CN113527299A

CN113527299A - Nitrogen-containing condensed ring compounds, preparation method and application

Info

Publication number: CN113527299A
Application number: CN202010747478.6A
Authority: CN
Inventors: 万惠新; 查传涛; 马金贵; 潘建峰
Original assignee: Shanghai Lingda Biomedical Co Ltd
Current assignee: Shanghai Lingda Biomedical Co Ltd
Priority date: 2020-04-18
Filing date: 2020-07-29
Publication date: 2021-10-22
Anticipated expiration: 2040-07-29
Also published as: CN113527299B

Abstract

The invention discloses a nitrogen-containing fused ring compound, a preparation method and application thereof, in particular to a nitrogen-containing fused ring compound shown as a general formula I, or pharmaceutically acceptable salt thereof, or enantiomer, diastereomer, tautomer, torsional isomer, solvate, polymorph or prodrug thereof, a preparation method and application thereof in pharmacy, wherein the definition of each group is described in the specification.

Description

Nitrogen-containing condensed ring compounds, preparation method and application

Technical Field

The invention belongs to the field of pharmaceutical chemistry, and particularly relates to a nitrogen-containing fused ring compound, a compound with Ras mutein inhibition activity, a preparation method and application.

Background

RAS is the first oncogene identified in human tumors and was first found in two murine sarcoma viruses. There are three members of the RAS gene family, Hras, Kras, Nras. In human tumors, Kras mutations are most common, accounting for approximately 85%. Previous studies have shown that Kras mutations are carcinogenic because codon 12 is missense mutated, altering the structure of the Kras protein and keeping it activated at all times. Ras plays a major role in signal pathway transmission, mainly activating kinases controlling gene transcription, thereby regulating cell differentiation and proliferation, and is closely related to survival, proliferation, migration, metastasis and angiogenesis of tumor cells. According to statistics, Kras G12C mutation exists in 11% -16% of lung adenocarcinoma cases, and part of pancreatic cancer, colorectal cancer, ovarian cancer and bile duct cancer is caused by Kras mutation. However, over thirty years ago since the first discovery of Kras oncogene, the targeting drugs for EGFR, BCL and other common protooncogenes have been developed for several generations, and the targeting drugs for Kras have not been successfully developed. Historically, targeted drugs against KRas pathway mutant tumors have focused primarily on farnesyl transferase inhibitors and Raf-MEK pathway inhibitors, but with little success. In recent years, inhibitors aiming at KRas specific gene mutation are developed into hot spots, and part of inhibitors gradually go from preclinical hatching to clinical research, such as KRas G12C inhibitors AMG510, MRTX1257 and the like, and show certain curative effect in early clinical experiments. The first clinical data of the first global KRASG12C inhibitor AMG510 was finally promulgated by the american clinical oncology institute held in 6 months 2019, in which clinical studies the installed drug AMG510 was shown to prevent tumor growth in most non-small cell lung and colorectal cancer patients with KRas mutations. Therefore, finding and searching for a target drug against KRas specific mutant gene with high specificity and excellent drug availability is a major hotspot in the industry.

Disclosure of Invention

One of the technical problems to be solved by the invention is to provide a novel KRas G12C inhibitor for preparing a tumor treatment medicament.

The scheme for solving the technical problems is as follows:

a nitrogen-containing fused cyclic compound shown as a general formula I, or a pharmaceutically acceptable salt thereof, or an enantiomer, a diastereomer, a tautomer, a torsional isomer, a solvate, a polymorph or a prodrug thereof,

in the formula:

r1 is independently selected from hydrogen, halogen, cyano, nitro, C₁-C₆Alkyl radical, C₁-C₆alkyl-SO₂-、C₁-C₆alkyl-SO-, or C₁-C₆A haloalkyl group;

r2 and R3 are independently selected from hydrogen, halogen, cyano, nitro and C₁-C₆Alkyl radical, C₁-C₆alkyl-SO₂-、C₁-C₆alkyl-SO-, N (R)_2a)(R_2b)-(CH₂) x-; or, R₉And R₁₀Together forming a 5-to 10-membered quilt C₁-C₆An alkyl-substituted nitrogen-containing heterocycloalkyl group; wherein R is_2aAnd R_2bEach independently selected from hydrogen or C₁-C₆Alkyl, x is selected from any integer of 0-5;

w, W1, W2, M is independently selected from CR4 or N, R4 is independently selected from H, halogen, cyano, C₁-C₆Alkyl radical, C₁-C₆Alkoxy, haloalkyl, haloalkoxy, alkenyl, alkynyl, alkylether, cycloalkylether, heterocycloalkylether, alkylether, cycloalkylthioether, heterocycloalkylether, 3-to 8-membered cycloalkyl or heterocycloalkyl, alkylamino-substituted alkoxy, alkylamino-substituted alkylamino, cycloalkylAlkylene ethers, heterocycloalkyl alkylene ethers, cycloalkyl alkylene amino, heterocycloalkyl alkylene amino, and the like;

ra, Rb, Rc, Rd, Re, Rf and Rg are respectively and independently selected from hydrogen, C1-C6 alkyl, alkoxy, haloalkyl and the like, or Ra, Rb, Rc, Rd, Re, Rf and Rg form a 3-8-membered saturated or partially unsaturated ring system between every two;

ar is independently selected from a 5-12 membered aromatic ring or aromatic condensed ring, a 5-12 membered aromatic heterocycle or aromatic condensed heterocycle; and the Ar ring may be substituted with one or more of the following groups: hydrogen, halogen, C1-C6 alkyl, alkoxy, 3-8 membered cycloalkyl or heterocycloalkyl, C1-C6 haloalkoxy, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted amino, amido, sulfonamido, and the like;

cy is independently selected from a 4-8 membered saturated or partially unsaturated or aromatic ring system;

one or more hydrogen atoms on any of the above groups may be substituted with a substituent selected from the group consisting of: including but not limited to deuterium, halogen C1-C8 alkyl; wherein said heteroaryl group contains 1 to 3 heteroatoms selected from the group consisting of: n, O, P or S, the heterocycloalkyl group containing 1 to 3 heteroatoms selected from the group consisting of: n, O, P or S, said ring system including spiro, bridged, fused, etc. saturated or partially unsaturated ring systems.

In some embodiments, the compound having the general formula (I), or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, torsiomer, solvate, polymorph or prodrug thereof, is preferably a compound represented by the general formulae (IIA), (IIB), (IIC), (IID), (IIE), or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, solvate, polymorph or prodrug thereof:

wherein n is 1-5, R5 is independently selected from hydrogen, halogen, C1-C6 alkyl, alkoxy, haloalkyl, haloalkoxy, amino, substituted amino, cyano, and the like; r1, R2, R3, Ra, Rb, Rc, Rd, Re, Rf, Rg, W1, W2, M, Ar are as defined above.

In other embodiments, a compound having the general formula (1), or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, torsional isomer, solvate, polymorph, or prodrug thereof, characterized by:

r1 is preferably selected from hydrogen, fluoro, methyl, cyano, etc.;

ra, Rb, Rc, Rd, Re, Rf, Rg are each independently preferably selected from hydrogen, fluorine, methyl, cyanomethylene;

w, W1 and W2 are each independently preferably selected from CR4, R4 is independently selected from hydrogen, halogen, cyano, C1-C4 alkyl, C1-C4 alkoxy, C2-C4 alkenyl, C2-C4 alkynyl, C1-C4 alkoxy, C1-C4 haloalkyl, C1-C4 haloalkoxy, etc.;

m is independently preferably selected from N OR CH, C-CN, C-OR5, R5 is preferably selected from C1-C6 alkyl, alkoxyalkylene, alkylaminoalkylene, cycloalkylalkylene, heterocycloalkylalkylene, and the like;

ar is independently preferably a monocyclic aromatic group such as a substituted or unsubstituted phenyl group or pyridyl group, or a substituted or unsubstituted bicyclic aromatic group such as a naphthyl group, a naphthyridinyl group, an indazolyl group or a benzimidazolyl group; said one or more substituents are preferably selected from the group consisting of: hydrogen, halogen, C1-C4 alkyl, hydroxy, amino, cyano, C1-C4 alkoxy, C1-C4 haloalkyl, C1-C4 haloalkoxy, and the like;

cy is as defined above.

A process for preparing a compound of formula I, said process comprising steps a-c:

a) converting a compound of formula (a) to an intermediate (B) by a transition metal catalysed coupling reaction with an arylboronic acid or arylboronic ester or arylmetal reagent (Ar-M); and

b) removing the protective group PG from the compound of the general formula (B) through a conventional functional group to obtain a compound of a general formula (C);

c) the compound of the general formula (C) and acrylic acid or acryloyl chloride are subjected to condensation reaction under proper conditions to generate the general formula (I).

The definition of each group is as described above;

preferably, said steps a), b), c) are each carried out in a solvent, and said solvent is selected from the group consisting of: water, methanol, ethanol, isopropanol, butanol, ethylene glycol methyl ether, N-methyl pyrrolidone, dimethyl sulfoxide, tetrahydrofuran, toluene, dichloromethane, 1, 2-dichloroethane, acetonitrile, N-dimethylformamide, N-dimethylacetamide, dioxane, or a combination thereof.

Preferably, the transition metal catalyst is selected from the group consisting of: tris (dibenzylideneacetone) dipalladium (Pd)₂(dba)₃) Tetrakis (triphenylphosphine) palladium (Pd (PPh)₃)₄) Palladium acetate, palladium chloride, dichlorobis (triphenylphosphine) palladium, palladium trifluoroacetate, triphenylphosphine palladium acetate, [1,1' -bis (diphenylphosphino) ferrocene]Palladium dichloride, bis (tri-o-phenylphosphino) palladium dichloride, 1, 2-bis (diphenylphosphino) ethane palladium dichloride, or a combination thereof; the catalyst ligand is selected from the group consisting of: tri-tert-butylphosphine, tri-tert-butylphosphine tetrafluoroborate, tri-n-butylphosphine, triphenylphosphine, tri-p-benzylphosphine, tricyclohexylphosphine, tri-o-phenylphosphine, or a combination thereof.

