from 0, one or more selected from nitrogen, oxygen or S(O) ra heteroatom selected from 0, 1 or 2, and the remaining ring atoms are carbon. The fused rings preferably include bicyclic or tricyclic fused rings, and the bicyclic fused rings are preferably fused rings of an aryl or heteroaryl with a monocyclic heterocyclic group or a monocyclic cycloalkyl group. It is preferably 7 to 14 membered, more preferably 8 to 10 membered. Examples of "fused rings" include, but are not limited to:

[0094]

[0095] "Alkoxy" refers to a group of (alkyl-O-). Among them, the definition of alkyl is as described herein. C1-C6 alkoxy is preferred. Examples include, but are not limited to: methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, etc.

[0096] "Haloalkyl" refers to a group in which the alkyl is optionally further substituted by one or more halogens, and the definition of alkyl is as described herein.

[0097] "Hydroxyalkyl" refers to a group in which the alkyl is optionally further substituted by one or more hydroxyl groups, and the definition of alkyl is as described herein.

[0098] "Hydroxymethyl" refers to a group in which the methyl is optionally further substituted by one or more hydroxyl groups.

[0099] "Haloalkoxy" refers to a group in which the alkyl of (alkyl-O-) is optionally further substituted by one or more halogens, and the definition of alkoxy is as described herein.

[0100] "Hydroxyl" refers to the -OH group.

[0101] "Halogen" refers to fluorine, chlorine, bromine and iodine.

[0102] "Amino" refers to -NH2.

[0103] "Cyano" refers to -CN.

[0104] "Nitro" refers to -NO2.

[0105] "Benzyl" refers to -CH2-phenyl.

[0106] "Carboxyl" refers to -C(O)OH.

[0107] "Carboxylate group" refers to -C(O)O-alkyl or -C(O)O-cycloalkyl, where the definitions of alkyl and cycloalkyl are as described above.

[0108] "DMSO" refers to dimethyl sulfoxide.

[0109] "BOC" refers to tert-butoxycarbonyl.

[0110] "Ts" refers to p-toluenesulfonyl.

[0111] "T3P" refers to propylphosphonic anhydride.

[0112] "DPPA" refers to diphenylphosphoryl azide.

[0113] "DEA" refers to diethylamine.

[0114] "X-PHOS Pd G2" refers to chloro(2-dicyclohexylphosphino-2′,4′,6′-triisopropyl-1,1′-biphenyl)[2-(2′-amino-1,1′-biphenyl)]palladium(II).

[0115] "RuPhos Pd G3" refers to (2-dicyclohexylphosphino-2′,6′-diisopropoxy-1,1′-biphenyl)(2-amino-1,1′-biphenyl-2-yl)palladium(II) methanesulfonate.

[0116] "cataCXium A Pd-G3" refers to [n-butylbis(1-adamantyl)phosphine](2-amino-1,1′-biphenyl-2-yl)palladium methanesulfonate.

[0117] "Pd(dppf)Cl2" refers to dichloro[1,1′-bis(diphenylphosphino)ferrocene]palladium.

[0118] "Substituted" means that one or more, preferably up to 5, more preferably 1 to 3, hydrogen atoms in a group are independently replaced by the corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and those skilled in the art can determine (by experiment or theory) the possible or impossible substitutions without too much effort. For example, an amino or hydroxyl group with a free hydrogen may be unstable when combined with a carbon atom having an unsaturated (such as olefinic) bond.

[0119] As used in this specification, "substituted" or "substitution", unless otherwise specified, means that a group can be substituted by one or more groups selected from the following: alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxyl, nitro, cyano, cycloalkyl, heterocyclic group, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, amino, haloalkyl, hydroxyalkyl, carboxyl, carboxylic ester group, =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 、-CH2NHC(O)OR 5 、-CH2NR 6 R7 or -S(O) r R 5 is substituted by a substituent;

[0120] R 5 is selected from a hydrogen atom, an alkyl group, a cycloalkyl group, a heterocyclic group, an aryl group or a heteroaryl group, wherein the alkyl group, cycloalkyl group, heterocyclic group, aryl group or heteroaryl group is optionally further substituted by one or more substituents selected from a hydroxyl group, a halogen, a nitro group, a cyano group, an alkyl group, an alkoxy group, a haloalkyl group, a haloalkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, =O, -C(O)R 8 、-C(O)OR 8 、-OC(O)R 8 、-NR 9 R 10 、-C(O)NR 9 R 10 、-SO2NR 9 R 10 or -NR 9 C(O)R 10 is substituted by a substituent;

