CN120136869A

CN120136869A - Tricyclic fused heterocyclic PDE3/4 dual inhibitors and preparation method and application thereof

Info

Publication number: CN120136869A
Application number: CN202311717507.4A
Authority: CN
Inventors: 贾振华; 王宏涛; 侯云龙; 齐慧; 秘尧; 郝媛媛; 童亚男; 李发光; 张朋飞; 郝梦安; 王晓航; 许铎
Original assignee: Shijiazhuang Yiling Pharmaceutical Co Ltd
Current assignee: Shijiazhuang Yiling Pharmaceutical Co Ltd
Priority date: 2023-12-13
Filing date: 2023-12-13
Publication date: 2025-06-13
Also published as: WO2025124460A1

Abstract

The invention provides a tricyclic fused heterocyclic PDE3/4 dual inhibitor and a preparation method and application thereof, in particular to a compound with a structural formula I or a pharmaceutically acceptable form thereof, wherein each substituent is defined as an instruction book respectively. The compound or the pharmaceutically acceptable form thereof can be used as a phosphodiesterase inhibitor and has higher activity.

Description

Tricyclic fused heterocyclic PDE3/4 dual inhibitor and preparation method and application thereof

Technical Field

The invention relates to a tricyclic fused heterocyclic phosphodiesterase inhibitor (especially PDE3/4 dual inhibitor) and a preparation method and application thereof.

Background

Phosphodiesterase (PDE) belongs to a superfamily of enzymes, comprising at least 11 families, 22 subtypes, involved in intracellular and extracellular information transfer and functional regulation. PDE catalyzes the hydrolytic opening of the intracellular second messengers cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), producing AMP and GMP, respectively.

The PDE3 family consists of 2 genes, PDE3A and PDE3B. PDE3 activity is mainly concentrated in alveolar macrophages, endothelial cells, platelets in the respiratory system. PDE3 is involved in the regulation of various physiological activities in the body, such as vasodilation smooth muscle, platelet aggregation resistance, thrombosis resistance, heart strengthening, cell proliferation resistance and the like, and excessive use of PDE3 inhibitors can cause adverse reactions such as hypotension, tachycardia and the like, thus greatly limiting the clinical application thereof.

PDE4 is an enzyme that specifically hydrolyzes cAMP, and its family consists of four subtypes PDE4A, PDE4B, PDE C and PDE4D, each of which has a corresponding gene coding and has a different cellular distribution and action, respectively. PDE4 is predominantly distributed in airway smooth muscle cells, inflammatory cells and immune cells, regulating intracellular cAMP levels. Currently, most of the clinical PDE4 inhibitors have a certain degree of side effects, such as nausea, vomiting and other gastrointestinal reactions, and even depression.

Given the limitations of the use of PDE3 or PDE4 inhibitors alone and the side effects of PDE inhibitors, dual inhibition of inhaled PDE3/4 appears to be a more attractive way to target key pathological features in the treatment of COPD and asthma. There is evidence that dual target PDE3/4 inhibitors administered by inhalation have synergistic inhibitory effects, including synergistic anti-inflammatory and bronchodilatory effects.

CN100415743C discloses a pyrimido [6,1a ] isoquinolin-4-one derivative:

The compounds of the general formula are useful as PDE inhibitors for the treatment of respiratory diseases such as asthma, have a longer duration of action than that of trequindox, and do not have the very bitter taste of trequindox.

CN112368281A discloses tricyclic compounds as PDE3/PDE4 dual inhibitors

The compounds of the general formula are useful in the preparation of medicaments for PDE3/PDE4 related diseases, especially in the treatment of e.g. Chronic Obstructive Pulmonary Disease (COPD).

There remains a need in the art for new PDE3/PDE4 inhibitors, in particular PDE3/PDE4 inhibitors with high activity and good potency.

Disclosure of Invention

It is an object of the present invention to provide novel compounds as PDE inhibitors.

It is another object of the present invention to provide a process for the preparation of said compounds.

It is a further object of the present invention to provide the use of said compounds.

It is a further object of the present invention to provide pharmaceutical compositions comprising said compounds and uses thereof.

It is another object of the present invention to provide intermediates for the preparation of said compounds.

It is another object of the present invention to provide a process for the preparation of said intermediates.

< First aspect >

The present invention provides a compound of formula I or a pharmaceutically acceptable form thereof,

Wherein:

Each R ₁、R₂ is independently selected from the group consisting of H, C _1-6 straight chain alkyl, C _3-6 branched chain alkyl, and C _3-6 cycloalkyl, the straight chain alkyl, branched chain alkyl, or cycloalkyl optionally further substituted with 0 to 4 substituents selected from D, F, cl, br, I, OH, = O, NH ₂、CN、COOH、C_1-4 alkyl, C _1-4 alkoxy;

Each R ₃、R₄、R₅ is independently selected from H, halogen, CN, C _1-6 alkoxy, C _1-6 straight chain alkyl, C _3-6 branched alkyl, and C _3-6 cycloalkyl, the alkoxy, straight chain alkyl, branched alkyl, or cycloalkyl optionally being further substituted with 0 to 4 substituents selected from D, F, cl, br, I, OH, = O, NH ₂、CN、COOH、C_1-4 alkyl, C _1-4 alkoxy;

L is selected from And n is 0, 1 or 2;k is 0, 1,2 or 3, H in L is optionally further substituted with 0 to 4 substituents selected from F, cl, br, I, OH, = O, NH ₂、CN、COOH、C_1-4 alkyl, C _1-4 alkoxy, wherein R ₉、R₁₀ is each independently selected from H, C _1-6 straight chain alkyl, C _3-6 branched alkyl, C _3-6 cycloalkyl, C _3-6 heterocycloalkyl, C _6-10 aryl, C _5-10 heteroaryl and COOCH ₃, and R ₉、R₁₀ is not simultaneously H, the heteroaryl contains 1 to 3 heteroatoms selected from N, O and S;

R ₆ is selected from Wherein R ₁₁ is selected from amino, C _1-6 alkoxy, C _3-12 cycloalkyl, C _6-10 aryl, C _5-10 heterocyclyl containing 1-3 heteroatoms selected from N, O and S, said amino, alkoxy, cycloalkyl, aryl, heteroaryl optionally being further substituted with 0 to 4 substituents selected from H, F, cl, br, I, OH, = O, NH ₂、CN、COOH、CF₃、C_1-4 alkyl, C _1-4 alkoxy, C _3-6 cycloalkyl;

Optionally, R ₆ forms with L

R ₇、R₈ are each independently selected from H, =o, halogen, NH ₂、CN、C_1-6 straight chain alkyl, C _3-6 branched alkyl, C _3-6 cycloalkyl, and

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, each R ₁、R₂ is independently selected from CH ₃、CHF₂、CD₃ or C _3-6 cycloalkyl.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, R ₁、R₂ is CD ₃, respectively.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, one of R ₁、R₂ is CH ₃ and the other is CHF ₂.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, each R ₃、R₄、R₅ is independently selected from CH ₃、i-Pr、OMe、CD₃ or halogen.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, R ₁、R₂ corresponds to a group or value shown in any one of the compounds of table 1, respectively.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, R ₅ is CH ₃, said CH ₃ is optionally further substituted with 0 to 3D, R ₃、R₄ is the same and is selected from CH ₃, I-Pr, OMe or halogen.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, R ₃、R₄、R₅ corresponds to a group or value shown in any one of the compounds of table 1, respectively.

According to some embodiments of the invention, in the compounds of formula I of the invention or a pharmaceutically acceptable form thereof, L isN is 0 or 1, preferably 0. Preferably, k is 0, 1 or 2. Preferably, one of R ₉、R₁₀ is H and the other is selected from H, C _1-6 straight chain alkyl, C _3-6 branched alkyl, C _3-6 cycloalkyl, C _3-6 heterocycloalkyl, C _6-10 aryl, C _5-10 heteroaryl containing 1 or 2 heteroatoms N, and COOCH ₃, H in L being optionally further substituted with 0 to 4 substituents selected from F, cl, br, I, OH, = O, NH ₂、CN、COOH、C_1-4 alkyl, C _1-4 alkoxy.

According to some embodiments of the invention, in the compounds of formula I of the invention or a pharmaceutically acceptable form thereof, L is

According to some embodiments of the invention, in the compounds of formula I of the invention or a pharmaceutically acceptable form thereof, L isN is 0 or 1, and k is 1.

According to some embodiments of the invention, in the compounds of formula I of the invention or a pharmaceutically acceptable form thereof, L isN is 2 and k is 2.

According to some embodiments of the invention, in the compounds of formula I of the invention or a pharmaceutically acceptable form thereof, L isN is 1, k is 1 or 2.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, L corresponds to a group or value as shown in any one of the compounds of table 1.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, R ₆ isWherein R ₁₁ is amino optionally further substituted with 0,1 or 2 substituents selected from F, cl, br, I, OH, = O, NH ₂、CN、COOH、CF₃、C_1-4 alkyl, C _1-4 alkoxy, C _3-6 cycloalkyl.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, R ₆ isWherein R ₁₁ is C _6-10 aryl or C _5-10 heterocyclyl, said heterocyclyl containing 1,2 or 3 heteroatoms N, said aryl, heteroaryl optionally being further substituted with 0, 1,2 or 3 substituents selected from H, F, cl, br, I, OH, = O, NH ₂、CN、COOH、CF₃、C_1-4 alkyl, C _1-4 alkoxy, C _3-6 cycloalkyl.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, R ₆ corresponds to a group or value shown for any of the compounds in table 1.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, R ₇ is H or CH ₃,R₈ is H or F.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, R ₇、R₈ corresponds to a group or value shown for any of the compounds in table 1.

According to some embodiments of the invention, in the compounds of formula I of the invention or pharmaceutically acceptable forms thereof, R ₁、R₂、R₃、R₄、R₅、R₆、R₇、R₈, L correspond to the groups or values shown in any one of the compounds of table 1, respectively.

According to some embodiments of the invention, in a compound of formula I of the invention or a pharmaceutically acceptable form thereof, the compound is selected from one or more of the compounds shown in table 1:

TABLE 1

According to some embodiments of the invention, the compound of formula I of the invention or a pharmaceutically acceptable form thereof is selected from pharmaceutically acceptable salts or co-crystals, stereoisomers, tautomers, deuterides, solvates, chelates, non-covalent complexes or prodrugs.

< Second aspect >

The invention also provides an intermediate compound which has a structure shown in a formula II:

Wherein R ₁、R₂、R₃、R₄、R₅、R₇、R₈ is as defined according to any one of the schemes of the < first aspect > compound of formula I or a pharmaceutically acceptable form thereof;

L ₁ is selected from R ₉、R₁₀, n, k are as defined according to any one of the schemes of the invention for compounds of formula I or a pharmaceutically acceptable form thereof according to the < first aspect >, R ₁₂、R₁₃ are each independently H, boc, cbz, SEM, fmoc, alloc, pht, OTs, PMB, bn, trt.

According to some embodiments of the invention, the intermediate compounds of the invention have the following structure:

One of R ₁₂、R₁₃ is H and the other is Boc, cbz, SEM, fmoc, alloc, pht, OTs, PMB, bn, trt.

< Third aspect >

The present invention also provides a process for preparing a compound according to the < first aspect > of the invention or a pharmaceutically acceptable form thereof. Synthetic routes may be designed to prepare the compounds of formula I or pharmaceutically acceptable forms thereof according to the chemical structure of the compounds of formula I or pharmaceutically acceptable forms thereof, with reference to methods known in the art.

According to some embodiments of the invention, the method of preparing a compound according to the invention < first aspect > or a pharmaceutically acceptable form thereof, according to the invention, comprises:

the intermediate compound of the < second aspect > of the present invention is subjected to a modification reaction at the carboxy terminus shown in formula II to prepare a compound having structural formula I.

According to some embodiments of the invention, the method of preparing a compound according to the invention < first aspect > or a pharmaceutically acceptable form thereof according to the invention further comprises a process of preparing an intermediate compound according to the invention < second aspect >.

According to some embodiments of the invention, the method of preparing a compound according to the invention < first aspect > or a pharmaceutically acceptable form thereof according to the invention comprises the steps as shown in any of the reaction schemes of examples 1-38.

< fourth aspect >

The present invention also provides a pharmaceutical composition comprising a compound of the < first aspect > of the invention or a pharmaceutically acceptable form thereof (preferably a pharmaceutically acceptable salt), together with a pharmaceutically acceptable carrier, excipient and/or one or more other therapeutic agents.

< Fifth aspect >

The invention also provides the use of a compound according to the < first aspect > of the invention or a pharmaceutically acceptable form thereof (preferably a pharmaceutically acceptable salt) or a pharmaceutical composition according to the < fourth aspect > of the invention in the manufacture of a formulation for inhibiting phosphodiesterase. Preferably, the phosphodiesterase comprises PDE3 and/or PDE4.

< Sixth aspect >

The invention also provides the use of a compound according to the < first aspect > of the invention or a pharmaceutically acceptable form thereof (preferably a pharmaceutically acceptable salt) or a pharmaceutical composition according to the < fourth aspect > of the invention in the manufacture of a medicament for the treatment of phosphodiesterase-related diseases.

The present invention also provides a method of treating a phosphodiesterase-related disorder comprising administering to a subject an effective amount of a compound according to the < first aspect > of the invention or a pharmaceutically acceptable form thereof (preferably a pharmaceutically acceptable salt) or a pharmaceutical composition according to the < fourth aspect > of the invention.

According to some embodiments of the invention, the phosphodiesterase comprises PDE3 and/or PDE4.

According to some embodiments of the invention, the phosphodiesterase-related disease comprises a respiratory disease such as asthma.

According to some embodiments of the invention, the subject is a mammal or a human, preferably the subject is a human.

The compound with the structural formula I or a pharmaceutically acceptable form thereof can be used as a phosphodiesterase inhibitor, has high-efficiency inhibitory activity on phosphodiesterase, especially PDE3 and/or PDE4, and has practical value.

Definition and description

The following terms and phrases used herein are intended to have the following meanings unless otherwise indicated. A particular term or phrase, unless otherwise specifically defined, should not be construed as being ambiguous or otherwise clear, but rather should be construed in a generic sense. When trade names are presented herein, it is intended to refer to their corresponding commercial products or active ingredients thereof.

Unless otherwise indicated, with solid wedge bondsAnd a wedge-shaped dotted bondRepresenting the absolute configuration of a solid centre by straight solid keysAnd straight dotted line keyRepresenting the relative configuration of the three-dimensional center by wavy linesSolid key representing wedge shapeOr wedge-shaped dotted bondOr by wave linesRepresenting straight solid keysAnd straight dotted line key

The term "pharmaceutically acceptable" is intended to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.

The term "pharmaceutically acceptable salt" or "pharmaceutically acceptable salt thereof" refers to salts of the compounds of the present invention, prepared from the compounds of the present invention which have a particular substituent found with a relatively non-toxic acid or base. When the compounds of the present invention contain relatively acidic functional groups, base addition salts may be obtained by contacting neutral forms of such compounds with a sufficient amount of a base in pure solution or in a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic ammonia or magnesium salts or similar salts. When the compounds of the present invention contain relatively basic functional groups, the acid addition salts may be obtained by contacting the neutral form of such compounds with a sufficient amount of an acid in pure solution or in a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include inorganic acid salts including, for example, hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, bicarbonate, phosphoric acid, phosphate-bicarbonate, phosphate-dihydrogen, sulfuric acid, bisulfate, hydroiodic acid, phosphorous acid, and the like, and organic acid salts including, for example, acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid, methanesulfonic acid, and the like, and salts of amino acids (e.g., arginine, and the like), and salts of organic acids such as glucuronic acid, and the like. Certain specific compounds of the invention contain basic and acidic functionalities that can be converted to either base or acid addition salts.

Pharmaceutically acceptable salts of the invention can be synthesized from the parent compound containing an acid or base by conventional chemical methods. Typically, such salts are prepared by reacting these compounds in free acid or base form with a stoichiometric amount of the appropriate base or acid in water or an organic solvent or a mixture of both.

The term "eutectic" means that the crystalline material comprises two or more distinct solids at room temperature, each containing different physical properties, such as structure, melting point, and heat of fusion.

The term "stereoisomer" (or "optical isomer") refers to a stable isomer that has a perpendicular plane of asymmetry due to at least one chiral factor (including chiral center, chiral axis, chiral plane, etc.), thereby enabling rotation of plane polarized light. The present invention also includes stereoisomers and mixtures thereof, due to the presence of asymmetric centers and other chemical structures in the compounds of the present invention which may lead to stereoisomers. Since the compounds of the present invention and salts thereof may include asymmetric carbon atoms, they can exist as single stereoisomers, racemates, mixtures of enantiomers and diastereomers. Typically, these compounds can be prepared in the form of a racemic mixture. However, if desired, such compounds can be prepared or isolated to give pure stereoisomers, i.e., single enantiomers or diastereomers, or mixtures enriched in single stereoisomers (purity. Gtoreq.98%, purity. Gtoreq.95%,. Gtoreq.93%,. Gtoreq.90%,. Gtoreq.88%,. Gtoreq.85% or. Gtoreq.80%). The individual stereoisomers of the compounds are prepared synthetically from optically active starting materials containing the desired chiral centers or by preparation of mixtures of enantiomeric products followed by separation or resolution, e.g., conversion to mixtures of diastereomers followed by separation or recrystallization, chromatography, use of chiral resolving agents, or direct separation of the enantiomers on chiral chromatographic columns. Starting compounds having specific stereochemistry are either commercially available or can be prepared according to the methods described herein and resolved by methods well known in the art. All stereoisomeric forms of the compounds of the invention are within the scope of the compounds of the invention unless otherwise indicated.

The term "tautomer" (or "tautomeric form") refers to structural isomers having different energies that can be converted to each other by a low energy barrier. If tautomerism is possible (e.g., in solution), chemical equilibrium of the tautomers can be achieved. For example, proton tautomers (or proton transfer tautomers) include, but are not limited to, interconversions by proton transfer, such as keto-enol isomerisation, imine-enamine isomerisation, amide-imine alcohol isomerisation, and the like. Unless otherwise indicated, all tautomeric forms of the compounds of the invention are within the scope of the compounds of the invention.

Unless otherwise indicated, compounds represented by the structural formula of the present invention may be in the form of a single stereoisomer or tautomer that is purified, or may be in the form of a mixture comprising a plurality of stereoisomers or tautomers.

The term "solvate" refers to a substance formed by the association of a compound of the invention, or a pharmaceutically acceptable salt thereof, with at least one solvent molecule by non-covalent intermolecular forces. Common solvates include, but are not limited to, hydrates, ethanolates, acetonates, and the like.

The term "chelate" is a complex having a cyclic structure, obtained by chelation of two or more ligands with the same metal ion to form a chelate ring.

The term "non-covalent complex" is formed by the interaction of a compound with another molecule, wherein no covalent bond is formed between the compound and the molecule. For example, recombination can occur by van der Waals interactions, hydrogen bonding, and electrostatic interactions (also known as ionic bonding).

The term "prodrug" refers to a derivative compound that is capable of providing a compound of the invention directly or indirectly after administration to a patient. Particularly preferred derivative compounds or prodrugs are compounds that, when administered to a patient, may increase the bioavailability of the compounds of the invention (e.g., are more readily absorbed into the blood) or promote delivery of the parent compound to the site of action (e.g., the lymphatic system). All prodrug forms of the compounds of the invention are within the scope of the invention unless otherwise indicated, and the various prodrug forms are well known in the art.

The term "independently" means that at least two groups (or ring systems) present in the structure that are the same or similar in value range may have the same or different meanings in the particular case. For example, the substituents X and Y are each independently hydrogen, halogen, hydroxy, cyano, alkyl or aryl, and when the substituent X is hydrogen, the substituent Y may be either hydrogen or halogen, hydroxy, cyano, alkyl or aryl, and similarly when the substituent Y is hydrogen, the substituent X may be either hydrogen or halogen, hydroxy, cyano, alkyl or aryl.

The terms "comprising" and "including" are used in their open, non-limiting sense.

"Optional" or "optionally" means that the subsequently described event or circumstance may but need not occur, and that the description includes instances where said event or circumstance occurs and instances where it does not.