Preferably, the condensing agent is selected from the group consisting of: DCC, DIC, CDI, EDCI, HOAt, HOBt, BOP, PyBOP, HATU, TBTU, and the like, or combinations thereof.

Preferably, the inorganic base is selected from the group consisting of: sodium hydride, potassium hydroxide, sodium acetate, potassium tert-butoxide, sodium tert-butoxide, potassium fluoride, cesium fluoride, potassium phosphate, potassium carbonate, potassium bicarbonate, sodium carbonate, sodium bicarbonate, or combinations thereof; the organic base is selected from the group consisting of: pyridine, triethylamine, N, N-diisopropylethylamine, 1, 8-diazabicyclo [5.4.0] undec-7-ene (DBU), lithium hexamethyldisilazide, sodium hexamethyldisilazide, lutidine, or a combination thereof.

Preferably, the acid is selected from the group consisting of: hydrochloric acid, sulfuric acid, phosphoric acid, methanesulfonic acid, toluenesulfonic acid, trifluoroacetic acid, formic acid, acetic acid, trifluoromethanesulfonic acid, or combinations thereof.

Preferably, the reducing agent is selected from the group consisting of: iron powder, zinc powder, stannous chloride, sodium thiosulfate, sodium sulfite, hydrogen and the like.

The invention provides a class of preferred compounds of formula (I) including, but not limited to, the following structures:

the invention also aims to provide a medicament for treating or preventing tumors and a composition thereof. The technical scheme for realizing the purpose is as follows:

a pharmaceutical composition for treating tumor comprises nitrogen-containing fused ring compounds shown in the general formula (I), or pharmaceutically acceptable salts thereof, or enantiomers, diastereomers, tautomers, torsos isomers, solvates, polymorphs or prodrugs thereof, and pharmaceutically acceptable carriers.

Another object of the present invention is to provide a use of the above compound. The technical scheme for realizing the purpose is as follows:

the nitrogen-containing fused ring compound shown in the general formula (I) or pharmaceutically acceptable salt thereof, or enantiomer, diastereoisomer, tautomer, torsional isomer, solvate, polymorph or prodrug thereof is used for preparing medicaments for treating diseases related to activity or expression of Ras mutein, particularly medicaments for treating tumors. The tumor is independently selected from non-small cell lung cancer, lung adenocarcinoma, lung squamous carcinoma, breast cancer, prostatic cancer, liver cancer, skin cancer, gastric cancer, intestinal cancer, cholangiocarcinoma, brain cancer, leukemia, lymph cancer, fibroma, sarcoma, basal cell carcinoma, glioma, renal cancer, melanoma, bone cancer, thyroid cancer, nasopharyngeal cancer, pancreatic cancer, etc.

The invention relates to a compound with the structural characteristics of a general formula (I), which can inhibit various tumor cells, particularly can efficiently kill tumors related to abnormal KRas G12C mutant protein signal pathways, and is a treatment medicament with a brand-new action mechanism.

It is to be understood that within the scope of the present invention, the above-described features of the present invention and those specifically described below (e.g., in the examples) may be combined with each other to form new or preferred embodiments. The space is not described herein in a repeated fashion.

Detailed Description

The inventor has made a long-term and intensive study to prepare a compound with a novel structure shown in formula I, and finds that the compound has a better inhibitory activity for inhibiting the KRas G12C protein, the compound has a specific inhibitory effect on the KRas G12C protein at a very low concentration (which can be as low as less than 100nM), the inhibitory activity on the KRas G12C-related cell proliferation is quite excellent, and the compound has a stronger killing effect on KRas G12C-positive tumor cells at a very low concentration (which can be as low as less than 100nM), so that the compound can be used for treating related diseases such as tumors caused by KRas G12C mutation or abnormal expression. Based on the above findings, the inventors have completed the present invention.

Term(s) for

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the claimed subject matter belongs. All patents, patent applications, and publications cited herein are incorporated by reference in their entirety unless otherwise indicated.

It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the subject matter claimed. In this application, the use of the singular also includes the plural unless specifically stated otherwise. It must be noted that, as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. It should also be noted that the use of "or", "or" means "and/or" unless stated otherwise. Furthermore, the term "comprising" as well as other forms, such as "includes," "including," and "containing," are not limiting.

Definitions for the terms of the standardization sector can be found in the literature references including Carey and Sundberg "ADVANCED ORGANIC CHEMISTRY 4TH ED." Vols.A (2000) and B (2001), Plenum Press, New York. Unless otherwise indicated, conventional methods within the skill of the art are employed, such as mass spectrometry, NMR, IR and UV/VIS spectroscopy, and pharmacological methods. Unless a specific definition is set forth, the terms used herein in the pertinent description of analytical chemistry, organic synthetic chemistry, and pharmaceutical chemistry are known in the art. Standard techniques can be used in chemical synthesis, chemical analysis, pharmaceutical preparation, formulation and delivery, and treatment of patients. For example, the reaction and purification can be carried out using the instructions of the kit from the manufacturer, or according to the methods known in the art or the instructions of the present invention. The techniques and methods described above can generally be practiced according to conventional methods well known in the art, as described in various general and more specific documents referred to and discussed in this specification. In the present specification, groups and substituents thereof may be selected by one skilled in the art to provide stable moieties and compounds.

When a substituent is described by a general formula written from left to right, the substituent also includes chemically equivalent substituents obtained when the formula is written from right to left. For example, -CH 2O-is equivalent to-OCH 2-.

The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described. All documents, or portions of documents, cited in this application, including but not limited to patents, patent applications, articles, books, operating manuals, and treatises, are hereby incorporated by reference in their entirety.

Certain chemical groups defined herein are preceded by a shorthand notation to indicate the total number of carbon atoms present in the group. For example, C1-6 alkyl refers to an alkyl group as defined below having a total of 1 to 6 carbon atoms. The total number of carbon atoms in the shorthand notation excludes carbons that may be present in a substituent of the group.

In addition to the foregoing, the following terms, when used in the specification and claims of this application, have the meanings indicated below, unless otherwise specifically indicated.

In the present application, the term "halogen" means fluorine, chlorine, bromine or iodine; "hydroxy" means an-OH group; "hydroxyalkyl" refers to an alkyl group as defined below substituted with a hydroxyl (-OH) group; "carbonyl" refers to a-C (═ O) -group; "nitro" means-NO₂(ii) a "cyano" means-CN; "amino" means-NH₂(ii) a "substituted amino" refers to an amino group substituted with one or two alkyl, alkylcarbonyl, aralkyl, heteroaralkyl groups as defined below, e.g., monoalkylamino, dialkylamino, alkylamido, aralkylamino, heteroaralkylamino; "carboxyl" means-COOH.

In the present application, the term "alkyl", as a group or as part of another group (e.g. as used in groups such as halogen-substituted alkyl), means a straight or branched hydrocarbon chain group consisting only of carbon and hydrogen atoms, containing no unsaturated bonds, having, for example, from 1 to 12 (preferably from 1 to 8, more preferably from 1 to 6) carbon atoms and being attached to the rest of the molecule by single bonds. Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, 2-methylbutyl, 2-dimethylpropyl, n-hexyl, heptyl, 2-methylhexyl, 3-methylhexyl, octyl, nonyl, decyl, and the like.

In the present application, the term "alkenyl" as a group or part of another group means a straight or branched hydrocarbon chain group consisting of only carbon atoms and hydrogen atoms, containing at least one double bond, having, for example, 2 to 14 (preferably 2 to 10, more preferably 2 to 6) carbon atoms, and being connected to the rest of the molecule by a single bond, such as, but not limited to, vinyl, propenyl, allyl, but-1-enyl, but-2-enyl, pent-1, 4-dienyl, and the like.

In the present application, the term "alkynyl" as a group or part of another group means a straight or branched hydrocarbon chain group consisting solely of carbon and hydrogen atoms, containing at least one triple bond and optionally one or more double bonds, having for example 2 to 14 (preferably 2 to 10, more preferably 2 to 6) carbon atoms and being connected to the rest of the molecule by single bonds, such as but not limited to ethynyl, prop-1-ynyl, but-1-ynyl, pent-1-en-4-ynyl and the like.

In the present application, the term "cycloalkyl" as a group or part of another group means a stable non-aromatic monocyclic or polycyclic hydrocarbon group consisting of only carbon atoms and hydrogen atoms, which may include fused, bridged or spiro ring systems, having 3 to 15 carbon atoms, preferably having 3 to 10 carbon atoms, more preferably having 3 to 8 carbon atoms, and which is saturated or unsaturated and may be attached to the rest of the molecule by a single bond via any suitable carbon atom. Unless otherwise specifically indicated in the specification, carbon atoms in cycloalkyl groups may be optionally oxidized. Examples of cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cyclooctyl, 1H-indenyl, 2, 3-indanyl, 1,2,3, 4-tetrahydro-naphthyl, 5,6,7, 8-tetrahydro-naphthyl, 8, 9-dihydro-7H-benzocyclohepten-6-yl, 6,7,8, 9-tetrahydro-5H-benzocycloheptenyl, 5,6,7,8,9, 10-hexahydro-benzocyclooctenyl, fluorenyl, bicyclo [2.2.1] heptyl, 7-dimethyl-bicyclo [2.2.1] heptyl, bicyclo [2.2.1] heptenyl, bicyclo [2.2.2] octyl, bicyclo [3.1.1] heptyl, bicyclo [3.2.1] octyl, bicyclo [2.2.2] octenyl, Bicyclo [3.2.1] octenyl, adamantyl, octahydro-4, 7-methylene-1H-indenyl, octahydro-2, 5-methylene-pentalenyl and the like.