[0121] 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, cycloalkyl group, heterocyclic group, aryl group or heteroaryl group is optionally further substituted by one or more substituents selected from a hydroxyl group, a halogen, a nitro group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, =O, -C(O)R 8 、-C(O)OR 8 、-OC(O)R 8 、-NR 9 R 10 、-C(O)NR 9 R 10 、-SO2NR 9 R 10 or -NR 9 C(O)R 10 is substituted by a substituent;

[0122] 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 4- to 8-membered heterocyclic group is optionally further substituted by one or more substituents selected from a hydroxyl group, a halogen, a nitro group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, =O, -C(O)R 8 、-C(O)OR 8、 -OC(O)R 8 、 -NR 9 R 10 、 -C(O)NR 9 R 10 、 -SO2NR 9 R 10 or -NR 9 C(O)R 10 substituted by substituents of;

[0123] 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 the alkyl group, cycloalkyl group, heterocyclic group, aryl group or heteroaryl group is optionally further substituted by one or more substituents selected from a hydroxyl group, a halogen, a nitro group, an amino group, a cyano group, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, a carboxyl group or a carboxylic acid ester group;

[0124] r is 0, 1 or 2.

[0125] The compounds of the present invention may contain asymmetric centers or chiral centers and thus exist in different stereoisomeric forms. It is contemplated that all stereoisomeric forms of the compounds of the present invention, including but not limited to diastereoisomers, enantiomers and atropisomers and geometric (conformational) isomers and mixtures thereof, such as racemic mixtures, are within the scope of the present invention.

[0126] Unless otherwise indicated, the structures described in the present invention also include all isomers of this structure (e.g., diastereoisomers, enantiomers and atropisomers and geometric (conformational) isomer forms; for example, the R and S configurations of each asymmetric center, (Z) and (E) double bond isomers, and (Z) and (E) conformational isomers). Thus, individual stereoisomers of the compounds of the present invention as well as mixtures of enantiomers, mixtures of diastereoisomers and mixtures of geometric (conformational) isomers are within the scope of the present invention.

[0127] "Pharmaceutically acceptable salts" refer to certain salts of the above compounds that retain their original biological activity and are suitable for pharmaceutical use. The pharmaceutically acceptable salts of the compounds represented by formula (I) can be metal salts, amine salts formed with suitable acids.

[0128] "Pharmaceutical composition" means a mixture containing one or more compounds described herein or their physiologically pharmaceutically acceptable salts or prodrugs and other chemical components, as well as other components such as physiologically pharmaceutically acceptable carriers and excipients. The purpose of the pharmaceutical composition is to facilitate administration to an organism, facilitate absorption of the active ingredient and thereby exert biological activity.

[0129] Synthesis Method of the Compounds of the Invention

[0130] To achieve the object of the present invention, the following technical solutions are adopted in the present invention:

[0131] A method for preparing a compound of general formula (I) or a stereoisomer, tautomer or pharmaceutically acceptable salt thereof according to the present invention comprises the following steps:

[0132]

[0133] The compound of general formula (IA) and the compound of general formula (IB) are subjected to a Suzuki coupling reaction under the action of a palladium catalyst and a basic reagent to obtain a compound of general formula (IC); the compound of general formula (IC) is further deprotected, and optionally further deprotected on ring B, to obtain a compound of general formula (I);

[0134] Wherein:

[0135] X is a leaving group, preferably chlorine;

[0136] PG is a protecting group, preferably tert-butoxycarbonyl;

[0137] M is selected from -B(OH)2, -BF3K or

[0138] Ring B, R 1 ~R 4 、Q, Y, m and n are as defined in general formula (I). Detailed implementation mode

[0139] The following examples are used to further describe the present invention, but these examples do not limit the scope of the present invention.

[0140] Examples

[0141] The examples give the preparation of representative compounds represented by formula (I) and related structure identification data. It must be noted that the following examples are for illustrating the present invention rather than limiting the present invention. 1 The 1H NMR spectra were measured on a Bruker instrument (400 MHz), and the chemical shifts are expressed in ppm. Tetramethylsilane was used as the internal standard (0.00 ppm). 1 Representation method of 1H NMR: s = singlet, d = doublet, t = triplet, m = multiplet, br = broadened, dd = doublet of doublets, dt = doublet of triplets. When coupling constants are provided, the unit is Hz.

[0142] The mass spectra were obtained by LC / MS instrument, and the ionization mode can be ESI or APCI.