The term "substituted" means that any one or more hydrogen atoms on a particular atom is substituted with a substituent, and may include heavy hydrogens and variants of hydrogens, provided that the valence of the particular atom is normal and the substituted compound is stable. When the substituent is oxygen (i.e., =o), it means that two hydrogen atoms are substituted. Oxygen substitution does not occur on the aromatic group. The term "optionally substituted" means that the substituents may or may not be substituted, and the types and numbers of substituents may be arbitrary on the basis that they can be chemically achieved unless otherwise specified.

When any variable (e.g., R) occurs more than once in the composition or structure of a compound, its definition in each case is independent. Thus, for example, if a group is substituted with 0 to 2R, the group may optionally be substituted with up to two R's, and R's in each case have independent options. Furthermore, combinations of substituents and/or variants thereof are only permissible if such combinations result in stable compounds.

When a variable in a formula is selected from the group consisting of a deletion, it is indicated as absent, e.g., when R in C-R is selected from the group consisting of a deletion, it is indicated that the structure is in fact C.

When one variable in the formula linking two groups is selected from a bond or is absent, it means that the two groups to which it is linked are directly linked, e.g., L in A-L-Z represents a bond or is absent, it means that the structure is actually A-Z.

When the recited substituents do not indicate which atom is attached to the substituted group, such substituents may be bonded through any atom thereof, for example, a phenyl group may be attached as a substituent to the substituted group through any carbon atom on the benzene ring.

Unless otherwise specified, the term "alkyl" is used to denote a straight or branched saturated hydrocarbon group, which may be mono-substituted (e.g., -CH ₂ F) or poly-substituted (e.g., -CF ₃), and may be monovalent (e.g., methyl), divalent (e.g., methylene), or multivalent (e.g., methine). Examples of alkyl groups include methyl (Me), ethyl (Et), propyl (e.g., n-propyl and isopropyl), butyl (e.g., n-butyl, isobutyl, s-butyl, t-butyl), pentyl (e.g., n-pentyl, isopentyl, neopentyl), and the like.

Unless otherwise specified, the term "alkylene" refers to a divalent straight or branched chain alkane group consisting of only carbon and hydrogen atoms, free of saturation, and linked to other fragments by two single bonds, respectively, including, but not limited to, methylene, 1-ethylene, 1, 2-ethylene, and the like. For example, "C _1-3 alkylene" refers to a saturated divalent straight or branched chain alkyl group containing from 1 to 3 carbon atoms.

Unless otherwise specified, "cycloalkyl" includes any stable cyclic or polycyclic hydrocarbon group, any carbon atom being saturated, either mono-or polysubstituted, and either monovalent, divalent or multivalent. Examples of such cycloalkyl groups include, but are not limited to, cyclopropyl, norbornyl, [2.2.2] bicyclooctane, [4.4.0] bicyclodecane, and the like.

Unless otherwise specified, the term "alkoxy" means an alkyl group attached to the remainder of the molecule through an oxygen atom, wherein the alkyl group has the meaning as described herein. Unless otherwise specified, C _1-5 alkoxy includes C1, C2, C3, C4, and C5 alkoxy. Examples of alkoxy groups include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, sec-butoxy, tert-butoxy, n-pentoxy and S-pentoxy. The alkoxy group may be optionally substituted with one or more substituents described herein.

The term "3-6 membered ring" means, unless otherwise specified, a saturated or unsaturated monocyclic ring, with or without heteroatoms, containing 3, 4, 5 or 6C, O, S, N atoms in the ring, which "3-6 membered ring" may be attached to the remainder of the formula by any one of the carbon atoms, or, if present, nitrogen atoms.

The term "amine (amino)" refers to-NH ₂, -NH (alkyl) or-N (alkyl) unless otherwise specified.

The term "aromatic ring" means a single ring of polyunsaturated aromatic alkanes, which may be mono-or polysubstituted, unless otherwise specified.

The term "4-6 membered heterocycloalkyl" means, unless otherwise specified, a saturated monovalent monocyclic hydrocarbon ring containing 3, 4 or 5 carbon atoms in the ring and one or more heteroatom groups selected from O, NR ^a, wherein R ^a represents a hydrogen atom or a C _1-6 alkyl group, and the "4-6 membered heterocycloalkyl" may be attached to the remainder of the molecule through any one of the carbon atoms, or, if present, a nitrogen atom.

Unless otherwise specified, the term "heteroaryl" refers to an aromatic ring containing one to four heteroatoms selected from one or more of N, O and S.

Unless otherwise specified, the term "heterocyclyl" refers to a saturated or partially saturated, monocyclic or polycyclic (such as bicyclic, for example: and ring, bridged ring, or spiro ring) whose ring atoms are made up of carbon atoms and at least one heteroatom selected from N, O and S, wherein the S atoms are optionally substituted to form S (=o), S (=o) ₂, or S (=o) (=nr ^x),R^x is independently selected from H or C _1-4 alkyl. For example, the heterocyclic group may be an oxiranyl group, an aziridinyl group, an azetidinyl group, an oxetanyl group, a tetrahydrofuranyl group, a dioxolyl group, a pyrrolidinyl group, a pyrrolidonyl group, an imidazolidinyl group, a pyrazolidinyl group, a tetrahydropyranyl group, a piperidinyl group, a piperazinyl group, a morpholinyl group, a thiomorpholinyl group, a dithianyl group, or a trithianyl group.

Unless otherwise specified, the term "aryl" refers to a monocyclic or fused polycyclic aromatic hydrocarbon group having a conjugated pi-electron system. For example, the term "C _6-10 aryl" as used herein refers to an aryl group having 6 to 10 carbon atoms. For example, aryl may be phenyl, naphthyl, anthracenyl, phenanthrenyl, acenaphthylenyl, azulenyl, fluorenyl, indenyl, pyrenyl, and the like.

Unless otherwise specified, the term "heteroaryl" refers to a monocyclic or fused polycyclic aromatic group having a conjugated pi-electron system, the ring atoms of which are made up of carbon atoms and at least one heteroatom selected from N, O and S. If valence requirements are met, the heteroaryl group may be attached to the remainder of the molecule through any one of the ring atoms. For example, the term "5-10 membered heteroaryl" as used in the present invention refers to heteroaryl groups having 5 to 10 ring atoms. For example, heteroaryl groups can be thienyl, furyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, triazolyl, thiadiazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl and its benzo derivatives, pyrrolopyridinyl, pyrrolopyrazinyl, pyrazolopyridinyl, imidazopyridinyl, pyrrolopyrimidinyl, pyrazolopyrimidinyl, purinyl, and the like.

Unless otherwise specified, the term "halogen" refers to fluorine (F), chlorine (Cl), bromine (Br) and iodine (I). The term "hydroxy" refers to-OH. The term "cyano" refers to-CN. The term "amino" refers to-NH ₂.

The term "pharmaceutical composition" refers to a mixture of one or more compounds or pharmaceutically acceptable forms of the present invention with other chemical components, wherein "other chemical components" refers to pharmaceutically acceptable carriers, excipients, and/or one or more other therapeutic agents, unless otherwise specified. By "carrier" is meant a material that does not cause significant irritation to the organism and does not abrogate the biological activity and properties of the administered compound. "excipient" refers to an inert substance that is added to a pharmaceutical composition to facilitate administration of a compound. Non-limiting examples include calcium carbonate, calcium phosphate, sugars, starches, cellulose derivatives (including microcrystalline cellulose), gelatin, vegetable oils, polyethylene glycols, diluents, granulating agents, lubricants, binders, and disintegrating agents.

The compounds of the present invention have good PDE inhibitory activity and have potential application value in the treatment of PDE-related diseases, in particular PDE3/PDE 4-related diseases.

Drawings

FIG. 1 is a single crystal diffraction pattern of compound 1-2 of the present invention.

Detailed Description

Embodiments of the present invention will be described in detail below with reference to examples, but it will be understood by those skilled in the art that the following examples are only for illustrating the present invention and should not be construed as limiting the scope of the present invention. The specific conditions are not noted in the examples and are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or apparatus used were conventional products commercially available without the manufacturer's attention.

The structure of the compound is determined by nuclear magnetic resonance or mass spectrometry. Nuclear magnetic resonance was measured using a BRUKER 400M nuclear magnetic instrument with the measurement solvent being deuterated dimethyl sulfoxide (DMSO-d ₆) or deuterated chloroform (CDCl ₃), the internal standard being Tetramethylsilane (TMS), the chemical shift (δ) being given in units of 10 ^-6 (ppm). Mass spectra were determined using Waters ACQUITY Arc/ACQUITY QDa or Thermo U3000-ISQ EC liquid chromatography.

High performance liquid chromatography analysis was performed using a Thermo U3000 high pressure liquid chromatograph. The high performance liquid phase preparation uses Hanbang DAC-50 or Shimadzu LC-20AP preparative chromatograph.

Reaction monitoring uses thin layer chromatography or liquid chromatography-mass spectrometry. The developing agent system used in the thin layer chromatography comprises a dichloromethane and methanol system, a petroleum ether and ethyl acetate system, and the volume ratio of the solvent is adjusted according to the polarity of the compound, or a small amount of triethylamine is added. The LC-MS uses Waters ACQUITY Arc/ACQUITY QDa or Thermo U3000-ISQ EC LC-MS.

Column chromatography generally uses 200-300 mesh silica gel as a carrier. The eluent system comprises a dichloromethane and methanol system, a petroleum ether and ethyl acetate system, and the volume ratio of the solvent is adjusted according to the polarity of the compound, or a small amount of triethylamine is added.

The following examples and comparative examples are not particularly described, the reaction temperature is room temperature (20 ℃ C. To 30 ℃ C.), and the solvents are dried and purified according to standard methods.

The comparative compounds are shown in Table 2.

Table 2, comparative example Compounds

Preparation of comparative example compound D1:

the preparation method comprises the following steps:

1.

To a solution of 2,4, 6-trimethylaniline (2.0 g,14.7mmol,1 eq.) and tert-butyl N- (3-bromopropyl) carbamate (14.1 g,59.1mmol,4 eq.) in DMF (20 mL,258.4 mmol) under nitrogen at room temperature was added K ₂CO₃ (6.1 g,44.3mmol,3 eq.) and KI (2.5 g,14.7mmol,1 eq.). The reaction was allowed to react overnight at 80 ℃. The reaction mixture was extracted with EA (3X 50 ml). The organic phases were combined, washed with saturated sodium chloride solution (2X 30 ml) and dried over anhydrous sodium sulfate. The mixture was filtered and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography, EA/PE (1/4), to give tert-butyl N- (3- [ (2, 4, 6-trimethylphenyl) amino ] propylcarbamate (2.2 g, yield 50.86%).

LCMS(ESI,m/z):[M+H]⁺=293.2

2.

To a solution of tert-butyl N- (3- [ (2, 4, 6-trimethylphenyl) amino ] propylcarbamate (1.2 g,4.1mmol,1 eq.) and 2-chloro-9, 10-dimethoxy-6, 7-dihydropyrimido [4,3-a ] isoquinolin-4-one (1.8 g,6.1mmol,1.5 eq.) in dioxane (20 mL), under nitrogen protection, cs ₂CO₃ (2.7 g,8.2mmol,2 eq.) and APhos Pd G ₃ (0.3 g,0.4mmol,0.1 eq.) were added, respectively, and 4- (di-tert-butylphosphono) -N, N-dimethylaniline (0.1 g,0.4mmol,0.1 eq.) were stirred at 100℃overnight, the reaction mixture was extracted with EA (3X 50 mL), the organic phase was combined, washed with saturated sodium chloride solution (2X 30 mL), and dried over anhydrous sodium sulfate after filtration, the filtrate was concentrated under reduced pressure, and the following a gradient of tert-butyl N, N-dimethylaniline (0.1 g,0.4mmol,0.1 eq.) was purified by a gradient of tert-butyl 4- (2-butylphosphono) and 4-N, N-dimethylaniline (0.1 mmol,0.1 eq.) was obtained, 100℃over a gradient of tert-butyl carbamate (10.10.10% aqueous chromatography, 10% to give tert-butylquinoline, 10-4-2-amino, 10% aqueous chromatography residue, 10% aqueous solution, 10% of three-phase chromatography, 200% aqueous phase, 200% chromatography, 4-phase, 3% aqueous phase, and 3% aqueous phase chromatography phase, 10% chromatography phase, and three-phase chromatography phase.

LCMS(ESI,m/z):[M+H]⁺=549.4

3.

To a solution of tert-butyl N- [3- ((9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl (2, 4, 6-trimethylphenyl) amino) propyl ] carbamate (290 mg,1 eq) in DCM (5 mL) under nitrogen protection at 0℃was added dropwise TFA (5 mL) -nitrogen protection at room temperature, the resulting residue was stirred for 2h, the reaction mixture was extracted with DCM (3X 20 mL) -the organic phases were combined, washed with saturated sodium chloride solution (2X 10 mL) and dried over anhydrous sodium sulfate, after filtration of the resulting mixture, the filtrate was concentrated under reduced pressure to give 2- [ (3-aminopropyl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (200 mg, crude product).

LCMS(ESI,m/z):[M+H]⁺=449.3

4.

To a solution of 2- [ (3-aminopropyl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6 h,7 h-pyrimido [4,3-a ] isoquinolin-4-one (200 mg,0.4mmol,1.0 eq.) and TEA (135 mg,1.3mmol,3 eq.) in DCM (5 mL) at 0 ℃ under nitrogen was added dropwise trimethylsilyl isocyanate (128.4 mg,1.1mmol,2.5 eq.). The resulting mixture was stirred under nitrogen at 50 ℃ for 2 days. The resulting residue was concentrated under reduced pressure. The crude product was purified by high performance liquid chromatography to give 3- ((9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl (2, 4, 6-trimethylphenyl) amino) propiurea (7.5 mg, yield 3.4%) under the following conditions (column: xbridge BEH Phenyl μm,19 x 250mm; mobile phase A: water (0.1% FA), mobile phase B: meOH; flow: 25mL/min, gradient: 43% B to 53% B; wavelength: 254nm/220nm; RT1 (min): 9.6)

LCMS(ESI,m/z):[M+H]⁺=492.25

¹H NMR(400MHz,Methanol-d₄)δ8.22(broad,1H),7.26(s,1H),7.01(s,2H),6.94(s,1H),6.24(s,1H),4.05(t,J＝6.4Hz,2H),3.91(s,6H),3.61(t,J＝7.8Hz,2H),3.45(t,J＝6.6Hz,2H),3.01–2.89(m,2H),2.29(s,3H),2.24(s,6H),1.88(s,2H).

Preparation of comparative example compound D2:

To a 50mL flask was added (2E) -9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) imino ] -3H,6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (1.0 g,2.55mmol,1.0 equiv.) and dichloromethane (20 mL) at room temperature, stirred, acetic anhydride (0.29 g,2.81mmol,1.1 equiv.) was added, and stirred under nitrogen at room temperature for 24h. After the reaction, the system was cooled to 0℃and 10ml of water was added dropwise, stirred for 1H, extracted three times with methylene chloride, the organic phases were combined, dried and distilled, and the compound N- (9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-2-yl) -N-m-methylacetamide (0.11 g) was obtained by purification on a silica gel column.

LCMS(ESI,m/z):[M+H]⁺=434.21

¹H NMR(400MHz,CDCl₃)δ7.72(s,1H),7.18(s,1H),6.94(s,2H),6.74(s,1H),4.19(t,J＝6.4Hz,2H),3.92(s,6H),2.95(t,J＝6.4Hz,2H),2.31(s,3H),2.12(s,6H),2.05(s,3H).

Preparation of comparative example compound D3:

(2E) -9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) imino ] -3H,6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (1.00 g,2.554mmol,1.0 eq) and 2-bromopropane (0.47g,3.831mmol,1.5eq)、Pd₂(dba)₃(0.23g,0.255mmol,0.1eq)、Cs₂CO₃(2.50g,7.662mmol,3.0eq) were dissolved in 1, 4-dioxane (20 ml) and reacted under nitrogen protection at stirring and elevated temperature to reflux for 12H, after the reaction was completed, water (50 ml) and ethyl acetate were added to extract (50 ml. Times.3), saturated sodium chloride (50 ml. Times.1) was washed, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 2- (isopropyl (methyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (7.8 g, 71.43%).

LCMS(ESI,m/z):[M+H]⁺=434.25

¹H NMR(400MHz,Methanol-d₄)δ7.08(s,2H),6.89(s,1H),6.58(s,1H),5.35(s,1H),4.82–4.74(m,1H),4.06(t,J＝6.4Hz,2H),3.86(s,3H),3.63(s,3H),2.93(t,J＝6.4Hz,2H),2.34(s,3H),2.14(s,3H),1.25(s,3H),1.23(s,3H).

Preparation of comparative example compound D4:

9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) amino ] -6h,7 h-pyrimidine [4,3-a ] isoquinoline-4-1 (200 mg,0.5mmol,1.0 eq.) is dissolved in solvent DMF (2 mL) and NaH (40 mg,1.0mmol,2.0 eq., 60% purity) is added at 0deg.C and the resulting mixture stirred under nitrogen for 0.5h. To the above mixture was added (2-bromoethyl) diethylamine hydrobromide (400 mg,1.5mmol,3.0 eq) and stirred at room temperature for 1h. The reaction was quenched by the addition of water (10 mL) at room temperature. The mixture was extracted with ethyl acetate (2X 50 mL), and the organic layer was washed with saturated brine (3X 100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. Column chromatography (EA/pe=1/2) purification gave 2- { [2- (diethylamino) ethyl ] (2, 4, 6-trimethylphenyl) amino } -9, 10-dimethoxy-6 h,7 h-pyrimidine [4,3-a ] isoquinoline-4-1 (37.2 mg, 14% yield).

LCMS(ESI):[M+H]⁺=491.25

¹H NMR(400MHz,Methanol-d₄)δ6.95–6.85(m,3H),6.77(s,1H),5.50(s,1H),4.44–4.35(m,2H),4.06–3.99(m,2H),3.87(s,3H),3.69(s,3H),2.99–2.90(m,4H),2.74(q,J＝7.2Hz,4H),2.28(s,3H),2.05(s,6H),1.16(t,J＝7.2Hz,6H).

Preparation of comparative example compound D5:

the preparation method comprises the following steps:

1.

9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) amino ] -6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (500 mg,1.2mmol,1.0 eq) and Cs ₂CO₃ (1248 mg,3.8mmol,3.0 eq) were added to a solution of DMF (10 mL) at room temperature under nitrogen, tert-butyl N- (2-bromoethyl) carbamate (715 mg,3.1mmol,2.5 eq) and Pd ₂(dba)₃ (116 mg,0.1mmol,0.1 eq) were added in portions. The resulting residue was stirred overnight at 100 degrees celsius under nitrogen. The reaction mixture was extracted with ethyl acetate (3X 50 mL). The combined organic phases were washed with saturated sodium chloride solution (2X 30 mL) and dried over anhydrous sodium sulfate. After the resulting mixture was filtered, the filtrate was concentrated under reduced pressure. The resulting residue was purified by column chromatography on silica gel, dichloromethane/methanol (20/1), to give tert-butyl N- [2- ((9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl (2, 4, 6-trimethylphenyl) amino) ethyl ] carbamate (200 mg, 29.29%).

LCMS(ESI):[M+H]⁺=535.2

2.

Tert-butyl N- [2- ((9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl (2, 4, 6-trimethylphenyl) amino) ethyl ] carbamate (180 mg,0.3mmol,1.0 eq.) is added to a solution of HCl in dioxane (4 mL, 4M) at room temperature under nitrogen, the reaction is stirred for 1h, the reaction mixture is basified with saturated aqueous sodium bicarbonate to pH=8, the reaction mixture is extracted with ethyl acetate (3X 20 mL.) the organic phases are combined, the solution is backwashed with saturated aqueous sodium chloride (2X 10 mL), dried over anhydrous sodium sulfate, the resulting mixture is filtered, and the filtrate is concentrated under reduced pressure to give 2- [ (2-aminoethyl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (160 mg, crude product).

LCMS(ESI):[M+H]⁺=435.2

3.