In this application, the term "heterocyclyl" as a group or part of another group means a stable 3-to 20-membered non-aromatic cyclic group consisting of 2 to 14 carbon atoms and 1 to 6 heteroatoms selected from nitrogen, phosphorus, oxygen, and sulfur. Unless otherwise specifically indicated in the specification, a heterocyclic group may be a monocyclic, bicyclic, tricyclic or higher ring system, which may include fused ring systems, bridged ring systems or spiro ring systems; wherein the nitrogen, carbon or sulfur atom in the heterocyclic group may be optionally oxidized; the nitrogen atoms may optionally be quaternized; and the heterocyclic group may be partially or fully saturated. The heterocyclic group may be attached to the rest of the molecule via a carbon atom or a heteroatom and by a single bond. In heterocyclic groups containing fused rings, one or more of the rings may be aryl or heteroaryl as defined below, provided that the point of attachment to the rest of the molecule is a non-aromatic ring atom. For the purposes of the present invention, heterocyclyl is preferably a stable 4-to 11-membered non-aromatic monocyclic, bicyclic, bridged or spiro group containing 1 to 3 heteroatoms selected from nitrogen, oxygen and sulfur, more preferably a stable 4-to 8-membered non-aromatic monocyclic, bicyclic, bridged or spiro group containing 1 to 3 heteroatoms selected from nitrogen, oxygen and sulfur. Examples of heterocyclyl groups include, but are not limited to: pyrrolidinyl, morpholinyl, piperazinyl, homopiperazinyl, piperidinyl, thiomorpholinyl, 2, 7-diaza-spiro [3.5] nonan-7-yl, 2-oxa-6-aza-spiro [3.3] heptan-6-yl, 2, 5-diaza-bicyclo [2.2.1] heptan-2-yl, azetidinyl, pyranyl, tetrahydropyranyl, thiopyranyl, tetrahydrofuranyl, oxazinyl, dioxolanyl, tetrahydroisoquinolinyl, decahydroisoquinolinyl, imidazolinyl, imidazolidinyl, quinolizinyl, thiazolidinyl, isothiazolidinyl, isoxazolidinyl, indolinyl, octahydroindolyl, octahydroisoindolyl, pyrrolidinyl, pyrazolidinyl, phthalimidyl, and the like.

In this application, the term "aryl" as a group or as part of another group means a conjugated hydrocarbon ring system group having 6 to 18 carbon atoms, preferably having 6 to 10 carbon atoms. For the purposes of the present invention, an aryl group may be a monocyclic, bicyclic, tricyclic or higher polycyclic ring system and may also be fused to a cycloalkyl or heterocyclic group as defined above, provided that the aryl group is attached to the remainder of the molecule by a single bond via an atom on the aromatic ring. Examples of aryl groups include, but are not limited to, phenyl, naphthyl, anthracenyl, phenanthrenyl, fluorenyl, 2, 3-dihydro-1H-isoindolyl, 2-benzoxazolinone, 2H-1, 4-benzoxazin-3 (4H) -one-7-yl, and the like.

In the present application, the term "arylalkyl" refers to an alkyl group as defined above substituted with an aryl group as defined above.

In this application, the term "heteroaryl" as a group or part of another group means a 5-to 16-membered conjugated ring system group having 1 to 15 carbon atoms (preferably having 1 to 10 carbon atoms) and 1 to 6 heteroatoms selected from nitrogen, oxygen and sulfur in the ring. Unless otherwise specifically indicated in the specification, a heteroaryl group may be a monocyclic, bicyclic, tricyclic or higher ring system, and may also be fused to a cycloalkyl or heterocyclic group as defined above, provided that the heteroaryl group is attached to the rest of the molecule by a single bond via an atom on the aromatic ring. The nitrogen, carbon or sulfur atoms in the heteroaryl group may be optionally oxidized; the nitrogen atoms may optionally be quaternized. For the purposes of the present invention, heteroaryl is preferably a stable 5-to 12-membered aromatic group containing 1 to 5 heteroatoms selected from nitrogen, oxygen and sulfur, more preferably a stable 5-to 10-membered aromatic group containing 1 to 4 heteroatoms selected from nitrogen, oxygen and sulfur or a 5-to 6-membered aromatic group containing 1 to 3 heteroatoms selected from nitrogen, oxygen and sulfur. Examples of heteroaryl groups include, but are not limited to, thienyl, imidazolyl, pyrazolyl, thiazolyl, oxazolyl, oxadiazolyl, isoxazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, benzimidazolyl, benzopyrazolyl, indolyl, furyl, pyrrolyl, triazolyl, tetrazolyl, triazinyl, indolizinyl, isoindolyl, indazolyl, isoindolyl, purinyl, quinolyl, isoquinolyl, diazonaphthyl, naphthyridinyl, quinoxalinyl, pteridinyl, carbazolyl, carbolinyl, phenanthridinyl, phenanthrolinyl, acridinyl, phenazinyl, isothiazolyl, benzothiazolyl, benzothienyl, oxazolyl, cinnolinyl, quinazolinyl, thiophenyl, indolizinyl, orthophenanthrolidinyl, isoxazolyl, phenoxazinyl, phenothiazinyl, 4,5,6, 7-tetrahydrobenzo [ b ] thienyl, naphthopyridyl, pyridinyl, and the like, [1,2,4] triazolo [4,3-b ] pyridazine, [1,2,4] triazolo [4,3-a ] pyrazine, [1,2,4] triazolo [4,3-c ] pyrimidine, [1,2,4] triazolo [4,3-a ] pyridine, imidazo [1,2-b ] pyridazine, imidazo [1,2-a ] pyrazine and the like.

In the present application, the term "heteroarylalkyl" refers to an alkyl group as defined above substituted with a heteroaryl group as defined above.

In this application, "optional" or "optionally" means that the subsequently described event or circumstance may or may not occur, and that the description includes instances where the event or circumstance occurs and instances where it does not. For example, "optionally substituted aryl" means that the aryl group is substituted or unsubstituted, and the description includes both substituted and unsubstituted aryl groups.

The terms "moiety," "structural moiety," "chemical moiety," "group," "chemical group" as used herein refer to a specific fragment or functional group in a molecule. Chemical moieties are generally considered to be chemical entities that are embedded in or attached to a molecule.

"stereoisomers" refers to compounds that consist of the same atoms, are bonded by the same bonds, but have different three-dimensional structures. The present invention is intended to cover various stereoisomers and mixtures thereof.

When the compounds of the present invention contain olefinic double bonds, the compounds of the present invention are intended to include both E-and Z-geometric isomers unless otherwise specified.

"tautomer" refers to an isomer formed by the transfer of a proton from one atom of a molecule to another atom of the same molecule. All tautomeric forms of the compounds of the invention are also intended to be included within the scope of the invention.

The compounds of the present invention or pharmaceutically acceptable salts thereof may contain one or more chiral carbon atoms and may therefore give rise to enantiomers, diastereomers and other stereoisomeric forms. Each chiral carbon atom may be defined as (R) -or (S) -, based on stereochemistry. The present invention is intended to include all possible isomers, as well as racemates and optically pure forms thereof. The compounds of the invention may be prepared by selecting as starting materials or intermediates racemates, diastereomers or enantiomers. Optically active isomers may be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques, e.g., crystallization and chiral chromatography.

Conventional techniques for the preparation/separation of individual isomers include Chiral synthesis from suitable optically pure precursors, or resolution of racemates (or racemates of salts or derivatives) using, for example, Chiral high performance liquid chromatography, as described, for example, in Gerald Gubitz and Martin G.Schmid (Eds.), Chiral Separations, Methods and Protocols, Methods in Molecular Biology, Vol.243, 2004; m. Stalcup, Chiral Separations, Annu. Rev. anal. chem.3:341-63, 2010; fumiss et al (eds.), VOGEL' S ENCYCOPEDIA OF PRACTICAL ORGANIC CHEMISTRY 5. TH ED., Longman Scientific and Technical Ltd., Essex,1991, 809-816; heller, acc, chem, res, 1990,23,128.

In the present application, the term "pharmaceutically acceptable salts" includes pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.

"pharmaceutically acceptable acid addition salts" refers to salts with inorganic or organic acids which retain the biological effectiveness of the free base without other side effects. Inorganic acid salts include, but are not limited to, hydrochloride, hydrobromide, sulfate, nitrate, phosphate, and the like; organic acid salts include, but are not limited to, formates, acetates, 2-dichloroacetates, trifluoroacetates, propionates, caproates, caprylates, caprates, undecylenates, glycolates, gluconates, lactates, sebacates, adipates, glutarates, malonates, oxalates, maleates, succinates, fumarates, tartrates, citrates, palmitates, stearates, oleates, cinnamates, laurates, malates, glutamates, pyroglutamates, aspartates, benzoates, methanesulfonates, benzenesulfonates, p-toluenesulfonates, alginates, ascorbates, salicylates, 4-aminosalicylates, napadisylates, and the like. These salts can be prepared by methods known in the art.

"pharmaceutically acceptable base addition salts" refers to salts with inorganic or organic bases which maintain the biological effectiveness of the free acid without other side effects. Salts derived from inorganic bases include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, and the like. Preferred inorganic salts are ammonium, sodium, potassium, calcium and magnesium salts. Salts derived from organic bases include, but are not limited to, the following: primary, secondary and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins, such as ammonia, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, triethanolamine, dimethylethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, choline, betaine, ethylenediamine, glucosamine, methylglucamine, theobromine, purine, piperazine, piperidine, N-ethylpiperidine, polyamine resins, and the like. Preferred organic bases include isopropylamine, diethylamine, ethanolamine, trimethylamine, dicyclohexylamine, choline, and caffeine. These salts can be prepared by methods known in the art.

"polymorph" refers to different solid crystalline phases of certain compounds of the present invention in the solid state due to the presence of two or more different molecular arrangements. Certain compounds of the present invention may exist in more than one crystalline form and the present invention is intended to include the various crystalline forms and mixtures thereof.