[0143] For thin-layer chromatography silica gel plates, Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plates are used. The specifications of the silica gel plates used in thin-layer chromatography (TLC) are 0.15 mm to 0.2 mm, and the specifications of the silica gel plates used for thin-layer chromatography separation and purification of products are 0.4 mm to 0.5 mm.

[0144] For column chromatography, silica gel with 200 - 300 mesh from Yantai Huanghai is generally used as the carrier.

[0145] In the following examples, unless otherwise specified, all temperatures are in Celsius. Unless otherwise specified, various starting materials and reagents are commercially available or synthesized according to known methods. Commercially available raw materials and reagents are used directly without further purification. Unless otherwise specified, commercially available manufacturers include but are not limited to Shanghai Haohong Biopharmaceutical Technology Co., Ltd., Shanghai Shaoyuan Reagent Co., Ltd., Shanghai Bide Pharmaceutical Technology Co., Ltd., Sigma-Aldrich (Shanghai) Co., Ltd., and Shanghai Linkai Pharmaceutical Technology Co., Ltd., etc.

[0146] CD3OD: Deuterated methanol.

[0147] CDCl3: Deuterated chloroform.

[0148] DMSO-d6: Deuterated dimethyl sulfoxide.

[0149] Unless otherwise specified in the examples, the solution in the reaction refers to an aqueous solution.

[0150] For the purification of compounds, the eluent systems of column chromatography and thin-layer chromatography are used, and the system is 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. The volume ratio of the solvents varies according to the polarity of the compound, and a small amount of acidic or basic reagents can also be added for conditioning, such as acetic acid or triethylamine, etc.

[0151] Room temperature: 20 °C - 30 °C.

[0152] Example 1

[0153] 4-(4-((1R,5S)-3,8-diazabicyclo[3.2.1]octan-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5-fluoronaphthalen-2-ol

[0154] 4-(4-((1R,5S)-3,8-Diazabicyclo[3.2.1]oct-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5-fluoronaphthalen-2-ol

[0155]

[0156] The first step

[0157] tert-butyl(1R,5S)-8-(2,7-dichloro-8-fluoropyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylate

[0158] (1R,5S)-8-(2,7-Dichloro-8-fluoropyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester

[0159] Dissolve 2,4,7-trichloro-8-fluoropyrido[4,3-d]pyrimidine 1a (1.1 g, 4.36 mmol) in dichloromethane (19.96 mL), add N,N-diisopropylethylamine (3.94 g, 30.50 mmol, 5.04 mL), cool down to -40 °C, add (1R,5S)-tert-butyl 3,8-diazabicyclo[3.2.1]octane-3-carboxylate 1b (924.96 mg, 4.36 mmol), and react at room temperature for 1 hour. After the reaction is completed, add water (10 mL), extract with dichloromethane (20 mL), wash the organic phase with saturated brine (20 mL × 3), dry over anhydrous sodium sulfate, filter, and purify the obtained residue by silica gel column chromatography (eluent: System A) to obtain the product (1R,5S)-8-(2,7-dichloro-8-fluoropyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester 1c (750 mg, 1.75 mmol), with a yield of 40.19%.

[0160] LC / MS: 427.9 [M+H] +

[0161] The second step

[0162] tert-butyl(1R,5S)-8-(7-chloro-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylate

[0163] (1R,5S)-8-(7-chloro-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester

[0164] Dissolve tert-butyl(1R,5S)-8-(2,7-dichloro-8-fluoropyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylate 1c (750 mg, 1.75 mmol), ((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methanol 1d (362.42 mg, 2.28 mmol), 4A molecular sieve (7.71 g, 17.51 mmol) and N,N-diisopropylethylamine (678.96 mg, 5.25 mmol, 868.24 μL) in 1,4-dioxane (3.28 mL), and heat the mixture to 90 °C for reaction for 3 hours. After the reaction is completed, filter the mixture. Wash the filtrate with saturated brine (100 mL × 3), dry it over anhydrous sodium sulfate, filter it, concentrate it under reduced pressure, and separate and purify the obtained residue by silica gel column chromatography (eluent: System B) to obtain the product tert-butyl(1R,5S)-8-(7-chloro-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylate 1e (500 mg, 907.40 μmol), with a yield of 51.82%.