2- [ (2-Aminoethyl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (100 mg,0.2mmol,1.0 eq.) and TEA (69 mg,0.6mmol,3.0 eq.) were added dropwise to a solution of DCM (5 mL) at room temperature under nitrogen. The residue obtained was stirred at room temperature under nitrogen for 1.5h. The reaction mixture was extracted with dichloromethane (3X 20 mL). The combined organic phases were backwashed with saturated brine (2X 50 mL) and dried over anhydrous sodium sulfate. After the resulting mixture was filtered, the filtrate was concentrated under reduced pressure. The crude product was purified by high performance liquid phase to give 2- ((9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl (2, 4, 6-trimethylphenyl) amino) ethylurea (26.4 mg, 24.02%) under the following conditions (column: YMC TRIART C: exRs 5m,19mm x 250mm; mobile phase A: water (10 mmol/L NH ₄HCO₃), mobile phase B: acetonitrile; flow rate: 25mL/min; gradient: 32%B to 55%B in 10min; wavelength: 254nm/220nm; RT1 (min): 9.17).

LCMS(ESI):[M+H]⁺=478.20

¹H NMR(400MHz,Methanol-d₄)δ7.11(s,2H),6.94(s,1H),6.70(s,1H),5.51(s,1H),4.15–4.08(m,2H),3.99–3.92(m,2H),3.89(s,3H),3.67(s,3H),3.44(t,J＝6.4Hz,2H),2.97(t,J＝6.4Hz,2H),2.36(s,3H),2.18(s,6H).

Preparation of comparative example compound D6:

(2E) -9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) imino ] -3H,6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (500 mg,1.28mmol,1.0 eq) and 1-bromobutane (263mg,1.92mmol,1.5eq)、Pd₂(dba)₃(119mg,0.13mmol,0.1eq)、Cs₂CO₃(1.25g,3.84mmol,3.0eq)、Xantphos(150mg,26mmol,0.2eq) were dissolved in 1, 4-dioxane (10 ml) and stirred under nitrogen atmosphere until reflux was effected for 12H, water (50 ml) and ethyl acetate were added to extract (50 ml. Times.3) and saturated sodium chloride (50 ml. Times.1) were washed, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by silica gel column to give 2- (butyl (m-methyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (121 mg, 21.1%).

LCMS(ESI):[M+H]⁺=448.26

¹H NMR(400MHz,CDCl₃)δ7.01(s,2H),6.71(s,1H),6.64(s,1H),5.34(s,1H),4.17(t,J＝6.4Hz,2H),3.93–3.85(m,5H),3.73(s,3H),2.90(t,J＝6.4Hz,2H),2.35(s,3H),2.16(s,6H),1.62–1.51(m,2H),1.40–1.27(m,2H),0.89(t,J＝8.0Hz).

Example 1, compounds 1-1 and Compounds 1-2

The preparation method comprises the following steps:

1.

Tert-butyl N- (2-bromopropyl) carbamate (0.91 g,3.8mmol,1.5 eq.) cesium carbonate (2.50 g, 7.662mmol,3 eq.) Pd ₂(dba)₃ (0.23 g,0.26mmol,0.1 eq.) and (E) -2- (methylsulfonylimino) -9, 10-dimethoxy-2, 3,6, 7-tetrahydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (1 g,2.54mmol,1.0 eq.) were dissolved in 20mL 1, 4-dioxane, protected with nitrogen and stirred overnight at 110 ℃. Concentrated under reduced pressure, dissolved in 10mL of water, extracted with ethyl acetate (3X 10 mL), and the organic layer was washed with 20mL of saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reverse phase column chromatography under the following conditions (column: C18; mobile phase A: water (10 mmol/L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: 30%B to 50%B in 20min; wavelength: 254 nm) to give tert-butyl (E) - (2- (2- (methylsulfonylimino) -9, 10-dimethoxy-4-oxo-6, 7-dihydro-2H-pyrimido [6,1-a ] isoquinolin-3 (4H) -yl) propyl) carbamate (500 mg, yield 35.6%).

LCMS(ESI):[M+H]⁺=549.06

2.

Tert-butyl (E) - (2- (2- (methylsulfonylimino) -9, 10-dimethoxy-4-oxo-6, 7-dihydro-2H-pyrimido [6,1-a ] isoquinolin-3 (4H) -yl) propyl) carbamate (500 mg,0.91mmol,1 eq.) was dissolved in 10ml of 1,4 dioxane solution (4M) of hydrochloric acid and stirred at room temperature for 1 hour. Concentrated under reduced pressure, dissolved with 10mL of water, extracted with ethyl acetate (3X 10 mL), the organic layer was washed with 20mL of saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 2- ((1-aminopropan-2-yl) (methanesulfonyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (300 mg, 73.3% yield).

LCMS(ESI):[M+H]⁺=449.16

3.

2- ((1-Aminopropan-2-yl) (methanesulfonyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (200 mg,0.45mmol,1.0 eq.) triethylamine (135 mg,1.34mmol,3 eq.) and trimethylsilane isocyanate (102.73 mg,0.9mol,2 eq.) were dissolved in 10mL of dichloromethane and stirred at room temperature for 2 hours. Concentrated under reduced pressure, dissolved in 10mL of water, extracted with ethyl acetate (3X 10 mL), the organic layer was washed with 20mL of saturated sodium chloride, dried over anhydrous Na ₂SO₄, filtered, and concentrated under reduced pressure. The crude product was purified by reverse phase column chromatography under the following conditions (column: xselect CSH C m,30mm X150 mm; mobile phase A: water (0.1% FA), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: 28%B to 38%B in 8 min; wavelength: 254nm/220 nm; RT1 (min): 5.85/7.65) to give 1- (2- ((9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-2-yl) (methanesulfonamido) propyl) urea (100 mg, yield 45.6%).

LCMS(ESI):[M+H]⁺=492.35

4.

The racemate (100 mg) was purified by Chiral-HPLC on a column of CHIRALPAK IE-3,4.6 x 50mm,3um, mobile phase a: DCM-HPLC, mobile phase B: ETOH: dcm=1:1, flow rate: 18mL/min, gradient: isocratic 20, wavelength: 220nm, rt1 (min): 17.4, rt2 (min): 21.8, sample solvent: meOH-HPLC, sample loading: 0.45mL, run number: 5. The product is obtained.

Compound 1-1 (isomer 1, RT1 (min): 17.4,13.8mg, 22.14% yield)

LCMS(ESI):[M+H]⁺=492.35

¹H NMR(400MHz,Methanol-d₄)δ7.14–7.11(d,J＝10.1Hz,2H),6.93(s,1H),6.64(s,1H),5.40(s,1H),4.49(broad,1H),4.20–4.04(m,2H),3.89(s,3H),3.66(s,3H),2.97(t,J＝6.5Hz,2H),2.37(s,3H),2.22(s,3H),2.10(s,3H),1.05(s,3H).

Compound 1-2 (isomer 2, R2 (min): 21.8,13.0mg, 22.0% yield)

LCMS(ESI):[M+H]⁺=492.35

¹H NMR(400MHz,Methanol-d₄)δ7.12(d,J＝10.1Hz,2H),6.93(s,1H),6.64(s,1H),5.40(s,1H),4.49(broad,1H),4.20–4.04(m,2H),3.89(s,3H),3.66(s,3H),2.97(t,J＝6.5Hz,2H),2.37(s,3H),2.22(s,3H),2.08(s,6H),1.05(s,3H).

The single crystal diffraction diagram of the compound 1-2 is shown in figure 1.

Example 2, compound 2

The preparation method comprises the following steps:

1.

9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) amino ] -6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (3.0 g,7.7mmol,1.0 eq.) and tert-butyl 3-bromopyrrolidine-1-carboxylate (2.3 g,9.2mmol,1.2 eq.) are dissolved in 1, 4-dioxane (100 mL), xantphos (0.9 g,1.5mmol,0.2 eq.), cesium carbonate (5.0 g,15.3mmol,2.0 eq.) and Pd ₂(dba)₃ (0.7 g,0.7mmol,0.1 eq.) are added. After stirring for 18h under nitrogen at 100 ℃, the resulting mixture was concentrated under reduced pressure. The residue was purified by reverse phase chromatography on a column, C18 silica gel column, mobile phase, acetonitrile, water, gradient 0% -100% for 30 min, detector, UV 254 nm to give tert-butyl 3- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) pyrrolidine-1-carboxylate (1.4 g, yield 32.6%).

LCMS(ESI):[M+H]⁺=561.3

2.

3- ({ 9, 10-Dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) pyrrolidine-1-carboxylic acid tert-butyl ester (300 mg,0.5mmol,1.0 eq.) was mixed with a solution of 1, 4-dioxane of hydrochloric acid (8 mL), and the resulting mixture was stirred at room temperature for 1h. The resulting mixture was concentrated under reduced pressure to give 9, 10-dimethoxy-2- [ pyrrolidin-3-yl (2, 4, 6-trimethylphenyl) amino ] -6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (300 mg, crude). The crude product was used directly in the next step without further purification.

LCMS(ESI):[M+H]⁺=461.2

3.

9, 10-Dimethoxy-2- [ pyrrolidin-3-yl (2, 4, 6-trimethylphenyl) amino ] -6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (100 mg,0.2mmol,1.0 eq.) is dissolved in methylene chloride (4 mL) at room temperature, triethylamine (109 mg,1.1mmol,5.0 eq.) and isocyano-trimethylsilane (38 mg,0.3mmol,1.5 eq.) are added. The resulting mixture was stirred at room temperature for 2h. The resulting mixture was concentrated under reduced pressure. The crude product was purified in reverse phase (column: YMC-Triart C8.5 mmol/L, mobile phase A: water (10 mmol/L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: rising from 32% B to 50% B in 7 min; wavelength: 254nm/220nm; RT1 (min): 6.23) to give 3- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) pyrrolidine-1-carboxamide (30.5 mg, yield 27.4%).

LCMS(ESI,m/z):[M+H]⁺=504.25

¹H NMR(400MHz,Methanol-d4)δ7.13(s,2H),6.92(s,1H),6.64(s,1H),5.41(s,1H),5.08–4.99(m,1H),4.19–4.03(m,3H),3.88(s,3H),3.66(s,3H),3.58–3.52(m,1H),3.29(t,J＝7.3Hz,1H),3.25–3.18(m,1H),2.96(t,J＝6.5Hz,2H),2.38(s,3H),2.29–2.22(m,1H),2.17–2.15(m,6H),1.77(s,1H).

Example 3, compound 3-1 and Compound 3-2

The preparation method comprises the following steps:

1.

Dimethoxyphenylethylamine (50.0 g,275.8mmol,1.0 eq.) and ethyl cyanoacetate were sequentially added to a single-necked flask, reacted at 100℃for 16h under nitrogen, then cooled to 70℃and ethanol (80 mL) was added. The filter cake was washed with ethanol (3×30 mL) and the remaining ethanol in the filter cake was dried to give 2-cyano-N- [2- (3, 4-dimethoxyphenyl) ethyl ] acetamide (50.0 g,73.0% yield).

LCMS(ESI,m/z):[M+H]⁺=249.2

2.

In a single vial was added 2-cyano-N- [2- (3, 4-dimethoxyphenyl) ethyl ] acetamide (50.0 g,201.3mmol,1.0 eq.) and phosphorus oxychloride (500 mL) in sequence and reacted overnight at 85℃under nitrogen. The solvent was dried, water (300 mL) was added, extracted with DCM (3X 400 mL), the combined organic phases were dried over anhydrous sodium sulfate, and the organic phases were dried to give 2- [ (1E) -6, 7-dimethoxy-3, 4-dihydro-2H-isoquinolin-1-ylidene ] acetonitrile (45.0 g, 97.0%).

LCMS(ESI,m/z):[M+H]⁺=231.2

3.

In a single flask, 2- [ (1E) -6, 7-dimethoxy-3, 4-dihydro-2H-isoquinolin-1-ylidene ] acetonitrile (45.0 g,195.4mmol,1.0 equivalent) and concentrated sulfuric acid (320 mL) were sequentially added, and the reaction was carried out at room temperature for 3 hours. The reaction was slowly poured into ice water (1L), the pH was adjusted to ph=7 with sodium hydroxide (4M) at 0 ℃, extracted with EtOAc (3×600 mL), the combined organic phases were dried over anhydrous sodium sulfate, and the organic phases were dried to give 2- [ (1E) -6, 7-dimethoxy-3, 4-dihydro-2H-isoquinolin-1-ylidene ] acetamide (40.0 g, 82.4%).

LCMS(ESI,m/z):[M+H]⁺=249.2

4.

2- [ (1E) -6, 7-dimethoxy-3, 4-dihydro-2H-isoquinolin-1-ylidene ] acetamide (40.0 g,161.1mmol,1.0 eq), ethanol (600 mL), and a solution of sodium ethoxide in ethanol (127 mL, 21%) were sequentially added to a single vial, reacted at 80℃for 0.5H, and then diethyl carbonate (57.1 g,483.3mmol,3.0 eq) was added at that temperature, and reacted at 80℃overnight. Cooled to room temperature, water (500 mL) was added, the pH was adjusted to 7 with dilute hydrochloric acid, the mixture was filtered, the filter cake was rinsed with ethanol (3 x100 mL), and the remaining ethanol in the filter cake was dried to give 9, 10-dimethoxy-3H, 6H, 7H-pyrimido [4,3-a ] isoquinoline-2, 4-dione (40.0 g, 90.5%).

LCMS(ESI,m/z):[M+H]⁺=275.2

5.

9, 10-Dimethoxy-3H, 6H, 7H-pyrimido [4,3-a ] isoquinoline-2, 4-dione (40.0 g,145.8mmol,1.0 eq.) and phosphorus oxychloride (240 mL) were added sequentially in a single flask and reacted overnight at 100℃under nitrogen. The solvent was dried, dissolved in water (500 mL), extracted with DCM (3X 500 mL) and the combined organic phases were dried over anhydrous sodium sulfate and the dried organic phases were dried to give 2-chloro-9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (30.0 g, 70.2%).

LCMS(ESI,m/z):[M+H]⁺=293.2

6.

In a single vial were added 2-chloro-9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (2.0 g,6.8mmol,1.0 eq.), isopropanol (50 mL), and trimethylaniline (4.6 g,34.1mmol,5.0 eq.) in sequence, and reacted overnight at 90℃under nitrogen. The solvent was dried, EA (20 mL) and saturated aqueous sodium bicarbonate (200 mL) were added, filtered, the filter cake was rinsed with EA (3X 50 mL), and the residual EA in the filter cake was dried to give (2E) -9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) imino ] -3H,6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (2.0 g, 74.8%).

LCMS(ESI,m/z):[M+H]⁺=392.2

7.

In a single vial, (2E) -9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) imino ] -3H,6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (2.0 g,5.1mmol,1.0 eq), ethyl 4-bromopentanoate (2.1 g,10.2mmol,2.0 eq), 1, 4-dioxane (100 mL), cesium carbonate (5.0 g,15.3mmol,3.0 eq), xantphos (0.6 g,1.0mmol,0.2 eq) and Pd ₂(dba)₃ (0.5 g,0.5mmol,0.1 eq) were sequentially added and reacted at 100℃for 12h under nitrogen. Normal phase purification with DCM/MeOH (10/1) as mobile phase afforded ethyl 4- ({ 9, 10-dimethoxy-4-oxo-6 h,7 h-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) valerate (500 mg,18.8% yield).

LCMS(ESI,m/z):[M+H]⁺=520.3

8.

A single-necked flask was charged with a solution of ethyl 4- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) valerate (500 mg,1.0mmol,1.0 eq.), methanol (50 mL), and sodium hydroxide (77 mg,2.0mmol,2.0 eq.) in water (10 mL) in this order, and reacted at room temperature for 3 hours. The solvent was dried, water (20 mL) was added, acid adjusted to ph=5 with dilute hydrochloric acid, extracted with EA (3×50 mL), the combined organic phases were dried over anhydrous sodium sulfate, and the dried organic phases gave 4- ({ 9, 10-dimethoxy-4-oxo-6 h,7 h-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) pentanoic acid (450 mg,95.1% yield).

LCMS(ESI,m/z):[M+H]⁺=492.2

9.

4- ({ 9, 10-Dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) pentanoic acid (300 mg,0.6mmol,1.0 eq), 1.4-dioxane (100 mL), triethylamine (182 mg,1.8mmol,3.0 eq) and DPPA (336 mg,1.2mmol,2.0 eq) were sequentially added to a three-necked flask, and reacted at room temperature for 1h under nitrogen, then heated to 100 ℃ and reacted for 1h, cooled to 80 ℃ and added with an aqueous tetrahydrofuran solution (1.4M) (100 mL), reacted at room temperature for 2h, dried the solvent, and reversed (column: X-Bridge BEH C18 μm, 30X 150mm; mobile phase A water (10 mmol/L NH ₄HCO₃), mobile phase B acetonitrile, flow rate 60mL/min mL/min, gradient 39%B to 48%B in 9min, wavelength 254nm/220nm, RT1 (min): 8) to give 3- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) butanourea (94 mg,30.46% yield).

LCMS(ESI,m/z):[M+H]⁺=506.3

10.

Crude 3- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) butanone (50 mg) was resolved by Chiral-HPLC (column: CHIRAL ART Cellulose-SB, 2X 25cm,5 μm; mobile phase A: mtBE (0.1% DEA) -HPLC; mobile phase B: meOH: DCM=1:1-HPLC; flow rate:20mL/min; gradient: isocratic; wavelength: 254nm; RT1 (min): 5.3; RT2 (min): 7.2;Sample Solvent:MeOH:DCM =1:1-HPLC; injection: 1mL;Number Of Runs:5) to give the product.

Compound 3-1 (isomer 1, RT1 (min): 5.3,17.6mg,34.78% yield)

LCMS(ESI,m/z):[M+H]⁺=506.50

¹H NMR(400MHz,Methanol-d4)δ7.16–7.08(m,2H),6.93(s,1H),6.64(s,1H),5.41(s,1H),4.59–4.55(m,1H),4.19–4.02(m,2H),3.89(s,3H),3.66(s,3H),3.38–3.34(m,1H),3.25–3.21(m,1H),2.97(t,J＝6.4Hz,2H),2.38(s,3H),2.34–2.30(m,1H),2.23(s,3H),2.08(s,3H),1.80–1.76(m,1H),0.99(d,J＝6.9Hz,3H).

Compound 3-2 (isomer 2, R2 (min): 7.2,17.8mg,34.78% yield)

LCMS(ESI,m/z):[M+H]⁺=506.50

¹H NMR(400MHz,Methanol-d₄)δ7.16–7.08(m,2H),6.93(s,1H),6.64(s,1H),5.41(s,1H),4.59–4.45(m,1H),4.19–4.02(m,2H),3.89(s,3H),3.66(s,3H),3.38–3.34(m,1H),3.25–3.21(m,1H),2.97(t,J＝6.4Hz,2H),2.38(s,3H),2.34–2.30(m,1H),2.23(s,3H),2.08(s,3H),1.80–1.76(m,1H),0.99(d,J＝6.9Hz,3H).

Example 4, compound 4-1 and Compound 4-2

The preparation method comprises the following steps:

1.

2,4, 6-trimethylaniline (3.0 g,22.2mmol,1.0 eq.) was dissolved in MeOH (30 mL) at room temperature in air, naBH ₃ CN (1.0 g,44.3mmol,2.0 eq.) and acetic acid (1 mL,17.4mmol,0.7 eq.) were added. The resulting mixture was stirred overnight at room temperature under nitrogen. The resulting mixture was concentrated under reduced pressure. The reaction was quenched with water at room temperature. The resulting mixture was extracted with ethyl acetate (2×100 mL). The combined organic phases were washed with saturated brine (3×200 mL) and dried over anhydrous sodium sulfate. The filtrate was concentrated under reduced pressure to give tert-butyl N- {3- [ (2, 4, 6-trimethylphenyl) amino ] cyclobutyl } carbamate (1.2 g, yield 16%).

LCMS(ESI):[M+H]+=305.1

2.

Tert-butyl N- {3- [ (2, 4, 6-trimethylphenyl) amino ] cyclobutyl } carbamate (1.2 g,3.9mmol,1.0 eq.) and 2-chloro-9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (1.3 g,4.7mmol,1.2 eq.) were added to isopropanol (15 ml) at room temperature. The reaction mixture was heated to 100 ℃ and stirred overnight under nitrogen at 100 ℃. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with petroleum ether/ethyl acetate (2/1) to give 2- [ (3-aminocyclobutyl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6 h,7 h-pyrimido [4,3-a ] isoquinolin-4-one (150 mg, yield 6%).

LC-MS(ESI):[M+H]⁺=461.2

3.