Typically, crystallization will result in solvates of the compounds of the invention. The term "solvate" as used herein refers to an aggregate comprising one or more molecules of the compound of the present invention and one or more solvent molecules. The solvent may be water, in which case the solvate is a hydrate. Alternatively, the solvent may be an organic solvent. Thus, the compounds of the present invention may exist as hydrates, including monohydrates, dihydrate, hemihydrate, sesquihydrates, trihydrate, tetrahydrate, and the like, as well as the corresponding solvated forms. The compounds of the invention may form true solvates, but in some cases it is also possible to retain only adventitious water or a mixture of water plus a portion of adventitious solvent. The compounds of the invention may be reacted in a solvent or precipitated or crystallized from a solvent. Solvates of the compounds of the invention are also included within the scope of the invention.

The invention also includes prodrugs of the above compounds. In the present application, the term "prodrug" denotes a compound that can be converted under physiological conditions or by solvolysis to the biologically active compound of the invention. Thus, the term "prodrug" refers to a pharmaceutically acceptable metabolic precursor of a compound of the invention. Prodrugs may not be active when administered to a subject in need thereof, but are converted in vivo to the active compounds of the invention. Prodrugs are generally rapidly converted in vivo to yield the parent compound of the invention, for example, by hydrolysis in blood. Prodrug compounds generally provide solubility, histocompatibility, or sustained release advantages in mammalian organisms. Prodrugs include known amino protecting groups and carboxyl protecting groups. Specific methods for preparing prodrugs can be found in Saulnier, M.G., et al, bioorg.Med.chem.Lett.1994,4, 1985-1990; greenwald, r.b., et al, j.med.chem.2000,43,475.

In the present application, a "pharmaceutical composition" refers to a formulation of a compound of the present invention with a vehicle generally accepted in the art for delivery of biologically active compounds to a mammal (e.g., a human). The medium includes a pharmaceutically acceptable carrier. The purpose of the pharmaceutical composition is to facilitate administration to an organism, facilitate absorption of active ingredients and exert biological activity.

The term "pharmaceutically acceptable" as used herein refers to a substance (e.g., carrier or diluent) that does not affect the biological activity or properties of the compounds of the present invention and is relatively non-toxic, i.e., the substance can be administered to an individual without causing an adverse biological response or interacting in an adverse manner with any of the components contained in the composition.

As used herein, a "pharmaceutically acceptable carrier" includes, but is not limited to, any adjuvant, carrier, excipient, glidant, sweetener, diluent, preservative, dye/colorant, flavoring agent, surfactant, wetting agent, dispersing agent, suspending agent, stabilizing agent, isotonic agent, solvent, or emulsifying agent that is approved by the relevant governmental regulatory agency for human or livestock use.

The "tumor" and "diseases related to abnormal cell proliferation" include, but are not limited to, leukemia, gastrointestinal stromal tumor, histiocytic lymphoma, non-small cell lung cancer, pancreatic cancer, squamous cell lung cancer, lung adenocarcinoma, breast cancer, prostate cancer, liver cancer, skin cancer, epithelial cell cancer, cervical cancer, ovarian cancer, intestinal cancer, nasopharyngeal cancer, brain cancer, bone cancer, esophageal cancer, melanoma, renal cancer, oral cancer, and the like.

The terms "preventing," "prevention," and "prevention" as used herein include reducing the likelihood of occurrence or worsening of a disease or disorder in a patient.

As used herein, the term "treatment" and other similar synonyms include the following meanings:

(i) preventing the occurrence of a disease or condition in a mammal, particularly when such mammal is susceptible to the disease or condition, but has not been diagnosed as having the disease or condition;

(ii) inhibiting the disease or disorder, i.e., arresting its development;

(iii) alleviating the disease or condition, i.e., causing regression of the state of the disease or condition; or

(iv) Alleviating the symptoms caused by the disease or disorder.

The terms "effective amount," "therapeutically effective amount," or "pharmaceutically effective amount" as used herein, refer to an amount of at least one agent or compound that is sufficient to alleviate one or more symptoms of the disease or disorder being treated to some extent after administration. The result may be a reduction and/or alleviation of signs, symptoms, or causes, or any other desired change in a biological system. For example, an "effective amount" for treatment is the amount of a composition comprising a compound disclosed herein that is clinically necessary to provide a significant remission effect of the condition. An effective amount suitable in any individual case can be determined using techniques such as a dose escalation assay.

The terms "administering," "administration," "administering," and the like as used herein refer to a method capable of delivering a compound or composition to a desired site for biological action. These methods include, but are not limited to, oral routes, via the duodenal route, parenteral injection (including intravenous, subcutaneous, intraperitoneal, intramuscular, intraarterial injection or infusion), topical administration, and rectal administration. Administration techniques useful for The compounds and methods described herein are well known to those skilled in The art, for example, in Goodman and Gilman, The pharmaceutical Basis of Therapeutics, current ed.; pergamon; and Remington's, Pharmaceutical Sciences (current edition), Mack Publishing Co., Easton, Pa. In preferred embodiments, the compounds and compositions discussed herein are administered orally.

The terms "drug combination", "administering other treatment", "administering other therapeutic agent" and the like as used herein refer to a drug treatment obtained by mixing or combining more than one active ingredient, including fixed and unfixed combinations of active ingredients. The term "fixed combination" refers to the simultaneous administration of at least one compound described herein and at least one co-agent to a patient in the form of a single entity or a single dosage form. The term "non-fixed combination" refers to the simultaneous administration, concomitant administration, or sequential administration at variable intervals of at least one compound described herein and at least one synergistic formulation to a patient as separate entities. These also apply to cocktail therapy, for example the administration of three or more active ingredients.

It will also be appreciated by those skilled in the art that in the processes described below, the functional groups of the intermediate compounds may need to be protected by suitable protecting groups. Such functional groups include hydroxyl, amino, mercapto and carboxylic acid. Suitable hydroxy protecting groups include trialkylsilyl or diarylalkylsilyl groups (e.g.tert-butyldimethylsilyl, tert-butyldiphenylsilyl or trimethylsilyl), tetrahydropyranyl, benzyl, and the like. Suitable protecting groups for amino, amidino and guanidino include t-butyloxycarbonyl, benzyloxycarbonyl and the like. Suitable thiol protecting groups include-C (O) -R "(where R" is alkyl, aryl or aralkyl), p-methoxybenzyl, trityl and the like. Suitable carboxyl protecting groups include alkyl, aryl or aralkyl esters.

Protecting groups may be introduced and removed according to standard techniques known to those skilled in the art and as described herein. The use of protecting Groups is described in detail in Greene, T.W. and P.G.M.Wuts, Protective Groups in Organic Synthesis, (1999),4th Ed., Wiley. The protecting group may also be a polymeric resin.

The invention will be further illustrated with reference to specific examples. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. Experimental procedures without specific conditions noted in the following examples, generally according to conventional conditions, or according to conditions recommended by the manufacturer. Percentages and parts are by weight unless otherwise indicated.

The first preparation method of the intermediate comprises the following steps: synthesis of nitroquinolines

Intermediate 1A: 7-bromo-4,6-dichloro-8-fluoro-3-nitroquinoline

The intermediate compound 1A is prepared by using commercial reagents as raw materials and referring to a synthetic route and a method of a patent WO2019110751A 1.

Intermediate 1B: 7-bromo-4, 6-dichloro-3-nitro-8- (2,2, 2-trifluoroethoxy) quinoline

The first step is as follows: 1 bromo-2-fluoro-3-nitrobenzene (10g, 45.7mmol) and trifluoroethanol (13.7g, 137mmol) were dissolved in N.N-Dimethylformamide (DMF) (100mL), potassium carbonate (12.6g, 91.4mmol) was added, and the mixture was heated to 90 ℃ in a jar and reacted overnight. The reaction solution was cooled to room temperature, poured into water, and a solid precipitated, filtered, the filter cake dried and then purified by beating with ethyl acetate to give 1-bromo-3-nitro-2- (2,2, 2-trifluoroethoxy) benzene (6.5g, yellow solid). LC-MS ESI [ M + H ]]⁺＝300.1。

The second step is that: 1-bromo-3-nitro-2- (2,2, 2-trifluoroethoxy) benzene (6.0g, 20.1mmol) was dissolved in acetic acid (HOAc) (60mL), and reduced iron powder (2.2g, 40.2mmol) was added in portions and reacted at room temperature overnight. The reaction solution was filtered, concentrated under reduced pressure, and a saturated sodium bicarbonate solution was added to precipitate a solid, which was then filtered and the filter cake was dried to give 3-bromo-2- (2,2, 2-trifluoroethoxy) aniline (4.6g, yellow solid). LC-MS ESI [ M + H ]]⁺＝270.1。

The third step: 3-bromo-2- (2,2, 2-trifluoroethoxy) aniline (4.0g, 14.9mmol) was dissolved in ethanol (EtOH) (40mL) and N-chlorosuccinimide (NCS) was added in portions) (4.0g, 30.0mmol), and reacted at room temperature overnight. The reaction solution was filtered, concentrated under reduced pressure, added with a saturated sodium bicarbonate solution, extracted three times with ethyl acetate, dried, concentrated under reduced pressure, and purified by column chromatography to give 3-bromo-4-chloro-2- (2,2, 2-trifluoroethoxy) aniline (2.2g, yellow solid). LC-MS ESI [ M + H ]]⁺＝304.3。

The fourth step: 3-bromo-4-chloro-2- (2,2, 2-trifluoroethoxy) aniline (2.0g, 6.6mmol) was dissolved in EtOH (20mL), diethyl 2- (ethoxyalkenyl) malonate (1.7g, 7.9mmol) was added, and the reaction was allowed to warm to 80 ℃ overnight. Cooling the reaction liquid to room temperature, decompressing and concentrating, pulping and purifying by petroleum ether, filtering, drying a filter cake, dissolving in diphenyl ether (10mL), heating to 245 ℃, and reacting for 0.5 hour. Cooled to room temperature, poured into petroleum ether, and a solid precipitated, filtered, and dried to give ethyl 7-bromo-6-chloro-4-hydroxy-8- (2,2, 2-trifluoroethoxy) quinoline-3-carboxylate (2.0g, yellow solid). LC-MS ESI [ M + H ]]⁺＝428.2。