[0165] LC / MS: 552.1[M + H] +

[0166] The third step

[0167] tert-butyl(1R,5S)-8-(8-fluoro-7-(8-fluoro-3-(methoxymethoxy)naphthalen-1-yl)-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylate

[0168] (1R,5S)-8-(8-Fluoro-7-(8-fluoro-3-(methoxymethoxy)naphthalen-1-yl)-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester

[0169] (1R,5S)-8-(7-Chloro-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester 1e (200 mg, 362.96 μmol), 2-(8-fluoro-3-(methoxymethoxy)naphthalen-1-yl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane 1f (168.79 mg, 508.14 μmol), potassium phosphate (231.14 mg, 1.09 mmol) and cataCXium A Pd-G3 (264.70 mg, 362.96 μmol) were dissolved in tetrahydrofuran (5 mL), protected by argon, and heated to 60 °C for reaction for 2 hours. After the reaction was completed, water (10 mL) was added, and the mixture was extracted with ethyl acetate (10 mL). The organic phase was washed with saturated brine (100 mL × 3), dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and the residue obtained was separated and purified by silica gel column chromatography (eluent: System A) to obtain the product (1R,5S)-8-(8-fluoro-7-(8-fluoro-3-(methoxymethoxy)naphthalen-1-yl)-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester 1g (180 mg, 249.73 μmol), with a yield of 68.80%.

[0170] LC / MS: 721.1 [M+H] +

[0171] The fourth step

[0172] 4-(4-((1R,5S)-3,8-diazabicyclo[3.2.1]octan-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5-fluoronaphthalen-2-ol

[0173] 4-(4-((1R,5S)-3,8-diazabicyclo[3.2.1]octan-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5-fluoronaphthalen-2-ol

[0174] Dissolve 1 g (180.00 mg, 249.73 μmol) of tert-butyl (1R,5S)-8-(8-fluoro-7-(8-fluoro-3-(methoxymethoxy)naphthalen-1-yl)-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylate in acetonitrile (5 mL). Add a 1,4-dioxane solution (5 mL) of 4 M HCl at 0 °C and react at room temperature for 0.5 h. After the reaction, add water (10 mL), extract with dichloromethane (20 mL × 2), combine the organic phases, wash the organic phases with saturated brine (100 mL × 3), dry over anhydrous sodium sulfate, filter, and concentrate under reduced pressure. The residue obtained is separated and purified by preparative liquid chromatography (separation column AKZONOBEL Kromasil; 250 × 21.2 mm ID.; 5 μm, 20 mL / min; mobile phase A: 0.05% TFA + H2O, mobile phase B: CH3CN) to obtain the product 4-(4-((1R,5S)-3,8-diazabicyclo[3.2.1]octan-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5-fluoronaphthalen-2-ol 1 (30 mg, 52.03 μmol), with a yield of 20.83%.

[0175] LC / MS: 577.1 [M+H] +

[0176] 11H NMR (500 MHz, DMSO-d6) δ 9.45 (s, 1H), 8.51 (m, J = 2.4 Hz, 1H), 8.32 (d, J = 2.2 Hz, 1H), 7.40 (t, J = 7.8 Hz, 1H), 7.16 (t, J = 2.2 Hz, 1H), 7.11 (t, J = 7.9 Hz, 1H), 7.04 (s, 1H), 5.43 - 5.37 (m, 1H), 4.40 (d, J = 10.8 Hz, 1H), 4.23 (d, J = 10.8 Hz, 1H), 3.83 (m, J = 4.3 Hz, 2H), 3.22 (m, J = 2.6 Hz, 1H), 3.17 - 3.03 (m, 4H), 2.79 (t, J = 12.1 Hz, 2H), 2.69 - 2.61 (m, 1H), 2.06 (m, J = 3.1 Hz, 1H), 2.01 - 1.94 (m, 2H), 1.94 - 1.88 (m, 2H), 1.88 - 1.65 (m, 6H).

[0177] Example 2

[0178] 4-(4-((1R,5S)-3,8-Diazabicyclo[3.2.1]octan-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5-ethynyl-6-fluoronaphthalen-2-ol

[0179] 4-(4-((1R,5S)-3,8-Diazabicyclo[3.2.1]octan-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5-ethynyl-6-fluoronaphthalen-2-ol

[0180]

[0181] The first step

[0182] tert-butyl(1R,5S)-8-(8-fluoro-7-(7-fluoro-3-(methoxymethoxy)-8-((triisopropylsilyl)ethynyl)naphthalen-1-yl)-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylate

[0183] (1R, 5S)-8-(8-fluoro-7-(7-fluoro-3-(methoxymethoxy)-8-((triisopropylsilyl)ethynyl)naphthalen-1-yl)-2-(((2R, 7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4, 3-d]pyrimidin-4-yl)-3, 8-diazabicyclo[3.2.1]octane-3-carboxylate tert-butyl ester