N1- (2, 4, 6-trimethylphenyl) cyclobutane-1, 3-diamine (150 mg,0.7mmol,1.0 eq.) was dissolved in DCM (1 ml) at room temperature in air and solid phosgene (193 mg,0.6mmol,2.0 eq.) was added to the solution. The resulting mixture was stirred under nitrogen at room temperature for 1 hour. Ammonia in methanol (15 mL, 7M) was added to the above mixture at room temperature). The resulting mixture was stirred at room temperature overnight. The resulting mixture was concentrated under reduced pressure. The residue was purified by reverse phase flash chromatography on a chromatographic column, C18 silica gel, mobile phase, acetonitrile water (10 mmol/L NH ₄HCO₃), 30% -47% gradient, 7 min, detector, UV 254 nm. The crude product was purified by pre-high performance liquid chromatography under the following conditions (column: CHIRAL ART Cellulose-SB, 2x 25cm,5 μm; mobile phase A: mtBE (0.1% DEA) -HPLC, mobile phase B: meOH: DCM=1:1-HPLC; flow rate: 20mL/min; gradient: isocratic; wavelength: 220nm; RT1 (min): 6.5; RT2 (min): 9.4;Sample Solvent: feed: 0.7mL; run number: 3) to give the product.

Compound 4-1 (isomer 1, RT1 (min): 6.5,16.3mg,9% yield)

LCMS(ESI):[M+H]⁺=504.30

¹H NMR(400MHz,Methanol-d₄)δ7.20(s,1H),7.05(s,2H),6.93(s,1H),6.11(s,1H),4.56–4.47(m,1H),4.28–4.15(m,1H),4.03(t,J＝6.2Hz,2H),3.90–3.88(m,6H),2.93(t,J＝6.2Hz,2H),2.78–2.66(m,2H),2.32(s,3H),2.23(s,6H),1.88–1.73(m,2H).

Compound 4-2 (isomer 2, R2 (min): 9.4,15.5mg,10% yield)

LCMS(ESI):[M+H]⁺=504.40

¹H NMR(400MHz,Methanol-d₄)δ7.26(s,1H),7.05(s,2H),6.95(s,1H),6.25(s,1H),5.05–4.90(m,1H),4.28–4.15(m,1H),4.04(t,J＝6.4Hz,2H),3.92(s,6H),2.95(t,J＝6.4Hz,2H),2.34–2.32(m,7H),2.22(s,6H).

Example 5, compound 5-1 and Compound 5-2

The preparation method comprises the following steps:

1.

2-chloro-9, 10-dimethoxy-6 h,7 h-pyrimidine [4,3-A ] isoquinoline-4-1 (200 mg,0.7mmol,1.0 eq.) and 2,4, 6-trimethylaniline (369 mg,2.7mmol,4.0 eq.) are dissolved in isopropanol (4 mL) and stirred under nitrogen at 90℃for 3 hours. The resulting mixture was concentrated under reduced pressure. Purification by column chromatography using DCM/MeOH (10/1) gave (2E) -9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) imine ] -3h,6H,7 h-pyrimidine [4,3-a ] isoquinoline-4-1 (100 mg, yield 37.39%).

LCMS(ESI,m/z):[M+H]⁺=392.1

2.

Tert-butyl 2-bromopropyl carbamate (304 mg,1.3mmol,1.0 eq.) and Cs ₂CO₃ (1.0 g,3.9mmol,3.0 eq.) were added to a mixed solution of (2E) -9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) imine ] -3h,6H,7H-pyrimido [4,3-a ] isoquinoline-1 (500 mg,1.3mmol,1.0 eq.) and dioxane (10 mL), pd ₂(dba)₃ (117 mg,0.1mmol,0.1 eq.) and Xantphos (74 mg,0.1mmol,0.1 eq.) were added under nitrogen. The resulting mixture was stirred under nitrogen at 80 ℃ for 6 hours. The resulting mixture was concentrated under reduced pressure. The reversed phase column chromatography purification is carried out under the conditions that the chromatographic column is C18 silica gel, mobile phase, water, meCN (0.1% FA), gradient of 30% -70%, 15min and ultraviolet of 254nm. Tert-butyl N- [2- ({ 9, 10-dimethoxy-4-oxo-6 h,7 h-pyrimidin [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] carbamate (400 mg, yield 57.08%) was obtained.

LCMS(ESI,m/z):[M+H]⁺=549.6

3.

A25 mL round bottom flask was charged with N- [2- ({ 9, 10-dimethoxy-4-oxo-6 h,7 h-pyrimidin [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] carbamate (100 mg,0.2mmol,1.0 eq.) and a solution of HCl in 1, 4-dioxane (2 mL, 4M). The reaction mixture was stirred under nitrogen at 25 ℃ for 1 hour. The resulting mixture was concentrated under reduced pressure. 2- [ (1-aminopropen-2-yl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6 h,7 h-pyrimidine [4,3-a ] isoquinolin-4-1 (80 mg, crude product) was obtained.

LCMS(ESI,m/z):[M+H]⁺=449.2

4.

2- [ (1-Aminopropane-2-yl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6 h,7 h-pyrimidine [4,3-a ] isoquinoline-4-1 (100 mg,0.2mmol,1.0 eq.) and isocyano-trimethylsilane (38 mg,0.3mmol,1.5 eq.) are stirred in DCM (5 mL) and TEA (60 mg,0.6mmol,3.0 eq.) is added dropwise under nitrogen at 25 ℃. The reaction mixture was stirred under nitrogen at 25 ℃ for 2 hours. The resulting mixture was concentrated under reduced pressure. Purifying by column chromatography, wherein the chromatographic column is C18 silica gel, the mobile phase is ACN water solution (0.1% NH ₃.H₂ O), the gradient is 25% -75%, 20min, and the ultraviolet is 254nm. This gave 2- ({ 9, 10-dimethoxy-4-oxo-6 h,7 h-pyrimidin [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propylurea (80 mg, 73.0% yield).

LCMS(ESI,m/z):[M+H]⁺=492.2

5.

Into a 50mL round bottom flask was charged 2- ({ 9, 10-dimethoxy-4-oxo-6 h,7 h-pyrimidin [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propiurea (100 mg,0.2mmol,1.0 eq.) and aqueous hydrobromic acid (5 mL). The resulting mixture was stirred under nitrogen at 120 ℃ for 3 hours. The mixture was extracted with ethyl acetate (2X 50 mL), and the organic layer was washed with saturated brine (3X 100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. 2- ({ 9, 10-dihydroxy-4-oxo-6 h,7 h-pyrimidin [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propylurea (80 mg, crude product) was obtained.

LCMS(ESI,m/z):[M+H]⁺=464.2

6.

2- ({ 9, 10-Dihydroxy-4-oxo-6 h,7 h-pyrimidin [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propylurea (100 mg,0.2mmol,1.0 eq) and K ₂CO₃ (149 mg,1.0mmol,5.0 eq) were added to DMF (1.5 mL) and ethyl 2-bromo-2, 2-difluoroacetate (44 mg,0.2mmol,1.0 eq) was added dropwise at room temperature. The reaction mixture was stirred under nitrogen at 55 ℃ for 1.5 hours. The resulting mixture was concentrated under reduced pressure. The reversed phase column chromatography purification is carried out under the conditions that the chromatographic column is C18 silica gel, the mobile phase, water, ACN (0.1% FA), gradient of 20% -70%, 20min and ultraviolet of 254nm. 2- { [9- (difluoromethoxy) -10-hydroxy-4-oxo-6 h,7 h-pyrimidin [4,3-a ] isoquinolin-2-yl ] (2, 4, 6-trimethylphenyl) amino } propylurea (10 mg, yield 9.03%) was obtained.

LCMS(ESI,m/z):[M+H]⁺=514.2

7.

2- { [9- (Difluoromethoxy) -10-hydroxy-4-oxo-6 h,7 h-pyrimidin [4,3-a ] isoquinolin-2-yl ] (2, 4, 6-trimethylphenyl) amino } propylurea (10 mg,0.02mmol,1.0 eq.) and K ₂CO₃ (4 mg,0.03mmol,1.5 eq.) were added dropwise at room temperature as solvent DMF (0.5 mL). The resulting mixture was stirred under nitrogen at 60 ℃ for 1.5 hours. The resulting mixture was concentrated under reduced pressure. Purifying by high performance liquid chromatography, wherein the column type is XB ridge BEH C18 μm,30 x 150mm, mobile phase A is water (10 mmol/LNH ₄HCO₃), mobile phase B is ACN, flow rate is 60mL/min, gradient is 40% B to 50% B for 8 min, wavelength is 254nm/220nm, and R1 (min) is 7.3 to obtain racemization product. The racemic product was purified by chiral high performance liquid chromatography (column: chiral ART cell-SB, 2X 25cm,5 μm; mobile phase A: meOH: DCM=1:1-HPLC, mobile phase B: mtBE (0.1% DEA) -HPLC; flow rate: 20ml/min; gradient: equidistant 50; wavelength: 220nm; RT1 (min): 4.4; RT2 (min): 10.7; sample solvent: meOH-HPLC; injection: 1 ml).

Compound 5-1 (isomer 1, RT1 (min): 4.4,1.54mg, 14.87% yield).

LCMS(ESI,m/z):[M+H]⁺=529.40.

¹H NMR(400MHz,Methanol-d4)δ7.20–7.05(m,3H),6.95–6.82(m,1H),6.80(s,1H),5.49(s,1H),4.49(s,1H),4.16–4.10(m,2H),3.72(s,3H),2.97(t,J＝6.4Hz,2H),2.37(s,3H),2.23(s,3H),2.10(s,3H),1.35–1.28(m,2H),1.06(s,3H).

Compound 5-2 (isomer 2, R2 (min): 10.7,2.19mg, 20.9% yield)

LCMS(ESI,m/z):[M+H]⁺=529.45

¹H NMR(400MHz,Methanol-d4)δ7.20–7.05(m,3H),6.95–6.82(m,1H),6.80(s,1H),5.49(s,1H),4.49(s,1H),4.16–4.10(m,2H),3.72(s,3H),2.96(t,J＝6.4Hz,2H),2.37(s,3H),2.22(s,3H),2.11(s,3H),1.35–1.28(m,2H),1.06(s,3H).

Example 6, compound 6

The preparation method comprises the following steps:

1.

Trimethylaniline (1.0 g,7.4mmol,1.0 eq.) urea N, N, N ', N' -tetramethyl-O- (7-azabenzotriazol-1-yl) hexafluorophosphate (3.4 g,8.8mmol,1.2 eq.) and N, N-diisopropylethylamine (3.9 mL,22.2mmol,3.0 eq.) were dissolved in 20mL dichloromethane and stirred at room temperature for 2 hours. Concentrated under reduced pressure, dissolved in 10mL of water, extracted with ethyl acetate (3X 10 mL), and the organic layer was washed with 20mL of saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by normal phase column chromatography under the following conditions (mobile phase A: petroleum ether, mobile phase B: ethyl acetate; flow rate: 60mL/min; gradient: 10%B to 50%B in 20min; wavelength: 254 nm) to give 2- (1, 3-dioxoisoindolin-2-yl) -N-methanesulfonylacetamide (600 mg, yield 25.17%).

LCMS(ESI):[M+H]⁺=323.14

2.

2- (1, 3-Dioxoisoindol-2-yl) -N- (2, 4, 6-trimethylphenyl) acetamide (600 mg,1.8mmol,1.0 eq.) 2-chloro-9, 10-dimethoxy-6, 7-dihydro-4H-pyrimidine [6,1-A ] isoquinolin-4-one (653 mg,2.2mmol,1.2 eq.) O, diphenyl O- (1-acetoxy-2, 2-trichloroethyl) phosphonate (118.40 mg,0.186mmol,0.1 eq.) methanesulfonic acid { [4- (N, N- (dimethylamino) phenyl ] di-tert-butylphosphino } (2 '-amino-1, 1' -biphenyl-2-yl) palladium (II) (49.39 mg,0.186mmol,0.1 eq.) and cesium carbonate (1.5 g,4.6mmol,2.5 eq.) were dissolved with 10mL of 1, 4-dioxane, nitrogen protection, 110℃stirred overnight, concentrated under reduced pressure, dissolved with 10mL of water, extracted with ethyl acetate (3X 20 mL), the organic layer washed with 20mL of saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, the crude product purified by reverse phase column chromatography under the following conditions (column: C18; mobile phase a water (10 mmol/L ammonium bicarbonate) mobile phase B acetonitrile, flow rate 60mL/min, gradient 30%B to 50%B in 20min, wavelength 254 nm) to give 2-amino-N- (9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-2-yl) -N-methanesulfonylacetamide (50 mg, yield 5.04%).

LCMS(ESI):[M+H]⁺=449.16

3.

2-Amino-N- (9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-2-yl) -N-methanesulfonylacetamide (20 mg,0.05mmol,1.0 eq.), triethylamine (14 mg,0.15mmol,3.0 eq.) and N, N' -carbonyldiimidazole (10.9 mg,0.07mmol,1.5 eq.) were dissolved in 2mL of dichloromethane, protected with nitrogen, and 2mL of a 1.4-dioxane solution of amine was added dropwise at 0℃and stirred at room temperature for 2 hours. The crude product was purified by reverse phase column chromatography under the following conditions (column: sunFire C18 5m,30mm X150 mm; mobile phase A: water (0.1% formic acid), mobile phase B: acetonitrile; flow rate: 60mL/min mL/min; gradient: 45%B to 59%B in 7 min; wavelength: 254nm/220nm; time: 5.92) to give N- (9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-2-yl) -N-methanesulfonyl-2-ureidoacetamide (2.9 mg, yield 13.10%).

LCMS(ESI,m/z):[M+H]⁺=475.20

¹H NMR(400MHz,DMSO-d6)δ8.45(s,1H),7.71(s,1H),7.18(s,1H),7.08-7.01(m,3H),4.77(s,2H),4.08(t,J＝6.6Hz,2H),3.86(s,3H),3.79(s,3H),2.97(t,J＝6.6Hz,2H),2.30(s,3H),2.12–2.10(s,6H).

Example 7, compound 7-1, compound 7-2 and Compound 7-3

The preparation method comprises the following steps:

1.

To methyl 2-bromocyclopropane-1-carboxylate (200 mg,1.1mmol,1.0 eq) and (2E) -9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) imino ] -3H,6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (437 mg,1.1mmol,1.0 eq) in 1, 4-dioxane (80 mL) was added cesium carbonate (1.1 g,3.4mmol,3.0 eq), xantphos (106 mg,0.2mmol,0.2 eq) and Pd ₂(dba)₃ (64 mg,0.1mmol,0.1 eq). After stirring at 100 ℃ under nitrogen for 12h, the resulting mixture was concentrated under reduced pressure. The residue was purified by column chromatography on silica eluting with methanol/dichloromethane (1/10) to give the crude product. The crude product was purified by chromatography (YMC TRIART C18. 18ExRs 5m,30 mm. Times.150 mm; mobile phase A: water (10 mmol/L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: 39% B to 62% B,10 min; wavelength: 254nm/220nm; RT1 (min): 8.33) to give methyl 2- [ (2E) -9, 10-dimethoxy-4-oxo-2- [ (2, 4, 6-trimethylphenyl) imino ] -6H, 7H-pyrimido [4,3-a ] isoquinolin-3-yl ] cyclopropane-1-carboxylate (100 mg, 18.3% yield).

LCMS(ESI):[M+H]⁺=490.2

2.

To methyl 2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) cyclopropane-1-carboxylate (160 mg,0.3mmol,1,0 eq.) in methanol (20 mL) was added an aqueous solution of sodium hydroxide (65 mg,1.6mmol,5.0 eq.) in water (5 mL) at room temperature. The mixture was stirred at 50 ℃ for 18h. After the completion of the reaction, the reaction solution was concentrated under reduced pressure. The resulting mixture was extracted with ethyl acetate (3×50 mL). The aqueous layer was acidified to ph=4 with 6M aqueous hydrochloric acid. The resulting mixture was extracted with dichloromethane (3×100 mL). The combined organic layers were washed with brine and dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated under reduced pressure to give 2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) cyclopropane-1-carboxylic acid (170 mg, crude product).

LCMS(ESI):[M+H]⁺=476.2

3.

To 2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) cyclopropane-1-carboxylic acid (170 mg,0.4mmol,1.0 eq.) in dichloromethane (10 mL) was added dropwise oxalyl chloride (0.1 mL,1.1mmol,3.0 eq.). The resulting mixture was stirred under nitrogen at 0 ℃ for 1h. The resulting mixture was concentrated under reduced pressure. The crude product was used directly without further purification. The crude product was dissolved in dichloromethane (1 mL) and added to a tetrahydrofuran solution of ammonia (60 mL,1.3 m) at 0 ℃. The resulting mixture was stirred at 0 ℃ for 1h. The resulting mixture was extracted with ethyl acetate (3×60 mL). The combined organic layers were washed with brine and dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated under reduced pressure to give 2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) cyclopropane-1-carboxamide (180 mg, crude product).

LCMS(ESI):[M+H]⁺=475.2

4.

A mixture of 2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) cyclopropane-1-carboxamide (180 mg,0.4mmol,1.0 eq.) in tetrahydrofuran (20 mL) was cooled to 0℃and sodium borohydride (143 mg,3.8mmol,10.0 eq.) was added in portions. Iodine granules (963 mg,3.8mmol,10.0 eq.) were dissolved in tetrahydrofuran (3 mL) and added to the resulting mixture. The resulting mixture was stirred at 70 ℃ for 18h. The reaction was quenched by the addition of water (30 mL) at room temperature. The resulting mixture was extracted with ethyl acetate (2×50 mL). The combined organic layers were washed with brine and dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by Prep-TLC (dichloromethane/methanol=10/1) to give 2- { [2- (aminomethyl) cyclopropyl ] (2, 4, 6-trimethylphenyl) amino } -9, 10-dimethoxy-6 h,7 h-pyrimido [4,3-a ] isoquinolin-4-one (45 mg, yield 25.8%).

LCMS(ESI):[M+H]⁺=461.2

5.

A mixture of 2- { [2- (aminomethyl) cyclopropyl ] (2, 4, 6-trimethylphenyl) amino } -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (45 mg,0.1mmol,1.0 eq), triethylamine (30 mg,0.3mmol,3.0 eq) and trimethylsilane isocyanate (23 mg,0.2mmol,2.0 eq) in DCM (5 mL) was stirred at room temperature for 3h. The resulting mixture was concentrated under reduced pressure. The crude product (45 mg) was purified by Prep-HPLC under the following conditions (Column: XB ridge BEH C18 OBD Prep Column 130,5m,19mm 250mm; mobile phase A: water (10 mmol/L ammonium bicarbonate), flow rate: 25mL/min; gradient: 45% B to 67% B10 min; wavelength: 254nm/220nm; RT1 (min): 7.93), to give [2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) cyclopropyl ] methyl urea (20 mg, 40.65% yield).

LCMS(ESI):[M+H]⁺=504.3

6.

Compound 7-1, isomer 1 (column: CHIRAL ART Amylose-SA, 2X 25cm,5 μm; mobile phase A: methanol: dichloromethane=1:1, mobile phase B: methyl tert-butyl ether (0.1% diethylamine) flow rate: 20mL/min; gradient: isocratic; wavelength: 254nm; RT1 (min): 6.7; sample solvent: methanol (0.1% diethylamine); sample loading: 1mL; run number: 3) gives the product (3.8 mg, 12.43% yield).

LCMS(ESI):[M+H]⁺=504.30

¹H NMR(400MHz,Methanol-d4)δ7.10(s,1H),7.07(s,1H),6.95(s,1H),6.72(s,1H),5.54(s,1H),4.25–4.06(m,2H),3.90(s,4H),3.68(s,3H),2.99(t,J＝6.6Hz,3H),2.48–2.40(m,1H),2.36(s,3H),2.23(s,3H),2.03(s,3H),1.43–1.31(m,2H).

Compound 7-2, isomer 2 (column: CHIRAL ART Amylose-SA, 2X 25cm,5 μm; mobile phase A: methanol: dichloromethane=1:1, mobile phase B: methyl tert-butyl ether (0.1% diethylamine) flow rate: 20mL/min; gradient: isocratic; wavelength: 254nm; RT2 (min): 10.9; sample solvent: methanol (0.1% diethylamine); sample loading: 1mL; run number: 3) gives the product (3.5 mg, 11.29% yield).