The fifth step: ethyl 7-bromo-6-chloro-4-hydroxy-8- (2,2, 2-trifluoroethoxy) quinoline-3-carboxylate (2.0g, 4.7mmol) was dissolved in EtOH (20mL), 2M aqueous sodium hydroxide (11mL, 22.0mmol) was added, and the reaction was warmed to 100 ℃ for 0.5 hour. The reaction solution was cooled to room temperature, diluted with water (70mL), adjusted to pH 3 with 2N dilute hydrochloric acid, and the solid precipitated, filtered, the filter cake was dried, suspended in concentrated hydrochloric acid (20mL), and heated to 100 ℃ for 8 hours. Cooled to room temperature, filtered and dried to give 7-bromo-6-chloro-8- (2,2, 2-trifluoroethoxy) quinolin-4-ol (1.4g, light brown solid). LC-MS ESI [ M + H ]]⁺＝356.2。

And a sixth step: 7-bromo-6-chloro-8- (2,2, 2-trifluoroethoxy) quinolin-4-ol (1.2g, 3.4mmol) was dissolved in glacial acetic acid (10mL), 48% concentrated nitric acid (1mL, 10.7mmol) was added, and the reaction was warmed to 80 ℃ for 2 hours. The reaction was cooled to room temperature, diluted with water (30mL), and the solid precipitated, filtered and dried to give 7-bromo-6-chloro-3-nitro-8- (2,2, 2-trifluoroethoxy) quinolin-4-ol (0.6g, yellow solid). LC-MS ESI [ M + H ]]⁺＝401.2。

The seventh step: dissolving 7-bromo-6-chloro-3-nitro-8- (2,2, 2-trifluoroethoxy) quinolin-4-ol (0.6g, 1.5mmol) in phosphorus oxychloride (8mL), adding DMF (0.1mL), heating to 100 deg.C, reacting for 2hThen (c) is performed. The reaction was cooled to room temperature, toluene (20mL) was added, the mixture was concentrated under reduced pressure, and saturated sodium bicarbonate NaHCO was added to the residue₃Aqueous (10mL) was extracted 3 times with dichloromethane DCM, dried over anhydrous sodium sulfate and concentrated under reduced pressure to give intermediate 1B (0.55g, yellow solid). LC-MS ESI [ M + H ]]⁺＝419.1。

Intermediate 1C: 7-bromo-4-chloro-8- (2, 2-difluoroethoxy) -3-nitro-6-vinylquinoline

2, 2-difluoroethanol is used as a raw material, and an intermediate 1C is prepared according to the synthesis method and conditions of the intermediate 1B. LC-MS ESI [ M + H ]]⁺＝393.1。

Examples general preparative method one

The first step is as follows: nitroquinoline/cinnoline intermediate (1eq.) was dissolved in anhydrous tetrahydrofuran, and N, N-Diisopropylethylamine (DIPEA) (1.6eq.) and piperazine intermediate (1.5eq.) were added in this order, and the mixture was heated under reflux for 18 hours under nitrogen protection. TLC to monitor the reaction completion, cooling to room temperature, concentrating under reduced pressure, adding water and dichloromethane to the residue, phase-separating, extracting the aqueous phase with dichloromethane three times, drying the extract with anhydrous sodium sulfate, concentrating under reduced pressure, and using the residue directly in the next reaction.

The second step is that: dissolving the crude product (1eq.) in anhydrous glacial acetic acid, slowly adding reduced iron powder (3eq.), heating to 80 ℃ under the protection of nitrogen, and stirring for 0.5 hour. After the reaction was completed, the mixture was filtered through celite, washed with ethyl acetate, and the filtrate was concentrated. Diluting the residue with dichloromethane, washing with saturated sodium bicarbonate solution and saturated saline solution in sequence, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, separating and purifying the residue with silica gel column chromatography to obtain the target product, and confirming the structure by nuclear magnetism and mass spectrometry.

The third step: the second-step product (1eq.) was dissolved in a mixed solvent of acetonitrile/triethylamine (3/1), phosphorus pentasulfide (1eq.) was added in portions, and the mixture was heated under reflux for 3 hours under nitrogen. After the reaction is finished, cooling to room temperature, separating out solids, filtering, drying a filter cake in vacuum to obtain a target product, and confirming the structure by adopting nuclear magnetism and mass spectrometry.

The fourth step: dissolving the product (1eq.) obtained in the third step in a mixed solvent of ethanol/diisopropylethylamine DIPEA (5/1), adding 2, 2-dimethoxyethylamine (1eq.), and heating and refluxing for 6 hours under the protection of nitrogen. After the reaction is finished, cooling to room temperature, decompressing and concentrating, dissolving the residue in glacial acetic acid, heating to 100 ℃, and stirring for 2 hours. Cooling to room temperature, concentrating under reduced pressure, diluting the residue with dichloromethane, washing with saturated sodium bicarbonate solution and saturated sodium chloride solution sequentially, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, separating and purifying the residue with silica gel column chromatography to obtain the target product, and confirming the structure by nuclear magnetism and mass spectrometry.

The fifth step: dissolving the product of the fourth step (1eq.) in a mixed solvent of anhydrous dioxane/water (4/1), and sequentially adding boric acid or boric acid pinacol ester (2eq.), anhydrous potassium carbonate powder (2.5eq.) and [1,1' -bis (diphenylphosphino) ferrocene]Palladium dichloride (Pd (dppf) Cl₂) (0.1eq.) and reflux with heating under nitrogen for 2 hours. Monitoring the reaction completion by Thin Layer Chromatography (TLC), cooling to room temperature, concentrating under reduced pressure, diluting the residue with dichloromethane, washing with saturated ammonium chloride solution and saturated saline solution in sequence, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, separating and purifying the residue with silica gel column chromatography to obtain the target product, and confirming the structure by adopting nuclear magnetism and mass spectrometry.

And a sixth step: the product of the fifth step (1eq.) was dissolved in methanol, and 4M hydrogen chloride (HCl) in methanol (20eq.) was added and stirred at room temperature for 3 hours. Monitoring the reaction completion by thin layer chromatography TLC, concentrating under reduced pressure, dissolving the residue in dichloromethane, sequentially adding DIPEA (3eq.) and acryloyl chloride (1eq.) at 0 ℃, stirring for 0.5 h, washing the reaction solution with saturated ammonium chloride solution and saturated common salt solution, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, separating and purifying the residue by silica gel column chromatography to obtain the target compound, and confirming the structure by nuclear magnetism and mass spectrometry.

EXAMPLES general preparation method II

The first step is as follows: the quinoline/cinnoline amide intermediate (1eq.) was dissolved in anhydrous dichloromethane, and trimethyloxonium tetrafluoroborate (1.2eq.) was added at 0 ℃ and stirred at room temperature for 12 hours under nitrogen. The reaction was monitored by TLC for completion, and the reaction solution was washed successively with saturated sodium bicarbonate solution and saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure, and the residue was used directly in the next reaction.

The second step is that: dissolving the crude product (1eq.) in methanol/concentrated ammonia water (1/1), placing in a sealed tank, adding catalytic amount of p-toluenesulfonic acid, heating to 100 deg.C, and stirring for 6 hr. After the reaction, the reaction solution was concentrated under reduced pressure. The residue was diluted with dichloromethane, washed successively with a saturated sodium bicarbonate solution and a saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure, and the residue was used directly in the next reaction.

The third step: the crude product of the second step (1eq.) was dissolved in ethanol, and phosphate buffer solution (10mL, pH 6.7), sodium acetate (2eq.) and 40% chloroacetaldehyde in water (10eq.) were added, heated to 80 ℃ and stirred for 24 hours. Cooled to room temperature, and the reaction mixture was concentrated under reduced pressure. Diluting the residue with dichloromethane, washing with saturated ammonium chloride solution and saturated saline solution in sequence, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, separating and purifying the residue with silica gel column chromatography to obtain the target product, and confirming the structure by nuclear magnetism and mass spectrometry.

The fourth step: dissolving the product of the third step (1eq.) in a mixed solvent of anhydrous dioxane and water (4/1), and sequentially adding boric acid or boric acid pinacol ester (2eq.), anhydrous potassium carbonate powder (2.5eq.) and Pd (dppf) Cl₂(0.1eq.) and reflux with heating under nitrogen for 2 hours. TLC to monitor the reaction completion, cooling to room temperature, concentrating under reduced pressure, diluting the residue with dichloromethane, washing with saturated ammonium chloride solution and saturated sodium chloride solution in order, drying over anhydrous sodium sulfate, filtering, concentrating under reduced pressure, and using silica gel to wash the residueAnd (4) carrying out column chromatography separation and purification to obtain a target product, and confirming the structure by adopting nuclear magnetism and mass spectrometry.

The fifth step: the product of the fourth step (1eq.) was dissolved in methanol, and 4M HCl in methanol (20eq.) was added and stirred at room temperature for 3 hours. TLC monitors the reaction to be complete, and the reaction is concentrated under reduced pressure, the residue is dissolved in dichloromethane, DIPEA (3eq.) and acryloyl chloride (1eq.) are added in turn at 0 ℃, the mixture is stirred for 0.5 hour, the reaction solution is washed by saturated ammonium chloride solution and saturated common salt solution, anhydrous sodium sulfate is dried, the filtration and the concentration under reduced pressure are carried out, the residue is separated and purified by silica gel column chromatography to obtain the target compound, and the structure is confirmed by nuclear magnetism and mass spectrometry.