[0184] It should be noted that the chemical name translation you provided may not be completely accurate in terms of chemical naming norms, but it is translated according to the requirements. For more accurate chemical name translations, it is recommended to consult professional chemical literature or relevant experts.(1R,5S)-8-(7-chloro-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolazin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester 1e (279.04 mg, 544.44 μmol), ((2-fluoro-6-(methoxymethoxy)-8-(4,4,5,5-tetramethyl -1,3,2-dioxaborolane-2-yl)naphthalen-1-yl)ethynyl)triisopropylsilane 2a (279.04 mg, 544.44 μmol), cesium carbonate (236.52 mg, 72592 μmol) and Pd(dppf)Cl2 (29.64 mg, 36.30 μmol) were dissolved in a mixed solvent of 1,4-dioxane (6 mL) and water (2 mL) under argon protection and heated to 110°C for 2 hours. After the reaction was completed, water (10 mL) was added, and the mixture was extracted with ethyl acetate (20 mL). The organic phase was washed with saturated brine (20 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (eluent: System A) to give the product (1R, 5S)-8-(8-fluoro-7-(7-fluoro-3-(methoxymethoxy)-8-((triisopropylsilyl)ethynyl)naphthalen-1-yl)-2-(((2R, 7aS)-2-fluorotetrahydro-1H-pyrrolazin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester 2b (100 mg, 110.97 μmol) with a yield of 30.57%.

[0185] LC / MS: 902.1[M+H] +

[0186] Step 2

[0187] tert-butyl(1R,5S)-8-(7-(8-ethynyl-7-fluoro-3-(methoxymethoxy)naphthalen-1-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotet rahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylate

[0188] (1R,5S)-8-(7-(8-Ethynyl-7-fluoro-3-(methoxymethoxy)naphthalen-1-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester

[0189] Dissolve (1R,5S)-8-(8-fluoro-7-(7-fluoro-3-(methoxymethoxy)-8-((triisopropylsilyl)ethynyl)naphthalen-1-yl)-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester 2b (100 mg, 110.97 μmol) in N,N-dimethylformamide (2 mL), add cesium fluoride (84.28 mg, 554.85 μmol), and react at room temperature for 0.5 h. After the reaction is completed, add water (10 mL), extract with ethyl acetate (10 mL), wash the organic phase with saturated brine (10 mL × 3), dry over anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain the crude product (1R,5S)-8-(7-(8-ethynyl-7-fluoro-3-(methoxymethoxy)naphthalen-1-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester 2c (80 mg, 107.41 μmol), with a yield of 96.79%.

[0190] LC / MS: 745.1 [M+H] +

[0191] The third step

[0192] 4-(4-((1R,5S)-3,8-diazabicyclo[3.2.1]octan-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5-ethynyl-6-fluoronaphthalen-2-ol

[0193] 4-(4-((1R,5S)-3,8-Diazabicyclo[3.2.1]oct-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5-ethynyl-6-fluoronaphthalen-2-ol

[0194] tert-Butyl (1R,5S)-8-(7-(8-ethynyl-7-fluoro-3-(methoxymethoxy)naphthalen-1-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylate 2c (80.00 mg, 107.41 μmol) was dissolved in acetonitrile (2 mL), 1,4-dioxane solution of 4 M HCl (1 mL) was added, and the reaction was carried out at room temperature for 1 hour. Concentrate under reduced pressure, and the obtained residue was separated and purified by preparative liquid chromatography (separation column AKZONOBEL Kromasil; 250×21.2 mm I.D.; 5 μm, 20 mL / min; mobile phase A: 0.05% TFA + H2O, mobile phase B: CH3CN) to obtain the product 4-(4-((1R,5S)-3,8-diazabicyclo[3.2.1]oct-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5-ethynyl-6-fluoronaphthalen-2-ol 2 (10 mg, 16.65 μmol), with a yield of 15.50%.

[0195] LC / MS: 601.1 [M+H] +

[0196] 1 1H NMR (500 MHz, DMSO-d) δ 9.01 (s, 1H), 8.27 (m, J = 3.6 Hz, 1H), 7.64 (d, J = 2.2 Hz, 1H), 7.28 (t, J = 8.3 Hz, 1H), 7.20 (t, J = 2.0 Hz, 1H), 7.16 (s, 1H), 5.52 - 5.38 (m, 1H), 4.43 - 4.25 (m, 2H), 4.21 (m, J = 7.9 Hz, 2H), 3.44 (s, 1H), 3.23 (t, J = 8.4 Hz, 2H), 3.02 (t, J = 12.1 Hz, 2H), 2.97 - 2.87 (m, 2H), 2.62 - 2.55 (m, 1H), 2.10 - 1.97 (m, 2H), 1.95 - 1.70 (m, 10H).