LCMS(ESI):[M+H]⁺=504.35

Compound 7-3, isomer 3 (Column: XB ridge BEH C18 OBD Prep Column 130,5m,19mm 250mm; mobile phase A: water (10 mmol/L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: 39% B to 61% B9 min; wavelength: 254nm/220nm; RT1 (min): 5.45) gives the product (20 mg, 40.6% yield).

LCMS(ESI):[M+H]⁺=504.35

¹H NMR(400MHz,Methanol-d4)δ7.10(s,1H),7.07(s,1H),7.02–6.89(m,2H),6.70(s,1H),5.57(s,1H),4.61(s,6H),4.27–4.04(m,2H),4.05–3.91(m,2H),3.90(s,3H),3.85–3.72(m,1H),3.68(s,3H),3.05–2.95(m,2H),2.92–2.87(m,1H),2.36(s,3H),2.31–2.14(m,8H).

Example 8, compound 8

Preparation steps

1.

After stirring N- (2-hydroxypropyl) carbamic acid tert-butyl ester (15.0 g,85.6mmol,1.0 eq) and TEA (26.0 g,256.8mmol,1.5 eq) in DCM (80 mL), DMAP (1.1 g,8.6mmol,0.1 eq) and TsCl (21.2 g,111.3mmol,1.3 eq) were added in portions at 0deg.C. The resulting mixture was stirred at room temperature for 2h. The resulting mixture was extracted with dichloromethane (3×200 mL). The organic layer was washed with brine and dried over anhydrous sodium sulfate. The filtrate after filtration was concentrated under reduced pressure and purified by silica gel column chromatography, PE/EA (5/1) eluted to give N- [2- [ (4-methylbenzenesulfonyl) oxy ] propyl ] carbamate (20.3 g, yield 71.9%).

LC-MS(ESI):[M+H]⁺=330.1

2.

To a solution of 9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) amino ] -6h,7 h-pyrimido [4,3-a ] isoquinolin-4-one (1.6 g,4.1mmol,1.0 eq.) in 1, 4-dioxane (20 mL) was added tert-butyl N- [2- [ (4-methylbenzenesulfonyl) oxy ] propyl ] carbamate (2.0 g,6.1mmol,1.5 eq.), cesium carbonate (2.7 g,8.2mmol,2.0 eq.), 4, 5-bis-diphenylphosphine-9, 9-dimethylxanthene (0.5 g,0.8mmol,0.2 eq.) and tris (dibenzylideneacetone) dipalladium (0.4 g,0.4mmol,0.1 eq.) and the mixture was sparged with nitrogen gas to 100 ℃. The mixture was stirred under nitrogen for 5h at 100 ℃. The reaction was extracted with ethyl acetate (3×200 mL) and water (150 mL). The organic phase was dried over anhydrous sodium sulfate, and then filtered and distilled. The residue obtained is purified by reverse phase column chromatography on a chromatographic column XB ridge BEH C18,5 μm,30 x 150mm, mobile phase A water (10 mmol/L ammonium bicarbonate), mobile phase B acetonitrile, flow rate 60mL/min, gradient 55%B to 65%B in 7min, wavelength 254nm/220nm, RT1 (min) 5.67/6.05. Tert-butyl N- [2- ({ 9, 10-dimethoxy-4-oxo-6 h,7 h-pyrimidin [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] carbamate (35.33 mg, yield 1.55%) was obtained.

LC-MS(ESI):[M+H]⁺=549.35

¹H NMR(400MHz,Methanol-d₄)δ7.11(d,J＝10.2Hz,2H),6.93(s,1H),6.64(s,1H),5.41(s,1H),4.57–4.39(m,1H),4.18–4.02(m,2H),3.89(s,3H),3.66(s,3H),3.55–3.48(m,1H),3.40(t,J＝8.2Hz,1H),2.97(t,J＝6.5Hz,2H),2.37(s,3H),2.22(s,3H),2.11(s,3H),1.46(s,9H),1.31(s,1H),1.08(d,J＝6.9Hz,3H).

Example 9, compound 9-1 and Compound 9-2

The preparation method comprises the following steps:

1.

2- ((1-aminoprop-2-yl) (methanesulfonyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (150 mg,0.33mmol,1.0 eq.) 3H-1,2, 3-triazole-4-carboxylic acid (56 mg,0.5mmol,1.5 eq.) 1- (3-dimethylaminopropyl) -3-ethylcarbodiamine (96 mg,0.5mmol,1.5 eq.) 1-hydroxybenzotriazole (67 mg,0.5mmol,1.5 eq.) was dissolved in 2ml DMF, nitrogen-protected, stirred at room temperature for 1 hour, concentrated under reduced pressure. The crude product was purified by reverse phase column chromatography under the following conditions (column: xselect CSH C m,30mmX 150mm; mobile phase A: water (0.1% FA), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: 27%B to 55%B in 9min; wavelength: 254nm/220nm; RT1 (min): 6.77/8.43) to give N- (2- ((9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-2-yl) (methanesulfonamido) propyl) -1H-1,2, 3-triazole-5-carboxamide (30 mg, 16.5% yield).

LCMS(ESI):[M+H]⁺=544.25

2.

The racemate (30 mg) was purified by Chiral-SFC under the following conditions (column: CHIRALPAK IH 3.25 cm,5um; mobile phase A: CO ₂; mobile phase B: methanol (0.1% 2M NH ₃ -MEOH), flow rate: 85mL/min; gradient: isocratic 35% B; column temperature (. Degree. C.); 35; pressure (bar): 100; wavelength: 220nm; RT1 (min): 2.92; RT2 (min): 4.42; sample solvent: DCM (0.1% 2M NH ₃ -MeOH) - - -HPLC; sample injection amount: 1 mL) to give the product

Compound 9-1 (isomer 1, R1 (min): 2.92,9.2mg, 28.58% yield)

LCMS(ESI):[M+H]⁺=544.25

¹H NMR(400MHz,Methanol-d₄)δ8.25(s,1H),7.15–7.11(m,2H),6.99–6.92(m,1H),6.65(s,1H),5.44(s,1H),4.74(s,1H),4.23–4.04(m,2H),3.95-3.88(m,4H),3.85–3.75(m,1H),3.67(s,3H),2.99(t,J＝6.4Hz,2H),2.38(s,3H),2.27(s,3H),2.13(s,3H),1.31(d,J＝4.0Hz,1H),1.11(d,J＝6.8Hz,3H).

Compound 9-2 (isomer 2, R2 (min): 4.42,8.8mg, yield 24.5%)

LCMS(ESI):[M+H]⁺=544.25

Example 10, compound 10-1 and Compound 10-2

The preparation method comprises the following steps:

1.

2- ((1-aminoprop-2-yl) (methanesulfonyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (150 mg,0.33mmol,1.0 eq.) imidazole-2-carboxylic acid (56 mg,0.5mmol,1.5 eq.) 1- (3-dimethylaminopropyl) -3-ethylcarbodiamine (96 mg,0.5mmol,1.5 eq.) 1-hydroxybenzotriazole (67 mg,0.5mmol,1.5 eq.) was dissolved in 2ml DMF, nitrogen-protected, stirred at room temperature for 1 hour and concentrated under reduced pressure. The crude product was purified by reverse phase column chromatography under the following conditions (column: xselect CSH C m,30mmX 150mm; mobile phase A: water (0.1% FA), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: 27% B to 55% B in9 min; wavelength: 254nm/220nm; RT1 (min): 6.77/8.43) to give N- (2- ((9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-2-yl) (methanesulfonamido) propyl) -1H-imidazole-2-carboxamide (70 mg, yield 44.0%).

LCMS(ESI):[M+H]⁺=544.25

2.

The racemate (70 mg) was purified by Chiral-SFC under the following conditions (column: CHIRAL ART Cellulose-SB, 3X 25cm,5 μm; mobile phase A: CO ₂, mobile phase B: meOH (0.1% 2M NH ₃ -MeOH), flow rate: 100mL/min, gradient: isocratic 25% B; column temperature (. Degree. C.): 35; pressure (bar): 100; wavelength: 220nm; RT1 (min): 9.4; RT2 (min): 10.72; sample solvent: meOH (0.1% DEA) - - - -HPLC; sample injection amount: 1 mL) to give the product

Compound 10-1 (isomer 1, RT1 (min): 9.4,24.9mg, 34.5% yield).

LCMS(ESI):[M+H]⁺=543.25

¹H NMR(400MHz,Methanol-d₄)δ7.21(s,3H),7.16–7.08(m,2H),6.94(s,1H),6.65(s,1H),5.43(s,1H),4.70(s,1H),4.21–4.02(m,2H),3.97–3.91(m,1H),3.89(s,4H),3.82–3.84(m,1H),3.67(s,3H),3.05–2.94(m,2H),2.38(s,3H),2.27(s,3H),2.14(s,3H),1.31(s,1H),1.15(d,J＝6.8Hz,3H).

Compound 10-2 (isomer 2, R2 (min): 10.72,24.4mg, 34.5% yield)

LCMS(ESI):[M+H]⁺=543.25

¹H NMR(400MHz,Methanol-d₄)δ7.21(s,3H),7.16–7.08(m,2H),6.94(s,1H),6.65(s,1H),5.43(s,1H),4.70(s,1H),4.21–4.02(m,2H),3.97–3.91(m,1H),3.89(s,4H),3.84–3.76(m,1H),3.67(s,3H),3.05–2.94(m,2H),2.38(s,3H),2.27(s,3H),2.14(s,3H),1.31(s,1H),1.15(d,J＝6.8Hz,3H).

Example 11, compound 11-1 and Compound 11-2

The preparation method comprises the following steps:

1.

2H-pyrazole-3-carboxylic acid (25 mg,0.2mmol,1.0 eq.) was dissolved in SOCl ₂ (1 mL) and stirred at room temperature under nitrogen for 2H. The resulting mixture was concentrated in vacuo and dried. The above mixture was dissolved in DCM (1 mL) and added dropwise under nitrogen to a solution of 2- [ (1-aminopropan-2-yl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6 h,7 h-pyrimido [4,3-a ] isoquinolin-4-one (100 mg,0.2mmol,1.0 eq.) and N, N-diisopropylethylamine (86 mg,0.7mmol,3.0 eq.) in DCM (5 mL) at 0 ℃. The reaction solution was stirred at room temperature under nitrogen for 2h. The reaction was quenched with water (5 mL) at room temperature, extracted with ethyl acetate (3×10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The reverse phase purification is adopted, and the conditions are that the chromatographic column is C18 silica gel, the mobile phase, ACN (0.1% FA) in water, the gradient of 10% -50% and the ultraviolet detector are adopted for 10min, and the wavelength is 254nm. Purification gave N- [2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimidin [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] -2H-pyrazole-3-carboxamide (40 mg, 32.0% yield).

2.

The crude product (40 mg) was purified by chromatography column CHIRALPAK IH X25 cm,5 μm, mobile phase A CO ₂, mobile phase B MEOH (0.1% DEA), flow rate 80mL/min, gradient 40% B, column temperature (C) 10 ℃, column head temperature (C) 10 ℃ 80mL/min, gradient 40% B isocratic, column temperature (C) 35, pressure (bar) 100, wavelength 220nm, RT1 (min) 2.73, RT2 (min) 4.17, sample solvent 4.8mL to give the product.

Compound 11-1 (isomer 1, R1 (min): 2.73,12.1mg, 9.6% yield)

LCMS(ESI):[M+H]⁺=543.55

¹H NMR(400MHz,Methanol-d₄)δ7.71(d,J＝2.4Hz,1H),7.13(d,J＝13.6Hz,2H),6.93(s,1H),6.78(d,J＝2.4Hz,1H),6.65(s,1H),5.43(s,1H),4.70(s,1H),4.16–4.10(m,2H),3.91–3.78(m,5H),3.67(s,3H),2.98(t,J＝6.4Hz,2H),2.38(s,3H),2.27(s,3H),2.13(s,3H),1.11(d,J＝6.8Hz,3H).

Compound 11-2 (isomer 2, R2 (min): 4.17,10.9mg, 26.6% yield)

LCMS(ESI):[M+H]⁺=543.35

¹H NMR(400MHz,Methanol-d₄)δ7.71(d,J＝2.4Hz,1H),7.18–7.12(m,2H),6.93(s,1H),6.78(d,J＝2.4Hz,1H),6.65(s,1H),5.43(s,1H),4.75–4.55(m,1H),4.16–4.10(m,2H),3.91–3.78(m,5H),3.67(s,3H),2.98(t,J＝6.4Hz,2H),2.38(s,3H),2.27(s,3H),2.13(s,3H),1.11(d,J＝6.8Hz,3H).

Example 12, compound 12-1 and Compound 12-2

The preparation method comprises the following steps:

1.

Palladium on carbon (10%, 2.90 g) was added to 1-benzyl-5-methyl-1, 2, 3-triazole-4-carboxylic acid ethyl ester (850 mg,3.5mmol,1.0 eq.) in 150mL of methanol in an autoclave. The mixture was stirred at room temperature under 20bar hydrogen pressure for 18 hours, filtered through celite and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE/EA (1/1) to give ethyl 5-methyl-1-hydro-1, 2, 3-triazole-4-carboxylate (260 mg, yield 48.4%).

LCMS(ESI,m/z):[M+H]⁺=156.1

2.

A mixture of 5-methyl-1-hydrogen-1, 2, 3-triazole-4-carboxylic acid ethyl ester (260 mg,1.7mmol,1.0 eq.) and sodium hydroxide (201 mg,5.1mmol,3.0 eq.) in methanol (10 mL) and water (10 mL) was stirred at 60℃for 18 hours. The resulting mixture was concentrated under reduced pressure. The mixture was acidified to ph=5 with 2 molar aqueous hydrochloric acid. The resulting mixture was concentrated under reduced pressure. The residue was purified by reverse phase chromatography on a chromatographic column, C18 silica gel, mobile phase, acetonitrile, water, gradient 0% -10% for 10min, detector, UV 254nm to give 5-methyl-3H-1, 2, 3-triazole-4-carboxylic acid (200 mg, 93.9% yield).

LCMS(ESI):[M+H]⁺=128.0

3.

2- [ (1-Aminopropan-2-yl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (100 mg,0.2mmol,1.0 eq.) and 5-methyl-3H-1, 2, 3-triazole-4-carboxylic acid (28.3 mg,0.2mmol,1.0 eq.) are dissolved in DMF (3 mL) and 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (64 mg,0.3mmol,1.5 eq.) and 1-hydroxybenzotriazole (45 mg,0.3mmol,1.5 eq.) are added at room temperature. The resulting mixture was stirred at room temperature for 2h. The resulting mixture was concentrated under reduced pressure to give a crude product. The crude product was purified in reverse phase (column: XBridge BEH Shield RP m,30 mm. Times.150 mm; mobile phase A: water (10 mmol/L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60ml/min; gradient: from 36% B to 54% B over 10 min; wavelength: 254nm/220 nm; RT1 (min): 6.18) to give N- [2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] -5-methyl-3H-1, 2, 3-triazole-4-carboxamide (30 mg, yield 24.1%).

LCMS(ESI):[M+H]⁺=558.3

4.

The crude product (30 mg) was purified by Chiral-HPLC under conditions of CHIRALPAK IH x 25cm,5 μm, mobile phase a: CO ₂, mobile phase B: methanol (0.1% diethylamine), flow rate 85mL/min, gradient: isocratic 35% B, column temperature (°c): 25 ℃, back pressure (bar): 100, wavelength: 254nm, rt1 (min): 2.85, rt2 (min): 4.5 sample solvent: methanol: dichloromethane = 1:1-HPLC, sample loading 1 mL) to give the product.

Compound 12-1 (isomer 1, R1 (min): 2.85,1.6mg, yield 5.14%)

LCMS(ESI,m/z):[M+H]⁺=558.35

¹H NMR(400MHz,Methanol-d₄)δ7.17–7.10(m,2H),6.93(s,1H),6.65(s,1H),5.43(s,1H),4.78–4.69(m,1H),4.20–4.04(m,2H),3.97–3.77(m,5H),3.67(s,3H),2.98(t,J＝6.5Hz,2H),2.59–2.52(m,3H),2.38(s,3H),2.27(s,3H),2.13(s,3H),1.11(d,J＝6.9Hz,3H).

Compound 12-2 (isomer 2, R2 (min): 4.5,3.2mg, yield 9.86%)

LCMS(ESI,m/z):[M+H]⁺=558.46

Example 13, compound 13-1 and Compound 13-2

The preparation method comprises the following steps:

1.

2- [ (1-Aminopropan-2-yl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (100 mg,0.2mmol,1.0 eq), 3H-imidazole-4-carboxylic acid (25 mg,0.2mmol,1.0 eq), HOBT (45 mg,0.3mmol,1.5 eq), EDCI (64 mg,0.3mmol,1.5 eq) were dissolved in DMF (5 mL) and stirred under nitrogen at room temperature for 2H. The reaction was quenched with water (10 mL) at room temperature. The aqueous layer was extracted with ethyl acetate (3×20 mL), and the organic layer was washed with saturated brine (3×50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. Reverse phase purification was performed using a chromatographic column, C18 silica gel, mobile phase, acetonitrile water (0.1% FA), gradient from 10% to 50% in 10 min, detector, UV 254nm. Purification gave N- [2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] -3H-imidazole-4-carboxamide (20 mg, yield 15.7%).

2.

The crude product (20 mg) was purified by Chiral-HPLC: CHIRALPAKIH x 25cm,5 μm, mobile phase a: CO ₂, mobile phase B: meOH: dcm=2:1 (0.1% 2m NH ₃ -MeOH), flow rate: 100mL/min, gradient: isocratic 18% B, column temperature (°c): 20 ℃, column head temperature (°c): 20 ℃ 100mL/min, gradient: isocratic 18% B, column temperature (°c): 35, back pressure (bar) 100, wavelength: 220nm, rt1 (min): 14.2, rt2 (min): 21.37;Sample Solvent:DCM —hplc, sample loading 2mL, to give the product.

Compound 13-1 (isomer 1, R1 (min): 14.2,2.04mg, 1.6% yield)

LCMS(ESI):[M+H]⁺=543.20

¹H NMR(400MHz,Methanol-d₄)δ7.75–7.70(m,2H),7.15–7.11(m,2H),6.98–6.92(m,1H),6.65(s,1H),5.43(s,1H),4.75–4.55(m,1H),4.15–4.09(m,2H),3.90(s,3H),3.85–3.80(m,2H),3.67(s,3H),2.98(t,J＝6.4Hz,2H),2.38(s,3H),2.33–2.27(m,3H),2.15(s,3H),1.16(d,J＝6.9Hz,3H).

Compound 13-2 (isomer 2, R2 (min): 21.37,2.72mg, 13.0% yield)

LCMS(ESI):[M+H]⁺=543.20

Example 14, compound 14

The preparation method comprises the following steps:

1.

2- [ (1-aminopropan-2-yl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (250 mg,0.6mmol,1.0 eq.), DCM (5 mL), DIEA (216 mg,1.7mmol,3.0 eq.), cyanoacetic acid (57 mg,0.7mmol,1.2 eq.) and HATU (318 mg,0.8mmol,1.5 eq.) were added sequentially to a single-port flask and reacted overnight at room temperature, the solvent was evaporated, and the reverse phase (column, C18 SILICA GEL; mobile phase, acetonitrile water (10 mmol/L NH ₄HCO₃), gradient 10% to 50% for 30 min; detection wavelength 254 nm) was purified to give 2-cyano-N- [2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl (2, 4, 6-trimethylphenyl) propyl ] acetamide (200 mg, 6%) in 200% yield.

LCMS(ESI,m/z):[M+H]⁺=516.2

2.

In a single vial was added 2-cyano-N- [2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] acetamide (200 mg,0.4mmol,1.0 eq.), 1.4-dioxane (30 mL), sodium acetate (95 mg,1.2mmol,3.0 eq.) and acetic anhydride (48 mg,0.5mmol,1.2 eq.) in sequence, and reacted overnight at 100 ℃. The solvent was dried and reversed phase (column: XBridge BEH Shield RP m,30 mm. Times.150 mm; mobile phase A: water (10 mmol/L NH ₄HCO₃), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: 21% B to 44% B9 min; wavelength: 254nm/220nm; RT1 (min): 6.82) to give (2Z) -2-cyano-N- [2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] -3-hydroxy-2-enamide (13.27 mg,6.1% yield).