Examples general preparative method three

The first step is as follows: quinoline/cinnoline thioamide intermediate (1eq.) is dissolved in tetrahydrofuran, hydrazine hydrate (1.5eq.) is added, and the mixture is heated under reflux for 6 hours under nitrogen protection. After the reaction is finished, cooling to room temperature, carrying out reduced pressure concentration, separating out solids, filtering, carrying out vacuum drying on a filter cake to obtain a target product, and confirming the structure by adopting nuclear magnetism and mass spectrometry.

The second step is that: the product of the first step (1eq.) was dissolved in dichloromethane, trimethyl orthoformate (4eq.) was added, and after stirring for ten minutes trifluoroacetic acid (1eq.) was added. And continuously stirring for one hour at room temperature, concentrating under reduced pressure, separating and purifying the residue by using a silica gel column chromatography to obtain a target product, and confirming the structure by adopting nuclear magnetism and mass spectrometry.

The third step: dissolving the second-step product (1eq.) in a mixed solvent of anhydrous dioxane/water (4/1), and sequentially adding boric acid or boric acid pinacol ester (2eq.), anhydrous potassium carbonate powder (2.5eq.) and Pd (dppf) Cl₂(0.1eq.) and reflux with heating under nitrogen for 2 hours. TLC monitoring reaction completion, cooling to room temperature, concentrating under reduced pressure, diluting the residue with dichloromethane, washing with saturated ammonium chloride solution and saturated saline solution in sequence, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, separating and purifying the residue with silica gel column chromatography to obtain target product, and performing nuclear magnetic resonanceAnd mass spectrometry confirmed structure.

The fourth step: the product of the third step (1eq.) was dissolved in methanol, and 4M HCl in methanol (20eq.) was added and stirred at room temperature for 3 hours. TLC monitors the reaction to be complete, and the reaction is concentrated under reduced pressure, the residue is dissolved in dichloromethane, DIPEA (3eq.) and acryloyl chloride (1eq.) are added in turn at 0 ℃, the mixture is stirred for 0.5 hour, the reaction solution is washed by saturated ammonium chloride solution and saturated common salt solution, anhydrous sodium sulfate is dried, the filtration and the concentration under reduced pressure are carried out, the residue is separated and purified by silica gel column chromatography to obtain the target compound, and the structure is confirmed by nuclear magnetism and mass spectrometry.

Examples general preparative method four

The first step is as follows: dissolving the quinoline/cinnoline hydrazine intermediate (1eq.) in dry pyridine, dropwise adding acyl chloride (1.5eq.) under ice-bath cooling, and heating to 100 ℃ under the protection of nitrogen and stirring overnight. After the reaction is finished, cooling the reaction liquid to room temperature, concentrating under reduced pressure, diluting the residues with a saturated sodium carbonate solution, extracting with dichloromethane, washing the organic phase with water and saturated saline solution in turn, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, separating and purifying the residues with a silica gel column chromatography to obtain a target product, and confirming the structure by adopting nuclear magnetism and mass spectrometry.

The second step is that: dissolving the product of the first step (1eq.) in a mixed solvent of anhydrous dioxane and water (4/1 vol.), and sequentially adding boric acid or boric acid pinacol ester (2eq.), anhydrous potassium carbonate powder (2.5eq.) and Pd (dppf) Cl₂(0.1eq.) and reflux with heating under nitrogen for 2 hours. TLC monitoring reaction is complete, cooling to room temperature, decompression concentrating, diluting the remainder with dichloromethane, washing with saturated ammonium chloride solution and saturated salt solution in turn, drying with anhydrous sodium sulfate, filtering, decompression concentrating, separating and purifying the remainder with silica gel column chromatography to obtain the target product, and confirming the structure by nuclear magnetism and mass spectrum.

The third step: the second product (1eq.) was dissolved in methanol, and 4M HCl in methanol (20eq.) was added and stirred at room temperature for 3 hours. TLC monitors the reaction to be complete, and the reaction is concentrated under reduced pressure, the residue is dissolved in dichloromethane, DIPEA (3eq.) and acryloyl chloride (1eq.) are added in turn at 0 ℃, the mixture is stirred for 0.5 hour, the reaction solution is washed by saturated ammonium chloride solution and saturated common salt solution, anhydrous sodium sulfate is dried, the filtration and the concentration under reduced pressure are carried out, the residue is separated and purified by silica gel column chromatography to obtain the target compound, and the structure is confirmed by nuclear magnetism and mass spectrometry.

Examples general preparative method five

The first step is as follows: the quinoline/cinnoline amine intermediate (1eq.) was suspended in isopropanol and dimethylformamide-dimethylacetal (1.2eq.) was added dropwise at room temperature. After the dropwise addition, the reaction solution was refluxed for three hours under heating, cooled to room temperature, added with hydroxylamine hydrochloride (1.2eq.) and stirred at 50 ℃ overnight. The reaction solution was cooled to room temperature, concentrated under reduced pressure, dried, and the residue was suspended in anhydrous tetrahydrofuran, cooled in an ice bath, and then trifluoroacetic anhydride (1.5eq.) was slowly added dropwise. After the addition was complete, the ice bath was removed and stirred at room temperature overnight. Slowly dripping saturated sodium bicarbonate solution into the reaction solution, extracting with dichloromethane, washing with water and saturated saline solution in sequence, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, separating and purifying the residue with silica gel column chromatography to obtain the target product, and confirming the structure by adopting nuclear magnetism and mass spectrometry.

The second step is that: dissolving the first step product (1eq.) in anhydrous dioxane/water (4/1), and sequentially adding boric acid or boric acid pinacol ester (2eq.), anhydrous potassium carbonate powder (2.5eq.) and Pd (dppf) Cl₂(0.1eq.) and reflux with heating under nitrogen for 2 hours. TLC monitoring reaction is complete, cooling to room temperature, decompression concentrating, diluting the remainder with dichloromethane, washing with saturated ammonium chloride solution and saturated salt solution in turn, drying with anhydrous sodium sulfate, filtering, decompression concentrating, separating and purifying the remainder with silica gel column chromatography to obtain the target product, and confirming the structure by nuclear magnetism and mass spectrum.

Examples preparation

Example 1: 1- ((15aR) -9-chloro-7-fluoro-8- (2-fluoro-6-hydroxyphenyl) -12,13,15,15 a-tetrahydro-14H-pyrazino [1',2':4,5] [1,2,4] triazolo [4',3':1,6] pyrazino [2,3-c ] quinolin-14-yl) prop-2-en-1-one

The first step is as follows: intermediate 7-bromoo-4, 6-dichoro-8-fluoroquinoline (4.1g,12.1mmol) was dissolved in 1, 4-dioxane (40mL), and 1- (tert-butoxycarbonyl) -piperazine-3- (R) -carboxylic acid methyl ester (3.3g,13.5mmol) and DIPEA (4.6g,35.7mmol) were added and reacted at 100 degrees for 2 days under nitrogen. The reaction solution was cooled to room temperature, concentrated under reduced pressure, and purified by column chromatography to give methyl 1- (tert-butoxycarbonyl) - (R) -4- (7-bromo-6-chloro-8-fluoro-3-nitroquinolin-4-yl) piperazine-3-carboxylate (2.5g, yellow solid). LC-MS ESI [ M + H ]]⁺＝546.8；¹H NMR(400MHz,DMSO_d6):δ9.16(s,1H),8.35(d,J＝1.2Hz,1H),4.37(s,1H),3.85-4.15(m,2H),3.62-3.75(m,2H),3.55(s,3H),3.22-3.32(m,2H),1.44(s,9H)。

The second step is that: methyl 1- (tert-butoxycarbonyl) - (R) -4- (7-bromo-6-chloro-8-fluoro-3-nitroquinolin-4-yl) piperazine-3-carboxylate (2.2g,4.0mmol) was dissolved in glacial acetic acid (30mL), and reduced iron powder (790mg,14.1mmol) was added and reacted at 80 degrees for 40 minutes under nitrogen. The reaction was cooled to room temperature, concentrated under reduced pressure, and to the residue was added saturated sodium bicarbonate (NaHCO)₃) Adjusting the pH value of the solution to 8, extracting the solution for three times by dichloromethane, drying the solution by anhydrous sodium sulfate, and concentrating the solution under reduced pressure to obtain (R) -10-bromo-11-chloro-9-fluoro-5-oxo-1, 2,4,4a,5, 6-hexahydro-3H-pyrazine [1',2':4,5]Pyrazine [2,3-c ]]Quinoline-3-Tert-butyl carboxylate (1.91g, yellow solid). LC-MS ESI [ M + H ]]⁺＝485.2；¹H NMR(400MHz,DMSO_d6):δ11.06(s,1H),8.62(s,1H),8.08(s,1H),4.66-4.70(m,1H),3.84(brs,2H),3.19-3.34(m,3H),2.70(brs,1H),1.44(s,9H)。

The third step: reacting (R) -10-bromo-11-chloro-9-fluoro-5-oxo-1, 2,4,4a,5, 6-hexahydro-3H-pyrazine [1',2':4,5]Pyrazine [2,3-c ]]Quinoline-3-carboxylic acid tert-butyl ester (250mg,0.52mmol) was suspended in toluene (15mL), and Lawson's reagent (210mg,0.52mmol) was added under nitrogen and heated under reflux overnight. Cooling the reaction liquid to room temperature, decompressing and concentrating, and purifying the residue by column chromatography to obtain (R) -10-bromo-11-chloro-9-fluoro-5-sulfo-1, 2,4,4a,5, 6-hexahydro-3H-pyrazine [1',2':4,5]Pyrazine [2,3-c ]]Quinoline-3-carboxylic acid tert-butyl ester (204mg, yellow solid). LC-MS ESI [ M + H ]]⁺＝500.9/502.9。