[0197] Example 3

[0198] 4-(4-((1R,5S)-3,8-diazabicyclo[3.2.1]octan-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5,7-difluorobenzo[d]thiazol-2-amine

[0199] 4-(4-((1R,5S)-3,8-diazabicyclo[3.2.1]octan-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5,7-difluorobenzo[d]thiazol-2-amine

[0200]

[0201] The first step

[0202] tert-butyl(1R,5S)-8-(7-(2-((tert-butoxycarbonyl)amino)-5,7-difluorobenzo[d]thiazol-4-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylate

[0203] (1R,5S)-8-(7-(2-((tert-butoxycarbonyl)amino)-5,7-difluorobenzo[d]thiazol-4-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylate tert-butyl ester

[0204] (1R,5S)-8-(7-chloro-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester 1e (100 mg, 181.48 μmol), (2-((tert-butoxycarbonyl)amino)-5,7-difluorobenzo[d]thiazol-4-yl)boronic acid 3a (89.86 mg, 272.22 μmol, prepared according to published patent US20200115375A1), sodium carbonate (57.71 mg, 544.44 μmol) and tetrakis(triphenylphosphine)palladium (20.97 mg, 18.15 μmol) were dissolved in 1,4-dioxane (5 mL), protected by argon, and heated to 110 °C for reaction for 5 hours. After the reaction was completed, the reaction solution was filtered, and the obtained residue was separated and purified by preparative liquid chromatography (separation column AKZONOBEL Kromasil; 250×21.2 mm I.D.; 5 μm, 20 mL / min; mobile phase A: 0.05% TFA + H2O, mobile phase B: CH3CN) to obtain the product (1R,5S)-8-(7-(2-((tert-butoxycarbonyl)amino)-5,7-difluorobenzo[d]thiazol-4-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester 3b (45 mg, 56.19 μmol), with a yield of 30.96%.

[0205] LC / MS: 800.9 [M+H] +

[0206] The second step

[0207] 4-(4-((1R,5S)-3,8-diazabicyclo[3.2.1]octan-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5,7-difluorobenzo[d]thiazol-2-amine

[0208] 4-(4-((1R,5S)-3,8-diazabicyclo[3.2.1]oct-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5,7-difluorobenzo[d]thiazol-2-amine

[0209] (1R,5S)-8-(7-(2-((tert-Butoxycarbonyl)amino)-5,7-difluorobenzo[d]thiazol-4-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-3-carboxylic acid tert-butyl ester 3b (30 mg, 37.46 μmol) was dissolved in dichloromethane (5 mL), and a 1,4-dioxane solution of 4 M HCl (5 mL) was added. The reaction was carried out overnight at room temperature. The reaction solution was concentrated under reduced pressure, and the residue obtained was separated and purified by preparative liquid chromatography (separation column AKZONOBEL Kromasil; 250×21.2 mm I.D.; 5 μm, 20 mL / min; mobile phase A: 0.05% TFA + H2O, mobile phase B: CH3CN) to obtain the product 4-(4-((1R,5S)-3,8-diazabicyclo[3.2.1]oct-8-yl)-8-fluoro-2-(((2R,7aS)-2-fluorotetrahydro-1H-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-7-yl)-5,7-difluorobenzo[d]thiazol-2-amine 3 (3 mg, 4.79 μmol), with a yield of 12.80%.

[0210] LC / MS: 600.9 [M+H] +

[0211] Biological evaluation

[0212] Test Example 1. Determination of the inhibitory activity of the compounds of the present invention against p-ERK1 / 2 in AGS cells

[0213] The following method was used to determine the inhibitory activity of the compounds of the present invention against p-ERK1 / 2 in AGS cells. This method used the Advanced phospho-ERK1 / 2 (Thr202 / Tyr204) kit (product number 64AERPEH) from Cisbio. The detailed experimental operations can refer to the kit instructions. AGS cells (containing the KRAS G12D mutation) were purchased from the Cell Resource Center of the Shanghai Institute of Life Sciences, Chinese Academy of Sciences.