LCMS(ESI,m/z):[M+H]⁺=558.25

¹H NMR(400MHz,Methanol-d4)δ7.15–7.05(m,2H),7.07–6.88(m,2H),6.63(s,1H),5.40(s,1H),4.68–4.59(m,1H),4.21–4.12(m,2H),4.11–4.05(m,1H),3.89(s,3H),3.66(s,3H),2.98(t,J＝6.5Hz,2H),2.37(s,3H),2.33(s,1H),2.25–2.23(m,6H),2.14(s,3H),1.09(d,J＝6.7Hz,3H).

Example 15, compound 15-1 and Compound 15-2

The preparation method comprises the following steps:

1.

Tert-butyl N- (2-hydroxybutyl) carbamate (500 mg,2.6mmol,1.0 eq) and TEA (1.1 mL,7.9mmol,3.0 eq) were dissolved in DCM (5 mL) under air at 0℃and TsCl (1.2 g,3.9mmol,1.5 eq) and DMAP (32 mg,0.3mmol,0.1 eq) were added, respectively. The resulting mixture was stirred under nitrogen at room temperature for 1.5 hours. The reaction was quenched with water at room temperature. The resulting mixture was extracted with dichloromethane (2×50 mL). The combined organic layers were washed with saturated brine (2×100 mL) and dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by reverse phase chromatography on a column, C18 silica gel, mobile phase, ACN water (10 mmol/L NH ₄HCO₃), gradient from 10% to 50% in 10min, detector, UV 254nm, to give tert-butyl N- {2- [ (4-methylbenzenesulfonyl) oxy ] butyl } carbamate (700 mg, 69% yield).

LCMS(ESI):[M+H]⁺=361.2

2.

To N, N-dimethylformamide (10 mL) were added Xanthos (295 mg,0.5mmol,0.2 equiv.) and Pd ₂(dba)₃ (233.mg, 0.3mmol,0.1 equiv.) under an air atmosphere at room temperature, tert-butyl 9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) amino ] -6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (1.0 g,2.5mmol,1.0 equiv.) and N- {2- [ (4-methylbenzenesulfonyl) oxy ] butyl } carbamate (1.3 g,3.8mmol,1.5 equiv.) and cesium carbonate (1.7 g,5.108mmol,2.0 equiv.). The resulting mixture was stirred overnight under nitrogen at 100 ℃. The reaction was quenched with water at room temperature. The resulting mixture was extracted with ethyl acetate (2×50 mL). The combined organic layers were washed with saturated brine (2×100 mL) and dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by reverse phase chromatography on a column, C18 silica gel, mobile phase, acetonitrile water (10 mmol/L NH ₄HCO₃), gradient of 50% -60%, 10 min, detector, UV 254nm. Tert-butyl N- [2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) butyl ] carbamate (330 mg, crude product) is finally obtained.

LCMS(ESI):[M+H]⁺=563.2

3.

Tert-butyl N- [2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) butyl ] carbamate (330 mg,0.6mmol,1.0 eq.) was dissolved in1, 4-dioxane (2 mL) under nitrogen at room temperature, and a solution of 1, 4-dioxane (2 mL, 4M) in hydrochloric acid was added. The resulting mixture was stirred under nitrogen at room temperature for 1.5 hours. The reaction was quenched with water at room temperature. The mixture was neutralized to ph=9 with saturated sodium carbonate solution. The resulting mixture was extracted with ethyl acetate (2×50 mL). The combined organic layers were washed with saturated brine (2×100 mL) and dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by Prep-TLC (DCM/meoh=20/1) to give 2- [ (1-aminobut-2-yl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6 h,7 h-pyrimido [4,3-a ] isoquinolin-4-one (150 mg, yield 49%).

LCMS(ESI):[M+H]⁺=463.2

4.

2- [ (1-Aminobutan-2-yl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (100 mg,0.2mmol,1 eq.) and triethylamine (65.mg, 0.6mmol,3.0 eq.) are dissolved in DCM (1 mL) and isocyano-trimethylsilane (37 mg,0.3mmol,1.5 eq.) is added under air at 0 ℃. The resulting mixture was stirred under nitrogen at room temperature for 2 hours. The resulting mixture was concentrated under reduced pressure. The crude product (50 mg) was purified by prep. liquid chromatography under conditions (column: XB ridge BEH C185, 19X 250mm; mobile phase A: water (10 mmol/L NH ₄HCO₃), mobile phase B: acetonitrile; flow rate: 25mL/min; gradient: from 41% B to 48% B over 10 min; wavelength: 254nm/220nm; RT (min): 7.25/8.32) to give the product.

Compound 15-1 (isomer 1, RT (min): 7.25,4.02mg, yield 4%)

LCMS(ESI):[M+H]+=506.25

¹H NMR(400MHz,Methanol-d4)δ7.15–7.06(m,2H),6.93(s,1H),6.63(s,1H),5.41(s,1H),4.21–4.02(m,3H),3.89(s,3H),3.66(s,4H),2.97(t,J＝8.0.Hz,2H),2.37(s,3H),2.21(s,3H),2.11(s,3H),1.66–1.52(m,2H),0.98(t,J＝7.4Hz,3H).

Compound 15-2 (isomer 2, RT (min): 8.32,4.02mg, yield 4%)

LCMS(ESI):[M+H]+=506.25

EXAMPLE 16 Compound 16

Preparation steps

1.

To a solution of 2- [ (1-aminopropan-2-yl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (200 mg,0.4mmol,1.0 eq.) and triethylamine (135 mg,1.3mmol,3.0 eq.) in methylene chloride (5 mL) at room temperature was added trimethylsilyl isocyanate (77 mg,0.7mmol,1.5 eq.). The resulting reaction mixture was stirred at room temperature for 2 hours. The reaction mixture was extracted with dichloromethane (3×20 mL). The organic phases were combined, backwashed with saturated brine and dried over anhydrous sodium sulfate. After the resulting mixture was filtered, the filtrate was concentrated under reduced pressure to give 2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propylurea (200 mg, crude product).

LCMS(ESI):[M+H]⁺=492.3

2.

To an 8mL sample bottle was added 2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propylurea (200 mg,0.4mmol,1.0 eq.), acetic acid (1 mL), acetic anhydride (1 mL), and hydrobromic acid (1 mL) at room temperature. The resulting reaction mixture was stirred at 120 ℃ overnight. The reaction mixture was basified with saturated aqueous sodium bicarbonate to ph=6. The reaction mixture was extracted with dichloromethane and methanol (3×50 mL). The combined organic phases were backwashed with saturated brine (3×50 mL) and dried over anhydrous sodium sulfate. After the resulting mixture was filtered, the filtrate was concentrated under reduced pressure to give 2- ({ 9, 10-dihydroxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propylurea (180 mg, yield 95.4%).

LCMS(ESI):[M+H]⁺=464.2

3.

To a solution of 2- ({ 9, 10-dihydroxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propylurea (150 mg,0.3mmol,1.0 eq.) in N, N-dimethylformamide (9 mL) was added potassium carbonate (268 mg,1.8mmol,6.0 eq.) and deuterated iodomethane (234 mg,1.5mmol,5.0 eq.). The resulting reaction mixture was stirred at room temperature overnight. The crude product obtained was purified by reverse phase Column chromatography to give 2- { [9, 10-bis (2H 3) methoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl ] (2, 4, 6-trimethylphenyl) amino } propanurea (25.92 mg, 15.52% yield) under the conditions of (Column: xbridge BEH Shield RP μm,30 x 150mm; mobile phase A: water (10 mmol/L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: 35%B to45%B in 7min; wavelength: 254nm/220nm; RT1 (min): 6.23).

LCMS(ESI):[M+H]⁺=498.30

¹H NMR(400MHz,Methanol-d₄)δ7.11(d,J＝10.6Hz,2H),6.93(s,1H),6.63(s,1H),5.40(s,1H),4.60(s,2H),4.49(s,1H),4.20–4.01(m,2H),2.97(t,J＝6.5Hz,2H),2.37(s,3H),2.22(s,3H),2.09(s,3H),1.04–0.96(m,3H).

Example 17, compound 17

To a solution of 2- [ (1-aminopropan-2-yl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6 h,7 h-pyrimido [4,3-a ] isoquinolin-4-one (60 mg,0.18mmol,1.0 eq.) in DCM (2 mL) was added triethylamine (41 mg,0.40mmol,3.0 eq.) and dimethyl dicarbonate (36 mg,0.27mmol,2.0 eq.) after stirring for two hours, the mixture was concentrated by rotary evaporation under reduced pressure. The crude product (80 mg) was purified by Prep-TLC normal phase, EA/pe=1/3 to give the product methyl N- [2- ({ 9, 10-dimethoxy-4-oxo-6 h,7 h-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] carbamate (23.01 mg, yield 32.94%).

LCMS(ESI):[M+H]⁺=507.25

¹H NMR(400MHz,Methanol-d₄)δ7.09(m,2H),6.90(s,1H),6.61(s,1H),5.38(s,1H),4.48–4.42(m,1H),4.20–4.02(m,2H),3.86(s,3H),3.64(s,6H),3.60–3.53(m,1H),3.48–3.38(m,1H),2.94(t,J＝6.5Hz,2H),2.35(s,3H),2.19(s,3H),2.09(s,3H),1.06(d,J＝6.9Hz,3H).

Example 18, compound 18

To a solution of benzoic acid (76 mg,0.62mmol,3.5 eq) in DMF (1 mL) was added 2- [ (1-aminoprop-2-yl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (80 mg,0.18mmol,1.0 eq.) 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (103 mg,0.53mmol,3.0 eq.) 1-hydroxybenzotriazole (72 mg,0.53mmol,3.0 eq.). The mixture was stirred for 2h. The mixture was concentrated under reduced pressure, and the crude product (100 mg) was purified by Prep-TLC, EA/pe=1/5 to give N- [2- ({ 9, 10-dimethoxy-4-oxo-6 h,7 h-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] benzamide) (18.91 mg, yield 19.11%).

LC-MS(ESI):[M+H]⁺=553.3

¹H NMR(400MHz,Methanol-d₄)δ7.92–7.87(m,2H),7.59–7.45(m,3H),7.17–7.08(m,2H),6.91(s,1H),6.63(s,1H),5.42(s,1H),4.69–4.60(m,1H),4.19–4.02(m,2H),3.93–3.82(m,4H),3.79–3.72(m,1H),3.64(s,3H),2.96(t,J＝6.5Hz,2H),2.36(s,3H),2.25(s,3H),2.11(s,3H),1.12(d,J＝7.0Hz,3H).

Example 19, compound 19

To a solution of 2-picolinic acid (41 mg,0.34mmol,1.0 eq) in DMF (1 mL) was added 2- [ (1-aminoprop-2-yl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (100 mg,0.22mmol,1.0 eq), 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (64 mg,0.34mmol,1.5 eq), 1-hydroxybenzotriazole (45 mg,0.34mmol,1.5 eq). The mixture was stirred for 2h. The mixture was concentrated under reduced pressure, and the crude product (100 mg) was purified by Prep-HPLC to give N- [2- ({ 9, 10-dimethoxy-4-oxo-6 h,7 h-pyrimido [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] pyridine-2-carboxamide (17.35 mg, yield 13.93%).

LCMS(ESI):[M+H]⁺=554.25

¹H NMR(400MHz,Methanol-d₄)δ8.69–8.60(m,1H),8.06(d,J＝7.7Hz,1H),8.02–7.90(m,1H),7.58–7.50(m,1H),7.15–7.09(m,2H),6.91(s,1H),6.62(s,1H),5.40(s,1H),4.83–4.76(m,1H),4.18–4.04(m,2H),3.86(s,3H),3.85(d,J＝6.4Hz,2H),3.64(s,3H),2.95(t,J＝6.5Hz,2H),2.36(s,3H),2.25(s,3H),2.12(s,3H),1.09(d,J＝6.9Hz,3H).

Example 20, compound 20

9, 10-Dimethoxy-2- [ (1-amino-2-propyl) (2, 4, 6-trimethylphenyl) amino ] -6h,7 h-pyrimidine [4,3-a ] isoquinoline-4-1 (70 mg,0.2mmol,1.0 eq.) and pyrimidine-4-carboxylic acid (58 mg,0.5mmol,3.0 eq.) are dissolved in DMF (2 mL). HOBT (63 mg,0.5mmol,3.0 eq.) and EDCI (90 mg,0.5mmol,3.0 eq.) were added to the above mixture at room temperature and stirred for 2h at room temperature. The crude product was purified by Prep-HPLC under the following conditions (column: XB ridge BEH C18. Mu.m, 30 x 150mm; mobile phase A: water (10 mmol/L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: rising from 40% B to 47% B over 7 min; wavelength: 254nm/220nm; RT1 (min): 6.45) to give N- [2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimidine [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] pyrimidine-4-carboxamide (19.00 mg, 21.34% yield).

LCMS(ESI):[M+H]⁺=555.35

¹H NMR(400MHz,Methanol-d₄)δ9.32(s,1H),9.03(d,J＝4.8Hz,1H),8.16–8.02(m,1H),7.18–7.10(m,2H),6.94(s,1H),6.64(s,1H),5.43(s,1H),4.60(s,1H),4.23–4.08(m,2H),3.89(d,J＝6.5Hz,5H),3.67(s,3H),2.98(t,J＝6.5Hz,2H),2.38(s,3H),2.27(s,3H),2.14(s,3H),1.23–0.98(m,3H).

Example 21, compound 21

9, 10-Dimethoxy-2- [ (1-amino-2-propyl) (2, 4, 6-trimethylphenyl) amino ] -6h,7 h-pyrimidine [4,3-a ] isoquinoline-4-1 (50 mg,0.1mmol,1.0 eq.) was dissolved in DCM (3 mL). To the above mixture were added isocyanatocyclopropane (14 mg,0.2mmol,1.5 eq.) and triethylamine (34 mg,0.3mmol,3.0 eq.) at room temperature, and stirred at room temperature for 2h. The crude product was purified by Prep-HPLC under the following conditions (column Xbridge BEH Shield RP, 5 μm, 19X 250mm; mobile phase A: water (10 mmol/L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: increasing from 36% B to 50% B in 7 min; wavelength: 254nm/220nm; RT1 (min): 6.05/6.5) to give 3-cyclopropyl-1- [2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimidine [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] urea (17.19 mg, yield 28.92%).

LCMS(ESI):[M+H]⁺=532.30

¹H NMR(400MHz,Methanol-d₄)δ7.15–7.05(m,2H),6.93(s,1H),6.63(s,1H),5.40(s,1H),4.57–4.49(m,1H),4.17–4.04(m,2H),3.88(s,3H),3.75–3.60(m,4H),3.55–3.45(m,1H),2.96(t,J＝6.5Hz,2H),2.57–2.48(m,1H),2.37(s,3H),2.23(s,3H),2.11(s,3H),1.09(d,J＝6.9Hz,3H),0.79–0.67(m,2H),0.57–0.46(m,2H).

Example 22, compound 22

9, 10-Dimethoxy-2- [ (1-amino-2-propyl) (2, 4, 6-trimethylphenyl) amino ] -6h,7 h-pyrimidine [4,3-a ] isoquinoline-4-1 (70 mg,0.2mmol,1.0 eq.) and triethylamine (47 mg,0.5mmol,3.0 eq.) were dissolved in solvent DCM (3 mL) at room temperature. N-methylcarbamoyl chloride (22 mg,0.2mmol,1.5 eq.) was added to the above mixture at 0℃and stirred at room temperature for 2h. The crude product was purified by Prep-HPLC under the following conditions (column: XB ridge BEH C18 μm,30 x 150mm; mobile phase A: water (10 mmol/L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: rising from 36% B to 48% B over 7 min; wavelength: 254nm/220nm; RT1 (min): 6.37) to give 1- [2- ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimidine [4,3-a ] isoquinolin-2-yl } (2, 4, 6-trimethylphenyl) amino) propyl ] -3-methylurea (16.82 mg, 21.25% yield).

LCMS(ESI):[M+H]⁺=506.30

¹H NMR(400MHz,Methanol-d₄)δ7.16–7.07(m,2H),6.93(s,1H),6.63(s,1H),5.39(s,1H),4.60(s,1H),4.46(t,J＝7.6Hz,1H),4.21–4.00(m,2H),3.95–3.85(m,4H),3.66(s,3H),3.05–2.80(m,5H),2.37(s,3H),2.21(s,3H),2.08(s,3H),1.02(d,J＝6.9Hz,3H).

Example 23, compound 23-1 and Compound 23-2

The preparation method comprises the following steps:

1.

4-bromo-2, 6-diisopropylaniline (5 g,20mmol,1.0 eq.) cesium carbonate (20.0 g,60mmol,3 eq.) a [1,1' -bis (diphenylphosphino) ferrocene ] palladium dichloride dichloromethane complex (1.6 g,2mmol,0.1 eq.) methylboronic acid (5.8 g,98mmol,5 eq.) and 2-dicyclohexylphosphine-2 ',6' -dimethoxybiphenyl (1.6 g,4mmol,0.2 eq.) were dissolved with 1, 4-dioxane (90 mL) and water (15 mL) and nitrogen-protected, 100 ℃ stirred overnight. Cooled to room temperature, dissolved in 100mL of water, extracted with ethyl acetate (3×100 mL), and the organic layer was washed with 100mL of saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by normal phase column chromatography under the following conditions (mobile phase A: dichloromethane, mobile phase B: methanol; flow rate: 100mL/min; gradient: 0%B to 10%B in 30min; wavelength: 254 nm) to give 2, 6-diisopropyl-4-methylaniline (2.1 g, yield 56.2%).

LCMS(ESI):[M+H]⁺=192.17

2.

A solution of 2, 6-diisopropyl-4-methylaniline (1 g,5.2mmol,1 eq.) and 2-chloro-9, 10-dimethoxy-6, 7-dihydro-4H-pyrimidin-6, 1-A isoquinolin-4-one (1.68 g,5.8mmol,1.1 eq.) in isopropanol (2 mL) was stirred overnight under nitrogen at 90 ℃. The resulting reaction solution was concentrated under reduced pressure, filtered, and the filter cake was collected and washed with ethyl acetate (2×10 ml) to give 2- [ (2, 6-diisopropyl-4-methylphenyl) amino ] -9, 10-dimethoxy-6 h,7 h-pyrimido [4,3-a ] isoquinolin-4-one (1.0 g, yield 42.7%).

LCMS(ESI):[M+H]⁺=448.25

3.

2- [ (2, 6-Diisopropyl-4-methylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (700 mg,1.6mmol,1 eq.) was dissolved in cesium carbonate (1.6 g,4.8mmol,3 eq.) and tert-butyl N- (2-bromopropyl) carbamate (560 mg,2.3mmol,1.5 eq.) with 1,4 dioxane (10 mL), nitrogen blanketed, and stirred overnight at 100 ℃. Cooled to room temperature, dissolved in 20mL of water, extracted with ethyl acetate (3X 10 mL), and the organic layer was washed with 20mL of saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reverse phase column chromatography under the following conditions (column: C18; mobile phase A: water (10 mmol/L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: 50% B to 80% B in 20min; wavelength: 254 nm) to give tert-butyl N- {2- [ (2, 6-diisopropyl-4-methylphenyl) ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl }) amino ] propyl } carbamate (600 mg, yield 63.4%).

LCMS(ESI):[M+H]⁺=605.35

4.

Tert-butyl N- {2- [ (2, 6-diisopropyl-4-methylphenyl) ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl }) amino ] propyl } carbamate (500 mg,0.91mmol,1 eq.) was dissolved in 10mL of 1,4 dioxane solution (4M) of hydrochloric acid and stirred at room temperature for 1 hour. Concentrated under reduced pressure, dissolved with 10mL of water, extracted with ethyl acetate (3X 10 mL), the organic layer was washed with 20mL of saturated sodium chloride, dried over anhydrous Na ₂SO₄, filtered, and concentrated under reduced pressure to give 2- ((1-aminopropan-2-yl) (2, 6-diisopropyl-4-methylphenyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (300 mg, 73.3% yield).

LCMS(ESI):[M+H]⁺=505.30

5.