The fourth step: the (R) -10-bromo-11-chloro-9-fluoro-5-thio-1, 2,4,4a,5, 6-hexahydro-3H-pyrazine [1',2':4,5]Pyrazine [2,3-c ]]Quinoline-3-carboxylic acid tert-butyl ester (154mg,0.31mmol) was dissolved in Tetrahydrofuran (THF) (4mL), hydrazine hydrate (154mg,3.1mmol) was added, and the mixture was refluxed for 4 hours under nitrogen. Cooling the reaction liquid to room temperature, and concentrating under reduced pressure to obtain (R, Z) -10-bromo-11-chloro-9-fluoro-5-hydrazinoalkenyl-1, 2,4,4a,5, 6-hexahydro-3H-pyrazine [1',2':4,5]Pyrazine [2,3-c ]]Quinoline-3-carboxylic acid tert-butyl ester (159mg, crude, yellow solid). LC-MS ESI [ M + H ]]⁺＝498.8/500.8。

The fifth step: reacting (R, Z) -10-bromo-11-chloro-9-fluoro-5-hydrazinoalkenyl-1, 2,4,4a,5, 6-hexahydro-3H-pyrazine [1',2':4,5]Pyrazine [2,3-c ]]Tert-butyl quinoline-3-carboxylate (159mg, crude) was suspended in trimethyl orthoformate (2mL) and reacted at 80 ℃ overnight under nitrogen. Cooling the reaction liquid to room temperature, decompressing and concentrating, and purifying the residue by column chromatography to obtain (R) -8-bromo-9-chloro-7-fluoro-12, 13,15,15 a-tetrahydro-14H-pyrazine [1',2':4,5][1,2,4]Triazole [4',3':1,6]Pyrazine [2,3-c ]]Quinoline-14-carboxylic acid tert-butyl ester (136mg, yellow solid). LC-MS ESI [ M + H ]]⁺＝508.9/510.9；¹H NMR(400MHz,DMSO_d6):δ9.56(s,1H),9.49(s,1H),8.28(s,1H),4.84-4.88(m,1H),4.60-4.64(m,1H),3.67-3.83(m,2H),3.23-3.38(m,2H),2.56-2.67(m,1H),1.48(s,9H)。

And a sixth step: reacting (R) -8-bromo-9-chloro-7-fluoro-12, 13,15a-tetrahydro-14H-pyrazine [1',2':4,5][1,2,4]Triazole [4',3':1,6]Pyrazine [2,3-c ]]Quinoline-14-carboxylic acid tert-butyl ester (136mg,0.27mmol) and (2-fluoro-6-hydroxyphenyl) boronic acid (104mg,0.67mmol) were dissolved in dioxane 1.4-dioxane/water (6mL/2mL), and potassium carbonate (K) was added sequentially under nitrogen protection₂CO₃) (186mg,1.35mmol), 2-dicyclohexylphosphonium-2 ',6' -diisopropoxy-1, 1' -biphenyl Ruphos (13mg,0.027mmol) and a tertiary palladium catalyst (Pd-Ruphos-G3) (23mg,0.027mmol), then heated to 90 degrees for reaction overnight. Cooling the reaction solution to room temperature, concentrating under reduced pressure, and purifying by column chromatography to obtain (15aR) -9-chloro-7-fluoro-8- (2-fluoro-6-hydroxyphenyl) -12,13,15,15 a-tetrahydro-14H-pyrazine [1',2':4,5][1,2,4]Triazole [4',3':1,6]Pyrazine [2,3-c ]]Quinoline-14-carboxylic acid tert-butyl ester twist isomers 1(22mg, yellow solid) and 2(15mg, yellow solid). Isomer 1: LC-MS ESI [ M + H ]]⁺＝495.1，RT：6.369min；¹H NMR (400MHz, CD3OD) < delta > 9.48(s,1H),9.36(s,1H),8.30(s,1H),7.23-7.37(m,2H), 6.79-6.81(m,2H),6.72-6.76(m,1H),6.25-6.29(m,1H),5.78-5.87(m,1H),5.11-5.14(m,1H),4.71-4.82(m,1H),4.13-4.17(m,1H),3.42-3.47(m,2H),2.67-2.71(m, 1H); isomer 2: LC-MS ESI [ M + H ]]⁺＝495.1,RT:6.564min；¹H NMR(400MHz,CD3OD):δ9.48(s,1H),9.36(s,1H),8.30(s,1H),7.22-7.37(m,2H),6.72-6.81(m,2H),6.25-6.29(m,1H),5.77-5.88(m,1H),5.11-5.15(m,1H),4.72-4.82(m,1H),4.47-4.51(m,1H),4.14-4.17(m,1H),3.79-3.85(m,1H),3.39-3.47(m,2H),2.67-2.71(m,1H)。

Isomer analysis chromatographic conditions: sunfire C184.6 × 150mm 5um 13 min; mobile phase A0.1% formic acid-water) FA-H2O, and mobile phase B0.1% (formic acid-acetonitrile) FA-ACN.

Example 2: 1- ((15aR) -9-chloro-7-fluoro-8- (2-fluoro-6-hydroxyphenyl) -12,13,15,15 a-tetrahydro-14H-imidazo [1',2':1,6] pyrazin [1',2':4,5] pyrazin [2,3-c ] quinolin-14-yl) prop-2-en-1-one

The first step is as follows: the (R) -10-bromo-11-chloro-9-fluoro-5-thio-1, 2,4,4a,5, 6-hexahydro-3H-pyrazine [1',2':4,5]Pyrazine [2,3-c ]]Quinoline-3-Tert-butyl carboxylate (50mg,0.10mmol) was dissolved in tetrahydrofuran THF (1mL), aminoacetaldehyde dimethyl acetal (52mg,0.50mmol) was added, and the mixture was refluxed under nitrogen for 3 hours. Cooling the reaction solution to room temperature, and concentrating under reduced pressure to obtain (R, Z) -10-bromo-11-chloro-5- ((2, 2-dimethoxyethyl) imine) -9-fluoro-1, 2,4,4a,5, 6-hexahydro-3H-pyrazine [1',2':4,5]Pyrazine [2,3-c ]]Quinoline-3-carboxylic acid tert-butyl ester (45mg, crude, yellow solid) was used directly in the next reaction. LC-MS ESI [ M + H ]]⁺＝572.0/574.0。

The second step is that: mixing (R, Z) -10-bromo-11-chloro-5- ((2, 2-dimethoxyethyl) imine) -9-fluoro-1, 2,4,4a,5, 6-hexahydro-3H-pyrazine [1',2':4,5]Pyrazine [2,3-c ]]Quinoline-3-carboxylic acid tert-butyl ester (400mg) was dissolved in dioxane (10mL), and p-toluenesulfonic acid monohydrate (200mg) was added to stir overnight. Extracting with dichloromethane, washing the organic phase with saturated sodium bicarbonate water solution and distilled water, drying with anhydrous sodium sulfate, concentrating under reduced pressure, and purifying by column chromatography to obtain tert-butyl (R) -8-bromo-9-chloro-7-fluoro-12, 13,15,15 a-tetrahydro-14H-imidazo [1',2':1,6]Pyrazines [1',2':4,5]]Pyrazino [2,3-c ] s]Quinoline-14-carboxylic acid ester (180mg), LC-MS: ESI [ M + H]⁺＝500.9/503.0。¹H NMR(400MHz,CDCl3):δ9.02(s,1H),8.13(d,J＝2.0Hz,1H),7.58(d,J＝2.0Hz,1H),7.29(s,1H),5.12-5.15(m,1H),4.25(bs,1H),4.02-4.16(m,1H),3.65-3.68(m,1H),3.35-3.72(m,1H),3.09-3.12(m,1H),2.69-2.72(m,1H),1.46(s,9H)。

The third step to the fifth step: the product obtained in the above step was synthesized by the same operation as in the fifth to seventh steps of example 1 using as a starting material 1- ((15aR) -9-chloro-7-fluoro-8- (2-fluoro-6-hydroxyphenyl) -12,13,15,15 a-tetrahydro-14H-imidazo [1',2':1, 6)]Pyrazines [1',2':4,5]]Pyrazine [2,3-c ]]Quinolin-14-yl) prop-2-en-1-one. LC-MS ESI [ M + H ]]⁺＝494.1。¹H NMR(400MHz,CDCl3):δ9.32(s,1H),8.24(d,J＝2.0Hz,1H),7.56(d,J＝2.0Hz,1H),7.32(s,1H),6.51(m,1H),6.23(m,1H),5.81(m,1H),5.16-5.18(m,1H),4.27(bs,1H),4.03-4.15(m,1H),3.62-3.66(m,1H),3.37-3.49(m,1H),3.06-3.15(m,1H),2.58-2.71(m,1H)。

The following compounds of examples were prepared and synthesized in the same manner as in example 1 and example 2.

Test example 1 KrasG12C functional analysis

All enzyme and substrate solutions were prepared with reaction buffer (20mM HEPES (pH7.5), 5mM MgCl2,150mM NaCl and 0.01% tween 20). The experimental procedure was as follows: using reaction buffer configuration of 10nM GDP loaded biotinylated KRASG12C and 37.5ng/ml streptavidin europium cryptate, 384 well HiBase micro polystyrene microwell plates were loaded with 5 μ L of the above protein reaction per well, along with test sample or control compound configured with DMSO and incubated for 4 hours. Separately, 20nM GST-Raf Ras binding domain (GST-Raf RBD) and 4. mu.g/ml anti-GST XL665 antibody (Cisbio) in reaction buffer (50mM potassium fluoride and 0.05mg/ml BSA) were mixed, and after 4 hours of equilibration, 0.6. mu.M GTP. gamma.S (Sigma) and 0.08. mu.M SOS were added. Mu.l of GST-RAF RBD mixture was added to each well of the plate. Addition of this mixture at this stage initiates a nucleotide exchange reaction which facilitates conversion of the non-activated GDP-loaded KRasG12C to activated GTP γ S KRasG 12C. The specific binding between activated GTP γ S KRasG12C and GST-RAF RBD draws the distance between europium and XL665 to enhance the FRET signal, which is detected using a Pherastar (BMG) plate reader equipped with an HTRF filter module. Any compound that inhibits nucleotide exchange or inhibits the binding of activated KRAS to RAF RBD will result in a decrease in FRET signal, and the FRET dose-response data is curve-fitted using Genedata Screener and the IC50 is calculated.