[0214] 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. 40,000 AGS cells were seeded per well in a 96-well plate with complete medium and cultured overnight in an incubator at 37°C with 5% CO2. The test compound was dissolved in DMSO to prepare a 10 mM stock solution, which was then diluted with F12K complete medium. 100 μL of F12K complete medium containing the test compound at the corresponding concentration was added to each well. The final concentration range of the test compound in the reaction system was 1000 nM - 0.015 nM. After culturing in the cell incubator for 3 hours, the cell supernatant was discarded, and the cells were washed with ice-cold PBS. Then, 50 μL of 1× cell phospho / total protein lysis buffer (component of the Advanced phospho-ERK1 / 2 kit) was added to each well for lysis. The 96-well plate was placed on ice for half an hour for lysis, and then the lysate was detected according to the instructions of the Advanced phospho-ERK1 / 2 (Thr202 / tyr204) kit. Finally, the fluorescence intensities at emission wavelengths of 620 nM and 665 nM were measured in a microplate reader in the TF-FRET mode at an excitation wavelength of 304 nM for each well, and the fluorescence intensity ratio of 665 / 620 for each well was calculated. By comparing with the fluorescence intensity ratio of the control group (0.1% DMSO), the percentage inhibition rate of the test compound at each concentration was calculated, and a non-linear regression analysis of the logarithm of the test compound concentration - inhibition rate was performed using GraphPad Prism5 software to obtain the IC 50 value.

[0215] The preferred compound of the present invention has a significant inhibitory effect on the p - ERK1 / 2 activity in AGS cells. The IC 50 of the preferred compound is < 500 nM, and the IC 50 of the more optimized compound is < 200 nM.

[0216] Test Example 2: Determination of the inhibitory effect of the compounds of the present invention on the proliferation of AsPC-1 cells

[0217] The following method was used to determine the effect of the compounds of the present invention on the proliferation of AsPC-1 (metastatic pancreatic adenocarcinoma) cells. AsPC-1 cells (containing the KRAS G12D mutation) were purchased from the Cell Resource Center of the Shanghai Institute of Life Sciences, Chinese Academy of Sciences, and cultured in RPMI 1640 medium containing 10% fetal bovine serum, 100 U penicillin, 100 μg / mL streptomycin, and 1 mM Sodium Pyruvate. Cell viability was determined by CellTiter- The assay was performed using a Luminescent Cell Viability Assay kit (Promega, catalog number G7573).

[0218] The experimental method was carried out according to the steps in the kit instruction manual, 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 the culture medium to prepare test samples, with the final concentration range of the compound being 1000 nM - 0.015 nM. Cells in the logarithmic growth phase were seeded into a 96-well cell culture plate at a density of 800 cells per well and cultured overnight in a 37 °C, 5% CO2 incubator. Subsequently, the test compound was added and the cells were cultured for an additional 120 hours. After the culture, 50 μL of CellTiter-Glo detection solution was added to each well, shaken for 5 minutes, and then left standing for 10 minutes. Subsequently, the luminescence values of each well of the samples were read on a microplate reader using the Luminescence mode. The percentage inhibition rate of the compound at each concentration point was calculated by comparing with the values of the control group (0.3% DMSO). Then, a non-linear regression analysis of the logarithm of the compound concentration - inhibition rate was performed in GraphPad Prism 5 software to obtain the IC 50 value.

[0219] The preferred compounds of the present invention have a significant inhibitory effect on the proliferation of AsPC-1 cells. The IC 50 of the preferred compounds is < 500 nM, and the IC 50 of the more optimized compounds is < 200 nM.

[0220] Test Example 3: Determination of the inhibitory ability of the compounds of the present invention on the interaction between KRAS G12D and RAF1 proteins

[0221] The following method was used to determine the ability of the compounds of the present invention to block the interaction between KRAS G12D:RAF1 proteins under in vitro conditions. This method uses a KRAS-G12C / SOS1 BINDING ASSAY KITS kit (63ADK000CB21PEG) from Cisbio. The detailed experimental procedures can refer to the kit instruction manual.

[0222] The experimental procedure is briefly described as follows: Prepare a 5X working solution concentration of Tag1-RAF1 and Tag2-KRAS-G12D proteins using diluent buffer (product number 62DLBDDF) for standby. The test compound is dissolved in DMSO to prepare a 10 mM stock solution, and then diluted with diluent buffer for standby. First, add 2 μL of the test compound (final concentration in the reaction system is 10000 nM - 0.1 nM) to the wells, then add 4 μL of the 5X working solution of Tag1-RAF1 and 4 μL of the 5X working solution of Tag2-KRAS-G12D, centrifuge and mix well, and let stand for 15 minutes; then add 10 μL of pre-mixed anti-Tag1-Eu 3+ and anti-Tag2-XL665, and incubate at room temperature for 4 hours; finally, use a microplate reader to measure the fluorescence intensities at emission wavelengths of 620 nm and 665 nm at an excitation wavelength of 304 nm in the TF-FRET mode for each well, and calculate the fluorescence intensity ratio of 665 / 620 for each well. By comparing with the fluorescence intensity ratio of the control group (0.1% DMSO), calculate the percentage inhibition rate of the test compound at each concentration, and perform a non-linear regression analysis of the logarithm of the test compound concentration - inhibition rate using GraphPad Prism 5 software to obtain the IC 50 value, as shown in Table 1 below.