2- ((1-Aminopropan-2-yl) (2, 6-diisopropyl-4-methylphenyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (300 mg,0.59mmol,1.0 eq.) triethylamine (180 mg,1.8mmol,3 eq.) and trimethylsilane isocyanate (103 mg,1.2mmol,2 eq.) were dissolved in 10mL DCM and stirred at room temperature for 2 hours. Concentrated under reduced pressure, dissolved in 10mL of water, extracted with ethyl acetate (3X 10 mL), the organic layer was washed with 20mL of saturated sodium chloride, dried over anhydrous Na ₂SO₄, filtered, and concentrated under reduced pressure. The crude product was purified by reverse phase column chromatography under the following conditions (column: XBIdge BEH C18. Mu.m, 30X 150mm; mobile phase A: water (10 mmol/L ammonium bicarbonate) mobile phase B: acetonitrile; flow rate: 60mL/min; gradient: 43% B to 55% B,8 min; wavelength: 254/220nm; RT1 (min): 5.77) to give 1- (2- ((2, 6-diisopropyl-4-methylphenyl) (9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-2-yl) amino) propyl) urea (150 mg, yield 45.0%).

6.

The racemate (150 mg) was purified by Chiral-SFC under the following conditions (column: CHIRALPAK IH,2 x 25cm,5 μm; mobile phase A: methanol: dichloromethane=1:1- -HPLC; mobile phase B: methyl tertiary ether (0.1% ethylenediamine) -HPLC; flow rate: 20mL/min; gradient: isocratic; column temperature (°C): 35; pressure (bar): 100; wavelength: 220nm; RT1 (min): 10; RT2 (min): 5; sample solvent: methanol- -HPLC; sample injection amount: 1 mL) to give the product.

Compound 23-1 (isomer 1, RT1 (min): 10,16.5mg, yield 6.4%).

LCMS(ESI):[M+H]⁺=548.30

¹H NMR(400MHz,Methanol-d₄)δ7.25–7.18(m,2H),6.94(s,1H),6.61(s,1H),5.45(s,1H),4.48(s,1H),4.15–4.08(m,2H),3.89(s,3H),3.63(s,3H),3.11–2.93(m,4H),2.75(s,1H),2.44(s,3H),1.36–1.25(m,8H),1.17(d,J＝6.8Hz,3H),1.02(d,J＝6.6Hz,4H).

Compound 23-2 (isomer 2, R2 (min): 5,16.42mg, yield 6.4%).

LCMS(ESI):[M+H]⁺=548.30

¹H NMR(400MHz,Methanol-d₄)δ7.26–7.18(m,2H),6.94(s,1H),6.61(s,1H),5.45(s,1H),4.48(s,1H),4.16–4.08(m,2H),3.89(s,3H),3.63(s,3H),3.11–2.93(m,3H),2.75(s,1H),2.44(s,3H),1.35–1.28(m,7H),1.17(d,J＝6.8Hz,3H),1.02(d,J＝6.6Hz,3H).

Example 24, compound 24-1 and Compound 24-2

The preparation method comprises the following steps:

1.

1, 3-difluoro-5-methyl-2-nitrobenzene (200 mg,1.1mmol,1 eq.) was dissolved in MeOH (5 mL) at room temperature in air, followed by the addition of sodium methoxide in methanol (3 mL, 60%). The reaction was carried out at room temperature overnight. After evaporation to dryness under reduced pressure, 50mL of water was added, 2X50mL of ethyl acetate was extracted, and the combined organic layers were washed with saturated brine (2X 100 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give 1, 3-dimethoxy-5-methyl-2-nitrobenzene (200 mg, crude product).

2.

1, 3-Dimethoxy-5-methyl-2-nitrobenzene (190 mg,0.9mmol,1 eq.) and H ₂ O (1 mL) were added to a solution of EtOH (5 mL) in air at room temperature, followed by NH ₄ Cl (154.mg, 3.mmol,3 eq.) and Fe (8.5 mg,0.2mmol,3 eq.). The reaction was allowed to react at 80 ℃ for two hours, the resulting mixture was filtered and the filter cake was washed with ethyl acetate (2 x30 mL). The reaction was quenched with water at room temperature. The resulting mixture was extracted with ethyl acetate (2×100 mL). The combined organic layers were washed with saturated sodium chloride (2×150 mL) and dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated under reduced pressure to give 2, 6-dimethoxy-4-methylaniline (150 mg, crude).

LCMS(ESI):[M+H]⁺=168.2

3.

2-Chloro-9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (273.mg, 0.9mmol,1.2 eq.) is dissolved in a solution of isopropanol (3 ml) in air at room temperature, 2, 6-dimethoxy-4-methylaniline (152.mg, 0.9mmol,1.2 eq.) is added. The resulting mixture was concentrated under reduced pressure to give 2- [ (2, 6-dimethoxy-4-methylphenyl) amino ] -9, 10-dimethoxy-6 h,7 h-pyrimido [4,3-a ] isoquinolin-4-one (300 mg, crude product).

LCMS(ESI):[M+H]⁺=424.2

4.

2- [ (2, 6-Dimethoxy-4-methylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (300 mg,0.7mmol,1 eq.) and tert-butyl N- (2-bromopropyl) carbamate (168.mg, 0.7mmol,1 eq.) and Cs ₂CO₃ (4635 mg,1.4mmol,2 eq.) are dissolved in a solution of 1, 4-dioxane (5 ml), xantPhos (81.mg, 0.1mmol,0.1 eq.) and Pd ₂(dba)₃ (64.mg, 0.1mmol,0.1 eq.) are added and the resulting mixture is stirred overnight under a nitrogen atmosphere at 100 ℃. The filtrate was concentrated under reduced pressure and the residue was purified by reverse phase flash chromatography on a chromatographic column, C18 silica gel, mobile phase, acetonitrile in water (10 mmol/L NH ₄HCO₃), gradient from 10% to 50% over 10min, detector, UV 254nm. Tert-butyl N- {2- [ (2, 6-dimethoxy-4-methylphenyl) ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl }) amino ] propyl } carbamate (270 mg,59% yield) was finally obtained.

LCMS(ESI):[M+H]⁺=581.2

5.

Tert-butyl N- {2- [ (2, 6-dimethoxy-4-methylphenyl) ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimido [4,3-a ] isoquinolin-2-yl }) amino ] propyl } carbamate (300 mg,0.6mmol,1.0 eq.) was dissolved in 1, 4-dioxane (2 mL), and a solution of 1, 4-dioxane (2 mL) in hydrochloric acid was added under nitrogen at room temperature. The resulting mixture was stirred under nitrogen at room temperature for 1.5 hours. The reaction was quenched with water at room temperature. The mixture was neutralized to ph=9 with saturated sodium carbonate solution. The resulting mixture was extracted with ethyl acetate (2×50 mL). The combined organic layers were washed with saturated brine (2×100 mL) and dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by Prep-TLC (CH ₂Cl₂/meoh=20/1) to give 2- [ (1-aminopropan-2-yl) (2, 6-dimethoxy-4-methylphenyl) amino ] -9, 10-dimethoxy-6 h,7 h-pyrimido [4,3-a ] isoquinolin-4-one (150 mg,49% yield).

LCMS(ESI):[M+H]⁺=481.2

6.

2- [ (1-Aminopropan-2-yl) (2, 6-dimethoxy-4-methylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (400 mg,0.8mmol,1 eq.) and TEA (0.4 mL,2.5mmol,3 eq.) were added to a solution of DCM (4 mL) and isocyano-trimethylsilane (191 mg,1.6mmol,2.0 eq.) was added and the resulting mixture stirred under nitrogen at room temperature for 2 hours and the reaction mixture concentrated under reduced pressure. The crude product (420 mg) was purified by high performance liquid chromatography under the following conditions (column: sunfire C18 5m,30 mm. Times.4.5 mm) mobile phase A: water (0.1% FA), mobile phase B: acetonitrile, flow rate: 60 mL/min, gradient: from 51% B to 74% B in 10min, wavelength: 254 nm/220 nm, RT1 (min): 6.03; 6.62), 1- (2- ((2, 6-dimethoxy-4-methylphenyl) (9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-2-yl) amino) propyl) urea (130 mg) was obtained, followed by purification by Chiral-HPLC under the following conditions (column: JW-CHIRALPAK IG-3,3.0 MeOH: 50mm;3um; mobile phase A: DCM=1:1- -HPLC, mobile phase B: mtBE (0.1% MeOH) - -DEA; 20 mL; 20 min; gradient: 5.2 mL; solvent: 220 mg; 5.2 mL: 5 min, etc., gradient: 2.1 mL: 5 mL; run times: 5 min).

Compound 24-1 (isomer 1, RT1 (min): 12,30.12mg,6.8% yield)

LCMS(ESI):[M+H]⁺=524.25

¹H NMR(400MHz,Methanol-d₄)δ6.92(s,1H),6.72–6.66(m,3H),5.53(s,1H),4.11–4.05(m,2H),3.89(s,3H),3.81(d,J＝10.1Hz,6H),3.69(s,3H),2.95(t,J＝6.5Hz,2H),2.47(s,3H).

Compound 24-2 (isomer 2, R2 (min) 16.5,44.03mg,10% yield)

LCMS(ESI):[M+H]⁺=524.25

Example 25, compound 25-1 and Compound 25-2

The preparation method comprises the following steps:

1.

A mixture of 2, 6-difluoro-4-methylaniline (1.0 g,6.9mmol,1 eq) and IPA (40 mL) of 2-chloro-9, 10-dimethoxy-6H, 7H-pyrimidine [4,3-a ] isoquinoline-4-1 (2.6 g,9.0mmol,1.3 eq) was stirred overnight under nitrogen at 90 ℃. The resulting mixture was concentrated under reduced pressure. Basified with saturated NaHCO ₃ to ph=8. The resulting mixture was extracted with EtOEt (3X 50 mL). The combined organic layers were washed with brine (2×100 mL) and dried over anhydrous Na ₂SO₄. The filtrate after filtration was concentrated under reduced pressure. Purification by silica gel column chromatography eluting with CH ₂Cl₂/MeOH (80/1-60/1) afforded 2- [ (2, 6-difluoro-4-methylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimidine [4,3-a ] isoquinoline-4-1 (2.0 g, 71.6%).

LCMS(ESI):[M+H]⁺=400.15

2.

2- [ (2, 6-Difluoro-4-methylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimidine [4,3-a ] isoquinolin-4-one (440 mg,1.1mmol,1 eq.) and tert-butyl N- (2-bromopropyl) carbamate (288.5 mg,1.2mmol,1.1 eq.) were stirred in DMF (15 mL) under nitrogen at 100deg.C overnight. The resulting mixture was concentrated under vacuum. Purification by column chromatography on silica gel eluting with CH ₂Cl₂/MeOH (10/1) gave N- {2- [ (2, 6-difluoro-4-methylphenyl) ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimidine [4,3-a ] isoquinolin-2-yl }) amino ] propyl carbamate tert-butyl (200 mg, 32.6%).

LCMS(ESI):[M+H]⁺=557.25

3.

N- {2- [ (2, 6-difluoro-4-methylphenyl) ({ 9, 10-dimethoxy-4-oxy-6H, 7H-pyrimidine [4,3-A ] isoquinolin-2-yl }) amino ] propyl } carbamate (200 mg,0.359mmol,1 equivalent) was stirred in 1, 4-dioxane (15 mL) under nitrogen at room temperature for 3h. The resulting mixture was concentrated under reduced pressure. As a result, 2- [ (1-aminopropen-2-yl) (2, 6-difluoro-4-methylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimidine [4,3-a ] isoquinolin-4-1 (150 mg, 91.45%). The crude product was used directly in the next step without further purification.

LCMS(ESI):[M+H]⁺=457.2

4.

2- [ (1-Aminopropen-2-yl) (2, 6-difluoro-4-methylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimidine [4,3-a ] isoquinolin-4-1 (50 mg,0.11mmol,1.0 eq.) and trimethylsilane isocyanate (12.6 mg,0.11mmol,1.0 eq.) are stirred in DCM (5 mL) at room temperature under nitrogen overnight. The resulting mixture was concentrated under reduced pressure. The crude product (60 mg) was purified by Prep-HPLC under the following conditions (column: XSelect CSH Fluoro Phenyl m,19mm x 250mm; mobile phase A: water (0.1% FA), mobile phase B: ACN; flow: 25mL/min; gradient: 42% B to 65% B,10 min; wavelength: 254nm/220nm; RT1 (min): 5.67) to give (2R) -2- [ (2, 6-difluoro-4-methylphenyl) ({ 9, 10-dimethoxy-4-oxo-6H, 7H-pyrimidine [4,3-a ] isoquinolin-2-yl }) amino ] propylurea (50 mg). Purification was then carried out by Chiral-HPLC under the following conditions (column: CHIRALPAK IH 2.25 cm,5 μm; mobile phase A: meOH: DCM=1:1-HPLC, mobile phase B: mtBE (0.1% DEA) -HPLC; flow: 20mL/min; gradient: equidistant; wavelength: 220nm; RT1 (min): 11.2; RT2 (min): 15.0; sample solvent: meOH: DCM=1:1-HPLC; injection: 1.5mL; running times: 5) to give the product.

Compound 25-1 (isomer 1, R1 (min): 11.2,13.41mg, 24.49%).

LCMS(ESI):[M+H]⁺=500.25

¹H NMR(400MHz,Methanol-d₄)δ6.90(s,1H),6.87(s,1H),6.85(s,1H),6.83(s,1H),5.70(t,J＝1.7Hz,1H),5.63(s,1H),4.00(t,J＝6.3Hz,2H),3.96–3.90(m,1H),3.88(s,3H),3.73(s,3H),3.65–3.56(m,1H),2.95(t,J＝6.2Hz,2H),2.35(s,3H),1.54(d,J＝6.9Hz,3H).

Compound 25-2 (isomer 2, R2 (min): 15.0,15.09mg, 21.59%)

LCMS(ESI):[M+H]⁺=500.25

¹H-NMR(400MHz,Methanol-d₄)δ6.89(s,1H),6.87(s,1H),6.86–6.84(m,1H),6.84–6.81(m,1H),5.70(t,J＝1.6Hz,1H),5.63(s,1H),4.00(t,J＝6.3Hz,2H),3.96–3.90(m,1H),3.88(s,3H),3.73(s,3H),2.97–2.93(m,2H),2.35(s,3H),1.54(d,J＝6.9Hz,3H).

Example 26, compound 26

Preparation steps

1.

In a single vial were added successively (2E) -9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) imino ] -3H,6H, 7H-pyrimidine [4,3-a ] isoquinolin-4-one (1.0 g,2.6mmol,1.0 eq.), dioxane (100 mL), cesium carbonate (1.7 g,5.1mmol,2.0 eq.) and tert-butyl (N- {4- [ (4-methylbenzenesulfonyl) oxy ] cyclohexyl } carbamate (1.4 g,3.8mmol,1.5 eq.) and reacted overnight at 100 ℃. The filtrate was filtered and dried and the resulting residue was purified by reverse phase column chromatography under conditions of column specification C18, mobile phase, water and acetonitrile, 10% to 100% gradient for 30 minutes, UV220 nm detector. Tert-butyl (4- ((9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimidin [6,1-a ] isoquinolin-2-yl) (methylsulfonyl) amino) cyclohexyl) carbamate (110 mg, 7.3%) was obtained.

LCMS(ESI,m/z):[M+H]⁺=589.3

2.

In a single vial was added tert-butyl (4- ((9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimidin [6,1-a ] isoquinolin-2-yl) (methylsulfonyl) amino) cyclohexyl) carbamate (100 mg,0.2mmol,1.0 eq.) and a solution of HCl in 1, 4-dioxane (4M, 15 mL) in sequence and reacted at room temperature for 2H. The solvent was dried to 2- [ (4-aminocyclohexyl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimidine [4,3-a ] isoquinolin-4-one (80 mg, 96.4%).

LCMS(ESI,m/z):[M+H]⁺=489.3

3.

2- [ (4-Aminocyclohexyl) (2, 4, 6-trimethylphenyl) amino ] -9, 10-dimethoxy-6H, 7H-pyrimidine [4,3-a ] isoquinolin-4-one (140 mg,0.3mmol,1.0 eq.) was dissolved in DCM (8 mL). Triethylamine (145 mg,1.5mmol,5.0 eq.) and trimethylsilyl isocyanate (40 mg,0.4mmol,1.2 eq.) were added to the above mixture at room temperature, and stirred at room temperature for 3h. The crude product was purified by Prep-HPLC under the following conditions (column: xbridge BEH Shield RP, 5 μm,19 x 250mm; mobile phase a: water (10 mmol/L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 25mL/min; gradient: decrease from 39% B to 4%B in 10 min; wavelength: 254nm/220 nm; rt1 (min): 9.4) to give 4- ({ 9, 10-dimethoxy-4-oxo-6 h,7 h-pyrimidin [4,3-a ] isoquinolin-2-yl } (2, 6-dimethylphenyl) amino) cyclohexylurea (35.71 mg, yield 22.81%).

LCMS(ESI):[M+H]+=532.25

¹H NMR(400MHz,Methanol-d₄)δ7.10(s,2H),6.92(s,1H),6.62(s,1H),5.39(s,1H),4.18–4.03(m,3H),3.88(s,4H),3.66(s,3H),2.96(t,J＝6.4Hz,2H),2.36(s,3H),2.21(s,6H),2.05–1.92(m,4H),1.88–1.80(m,2H),1.75–1.62(m,2H).

Example 27, compound 27

Preparation steps

1.

In a single vial were added successively (2E) -9, 10-dimethoxy-2- [ (2, 4, 6-trimethylphenyl) imino ] -3H,6H, 7H-pyrimido [4,3-a ] isoquinolin-4-one (1.0 g,2.6mmol,1.0 eq.), 1.4-dioxane (100 mL), cesium carbonate (2.5 g,7.7mmol,3.0 eq.) and tert-butyl 4- [ (4-methylbenzenesulfonyl) oxy ] piperidine-1-carboxylate (1.0 g,2.8mmol,1.1 eq.) and reacted overnight at 100 ℃. The filtrate was filtered and concentrated and the resulting residue was purified by reverse phase column chromatography with a gradient of 10% to 100% of water and acetonitrile for 30min, UV220 nm detector to give tert-butyl 4- ((9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-2-yl) (formamidino) amino) piperidine-1-carboxylate (120 mg, yield 8.17%).

LCMS(ESI,m/z):[M+H]⁺=575.3

2.

In a single vial was added tert-butyl 4- ((9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-2-yl) (formamidino) amino) piperidine-1-carboxylate (100 mg,0.2mmol,1.0 eq.) and 1, 4-dioxane (17 mL) of hydrogen chloride in sequence and reacted at room temperature for 2H. Concentration of the solvent afforded 2- (methylsulfonyl (piperidin-4-yl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (80 mg, 96.9% yield).

LCMS(ESI,m/z):[M+H]⁺=475.3

3.

To a solution of 2- (methylsulfonyl (piperidin-4-yl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (640 mg,2.2mmol,1.0 eq.) in methylene chloride (20 mL) was added triethylamine (640 mg,6.3mmol,3.0 eq.) and trimethylsilyl isocyanate (264 mg,3.2mmol,1.5 eq.). The mixture was stirred at room temperature for 2h. After concentrating the mixture by spin evaporation under reduced pressure, the residue obtained was purified by reverse phase column chromatography under the conditions of XBridge BEH Shield RP, 5 μm, 19X 250mm, mobile phase A: water (10 mmol/L ammonium bicarbonate), mobile phase B: acetonitrile, flow rate: 25mL/min, gradient: 35%B to 40%B in 10min, wavelength: 254nm/220nm, RT1 (min): 8.57 to give 4- [ (2E) -9, 10-dimethoxy-4-oxo-2- [ (2, 4, 6-trimethylphenyl) imino ] -6H, 7H-pyrimido [4,3-a ] isoquinolin-3-yl ] piperidine-1-carboxamide) (39.74 mg, yield 3.63%).

LCMS(ESI):[M+H]⁺=518.40

¹H NMR(400MHz,Methanol-d₄)δ7.10(s,2H),6.92(s,1H),6.62(s,1H),5.39(s,1H),4.73–4.62(m,1H),4.18–3.97(m,4H),3.88(s,3H),3.66(s,3H),3.02–2.90(m,4H),2.36(s,3H),2.17(s,8H),1.43–1.29(m,2H).