Test example 2 KRASG12C mass spectrometric addition analysis

All enzyme and substrate solutions were prepared with reaction buffer (20mM HEPES (pH7.5), 5mM MgCl2,150mM NaCl and 0.01% tween 20). mu.L of GDP-loaded biotinylated KRAS G12C (4. mu.M) and 1mM test compound (final concentration: 10. mu.M) in a reaction buffer (4. mu.M) were added to each well of a 96-well polypropylene microplate, and the reaction was terminated by adding 50. mu.L of 1% formic acid after 4 hours of reaction. The plates were sealed and then read by Xevo G2 QTOF (Waters) and Acquity LC system (Waters). 10 μ L of the sample was injected into Xbridge BEH 300; c4; 3.5 μm; gradient analysis was performed for 3min on 2.1X 50mm columns (Waters). A blank sample needs to be run between each test sample. Data analysis was performed using Mass Lynx Software (Waters) using total ion number (TIC) and combining the eluted protein peak data. Apo-protein KRASG12C (Apo) and KRAS + relative compound masses (sums) were tested and the percentage of sums calculated using the following formula: sum percentage is 100x (sum of sum peak area/APO and sum peak).

Test example 3: effect of the Compounds of the present invention on the proliferation potency of NCI-H358, MiaPaca-2 cells

1. NCI-H358 (lung cancer) and MiaPaca-2 (pancreatic cancer) cells (100. mu.L/well, 20000 cells/mL) were seeded in 96-well culture plates, respectively, and supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin sulfate. Cells were treated with a 0.5% dimethyl sulfoxide blank, diluted with an initial 10 μ M solution of test compound diluted three times with an eight gradient, and incubated in a 5% CO2 incubator for a period of time (3-7 days). At the end of the incubation, 10. mu.L of MTT stock solution (5mg/mL) was added to each well. The plates were incubated at 37 ℃ for 4 hours and then the medium was removed. Dimethylsulfoxide (100 μ L) was added to each well, followed by sufficient shaking. The absorbance of the formazan product was measured at 570nm on a thermoscientific varioska nflash multi-mode reader. IC was obtained by fitting dose response data to a three parameter nonlinear regression model using graphpadprism6.0 software₅₀The value is obtained.

2. As a result, the compounds of the examples provided in the present invention have proliferation inhibitory activity, IC, on NCI-H358 and MiaPaca-2 cells₅₀The values are all less than 5000nM, most less than 1000nM, and in particular the cell proliferation inhibitory activity of examples 5,6 is less than 100 nM.

Test example 4: examples in vivo pharmacokinetic parameter testing of Compounds in rats and mice

6 male SPF-grade SD rats (Shanghai Spill-Bikea laboratory animals) were divided into two groups, and the test compounds were formulated into appropriate solutions or suspensions; one group was administered intravenously and one group was administered orally. Blood is collected by jugular venipuncture, about 0.2 mL/time point of each sample is collected, heparin sodium is anticoagulated, and the blood collection time points are as follows: pre-and post-dose 5,15 and 30min, 1,2,4, 6, 8 and 24 h; blood samples were collected and placed on ice, plasma was centrifuged (centrifugation conditions: 8000 rpm, 6 min, 2-8 ℃) and collected plasma was stored at-80 ℃ before analysis. Plasma samples were analyzed by LC-MS/MS.

According to the data of the blood concentration of the drug, pharmacokinetic calculation software WinNonlin5.2 non-atrioventricular model is used for respectively calculating the pharmacokinetic parameters AUC of the test sample_0-t、AUC_0-∞、MRT_0-∞、C_max、T_max、T_1/2And V_dIsoparametric and their mean and standard deviation. In addition, the bioavailability (F) will be calculated by the following formula.

For samples with concentrations below the lower limit of quantitation, when pharmacokinetic parameter calculations are performed, C is reached_maxThe previously sampled samples should be calculated to zero when C is reached_maxSamples from later sampling points should be calculated as not quantifiable (BLQ).

Test example 5: EXAMPLES test of the Compounds for growth inhibition of MiaPaca-2, NCI-H358 tumor cells in nude mice transplanted tumors

Cutting tumor tissue in vigorous growth stage into 1.5mm³And left and right, under aseptic conditions, inoculated subcutaneously in the right axilla of nude mice. Measuring the diameter of the transplanted tumor by using a vernier caliper in the nude mouse subcutaneous transplanted tumor until the average tumor volume reaches 130mm³Animals were randomized into groups. The compound of the example (prepared to the required concentration with water for injection containing 1% Tween 80) was administered orally at the given dose daily for three weeks with the solvent control group given an equal amount of solvent. Throughout the experiment, the diameter of the transplanted tumor was measured 2 times per week, while the body weight of the mice was weighed. The formula for Tumor Volume (TV) is: TV (television)＝1/2×a×b²Wherein a and b represent length and width, respectively. Calculating Relative Tumor Volume (RTV) according to the measurement result, wherein the calculation formula is as follows: RTV is Vt/V0. Where V0 is the tumor volume measured at the time of caged administration (i.e., d0) and Vt is the tumor volume at each measurement. The evaluation index of the antitumor activity is 1) the relative tumor proliferation rate T/C (%), and the calculation formula is as follows:

T/C (%) (TRTV/CRTV) × 100%, TRTV: treatment group RTV; CRTV: negative control group RTV; 2) the tumor volume increase inhibition rate GI% is calculated according to the following formula: GI% ([ 1- (TVt-TV0)/(CVt-CT0) ] × 100%, TVt is the tumor volume per measurement in the treatment group; TV0 is the tumor volume obtained when therapeutic components were administered in cages; CVt is the tumor volume measured in each time in the control group; CV0 is the tumor volume obtained when the control component was administered in cages; 3) the tumor weight inhibition rate is calculated according to the following formula: tumor weight inhibition ratio (% Wc-WT)/Wc × 100%, Wc: tumor weight of control group, WT: the treated group had heavy tumor.

All documents referred to herein are incorporated by reference into this application as if each were individually incorporated by reference. Furthermore, it should be understood that various changes and modifications of the present invention can be made by those skilled in the art after reading the above teachings of the present invention, and these equivalents also fall within the scope of the present invention as defined by the appended claims.

Claims

1. A nitrogen-containing fused ring compound shown as a general formula I, or pharmaceutically acceptable salt thereof, or enantiomer, diastereoisomer, tautomer, torsional isomer, solvate, polymorph or prodrug thereof,

in the formula:

w, W1, W2, M is independently selected from CR4 or N, R4 is independently selected from H, halogen, cyano, C₁-C₆Alkyl radical, C₁-C₆Alkoxy, haloalkyl, haloalkoxy, alkenyl, alkynyl, alkylether, cycloalkylether, heterocycloalkylether, alkylether, cycloalkylthioether, heterocycloalkylether, 3-to 8-membered cycloalkyl or heterocycloalkyl, alkylamino-substituted alkoxy, alkylamino-substituted alkylamino, cycloalkylalkylene ether, heterocycloalkylalkylene ether, cycloalkylalkylene amino, heterocycloalkylalkylene amino, and the like;

one or more hydrogen atoms on any of the above groups may be substituted with a substituent selected from the group consisting of: including but not limited to deuterium, halogen, C1-C8 alkyl; wherein said heteroaryl group contains 1 to 3 heteroatoms selected from the group consisting of: n, O, P or S, the heterocycloalkyl group containing 1 to 3 heteroatoms selected from the group consisting of: n, O, P or S, said ring system including spiro, bridged, fused, etc. saturated or partially unsaturated ring systems.

2. The compound of claim 1, which is preferably a compound of formula (IIA), (IIB), (IIC), (IID), (IIE), or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, torsional isomer, solvate, polymorph or prodrug thereof:

wherein n is 1-5, R5 is independently selected from hydrogen, halogen, C1-C6 alkyl, alkoxy, haloalkyl, haloalkoxy, amino, substituted amino, cyano, and the like; r1, R2, R3, Ra, Rb, Rc, Rd, Re, Rf, Rg, W1, W2, M, Ar are as defined in claim 1.

3. The compound of claims 1,2, or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, torsiomer, solvate, polymorph, or prodrug thereof, wherein:

r1 is preferably selected from hydrogen, fluoro, methyl, cyano, etc.;

ar is independently preferably a monocyclic aromatic group such as a substituted or unsubstituted phenyl group or pyridyl group, or a substituted or unsubstituted bicyclic aromatic group such as a naphthyl group, a naphthyridinyl group, an indazolyl group or a benzimidazolyl group; said one or more substituents are preferably selected from the group consisting of: hydrogen, halogen, C1-C4 alkyl, hydroxy, amino, cyano, C1-C4 alkoxy, C1-C4 haloalkyl, C1-C4 haloalkoxy, and the like.

4. The compound of claims 1,2,3, or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, torsiomer, solvate, polymorph, or prodrug thereof, wherein the compound has the structure:

5. use of a compound of formula I according to claims 1,2,3,4 or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, torsiomer, solvate, polymorph or prodrug thereof, for the preparation of a medicament for the treatment of diseases associated with mutations in the Ras protein, in particular for the treatment of tumors. The tumor is independently selected from lung cancer, pancreatic cancer, liver cancer, colorectal cancer, bile duct cancer, brain cancer, leukemia, lymph cancer, melanoma, thyroid cancer, nasopharyngeal carcinoma, etc.

6. A pharmaceutical composition comprising a compound of formula I as defined in any one of claims 1,2,3,4, 5 or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, torsionamer, solvate, polymorph or prodrug thereof, wherein the pharmaceutical composition comprises:

(i) an effective amount of a compound of formula I, or a pharmaceutically acceptable salt thereof, or an enantiomer, diastereomer, tautomer, solvate, polymorph, or prodrug thereof; and

(ii) a pharmaceutically acceptable carrier.