[0223] Table 1 Data on the inhibitory ability of the compounds of the present invention on the interaction between KRAS G12D and RAF1 proteins

[0224] Example Number <![CDATA[IC 50 (nM)]]> 1 141 2 317 3 331

[0225] Conclusion: The compounds of the present invention have good inhibitory ability on the interaction between KRAS G12D and RAF1 proteins.

[0226] Test Example 4: Determination of the inhibitory effect of the compounds of the present invention on the proliferation of AGS cells

[0227] The following method is used to determine the effect of the compounds of the present invention on the proliferation of AGS (human gastric adenocarcinoma) cells. AGS cells (containing the KRAS G12D mutation) were purchased from the Cell Resource Center of the Shanghai Institute of Life Sciences, Chinese Academy of Sciences, and cultured in F-12K medium containing 10% fetal bovine serum, 100 U penicillin, and 100 μg / mL streptomycin. Cell viability was determined using the CellTiter- Luminescent Cell Viability Assay kit (Promega, product number G7573).

[0228] The experimental method was carried out according to 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 the culture medium to prepare test samples, with the final concentration range of the compound being 1000 nM - 0.015 nM. Cells in the logarithmic growth phase were seeded into a 96-well cell culture plate at a density of 500 cells per well and cultured overnight in an incubator at 37 °C and 5% CO2. Subsequently, the test compound was added and the cells were cultured for another 72 hours. After the culture was completed, 50 μL of CellTiter-Glo assay reagent was added to each well. After shaking for 5 minutes and standing for 10 minutes, the luminescence values of each well of the samples were read on a microplate reader using the Luminescence mode. By comparing with the values of the control group (0.3% DMSO), the percentage inhibition rate of the compound at each concentration point was calculated. Then, a non-linear regression analysis of the logarithm of the compound concentration - inhibition rate was performed using GraphPad Prism 5 software to obtain the IC50 value of the compound inhibiting cell proliferation, as shown in Table 2.

[0229] Table 2 IC50 data of the compounds of the present invention for inhibiting the proliferation of AGS cells

[0230] Example Number <![CDATA[IC 50 (nM)]]> 1 68 2 93

[0231] Conclusion: The compounds of the present invention have a good inhibitory effect on the proliferation of AGS cells.

Claims

1. A compound of formula (II) or a pharmaceutically acceptable salt thereof: Wherein: Ring B is selected from naphthyl or benzothiazolyl; Y is selected from -O-; R c is a halogen, R 1 For R 2 selected from a hydrogen atom; R 4 identical or different, each independently selected from a hydrogen atom, fluorine, a hydroxyl group, an amino group; m is selected from 0, 1, 2, 3 or 4; n is selected from 0, 1, 2 or 3.

2. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein selected from the following groups:

3. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 2, wherein R c is fluorine.

4. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein the compound is:

5. A pharmaceutical composition comprising an effective dose of the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4, and a pharmaceutically acceptable carrier, excipient or a combination thereof.

6. Use of the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4 or the pharmaceutical composition according to claim 5 in the preparation of a KRas G12D inhibitor.

7. Use of the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4 or the pharmaceutical composition according to claim 5 in the preparation of a medicament for treating a disease mediated by KRas G12D mutation, wherein the disease mediated by KRas G12D mutation is selected from cancers, and the cancers are selected from cardiac myxoma, lung cancer, gastric cancer, colorectal cancer, pancreatic cancer, prostate cancer, bladder cancer, hepatocellular carcinoma, cholangiocarcinoma, chondrosarcoma, multiple myeloma, uterine cancer, cervical cancer, seminoma, malignant melanoma, cutaneous squamous cell carcinoma, adrenocortical neuroblastoma, myeloid leukemia, acute lymphoblastic leukemia or glioblastoma.

8. The use according to claim 7, wherein the cancers are selected from pancreatic cancer, colorectal cancer and lung cancer.

9. The use according to claim 7, wherein the cancer is rectal cancer.

10. The use according to claim 7, wherein the lung cancer is selected from non-small cell lung cancer or small cell lung cancer.

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