EXAMPLE 28 Compound 28

The crude product (150 mg) was purified by Chiral-SFC under the following conditions (column: CHIRALPAK IH, 2X 25cm,5 μm; mobile phase A: methanol: dichloromethane=1:1- -HPLC; mobile phase B: methyl tert-ether (0.1% ethylenediamine) -HPLC; flow rate: 20mL/min; gradient: isocratic; wavelength: 220nm; RT1 (min): 8; RT2 (min): 13.0; sample solvent: methanol- -HPLC; sample volume: 1 mL) to give 1- (3- ((2, 6-diisopropyl-4-methylphenyl) (9, 10-dimethoxy-4-oxo-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-2-yl) amino) cyclobutyl) urea (13.00 mg, 14.9% yield).

LCMS(ESI):[M+H]⁺=560.30

¹H NMR(400MHz,Methanol-d₄)δ7.21(s,2H),6.93(s,1H),6.64(s,1H),5.46(s,1H),4.51–4.44(m,1H),4.14–4.06(m,2H),3.97–3.88(m,1H),3.89(s,3H),3.64(s,3H),2.94-2.90(m,4H),2.74-2.78(m,2H),2.44(s,3H),2.20–2.08(m,2H),1.30(d,J＝6.8Hz,7H),1.11(d,J＝6.8Hz,6H).

Example 29, compound 29

Compound 29 (isomer 1, R1 (min): 2.85,3.2mg, 10.3% yield)

LCMS(ESI,m/z):[M+H]⁺=558.35

Example 30, compound 30

To a 25mL single vial was added 2- ((1-aminoprop-2-yl) (methyltrimethyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (100 mg,0.22 mmol), 6-methylpyridine-2-carboxylic acid (45.3 mg,0.33 mmol), EDCI (63.3 mg,0.33 mmol), HOBT (44.6 mg,0.33 mmol) and DMF (2 mL) at room temperature, and stirred for 24H at room temperature. After the reaction, the reaction system was filtered and purified by reverse phase Flash (mobile phase A:0.5% aqueous FA solution, mobile phase B: acetonitrile; flow rate: 40mL/min; gradient: 20 minutes from 0% B to 55% B;50% B, wavelength: 254nm/220 nm;) to give the product (19.2 mg, 10.3%).

LC-MS(ESI):[M+H]⁺=568.2

¹H NMR(400MHz,CDCl₃)8.76-8.74(m,1H),7.86(d,J＝8.0Hz,1H),7.63(t,J＝12.0Hz,1H),7.19-7.16(m,1H),6.91(d,J＝8.0Hz,1H),6.62(s,1H),6.52(s,1H),5.22(s,1H),4.96-4.95(m,1H),4.16–4.10(m,2H),3.83(s,3H),3.82-3.75(m,2H),3.63(s,3H),2.84(t,J＝8.0Hz,2H),2.54(s,3H),2.26(s,3H),2.17(s,3H),2.05(s,3H),1.05(d,J＝8.0Hz,3H).

Example 31, compound 31

To a 25mL single vial was added 2- ((1-aminoprop-2-yl) (methyltrimethyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (150 mg,0.33 mmol), pyridine 5-methyl-2-carboxylate (68.6 mg,0.50 mmol), EDCI (95.9 mg,0.50 mmol), HOBT (67.6 mg,0.50 mmol) and DMF (3 mL) and stirred at room temperature for 24H. After the reaction, the reaction system was filtered and purified by reverse phase Flash (mobile phase A:0.5% aqueous FA solution, mobile phase B: acetonitrile; flow rate: 40mL/min; gradient: 20 minutes from 0% B to 55% B;50% B, wavelength: 254nm/220 nm;) to give the product (37.5 mg, 19.7%).

LC-MS(ESI):[M+H]⁺=568.2

¹H NMR(400MHz,CDCl₃)8.81(s,1H),8.35(s,1H),7.95(d,J＝8.0Hz,1H),7.54-7.51(m,1H),6.93(d,J＝8.0Hz,1H),6.62(s,1H),6.52(s,1H),5.21(s,1H),4.92-4.87(m,1H),4.17–4.07(m,1H),3.83(s,3H),3.82-3.75(m,2H),3.63(s,3H),2.84(t,J＝8.0Hz,2H),2.31(s,3H),2.25(s,3H),2.17(s,3H),2.05(s,3H),1.09(d,J＝8.0Hz,3H).

Example 32, compound 32

To a 25mL single vial was added 2- ((1-aminoprop-2-yl) (methyltrimethyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (150 mg,0.33 mmol), 4-methyl-2-pyridinecarboxylic acid (68.6 mg,0.50 mmol), EDCI (95.9 mg,0.50 mmol), HOBT (67.6 mg,0.50 mmol) and DMF (3 mL) at room temperature, and stirred for 24H at room temperature. After the reaction, the reaction system was filtered and purified by reverse phase Flash (mobile phase A:0.5% aqueous FA solution, mobile phase B: acetonitrile; flow rate: 40mL/min; gradient: 20 minutes from 0% B to 55% B;50% B, wavelength: 254nm/220 nm;) to give the product (32.3 mg, 17.3%).

LC-MS(ESI):[M+H]⁺=568.2

¹H NMR(400MHz,CDCl₃)8.87(s,1H),8.37(d,J＝8.0Hz,1H),8.21(s,1H),7.91(s,1H),7.15(d,J＝8.0Hz,1H),6.91(d,J＝12Hz,1H),6.62(s,1H),6.51(s,1H),5.23(s,1H),4.87-4.82(m,1H),4.19–4.07(m,2H),3.83(s,3H),3.82-3.75(m,2H),3.64(s,3H),2.85(t,J＝8.0Hz,2H),2.34(s,3H),2.26(s,3H),2.17(s,3H),1.98(s,3H),1.07(d,J＝4.0Hz,3H).

Example 33, compound 33

To a 25mL single vial was added 2- ((1-aminoprop-2-yl) (methyltrimethyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (150 mg,0.33 mmol), 3-methyl-2-pyridinecarboxylic acid (68.6 mg,0.50 mmol), EDCI (95.9 mg,0.50 mmol), HOBT (67.6 mg,0.50 mmol) and DMF (3 mL) at room temperature, and stirred for 24H at room temperature. After the reaction, the reaction system was filtered and purified by reverse phase Flash (mobile phase A:0.5% aqueous FA solution, mobile phase B: acetonitrile; flow rate: 40mL/min; gradient: 20 minutes from 0% B to 55% B;50% B, wavelength: 254nm/220 nm;) to give the product (12.8 mg, 6.8%).

LC-MS(ESI):[M+H]⁺=568.2

¹H NMR(400MHz,CDCl₃)8.86(s,1H),8.35(d,J＝4.0Hz,1H),7.47(d,J＝8.0Hz,1H),7.22-7.20(m,1H),6.91(d,J＝8.0Hz,1H),6.62(s,1H),6.52(s,1H),5.22(s,1H),4.96-4.91(m,1H),4.19–4.03(m,2H),3.83(s,3H),3.82-3.75(m,2H),3.63(s,3H),2.84(t,J＝8.0Hz,2H),2.64(s,3H),2.26(s,3H),2.18(s,3H),2.02(s,3H),1.08(d,J＝4.0Hz,3H).

Example 34, compound 34

To a 25mL single vial was added 2- ((1-aminoprop-2-yl) (methyltrimethyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (100 mg,0.22 mmol), pyrimidine-2-carboxylic acid (40.9 mg,0.33 mmol), EDCI (63.3 mg,0.33 mmol), HOBT (44.6 mg,0.33 mmol) and DMF (2 mL) and stirred at room temperature for 24H. After the reaction, the reaction system was filtered and purified by reverse phase Flash (mobile phase A:0.5% aqueous FA solution, mobile phase B: acetonitrile; flow rate: 40mL/min; gradient: 20 minutes from 0% B to 55% B;50% B, wavelength: 254nm/220 nm;) to give the product (20 mg, 10.9%).

LC-MS(ESI):[M+H]⁺=555.2

¹H NMR(400MHz,DMSO-d6)9.56(t,J＝8.0Hz,1H),9.03-9.00(m,2H),8.16(s,1H),7.87(s,1H),7.74-7.71(m,1H),7.12-7.10(m,2H),7.02(s,1H),4.74–4.70(m,1H),4.16(s,3H),4.15-4.10(m,2H),3.98-3.95(m,2H),3.89(s,3H),3.05-3.00(m,2H),2.31(s,3H),2.19(s,3H),2.03(s,3H),0.83(d,J＝8.0Hz,3H).

Example 35, compound 35

To a 25mL single vial was added 2- ((1-aminoprop-2-yl) (methyltrimethyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (100 mg,0.22 mmol), 5-methylpyrimidine-2-carboxylic acid (45.6 mg,0.33 mmol), EDCI (63.3 mg,0.33 mmol), HOBT (44.6 mg,0.33 mmol) and DMF (2 mL) at room temperature, and stirred for 24H at room temperature. After the reaction, the reaction system was filtered and purified by reverse phase Flash (mobile phase A:0.5% aqueous FA solution, mobile phase B: acetonitrile; flow rate: 40mL/min; gradient: 20 minutes from 0% B to 55% B;50% B, wavelength: 254nm/220 nm;) to give the product (27.6 mg, 14.7%).

LC-MS(ESI):[M+H]⁺=569.2

¹H NMR(400MHz,CDCl₃)9.28(s,1H),8.64(s,1H),6.91(d,J＝8.0Hz,1H),6.62(s,1H),6.51(s,1H),5.22(s,1H),4.93-4.90(m,1H),4.13–4.10(m,2H),3.83(s,3H),3.82-3.75(m,2H),3.63(s,3H),2.85-2.83(m,2H),2.31(s,3H),2.26(s,3H),2.16(s,3H),2.04(s,3H),1.05(d,J＝8.0Hz,3H).

Example 36, compound 36

To a 25mL single vial was added 2- ((1-aminoprop-2-yl) (methyltrimethyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (100 mg,0.22 mmol), pyrazine-2-carboxylic acid (40.9 mg,0.33 mmol), EDCI (63.3 mg,0.33 mmol), HOBT (44.6 mg,0.33 mmol) and DMF (2 mL) and the mixture was stirred at room temperature for 24H. After the reaction, the reaction system was filtered and purified by reverse phase Flash (mobile phase A:0.5% aqueous FA solution, mobile phase B: acetonitrile; flow rate: 40mL/min; gradient: 20 minutes from 0% B to 55% B;50% B, wavelength: 254nm/220 nm;) to give the product (9.5 mg, 5.2%).

LC-MS(ESI):[M+H]⁺=555.2

¹H NMR(400MHz,DMSO-d6)9.51(t,J＝8.0Hz,1H),9.13-9.12(m,1H),8.92(d,J＝4.0Hz,1H),8.78-8.77(m,1H),7.98(s,1H),7.60(s,1H),7.10-7.08(m,1H),7.05(s,1H),7.02(s,1H),4.74–4.70(m,1H),4.08(s,3H),4.07-4.05(m,2H),3.98-3.95(m,2H),3.89(s,3H),3.05-3.00(m,2H),2.31(s,3H),2.19(s,3H),2.03(s,3H),0.83(d,J＝8.0Hz,3H).

Example 37, compound 37

To a 25mL single vial was added 2- ((1-aminoprop-2-yl) (methyltrimethyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (100 mg,0.22 mmol), 6-methylpyrazine-2-carboxylic acid (45.6 mg,0.33 mmol), EDCI (63.3 mg,0.33 mmol), HOBT (44.6 mg,0.33 mmol) and DMF (2 mL) at room temperature, and stirred for 24H at room temperature. After the reaction, the reaction system was filtered and purified by reverse phase Flash (mobile phase A:0.5% aqueous FA solution, mobile phase B: acetonitrile; flow rate: 40mL/min; gradient: 20 minutes from 0% B to 55% B;50% B, wavelength: 254nm/220 nm;) to give the product (29.3 mg, 15.6%).

LC-MS(ESI):[M+H]⁺=569.2

¹H NMR(400MHz,CDCl₃)9.17(s,1H),8.97(s,1H),8.59(s,1H),7.00(d,J＝8.0Hz,2H),6.70(s,1H),6.60(s,1H),5.31(s,1H),5.08-5.07(m,1H),4.23–4.17(m,2H),3.92(s,3H),3.92-3.85(m,2H),3.72(s,3H),2.94-2.91(m,2H),2.68(s,3H),2.34(s,3H),2.25(s,3H),2.12(s,3H),1.11(d,J＝8.0Hz,3H).

Example 38, compound 38

To a 25mL single vial was added 2- ((1-aminoprop-2-yl) (methyltrimethyl) amino) -9, 10-dimethoxy-6, 7-dihydro-4H-pyrimido [6,1-a ] isoquinolin-4-one (150 mg,0.33 mmol), 5-methylpyrazine-2-carboxylic acid (69.1 mg,0.50 mmol), EDCI (95.8 mg,0.50 mmol), HOBT (67.6 mg,0.50 mmol) and DMF (3 mL) at room temperature, and stirred for 24H at room temperature. After the reaction, the reaction system was filtered and purified by reverse phase Flash (mobile phase A:0.5% aqueous FA solution, mobile phase B: acetonitrile; flow rate: 40mL/min; gradient: 20 minutes from 0% B to 55% B;50% B, wavelength: 254nm/220 nm;) to give the product (50.2 mg, 26.8%).

LC-MS(ESI):[M+H]⁺=569.2

¹H NMR(400MHz,CDCl₃)9.23(s,1H),8.48(s,1H),8.59(s,1H),7.00(d,J＝12.0Hz,2H),6.70(s,1H),6.60(s,1H),5.31(s,1H),4.98-4.93(m,1H),4.23–4.17(m,2H),3.92(s,3H),3.92-3.85(m,2H),3.72(s,3H),2.94-2.91(m,2H),2.65(s,3H),2.35(s,3H),2.25(s,3H),2.12(s,3H),1.14(d,J＝8.0Hz,3H).

PDE enzyme Activity inhibition assay

The compounds of each example were tested for PDE enzyme activity inhibition assay by the FP method and RPL-554 of example 1 of CN100415743C was used as a positive control.

PDE enzyme and Substrate (FAM-cyclic adenosine monophosphate/cyclic adenosine monophosphate) solutions were prepared in reaction buffer (1X IMAP Reaction Buffer containing, 0.1% BSA with 1mM DTT added). The positive control was diluted 3-fold at an initial PDE concentration of 1/10. Mu.M, 10+0dose. 0.05. Mu.L of compound in 100% DMSO was delivered to 384 well plates (Corning 4514) by acoustic liquid delivery technique (Echo 655), centrifuged at 1000rpm for 1 min, 2.5. Mu.L of PDE enzyme solution was transferred to 384 well plates and centrifuged at 1000rpm for 1 min, incubated at 25℃for 10min, 2.5. Mu.L of Sub solution was transferred to 384 well plates, centrifuged at 1000rpm for 1 min, incubated at 25℃for 60min, 15. Mu.L of binding agent mixture was transferred to 384 well plates and centrifuged at 1000rpm for 1 min, incubated at 25℃for 60min, and finally FP signal was read with BMG (PHERAstar FSX). IC50 values and nonlinear regression curve fits were obtained using GRAPHPAD PRISM software.

Table 3 shows the values of the IC ₅₀ for the enzyme activity experiments for the compounds of the comparative examples and examples. A smaller value for IC ₅₀ indicates a smaller amount of compound required for half inhibition of the different PDE enzymes, indicating a stronger inhibitory activity.

TABLE 3 Table 3

The conclusion is that the compound has high-efficiency inhibition activity on PDE3/4, is superior to the prior art, and has good development prospect.

The foregoing is merely illustrative of the embodiments of this invention and it will be appreciated by those skilled in the art that variations and modifications may be made without departing from the principles of the invention, and it is intended to cover all modifications and variations as fall within the scope of the invention.

Claims

1. A compound having structural formula I or a pharmaceutically acceptable form thereof selected from a pharmaceutically acceptable salt or co-crystal, stereoisomer, tautomer, deuterate, solvate, chelate, non-covalent complex, or prodrug thereof:

Wherein:

Optionally, R ₆ forms with L

2. The compound of claim 1, or a pharmaceutically acceptable form thereof, wherein:

Each R ₁、R₂ is independently selected from CH ₃、CHF₂、CD₃ or C _3-6 cycloalkyl;

Preferably, R ₁、R₂ is CD ₃, respectively;

Preferably, one of R ₁、R₂ is CH ₃ and the other is CHF ₂.

3. The compound or pharmaceutically acceptable form thereof according to claim 1 or 2, wherein:

Each R ₃、R₄、R₅ is independently selected from CH ₃、i-Pr、OMe、CD₃ or halogen;

Preferably, R ₅ is CH ₃, said CH ₃ is optionally further substituted with 0 to 3D, R ₃、R₄ is the same and is selected from CH ₃, i-Pr, OMe or halogen.

4. A compound according to any one of claims 1-3, or a pharmaceutically acceptable form thereof, wherein L is selected from one of the following schemes:

L is N is 0 or 1, preferably 0;

Preferably, k is 0, 1 or 2;

Preferably, one of R ₉、R₁₀ is H and the other is selected from H, C _1-6 straight chain alkyl, C _3-6 branched chain alkyl, C _3-6 cycloalkyl, C _3-6 heterocycloalkyl, C _6-10 aryl, C _5-10 heteroaryl and COOCH ₃, said heteroaryl containing 1 or 2 heteroatoms N, H in said L being optionally further substituted with 0 to 4 substituents selected from F, cl, br, I, OH, = O, NH ₂、CN、COOH、C_1-4 alkyl, C _1-4 alkoxy;

L is

L isN is 0 or 1, k is 1;

L is N is 0 or 1, k is 1;

L is N is 2, k is 2;

L is N is 1, k is 1 or 2, or

L isN is 2 and k is 2.

5. The compound of any one of claims 1-4, or a pharmaceutically acceptable form thereof, wherein R ₆ is selected from one of the following schemes:

r ₆ is Wherein R ₁₁ is amino optionally further substituted with 0,1 or 2 substituents selected from F, cl, br, I, OH, = O, NH ₂、CN、COOH、CF₃、C_1-4 alkyl, C _1-4 alkoxy, C _3-6 cycloalkyl;

r ₆ is Wherein R ₁₁ is C _6-10 aryl or C _5-10 heterocyclyl, said heterocyclyl containing 1,2 or 3 heteroatoms N, said aryl, heteroaryl optionally being further substituted with 0, 1,2 or 3 substituents selected from H, F, cl, br, I, OH, = O, NH ₂、CN、COOH、CF₃、C_1-4 alkyl, C _1-4 alkoxy, C _3-6 cycloalkyl.

6. The compound of any one of claims 1-5, or a pharmaceutically acceptable form thereof, wherein R ₇ is H or CH ₃,R₈ is H or F.

7. The compound or pharmaceutically acceptable form thereof according to claim 1, wherein the compound is selected from one or more of the compounds shown in table 1.

8. An intermediate compound having a structure represented by formula II:

wherein R ₁、R₂、R₃、R₄、R₅、R₇、R₈ is as defined in any one of claims 1 to 7;

L ₁ is selected from R ₉、R₁₀, n, k are as defined in any one of claims 1 to 7, R ₁₂、R₁₃ are each independently H, boc, cbz, SEM, fmoc, alloc, pht, OTs, PMB, bn, trt;

Preferably, the intermediate compound is

9. A process for preparing the compound of any one of claims 1-7, or a pharmaceutically acceptable form thereof, comprising:

a modification of the R ₁₂、R₁₃ terminus of the intermediate compound of claim 8, shown in formula II, to produce a compound of formula I.

10. The process of claim 9, further comprising a process for preparing the intermediate compound of claim 8.

11. A pharmaceutical composition comprising a compound according to any one of claims 1 to 7 or a pharmaceutically acceptable form thereof, together with a pharmaceutically acceptable carrier, excipient and/or one or more other therapeutic agents.

12. Use of a compound according to any one of claims 1 to 7 or a pharmaceutically acceptable form thereof or a pharmaceutical composition according to claim 11 in the preparation of a formulation for inhibiting phosphodiesterase.

13. Use of a compound according to any one of claims 1 to 7 or a pharmaceutically acceptable form thereof or a pharmaceutical composition according to claim 11 in the manufacture of a medicament for the treatment of phosphodiesterase-related diseases;

preferably, wherein the phosphodiesterase-related disease comprises a respiratory disease such as asthma.